TRENDS IN BIOSCIENCES 6-1, 2013 EDITION

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Trends in Biosciences 

A Quaterly International Scientific Journal 


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Dr. Randy Gaugler, Director, Centre for Vector Biology, Rutgers University, USA 

Dr. S.B. Sharma, Director, Plant Security, South Perth, Australia 

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Advisory Board 

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Dr. Absar Ahmad, Senior Scientist, National Chemical Laboratory, Pune 

Dr. N.P. Singh, Coordinator, AICRP Chickpea, IIPR, Kanpur 

Dr. Raman Kapoor, Head, Dept. of Biotechnology, Indian Sugarcane Research Institute, Lucknow 

Dr. S.K. Jain, Coordinator, AICRP Nematode and IARI, New Delhi 

Dr. Sanjeev Gupta, Coordinator, MULLaRP, IIPR, Kanpur 

Dr. Naimuddin, Sr. Scientist (Plant Pathology), IIPR, Kanpur 

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Trends in Biosciences 


CONTENTS 

MINI REVIEW 

1. Biodiversity and Its Responsibility in Biotechnology : A Review 1 

V.K. Mishra and D.K. Dwivedi 

2. Physiological, Biochemical and Molecular Changes during High Temperature in Plants – A Review 5 

Kshitij Kumar and I.U. Rao 

3. Knowledge and Gaps for Herbal Zinc in Relation to Their Role in Regulation of Male Fertility : A Review 14 

Manish Mathur 

4. Insectivorous Plants and Their Trapping Mechanism: A Review 19 

Murali, S. and Narasa Reddy, G. 

RESEARCH PAPERS 

5. Expression and Immunogenicity of E2 Glycoprotein Gene of Classical Swine Fever Virus Cloned in 23 

Eukaryotic Expression Vector 

Nitin Sharma, Purshotam Kaushik and Anant Rai 

6. Morphological and Anatomical Aspects of Callogenesis and Somatic Embryogenesis of Date Palm 28 

(Phoenix dactylifera L.) 

Akshaya Bhati and Atul Chandra 

7. Successful Therapeutic Management of Snake Bite in A Dog 31 

Sonal Shrivastava, Debosri Bhowmick and P.C. Shukla 

8. Biology and Feeding Potential of Cheilomenus sexmaculata (Fab.) on Bean Aphid (Aphis craccivora Koch) 33 

and Mustard Aphid (Liphaphis erysimi L.) 

Akshay Kumar, C.S. Prasad and G.N. Tiwari 

9. Survey, Vector Relationships and Host Range Studies of Tomato Leaf Curl Karnataka Virus Causing 36 

Sunflower Leaf Curl Disease 

Vanitha, L.S., Rangaswamy, K.T., Govindappa, M.R., Manjunatha, L., Shivakumar S. Chinchure 

and Govardhana, M. 

10. Losses Caused Due to Alternaria Blight Disease in the Grain Yield of Pigeonpea 40 

Laxman Prasad Balai and R.B. Singh 

11. Relative Performance of Resource Conservation Technologies in Maize Based Cropping System Under 43 

Temperate Kashmir 

Fayaz Ahmed Bahar 

12. Influence of Deltamethrin and Achook ® on Activities of Phosphatases in the Nervous Tissue of Zebrafish, 46 

Danio rerio 

Dilip Kumar Sharma and Badre Alam Ansari 

13. Herbal Rumenotoric Drugs and Their Effect on Digestion and Growth Performance of Crossbred Calves 50 

R.D. Gautam, D.P. Singh, Ram Niwas and Abed M. Albial 

14. Studies on Nutritional Quality of Idli Fortified with Whey Protein Concentrates 54 

Alka Yadav, Ranu Prasad and Ramesh Chandra 

15. Evaluation and Assessment of Ground Water of Shivpuri Town in Madhya Pradesh 56 

Chandana Jain and Ritu Mendiratta 

16. Spatial and Modular Variability in Phytosterol Composition in Tribulus terrestris 59 

Manish Mathur 

17. Performance Evaluation of Commercial Reapers for Finger Millet Harvesting 63 

H.G. Ashoka, G.M. Prashantha, E.G. Ashok and M.V. Channa Byregowda 

18. Induced Mutation Through Gamma Irradiation at Different Doses to Create Genetic Variability and Study 65 

the Improvement in Yield and Yield Attributes of Genotype HD 2867 

Shubhra Singh, Ram, M., S. Marker, Abrar Yasin, B. and Akhilesh Kumar

19. Assesment of Genetic Divergence in Chickpea Genotypes (Cicer arietinum L.) 68 

Sudhanshu Jain and Y.M. Indapurkar 

20. Phytotoxic Effect of Asclepias curasavica Linn on Metabolism of Parthenium hysterophorus L 70 

T.G. Nagaraja and A.H. Pudale 

21. Serological Detection and Host Range Studies of Peanut Bud Necrosis Virus Infecting Tomato 73 

Mallu Govardhana, Gopinath, K., Vanitha, L.S., Eswar Reddy Maddi and Manjunatha, L. 

22. Influence of Genotypes on Consumability of the Leaves, Growth and Development of Spodoptera litura (Fab.) 76 

Pradyumn Singh, N.S. Bhadauria and Pooja Chauhan 

23. Diversity Pattern of Beetles (Insecta: Coleoptera) In and Around the Historic Joysagar Tank of Assam, India 78 

I. Rahman, C. Sonowal and M. Nath 

24. Effect of Ginger and Mint Extract on Microflora of Yoghurt Culture 80 

Dinker Singh and Swashankh Kumar 

25. Anti-Fungal Evaluation of Calotropis Leaf Extract against Some Plant Pathogenic Fungi 82 

Rajni Singh Sasode and Pradyumn Singh 

26. Studies on Status of Leaf Spot of Groundnut in Chunar and Nearby Areas of Mirzapur District of Uttar Pradesh 86 

Bharat Chandra Nath, R.B. Singh and Laxman Prasad Balai 

27. Studies on the Effect of Scent Components on the Somatic Cells of Allium sativum L 88 

Ch. Srinivasulu 

28. A Study on the Quality of Milk in Gwalior 93 

Chandana Jain and Ritu Mediratta 

29. Prediction of Lifetime Milk Production from Early Lactation Traits in Crossbred Cattle 95 

Hemant Kumar and B.K. Hooda 

30. Fungal Growth on C1 Compounds: A Study of the Amino Acid Composition of a Methanol-utilizing Strain of 97 

Trichoderma viride 01 PP 

Mohd. Shahid, Mukesh Srivastva, Anuradha Singh and Neelam Pathak 

31. Varietal Evaluation of Cauliflower (Brassica oleracea var. botrytis L.) In Allahabad Agro – climatic Condition 99 

Mukesh Yadav, V.M. Prasad and Chandan Singh Ahirwar 

32. Screening of Chickpea Genotypes for Resistant against Pod Borer, Helicoverpa armigera Hubn. 101 

Jeewesh Kumar, D.C. Singh, A.P. Singh and S.K. Verma 

33. Knowledge Level of Farmers About Recommended Cultivation Practices of Groundnut in Panchayat Samiti 104 

Govindgarh of District Jaipur (Rajasthan) 

H.N. Verma and J.P. Yadav 

34. Stability Studies on Sewage Irrigation Effect on Grain Yield and Protein-content in Mungbean 106 

[Vigna radiata (L.) Wilczek 

Dyuti Pandey, P.S. Shukla and D.N. Shukla 

35. In Vitro Screening and Identification of Putative Sunflower (Helianthus annus L.) Transformants 108 

Expressing ECNAC1 Gene by Salt Stress Method 

K.C. Manjunath, A. Mahadeva, Rohini Sreevathsa, N. Ramachandra Swamy and T.G. Prasad 

36. Efficacy of Different Herbicides for Weed Control in Transplanted Rice (Oryza sativa. L) Under Vertisols 112 

of Rajasthan 

Chandra Prakash, R.K. Shivran, N.R. Koli and J.C. Sharma 

37. Bio-efficacy of Herbicide Combination on Weed Control and Yield Performance of Transplanted 115 

Rice (Oryza sativa. L) 

Chandra Prakash, R.K. Shivran, N.R. Koli and J.C. Sharma 

SHORT COMMUNICATIONS 

38. Genetic Variability, Heritability and Correlation Studies in Wheat Genotypes (Triticum aestivum L.) 118 

Ekta Singh, Ramteke P.W., M. Ram, B.A. Wani and Sadhna Singh 

39. Screening of Horsegram (Macrotyloma uniflorum) Genotypes against Horsegram Yellow Mosaic 120 

Virus (HgYMV) Disease in Karnataka 

Prema, G.U., Rudraswamy, P., Nagaraju and Rangaswamy, K.T. 

40. Evaluation of Garlic Genotypes Under Temperate Conditions of Kashmir Valley 122 

Mushtaq, A. Chatoo, Parvaze, A. Sofi, Khalid, R. Dar, Angrez Ali and Tasaduk Shafi 

Author Index Vol. 5 (1-4) 2012 124 

Subject Index Vol. 5 (1-4) 2012 128 

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Trends in Biosciences 6 (1): 1-4, 2013 


Biodiversity and Its Responsibility in Biotechnology : A Review 

V.K. MISHRA 1 AND D.K. DWIVEDI 

Department of Plant Molecular Biology and Genetic Engineering, N.D. University of Agriculture and 

Technology, Kumarganj, Faizabad 224 229 (U.P.) 

e-mail: vinay.mishra111@gmail.com 1 

ABSTRACT 

Biodiversity is threatened by agriculture as a whole, and 

particularly also by traditional methods of agriculture. 

Knowledge-based agriculture, including GM crops, can reduce 

this threat in the future. The introduction of no-tillage practices, 

which are beneficial for soil fertility, has been encouraged by 

the rapid spread of herbicide tolerant soybeans in the USA. 

The replacement of pesticides through Bt crops is advantageous 

for the nontarget insect fauna in test-fields. Biodiversity 

differences can mainly be referred to as differences in herbicide 

application management. Biotechnology, while controversial 

particularly in agricultural applications, has the potential to 

improve sustainability in several ways and is expected, thereby, 

to help maintain natural as well as agricultural biodiversity. 

This paper results from the combined contributions of scientists, 

industrialists , and go vernmental a nd public intere st 

organisations across Asia. 

Key words 

Biodiversity, GM crops, Ecosystem diversity, in-situ, 

Agro diversity 

Biodiversity is the total richness of biological variation 

in the all the species of plants, animals and microorganism 

and the variation between them. Biodiversity is important for 

several very different reasons: the intrinsic value of species 

in the wild, the many varieties of plants, animals and microorganisms 

used worldwide for farming and other human 

activities, as a genetic resource in healthcare, agriculture and 

food production, as well as for aesthetic and recreational 

purposes. Biotechnology involves the use of all life forms for 

human welfare. Therefore, extinction of wild species and 

destruction of ecosystems has been a major concern of policy 

makers and biotechnologists alike. 

It is estimated that approximately 5-30 million species 

existing in the living forms. About 6% of the identified species 

live in boreal or polar latitudes, 59% in the temperate zones 

and the remaining 35% in tropics. It include 300000 species of 

green plants and fungi, 360000 species of micro-organism, 

800000 species of insect and 40000 species of vertebrates. 

Biotechnology has the clear potential to save native 

untouched land, particularly in tropical rain forests, by 

reducing or eliminating the pressure to bring more land into 

cultivation. Tropical insect species are difficult to identify 

because most are confined to a single type of tropical forest 

or even to a particular tree species, which may itself have only 

a local distribution. Additionally, the species may have limited 

Dr. Vinay Kumar Mishra has done Ph.D. 

in Agriculture Biotechnology from Narendra 

Deva University of Agriculture & 

Technology, Kumarganj, Faizabad (U.P.). 

Presently he is working as lecturer in (Agri. 

Biotechnology), Division of Crop Sciences, 

Mahatma Gandhi Chitrakoot Gramodaya 

Vis hwavidyalaya, Chitrakoot, Satna, 

(M.P.). He attaineded six International and 

National Seminars and pub lished 11 

research papers, 2 review articles, 16 abstract papers and 16 

Hindi articles. 

Dr. D.K. Dwivedi has done Ph.D. and Post 

Doctoral Fellow from International Rice 

Research Institute, Philippines. Presently 

he is working as Associate Professor (Plant 

Biotechnology) Department of Plant 

Molecular Biolo gy and Genetic 

Engineering, N.D. University of Agriculture 

and Tec hnology, Narendra Nagar 

(Kumarganj), Faizabad, U.P., India. His 

professional experience is about more than 

24 years and to his credit 111 research 

papers includeing book chapters, bulletins and manuals. Dr. 

Dwivedi released 4 varieties namely Narendra Salinity Hybrid 

Rice 3, Narendra Hybrid Rice 2, UPRI 95-167 TGMS line and 

UPRI 95-140 TGMS line. He is associated in 4 projects as PI/ 

Co-PI and awarded Best Research Paper at Symposium on 

Molecular Approaches for Crop Improvement (7-8 Feb., 2007), 

NDUAT, Faizabad, India and also, Rockefeller Foundation Postdoctoral 

Fellowship. 

dispersal ability. Some 470-495 million species are now 

extinction is a fact of life. 

Types of biodiversity 

Biodiversity includes (1) genetic diversity, (2) species 

diversity and (3) ecosystem diversity. 

(A) Genetic Diversity 

Genetic diversity is usually important not only for the 

survival and evolution of species, it is also important to people 

for breeding improved crop plants with higher yields. This is 

because genetic erosion of several crops has already occurred, 

leading to the world’s dependence for food on just a few 

species. 

Economic Potential and Conservation of Medicinal and

2 Trends in Biosciences 6 (1), 2013 

Aromatic Plants from Brazil Approximately 2/3 of the biological 

diversity of the world is estimated to be in tropical zones, 

mainly in developing countries. With nearly 55,000 native 

species, Brazil is considered the country with the greatest 

biodiversity on the planet. These species are distributed over 

six major distinct biomes: Amazon (30,000); Cerrado (7,000); 

Caatinga (4,000); Atlantic rainforest (16,000), Pantanal (10,000) 

and the subtropical forest (3,000) India is a major center of 

origin and diversity of crop and medicinal plants. India poses 

out 20,000 species of higher plants, one third of it being 

endemic and 500 species are categorized to have medicinal 

value (Krishnan, et al., 2011). 

(B) Species Diversity 

This diversity includes the variation of species within 

the particular region. Each of these biological communities’ 

represents and adaptation of plants to particular regimes of 

climate, soil and other aspects of the environment. Global 

biodiversity is frequently expressed as the total number of 

species currently living on Earth, i.e., its species richness. 

Between about 1.5 to 1.75 million species have been discovered 

and scientifically described (LeCointre and Guyader, 

2001; Cracraft, 2002). Species richness and species evenness 

are probably the most frequently used measures of the total 

biodiversity of a region. Species diversity is also described in 

terms of the phylogenetic diversity, or evolutionary 

relatedness, of the species present in an area. 

(C) Ecosystem Diversity 

An ecosystem is a community plus the physical 

environment that it occupies at a given time. An ecosystem 

can exist at any scale, for example, from the size of a small tide 

pool up to the size of the entire biosphere. However, lakes, 

marshes, and forest stands represent more typical examples 

of the areas that are compared in discussions of ecosystem 

diversity. Ecosystem diversity refers to the diversity of a place 

at the level of ecosystems. The term differs from biodiversity, 

which refers to variation in species rather than ecosystems. 

Table 1. Gene banks of various crops in India 

Crop species 

Location of gene bank centre 

Wheat 

DWR, Karnal (Haryana) 

Rice 

CRRI, Cuttack (Orissa) 

Potato 

CPRI, Shimla (Himachal Pradesh) 

Cotton 

CICR, Nagpur (Maharashtra) 

Sugarcane 

SBI, Coimbatore (Tamil Nadu) 

Tobacco 

CTRI, Rajahmundry (Andhra Pradesh) 

Pulses 

IIPR, Kanpur, (Uttar Pradesh) 

Forage Crops 

IGFRI, Jhansi 

Plantation Crops 

CPCRI,Kasargod, (Kerala) 

Tuber Crops (except Potato) CTCRI, Trivandrum (Kerala) 

Oilseed Crops 

DOR, Hyderabad (Andhra Pradesh) 

Horticultural Crops IIHR, Bangalore (Karnataka) 

Sorghum 

NRC Sorghum, Hyderabad (Andhra 

Pradesh) 

Groundnut 

NRC Groundnut, Junagadh (Gujarat) 

Soybean 

NRC Soybean, Indore (Madhya Pradesh) 

Maize 

IARI, New Delhi 

Ecosystem diversity can also refer to the variety of 

ecosystems present in a biosphere, the variety of species and 

ecological processes that occur in different physical settings. 

Environmental disturbance on a variety of temporal and 

spatial scales can affect the species richness and, 

consequently, the diversity of an ecosystem. Ecosystems may 

be classified according to the dominant type of environment, 

or dominant type of species present; for example, a salt marsh 

ecosystem, a rocky shore intertidal ecosystem, a mangrove 

swamp ecosystem. Because temperature is an important aspect 

in shaping ecosystem diversity, it is also used in ecosystem 

classification (e.g., cold winter deserts, versus warm deserts) 

(Udvardy, 1975). 

Genetic modification and the sourcing of genes 

In vitro technology formed a prerequisite for the 

development of genetic modification of plants. Genetic 

modification of plants combines techniques for plant tissue 

culture, techniques for cloning, in vitro amplification (PCR), 

and transfer of DNA, either by the use of the soil bacterium 

Agrobacterium tumefaciens as a vector, through 

electroporation or by the use of a particle gun. It is based on 

the ability to change the genetic constitution of a single cell 

and regenerate a new plant from that single cell. The 

technology can be used to transfer a gene from a wild relative 

simply to speed up breeding, but it can also be used for the 

transfer of genes into the plant gene pool that could not be 

introduced into plant genomes by other means. 

Molecular marker technology has been used to identify 

genotypes and to confirm intergeneric hybrids between 

papaya and related Vasconcellea species. Molecular maps have 

been developed for some species. Genetic diversity within 

populations and between related wild species has been 

determined for some fruits. Specific DAF markers have been 

identified for sex determination in papaya; a SCAR marker has 

been developed to identify dwarfism in bananas; and, a CAPS 

marker to identify a PRSV-P resistant gene in highland papaya. 

The major application in genomics has been the rapid progress 

in sequencing the papaya genome in Hawaii (Drew, 2008). 

In 1999 more than 70 transgenic crop varieties were 

registered for commercial cultivation. The crops involved 

include corn, soybean, rapeseed, tomato, tobacco, potato, 

chicory, papaya, pumpkin and clover. In 2005, biotech soybean 

continued to be the principal biotech crop, occupying 54.4 

million hectares (60% of global biotech area), followed by 

maize (21.2 million hectares at 24%), cotton (9.8 million hectares 

at 11%) and canola (4.6 million hectares at 5% of global biotech 

crop area). During the first decade, 1996 to 2005, herbicide 

tolerance has consistently been the dominant trait followed 

by insect resistance and stacked genes for the two traits (James, 

2005). In the future diversification may be expected not only 

from the private sector, but also from the (international) public 

sector. The CGIAR Challenge programme Generation focuses 

on drought tolerance in cereals, legumes and clonal crops.

MISHRA & DWIVEDI, Biodiversity and its responsibility in biotechnology: A review 3 

Genetic modification efforts of the M.S. Swaminathan 

Foundation regard combating drought and salinity in locally 

adapted rice and wheat varieties, whereas the Indian Initiative 

on Crop Biofortification focuses on bio-availability and bioefficacy 

with respect to iron, zinc and phosphor in rice, wheat 

and maize. 

Native biodiversity and biotechnology 

Biodiversity in the wild has been massively reduced in 

the industrialised countries over many centuries and about 

half the tropical rain forests have already been destroyed. 

Yields of cereals in LDCs have gone up very considerably in 

the last forty years primarily as the result of the Green 

Revolution. However, the annual growth increases in cereal 

yields in LDCs have slowed down from about 3% during 1967- 

82 to about 1% per year from 1993 onwards. This lowering of 

the rate at which yields have increased means that productivity 

will probably not keep up with the demands of increased 

populations. The consequences for biodiversity are 

devastating as more land will be required for farming which 

will primarily come from areas with high native biodiversity, 

and in particular tropical rain forests. 

The single most promising way to avoid habitat 

destruction is to increase farm yields in a process that has 

been called the “Second Green Revolution”. Several 

components will be required including training and education 

of farmers (in particular of women who do most of the farm 

work in LDCs), more favourable economic and political 

climates, availability of farm credit schemes, etc. In addition, 

technical contributions will be necessary and, in particular, 

improved seed produced either by traditional crop breeding 

or by modern biotechnology. Reliance will have to be more on 

the latter since traditional breeding appears to have reached a 

plateau in yield and is slower, less precise and only feasible 

when interbreeding is possible. So agricultural biotechnology, 

which is viewed frequently in the public debate as harmful to 

biodiversity, is, paradoxically, likely to contribute to conserving 

it. A more limited concern that largely affects Northern Europe 

is the conservation of native plants and animals, in particular 

birds, in farmed areas. Their habitats are fields, hedges, 

roadsides and fallow land where they depend for food on 

insects and seeds produced by weeds in or near crops. 

Computer models suggest that more intense weed control 

measures may lead to smaller amounts of seeds being available 

to birds. This depends far more on weed management regimes 

rather than on transgenic plants. Herbicide tolerant beets allow 

farmers to control weeds later by treating after the seedlings 

have emerged. The more efficient methods may allow setting 

aside of more land. Setting aside more farmland requires 

financial incentives. 

Impact of agricultural biotechnology on biodiversity 

With the introduction of GM crops, concern has been 

raised that overall genetic diversity within crop species will 

decrease because breeding programs will concentrate on a 

smaller number of high value cultivars. However, several 

studies have specifically focused on this subject and they 

have concluded that the introduction of transgenic cultivars 

in agriculture has not significantly affected levels of genetic 

diversity within crop species. For example Sneller, 2003 looked 

at the genetic structure of the elite soybean population in 

North America, using coefficient of parentage (CP) analysis. 

The introduction of herbicide-tolerant cultivars with the 

Roundup Readyw [Monsanto http://www.monsanto.com/ 

monsanto/layout/ products/productivity/ roundup/ 

default.asp) trait was shown to have had little effect on 

soybean genetic diversity because of the widespread use of 

the trait in many breeding programs. Only 1% of the variation 

in CP among lines was related to differences between 

conventional and herbicide-tolerant lines, whereas 19% of 

the variation among northern lines and 14% of the variation 

among southern lines was related to differences among the 

lines from different companies and breeding programs. 

Similarly, when Bowman, et al., 2003 examined genetic 

uniformity among cotton varieties in the USA, they found 

that genetic uniformity had not changed significantly with 

the introduction of transgenic cotton cultivars. Genetic 

uniformity actually decreased by 28% over the period of 

introduction of transgenic cultivars. In conclusion, 

biotechnology represents a tool for enhancing genetic 

diversity in crop species through the introduction of novel 

genes. This does not aim at the single transgene inserted, but 

is based on the fact that beneficial characters can now be 

inserted in a variety of crops that have been neglected because 

of the limitations of traditional breeding methods, which failed 

to enhance the traits. There is great potential to achieve drought 

tolerance in vegetables (Slabbert, 2004) and to avoid postharvest 

losses in African grain staple crops. 

Impact of biotechnology on agro biodiversity 

Biotechnology has provided tools to more effectively 

maintain and utilize genetic resources ex situ as well as onfarm, 

and has thus contributed positively to the conservation 

of genetic resources. A limited contribution of biotechnology 

to the development of novel genetic resources in agroecosystems 

may also emerge in the near future, as a result of 

introgression of novel traits by means of genetic modification. 

However, the increasing role of biotechnology also forms a 

potential threat to the survival of agrobiodiversity in the 

environment, whether this is in the farmer’s field (on-farm), or 

in natural ecosystems (in situ). The impact of biotechnology 

on agrobiodiversity retains several components, which are 

very different in nature but do closely interrelate. While new 

technological developments take place at a rapid pace, their 

environmental and social implications are not well understood, 

let alone under control. Biological, socio-economic and 

cultural effects can be distinguished, and the various 

biotechnological applications differ strongly in the degree


and direction of their effect. Similarly, technological risks can 

be distinguished into two types: technology inherent risks 

concern the risks to human health, ecology, and the 

environment, and technology derived risks which are the result 

of the specific use of the technology, either reducing or 

enhancing the poverty gap, reducing or protecting 

biodiversity, and centralising or decentralising the control over 

and access to technology and its products. 

The risk of biodiversity loss is evident if we continue to 

experience the current trend for predatory and unregulated 

exploitation of living natural resources. Biodiversity has value 

for Science, and this value transcends merely subjective 

arguments. Brazil has a significant importance in biodiversity 

conservation due to the magnitude of its biological diversity. 

The Pantanal is a wetland system with a variety of ecosystems 

due to the seasonal flooding, and is recognized as one of the 

most important biodiversity biomes (Alho and Gonçalves, 

2005). It is well recognized that Brazil is important in the 

scientific value of biodiversity because of the size of its 

territory, the diversity of its biomes, the size of its river system, 

the concept of a mega diverse country, and the biodiversity 

hotspots identified, with a large and continuous continental 

biota. The estimated size of known or recorded Brazilian biota 

is around 200,000 species, representing about 10% of the 

world’s known number of species. These numbers may 

increase dramatically considering that the Neotropics 

constitutes, so far, one of the least studied regions. There are 

over 55,000 species of flora (without fungi species), 

representing nearly 20% of the world’s flora. Brazilian biomes 

hold an immense number of terrestrial invertebrates, including 

26,000 Lepidoptera, 12,000 Hymenoptera, 30,000 Coleoptera 

and so on. The number of amphibians (765 species) makes the 

country a leader in this taxon’s diversity. Reptiles comprise 

600 species, including 350 species of snakes (Lewinshon and 

Prado, 2006). 

The issues involved in the interaction between 

biodiversity and biotechnology have far-reaching 

consequences and need to be subject to an open and 

knowledge-based dialogue in society. The dialogue needs to 

include many different stakeholders, including farmers of 

developing countries, diverse scientists, policy makers and 

communicators. Cultural values involved in farming and food 

production needs to be taken into consideration, just as much 

as the emotional side of eating and drinking. 

A significant aspect of ethical behavior is openness. 

Transparent information is required both from scientists in 

non-commercial settings as well as from industry. Several large 

projects in biotechnology like HUGO, the Human Genome 

Organization, have an “ELSI” component, dealing with ethical, 

legal and societal implications of biotechnology. This shows 

that some scientists realize that science cannot be seen as a 

human activity taking place in a void, without any connection 

to social and political realities. Ethicists stress that it is the 

responsibility of scientists to be actively concerned with these 

issues and that they should take special responsibility in 

communicating with non-specialists to explain what they know 

as well as what they don’t know. 

The analysis focuses on (a) the political climate around 

GEs having been spread from Europe around the world; (b) 

the legal and trade consequences connected to regulation 

and political climate; (c) GMO over-regulation making use of 

GEs for the public sector inaccessible for cost and time 

reasons; (d) the financial support to professional anti-GElobby 

groups and (e) poor support for agricultural research in 

general (Potrykus Ingo, 2010). 

Preservation of the genetic diversity present in crop 

species is an important need being addressed by various public 

and private programs. In this respect, biotechnology can be a 

valuable tool for introducing novel (alien or non-alien) genes 

into underused crop traits and crop species. Furthermore, the 

development and introduction of GM crop varieties does not 

represent any greater risk to crop genetic diversity than the 

breeding programs associated with conventional agriculture. 

After all, the overall performance of a plant and the quality 

and quantity of its product is the result of thousands of genes 

and the genetic background is almost always more important 

for the questions dealt with in this review than a single 

transgene. 

LITERATURE CITED 

Alho, C.J.R. and Gonçalves, H.C. 2005. Biodiversidade do Pantanal. 

Ecologia e Conservacaoo. Campo Grande: Editora UNIDERP. ISBN: 

858739294-8. pp.142. 

Bowman, D.T., May, O.L. and Creech, J.B. 2003. Genetic uniformity 

of the US upland cotton crop since the introduction of transgenic 

cottons. Crop Sci., 43:515–518. 

Brown, T. and Johnes, G. 2003. New ways with old wheat – Part I, 

Archaeology University of Sheffield. 

Cracraft, C. 2002. Species concepts and speciation analysis. Ornithology 

1:159-187. 

Drew, R.A. 2008. Applications of Biotechnology to Tropical Fruit Crops 

in Australia and Worldwide. Acta Horticulturae, No. 787. 

James, C. 2005. Global Status of Commercialized Biotech/GM Crops: 

2004”, ISAAA (www.isaaa.org). 

Krishnan, P.N., Decruse, S.W. and Radha, R.K. 2011. Conservation of 

Medicinal Plants of Western Ghats, India and its Sustainable 

Utilization through in vitro Technology. In Vitro Cellular and 

Developmental Biology: Plant., 47(1):110-122. 

Lecointre, G. and Guyader, H.L. 2001. Classification phylogenetique 

du vivant. Paris, France: Belin. 

Lewinshon, T.M. and Prado, P.I. 2006. How many species are there in 

Brazil? Conservation Biology, 19(3): 619-624. 

Potrykus, I. 2010. Constraints to Biotechnology Introduction for 

Poverty Alleviation. New Biotechnology, 27(5):447-448. 

Slabbert, R. 2004. Drought tolerance, traditional crops and 

biotechnology: breeding towards sustainable development. South 

African J. Botany, 70: 116-123. 

Sneller, C.H. 2003. Impact of transgenic genotypes and subdivision on 

diversity within elite North American soybean germplasm. Crop 

Sci., 43: 409-414. 

Udvardy, M.D.F. 1975. A classification of the biogeographical provinces 

of the world. Gland, Switzerland: International Union for the 

Conservation of Nature and Natural Resources. 

Recieved on 12-08-2012 Accepted on 02-12-2012



Physiological, Biochemical and Molecular Changes during High Temperature in 

Plants – A Review 

KSHITIJ KUMAR AND I.U. RAO 

Department of Botany University of Delhi, Delhi 110 007 

e-mail: patelbiotech@gmail.com 

ABSTRACT 

Heat stress is often defined as the rise in temperature beyond 

a threshold level for a period of time sufficient to cause 

irreversible damage to plant growth and development. In 

general, a transient elevation in temperature, usually 10-15 

o 

C above ambient, is considered heat shock or heat stress. 

However, heat stress is a complex function of intensity 

(temperature in degrees), duration, and rate of increase in 

temperature. The extent to which it occurs in specific climatic 

zo nes depe nds on the proba bility and period o f high 

temperatures occurring during the day and/or the night. Heat 

tolerance is generally defined as the ability of the plant to 

grow and produce economic yield under high temperatures. 

However, while some res earc hers believe tha t night 

temperatures are major limiting factors others have argued 

that day and night temperatures do not affect the plant 

independently, and that the diurnal mean’ temperature is a 

better predictor of plant response to high temperature with day 

temperature having a secondary role. 

Key word 

Heat stress, Temperature, Heat shock Proteins 

Heat stress due to high ambient temperatures is a serious 

threat to crop production worldwide. Gaseous emissions due 

to human activities are substantially adding to the existing 

concentrations of greenhouse gases, particularly CO2, 

methane, chlorofluorocarbons and nitrous oxides. Different 

global circulation models predict that greenhouse gases will 

gradually increase world’s average ambient temperature. 

According to a report of the Intergovernmental Panel on 

Climatic Change (IPCC), global mean temperature will rise 0.3 

?C per decade reaching to approximately 1 and3 ?C above the 

present value by years 2025 and 2100, respectively, and leading 

to global warming. Rising temperatures may lead to altered 

geographical distribution and growing season of agricultural 

crops by allowing the threshold temperature for the start of 

the season and crop maturity to reach earlier. At very high 

temperatures, severe cellular injury and even cell death may 

occur within minutes, which could be attributed to a 

catastrophic collapse of cellular organization (Sch¨offl, et al., 

1999). At moderately high temperatures, injuries or death may 

occur only after long-term exposure. Direct injuries due to 

high temperatures include protein denaturation and 

aggregation, and increased fluidity of membrane lipids. Indirect 

or slower heat injuries include inactivation of enzymes in 

chloroplast and mitochondria, inhibition of protein synthesis, 

Mr Kshitij Kumar is a research Scholar 

at Department of Plant Molecular 

Biology Narendra Deva University of 

Agriculture & Technology Narendra 

Nagar (Kumarganj) Faizabad Uttar 

Pradesh India. Mr Kshitij Kumar has 

completed M.Sc. (Agriculture) in 

Agricultural Biotechnology from 

Narendra Deva University of 

Agriculture & Technology Narendra Nagar (Kumarganj) 

Faizabad, Uttar Pradesh, India. He has completed his M.Phil. 

(Botany) from Department of Botany University of Delhi 

under supervision of Prof. I.U. Rao. 

protein degradation and loss of membrane integrity (Howarth, 

2005). 

Effect of temperature at cellular level 

Dr. I.U. Rao is a ex-Professor of Botany 

at University of Delhi, Delhi. She has 

three decades experience of teaching 

and research and guiding several 

M.Phil. and Ph.D. students. She was 

Head, Dean and Proctor of University 

of Delhi. She has published many 

national and International papers in 

reputed journals. She has been involved 

in several research projects. 

At very high temperatures, severe cellular injury and 

even cell death may occur within minutes, which could be 

attributed to a catastrophic collapse of cellular organization 

(Schoffl et. al., 1999). At moderately high temperatures, injuries 

or death may occur only after long-term exposure. Direct 

injuries due to high temperatures include protein denaturation 

and aggregation, and increased fluidity of membrane lipids. 

Indirect or slower heat injuries include inactivation of enzymes 

in chloroplast and mitochondria, inhibition of protein 

synthesis, protein degradation and loss of membrane integrity 

(Howarth, 2005). Heat stress also affects the organization of 

microtubules by splitting and/or elongation of spindles, 

formation of microtubule asters in mitotic cells, and elongation 

of phragmoplast microtubules (Smertenko, et. al., 1997). These 

injuries eventually lead to starvation, inhibition of growth,


Table 1. 

Genetically engineered alterations in the temperature stress tolerance of plants 

Functions of transformed genes Host plants Effects on the enhancement of temperature Remarks References 

Heat shock proteins 

Heat shock transcription factor (ATHSF1) Arabidopsis High temperatures (enhanced) Heat-inducible Lee, et al., 1995. 

Heat shock protein (Hsp70) Arabidopsis High temperatures(lessened) antisense expression Lee JH, Sch¨offl F. 1996. 

Small heat shock protein (Hsp17.7) Carrot High temperatures (enhanced/lessened) Constitutive Malik, et al., 1999 

reduced ion flux, production of toxic compounds and reactive 

oxygen species (ROS) (Howarth, 2005). 

Effect of temperature at molecular level 

Immediately after exposure to high temperatures and 

perception of signals, changes occur at the molecular level 

altering the expression of genes and accumulation of 

transcripts, thereby leading to the synthesis of stress-related 

proteins as a stress-tolerance strategy (Iba, 2002). Expression 

of heat shock proteins (HSPs) is known to be an important 

adaptive strategy in this regard (Wahid, et. al., 2007). The 

HSPs, ranging in molecular mass from about 10 to 200 kDa, 

have chaperone-like functions and are involved in signal 

transduction during heat stress (Schoffl et. al., 1999). The 

tolerance conferred by HSPs results in improved physiological 

phenomena such as photosynthesis, assimilate partitioning, 

water and nutrient use efficiency, and membrane stability 

(Momcilovic & Ristic, 2007). 

Heat-stress threshold 

A threshold temperature refers to a value of daily mean 

temperature at which a detectable reduction in growth begins. 

Upper and lower developmental threshold temperatures have 

been determined for many plant species through controlled 

laboratory and field experiments. A lower developmental 

threshold or a base temperature is one below which plant 

growth and development stops. Similarly, an upper 

developmental threshold is the temperature above which 

growth and development ceases. Knowledge of lower 

threshold temperatures is important in physiological research 

as well as for crop production. Base threshold temperatures 

vary with plant species, but for cool season crops, 0°C is 

often the best-predicted base temperature (Miller et. al., 2001). 

Cool season and temperate crops often have lower threshold 

temperature values compared to tropical crops. Upper 

threshold temperatures also differ for different plant species 

and genotypes within species. However, determining a 

consistent upper threshold temperature is difficult because 

the plant behavior may differ depending on other 

environmental conditions (Miller et. al., 2001). In tomato, for 

example, when the ambient temperature exceeds 35°C, its seed 

germination, seedling and vegetative growth, flowering and 

fruit set, and fruit ripening are adversely affected. 

High temperature induces modifications in plants which 

may be direct as on existing physiological processes or indirect 

in altering the pattern of development. These responses may 

differ from one phenological stage to another. For example, 

long-term effects of heat stress on developing seeds may 

include delayed germination or loss of vigor, ultimately leading 

to reduced emergence and seedling establishment. Under 

diurnally varying temperatures, coleoptile growth in maize 

reduced at 40 o C and ceased at 45 o C (Weaich et. al., 1996). 

High temperatures caused significant declines in shoot dry 

mass, relative growth rate and net assimilation rate in maize, 

pearl millet and sugarcane, though leaf expansions were 

minimally affected (Wahid, 2007). Major impact of high 

temperatures on shoot growth is a severe reduction in the 

first internode length resulting in premature death of plants 

(Wahid et. al., 2007). For example, sugarcane plants grown 

under high temperatures exhibited smaller internodes, 

increased tillering, early senescence, and reduced total 

biomass (Wahid et. al., 2007). 

Heat stress, singly or in combination with drought, is a 

common constraint during anthesis and grain filling stages in 

many cereal crops of temperate regions. For example, heat 

stress lengthened the duration of grain filling with reduction 

in kernel growth leading to losses in kernel density and weight 

by up to 7% in spring wheat (Guilioni et. al., 2003). Similar 

reductions occurred in starch, protein and oil contents of the 

maize kernel (Wilhelm et. al., 1999) and grain quality in other 

cereals under heat stress (Maestri et. al., 2002). In wheat, 

both grain weight and grain number appeared to be sensitive 

to heat stress, as the number of grains per year at maturity 

declined with increasing temperature (Ferris et. al., 1998). 

Growth chamber and greenhouse studies suggest that 

high temperature is most deleterious when flowers are first 

visible and sensitivity continues for 10-15 days. Reproductive 

phases most sensitive to high temperature are gametogenesis 

(8-9 days before anthesis) and fertilization (1-3 days after 

anthesis) in various plants (Foolad, 2005). 

Effect of temperature on anatomy 

Although limited details are available, anatomical 

changes under high ambient temperatures are generally similar 

to those under drought stress. At the whole plant level, there 

is a general tendency of reduced cell size, closure of stomata 

and curtailed water loss, increased stomatal and trichomatous 

densities, and greater xylem vessels of both root and shoot 

(Anon et. al., 2004). In grapes (Vitis vinifera), heat stress 

severely damaged the mesophyll cells and increased 

permeability of plasma membrane (Zhang et. al., 2005). With 

the onset of high temperature regime, Zygophyllum qatarense 

produced polymorphic leaves and tended to reduce 

transpirational water loss by showing bimodal stomatal

KUMAR AND RAO, Physiological, Biochemical and Molecular Changes during High Temperature in Plants–A Review 7 

behavior (Sayed, 1996). At the subcellular level, major 

modifications occur in chloroplasts, leading to significant 

changes in photosynthesis. For example, high temperatures 

reduced photosynthesis by changing the structural 

organization of thylakoids (Karim et. al., 1997). Studies have 

revealed that specific effects of high temperatures on 

photosynthetic membranes result in the loss of grana stacking 

or its swelling. In response to heat stress, chloroplasts in the 

mesophyll cells of grape plants became round, the stroma 

lamellae became swollen, and the contents of vacuoles formed 

clumps, whilst the cristae were disrupted and mitochondria 

became empty (Zhang et. al., 2005). 

Effect of temperature on phenology 

Observation of changes in plant phenology in response 

to heat stress can reveal a better understanding of interactions 

between stress atmosphere and the plant. Different 

phenological stages vary in their sensitivity to high 

temperature; however, this depends on species and genotype 

as there are great inter-and intra-specific variations (Howarth, 

2005). Heat stress is a major factor affecting the rate of plant 

development, which may be increasing to a certain limit and 

decrease afterwards (Howarth, 2005). 

Vulnerability of species and cultivars to high 

temperatures may vary with the stage of plant development, 

but all vegetative and reproductive stages are affected by 

heat stress to some extent. During vegetative stage, for 

example, high day temperature can damage leaf gas exchange 

properties. During reproduction, a short period of heat stress 

can cause significant increases in floral buds and crop 

production under high temperatures. 

Effect of temperature on physiology 

Plant water status is most important variable under 

changing ambient temperatures (Mazorra et. al., 2002). In 

general, plants tend to maintain stable tissue water status 

regardless of temperature when moisture is ample; however, 

high temperatures severely impair this tendency when water 

is limiting (Wahid et. al., 2007). Under field conditions, high 

temperature stress is frequently associated with reduced water 

availability (Simoes-Araujo et. al., 2003). In Lotus creticus, 

for example, elevated night temperatures caused a greater 

reduction in leaf water potential of water-stressed as compared 

to non-stressed plants (Anon et. al., 2004). In sugarcane, leaf 

water potential and its components were changed upon 

exposure to heat stress even though the soil water supply 

and relative humidity conditions were optimal, implying an 

effect of heat stress on root hydraulic conductance (Wahid & 

Close, 2007). Similarly, in tomato, heat stress perturbed the 

leaf water relations and root hydraulic conductivity (Morales 

et. al., 2003). 

A key adaptive mechanism in many plants grown under 

abiotic stresses, including salinity, water deficit and extreme 

temperatures, is accumulation of certain organic compounds 

of low molecular mass, generally referred to as compatible 

osmolytes (Sakamoto & Murata, 2002). Under stress, different 

plant species may accumulate a variety of osmolytes such as 

sugars and sugar alcohols (polyols), proline, tertiary and 

quaternary ammonium compounds, and tertiary sulphonium 

compounds (Sairam & Tyagi, 2004). Accumulation of such 

solutes may contribute to enhanced stress tolerance of plants, 

as briefly described below. 

Glycinebetaine (GB), an amphoteric quaternary amine, 

plays an important role as a compatible solute in plants under 

various stresses, such as salinity or high temperature 

(Sakamoto & Murata, 2002). Capacity to synthesize GB under 

stress conditions differs from species to species (Ashraf & 

Foolad, 2007). For example, high level of GB accumulation 

was reported in maize (Quan et. al, 2004) and sugarcane 

(Wahid & Close, 2007) due to desiccating conditions of water 

deficit or high temperature. 

Like GB, proline is also known to occur widely in higher 

plants and normally accumulates in large quantities in response 

to environmental stresses (Kavi Kishore et. al., 2005). In 

assessing the functional significance of accumulation of 

compatible solutes, it is suggested that proline or GB synthesis 

may buffer cellular redox potential under heat and other 

environmental stresses (Wahid & Close, 2007). Similarly, 

accumulation of soluble sugars under heat stress has been 

reported in sugarcane, which entails great implications for 

heat tolerance (Wahid & Close, 2007). Under high 

temperatures, fruit set in tomato plants failed due to the 

disruption of sugar metabolism and proline transport during 

the narrow window of male reproductive development (Sato 

et. al., 2006). 

Among other osmolytes, g 7-4-aminobutyric acid 

(GABA), a non-protein amino acid, is widely distributed 

throughout the biological world to act as a compatible solute. 

GABA is synthesized from the glutamic acid by a single step 

reaction catalyzed by glutamate decarboxylase (GAD). An 

acidic pH activates GAD, a key enzyme in the biosynthesis of 

GABA. Episodes of high temperatures increase the cytosolic 

level of Ca, which leads to calmodulin-mediated activation of 

GAD (Taiz abd Zeiger, 2006). Several other studies show that 

various environmental stresses increase GABA accumulation 

through metabolic or mechanical disruptions, thus leading to 

cytosolic acidification. 

Effect of temperature on photosynthesis 

Increasing leaf temperature and photosynthetic photon 

flux density influence thermo tolerance adjustments of PSII, 

indicating their potential to optimize photosynthesis under 

varying environmental conditions as long as the upper thermal 

limits do not exceed (Marchand et. al., 2005). In tomato 

genotypes, differing in their capacity for thermotolerance as 

well as in sugarcane, an increased chlorophyll a/b ratio and a


decreased chlorophyll and carotenoids ratio were observed 

in the tolerant genotypes under high temperatures, indicating 

that these changes were related to thermotolerance of tomato 

(Wahid & Ghazanfar, 2006). Furthermore, under high 

temperatures, degradation of chlorophyll a and b was more 

pronounced in developed leaves compared to developing 

leaves (Karim et. al., 1999). Such effects on chlorophyll or 

photosynthetic apparatus were suggested to be associated 

with the production of active oxygen species (Camejo et. al., 

2005). 

PSII is highly thermolabile, and its activity is greatly 

reduced or even partially stopped under high temperatures 

(Camejo et. al., 2005), which may be due to the properties of 

thylakoid membranes where PSII is located (McDonald & 

Paulsen, 1997). Heat stress may lead to the dissociation of 

Oxygen Evolving Complex (OEC), resulting in an imbalance 

between the electron flow from OEC toward the acceptor side 

of PSII in the direction of PSI reaction center (De Ronde et. 

al., 2004). Heat stress causes dissociation of manganese (Mn)- 

stabilizing 33-kDa protein at PSII reaction center complex 

followed by the release of Mn atoms (Yamane et. al., 1998). 

High temperature influences the photosynthetic 

capacity of C 3 

plants more strongly than in C 4 

plants. It alters 

the energy distribution and changes the activities of carbon 

metabolism enzymes, particularly the rubisco, thereby altering 

the rate of RuBP regeneration by the disruption of electron 

transport and inactivation of the oxygen evolving enzymes of 

PSII (Salvucci & Crafts-Brandner, 2004). Heat shock reduces 

the amount of photosynthetic pigments (Todorov et. al., 2003), 

soluble proteins, rubisco binding proteins (RBP) and large- 

(LS) and small-subunits (SS) of rubisco in darkness but 

increases them in light, indicating their roles as chaperones 

and HSPs (Wahid et. al., 2007). 

In any plant species, the ability to sustain leaf gas 

exchange under heat stress has a direct relationship with heat 

tolerance. During the vegetative stage, high day temperature 

can cause damage to compensated leaf photosynthesis, 

reducing CO 2 

assimilation rates (Wahid et. al., 2007). Increased 

temperatures curtail photosynthesis and increase CO 2 

transfer 

conductance between intercellular spaces and carboxylation 

sites. Stomatal conductance (gs) and net photosynthesis (Pn) 

are inhibited by moderate heat stress in many plant species 

due to decreases in the activation state of rubisco (Morales 

et. al., 2003). Although with an increase in temperature, rubisco 

catalytic activity increases, a low affinity of the enzyme for 

CO 2 

and its dual nature as an oxygenase limits the possible 

increases in Pn. For example, in maize the Pn was inhibited at 

leaf temperatures above 38 °C and inhibition was much more 

severe when temperature was increased abruptly rather than 

gradually. However, this inhibition was independent of 

stomatal response to high temperature (Crafts-Brander & 

Salvucci, 2002). Despite observed negative effects of high 

temperature, the optimum temperature for leaf photosynthesis 

is likely to increase with elevated levels of atmospheric CO 2 

. 

Assimilate partitioning 

It was determined that photosynthesis had a broad 

temperature optimum from 20 to 30°C, however, it declined 

rapidly at temperatures above 30°C. The rate of 14 C assimilate 

movement out of the flag leaf (phloem loading), was optimum 

around 30°C, however, the rate of movement through the stem 

was independent of temperature from 1 to 50°C. It was 

concluded that, at least in wheat, temperature effects on 

translocation result indirectly from temperature effects on 

source and sink activities. From such results, increased 

mobilization efficiency of reserves from stem or other plant 

parts has been suggested as a potential strategy to improve 

grain filling and yield in wheat, under heat stress. This 

suggestion, however, is based on present limited knowledge 

of physiological basis of assimilate partitioning under high 

temperature stress. Further investigation in this area may lead 

to improved crop production efficiency under high 

temperature stress. 

Effect of temperature on cell membrane thermostability 

Sustained function of cellular membranes under stress 

is pivotal for processes such as photosynthesis and 

respiration (Blum, 1988). Heat stress accelerates the kinetic 

energy and movement of molecules across membranes thereby 

loosening chemical bonds within molecules of biological 

membranes. This makes the lipid bilayer of biological 

membranes more fluid by either denaturation of proteins or an 

increase in unsaturated fatty acids (Savchenko et. al., 2002). 

The integrity and functions of biological membranes are 

sensitive to high temperature, as heat stress alters the tertiary 

and quaternary structures of membrane proteins. Such 

alterations enhance the permeability of membranes, as evident 

from increased loss of electrolytes. The increased solute 

leakage, as an indication of decreased cell membrane 

thermostability (CMT), has long been used as an indirect 

measure of heat stress tolerance in diverse plant species, 

including soybean (Martineau et. al., 1979), potato and tomato 

(Chen et. al., 1982), wheat (Blum et. al., 2001), cotton (Ashraf 

et. al., 1994), sorghum (Marcum, 1998), cowpea (Ismail & Hall, 

1999) and barley (Wahid et. al., 2007). Electrolyte leakage is 

influenced by plant/tissue age, sampling organ, developmental 

stage, growing season, degree of hardening and plant species. 

In maize, injuries to plasmalemma due to heat stress were much 

less severe in developing than in mature leaves (Karim et. al., 

1997, 1999). 

Effect of temperature on hormonal changes 

Abscisic acid (ABA) and ethylene (C 2 

H 4 

), as stress 

hormones, are involved in the regulation of many 

physiological properties by acting as signal molecules. 

Different environmental stresses, including high temperature, 

result in increased levels of ABA. For example, recently it was


determined that in creeping bentgrass (Agrostis palustris), 

ABA level did not rise during heat stress but it accumulated 

upon recovery from stress suggesting a role during the latter 

period (Larkindale & Huang, 2005). However, the action of 

ABA in response to stress involves modification of gene 

expression. Analysis of ABA-responsive promoters revealed 

several potential cis- and trans-acting regulatory elements 

(Swamy & Smith, 1999). ABA mediates acclimation/ adaptation 

of plants to desiccation by modulating the up- or downregulation 

of numerous genes (Xiong et. al., 2002). Under 

field conditions, where heat and drought stresses usually 

coincide, ABA induction is an important component of 

thermotolerance, suggesting its involvement in biochemical 

pathways essential for survival under heat-induced 

desiccation stress (Maestri et. al., 2002). Other studies also 

suggest that induction of several HSPs (HSP70) by ABA may 

be one mechanism whereby it confers thermotolerance (Pareek 

et. al., 1998). More so, heat shock transcription factor 3 acts 

synergistically with chimeric genes with a small HSP promoter, 

which is ABA inducible (Rojas, et. al., 1999). 

Heat stress changes ethylene production differently in 

different plant species (Arshad and Frankenberger, 2002). For 

example, while ethylene production in wheat leaves was 

inhibited slightly at 35°C and severely at 40°C, in soybean, 

ethylene production in hypocotyls increased by increasing 

temperature up to 40°C and it showed inhibition at 45°C. 

Despite the fact that ACC accumulated in both species at 

40°C, its conversion into ethylene occurred only in soybean 

hypocotyls but not in wheat. Wheat leaves transferred to 18 

°C followed by a short exposure to 40°C showed an increase 

in ethylene production after 1 h lag period, possibly due to 

conversion of accumulated ACC to ethylene during that period 

(Tan, et. al., 1988). Similarly, creeping bentgrass showed 

ethylene production upon recovery, but not when under heat 

stress (Larkindale and Huang, 2005). Temperatures up to 35°C 

have been shown to increase ethylene production and 

ripening of propylene-treated kiwifruit, but temperature above 

35°C inhibits ripening by inhibiting ethylene production, 

although respiration continues until the tissue disintegration 

(Antunes and Sfakiotakis, 2000). 

Among other hormones, salicylic acid (SA) has been 

suggested to be involved in heat-stress responses elicited by 

plants. Salicylic acid is an important component of signaling 

pathways in response to systemic acquired resistance (SAR) 

and the hypersensitive response (HR) (Kawano, et. al., 1998). 

SA stabilizes the trimers of heat shock transcription factors 

and aids them to bind heat shock elements to the promoter of 

heat shock related genes. Long-term thermotolerance can be 

induced by SA, in which both Ca 2+ homeostasis and 

antioxidant systems are thought to be involved (Wang and 

Li, 2006). Sulphosalicylic acid (SSA), a derivative of SA, 

treatment can effectively remove H 2 

O 2 

and increase heat 

tolerance. In this regard, catalase (CAT) plays a key role in 

removing H 2 

O 2 

in cucumber (Cucumus sativus) seedlings 

treated with SSA under heat stress. In contrast, while 

glutathione peroxidase (GPX), ascorbate peroxidase (APX) 

and glutathione reductase (GR) showed higher activities in all 

SSA treatments under heat stress, these were not key enzymes 

in removing H 2 

O 2 

(Wahid, et. al., 2007). 

Secondary metabolites 

Carotenoids are widely known to protect cellular 

structures in various plant species irrespective of the stress 

type (Wahid, 2007). For example, the xanthophyll cycle (the 

reversible interconversion of two particular carotenoids, 

violaxanthin and zeaxanthin) has evolved to play this essential 

role in photoprotection. Since zeaxanthin is hydrophobic, it is 

found mostly at the periphery of the light-harvesting 

complexes, where it functions to prevent peroxidative damage 

to the membrane lipids triggered by ROS (Horton, 2002). Recent 

studies have revealed that carotenoids of the xanthophyll 

family and some other terpenoids, such as isoprene or a- 

tocopherol, stabilize and photoprotect the lipid phase of the 

thylakoid membranes (Velikova, et. al., 2005). When plants 

are exposed to potentially harmful environmental conditions, 

such as strong light and/or elevated temperatures, the 

xanthophylls including violaxanthin and zeaxanthin partition 

between the light-harvesting complexes and the lipid phase 

of the thylakoid membranes. The resulting interaction of the 

xanthophyll molecules and the membrane lipids brings about 

a decreased fluidity (thermostability) of membrane and a 

lowered susceptibility to lipid peroxidation under high 

temperatures (Havaux, 1998). 

Phenolics, including flavonoids, anthocyanins, lignins, 

etc., are the most important class of secondary metabolites in 

plants and play a variety of roles including tolerance to abiotic 

stresses (Wahid, 2007). Studies suggest that accumulation of 

soluble phenolics under heat stress was accompanied with 

increased phenyl ammonia lyase (PAL) and decreased 

peroxidase and polyphenol lyase activities (Rivero, et. al., 

2001). Anthocyanins, a subclass of flavonoid compounds, 

are greatly modulated in plant tissues by prevailing high 

temperature; low temperature increases and elevated 

temperature decreases their concentration in buds and fruits 

(Sachray, et. al., 2002). For example, high temperature 

decreases synthesis of anthocyanins in reproductive parts of 

red apples (Tomana and Yamada, 1988), chrysanthemums 

(Shibata, et. al., 1988) and asters (Sachray, et. al., 2002). One 

of the causes of low anthocyanin concentration in plants at 

high temperatures is a decreased rate of its synthesis and 

stability (Sachray, et. al., 2002). On the other hand, vegetative 

tissues under high temperature stress show an accumulation 

of anthocyanins including rose and sugarcane leaves (Wahid 

and Ghazanfar, 2006). It has been suggested that in addition 

to their role as UV screen, anthocyanins serve to decrease 

leaf osmotic potential, which is linked to increased uptake


and reduced transpirational loss of water under environmental 

stresses including high temperature (Chalker-Scott, 2002). 

These properties may enable the leaves to respond quickly to 

changing environmental conditions. 

Superoxide radical is regularly synthesized in the 

chloroplast and mitochondrion and some quantities are also 

produced in microbodies. The scavenging of O 2- by superoxide 

dismutase (SOD) results in the production of H 2 

O 2 

, which is 

removed by APX or CAT. However, both O 2- and H 2 

O 2 

are not 

as toxic as the (OH - ), which is formed by the combination of 

O 2- and H 2 

O 2 

in the presence of trace amounts of Fe 2+ and Fe 3+ 

by the Haber-Weiss reaction. The OH - can damage chlorophyll, 

protein, DNA, lipids and other important macromolecules, thus 

fatally affecting plant metabolism and limiting growth and yield 

(Sairam & Tyagi, 2004). 

Plants have developed a series of both enzymatic and 

non-enzymatic detoxification systems to counteract AOS, 

thereby protecting cells from oxidative damage (Sairam and 

Tyagi, 2004). For example, over expression of SOD in plants 

affects a number of physiological phenomena, which include 

the removal of H 2 

O 2 

, oxidation of toxic reductants, 

biosynthesis and degradation of lignin in cell walls, auxin 

catabolism, defensive responses to wounding, defense against 

pathogen or insect attack, and some respiratory processes 

(Seandalios, 1993). More specifically, expression and activation 

of APX is related to the appearance of physiological injuries 

caused in plants by thermal stress (Mazorra et. al., 2002). 

Heat shock proteins 

Synthesis and accumulation of specific proteins are 

ascertained during a rapid heat stress, and these proteins are 

designated as HSPs. Increased production of HSPs occurs 

when plants experience either abrupt or gradual increase in 

temperature (Schoffl, et. al., 1999). Induction of HSPs seems 

to be a universal response to temperature stress, being 

observed in all organisms ranging from bacteria to human 

(Vierling, 1991). Plants of arid and semi-arid regions may 

synthesize and accumulate substantial amounts of HSPs. 

Certain HSPs are also expressed in some cells under cyclic or 

developmental control (Hopf, et. al., 1992). In this case, the 

expression of HSPs is restricted to certain stages of 

development, such as embryogenesis, germination, pollen 

development and fruit maturation (Prasinos, et. al., 2005). 

Three classes of proteins, as distinguished by molecular 

weight, account for most HSPs, viz., HSP90, HSP70 and low 

molecular weight proteins of 15-30 kDa. The special 

importance of small HSPs in plants is suggested by their 

unusual abundance and diversity. The proportions of these 

three classes differ among plant species. HSP70 and HSP90 

mRNAs can increase ten-fold, while low molecular weight 

(LMW) HSPs can increase as 200-fold. Other proteins, such 

as 110 kDa polypeptides and ubiquitin, though less important, 

are also considered to be HSPs (Feussner, et. al., 1997). All 

small-HSPs in plants are encoded by six nuclear gene families, 

each gene family corresponding to proteins found in distinct 

cellular compartments like cytosol, chloroplast, endoplasmic 

reticulum (ER), mitochondria and membranes. Some nuclearencoded 

HSPs accumulate in the cytosol at low (27°C) and 

high (43°C) temperatures, but they accumulate in chloroplast 

at moderate (~37°C) temperatures (Waters, et. al., 1996). The 

gene for a nuclear-encoded HSP, Hsa32, encoding a 32 kDa 

protein, has been cloned in tomato (Liu, et. al., 2006). 

The mechanism by which HSPs contribute to heat 

tolerance is still enigmatic though several roles have been 

ascribed to them. Many studies assert that HSPs are molecular 

chaperones insuring the native configuration and functionality 

of cell proteins under heat stress. There is considerable 

evidence that acquisition of thermotolerance is directly related 

to the synthesis and accumulation of HSPs (Bowen, et. al., 

2002). For instance, HSPs provide for new or distorted proteins 

to fold into shapes essential for their normal functions. They 

also help shuttling proteins from one compartment to another 

and transporting old proteins to “garbage disposals” inside 

the cell. Among others, HSP70 has been extensively studied 

and is proposed to have a variety of functions such as protein 

translation and translocation, proteolysis, protein folding or 

chaperoning, suppressing aggregation, and reactivating 

denatured proteins (Zhang, et. al., 2005). Recently, dual role 

of LMW HSP21 in tomato has been described as protecting 

PSII from oxidative damage and involvement in fruit color 

change during storage at low temperatures (Neta-Sharir, et. 

al., 2005). 

In many plant species, thermotolerance of cells and 

tissues after a heat stress is pretty much dependent upon 

induction of HSP70, though HSP101 has also been shown to 

be essential (Schoffl, et. al., 1999). One hypothesis is that 

HSP70 participates in ATP-dependent protein unfolding or 

assembly/disassembly reactions and it prevents protein 

denaturation during heat stress (Iba, 2002). Evidence for the 

general protective roles of HSPs comes from the fact that 

mutants unable to synthesize them or the cells in which HSP70 

synthesis is blocked or inactivated are more susceptible to 

heat injury (Burke, 2001). Heat sensitivity was associated with 

reduced capacity of bentgrass variants to accumulate 

chloroplastic HSPs (Wang and Luthe, 2003). The level of 

HSP22, a member of the plant small HSP super-family, remained 

high under continuous heat stress (Lund, et. al., 1998). LMW- 

HSPs may play structural roles in maintaining cell membrane 

integrity. Localization of LMW-HSPs in chloroplast membranes 

further suggested that these proteins protect the PSII from 

adverse effects of heat stress and play a role in photosynthetic 

electron transport (Wahid, et. al., 2007). 

Other heat induced proteins 

Besides HSPs, there are a number of other plant proteins, 

including ubiquitin (Sun and Callis, 1997), cytosolic Cu/Zn-


SOD (Herouart and Inze, 1994) and Mn-POD (Brown et. al., 

1993), whose expressions are stimulated up during heat stress. 

For example, in Prosopis chilensis and soybean under heat 

stress, ubiquitin and conjugated-ubiquitin synthesis during 

the first 30min of exposure emerged as an important mechanism 

of heat tolerance (Ortiz and Cardemil, 2001). Some studies 

have shown that heat shock induces Mn-peroxidase, which 

plays a vital role in minimizing oxidative damages (Iba, 2002). 

In a study on Chenopodium murale, when leaf proteins extracts 

from thylakoid and stromal fractions were subjected to heat 

stress it was determined that Cu/Zn-SOD from stromal fraction 

was more heat tolerant than Cu/Zn-SOD from thylakoid, and 

this was responsible for chloroplastic stability under heat 

stress (Khanna-Chopra and Sabarinath, 2004). In another 

study, a number of osmotin like proteins induced by heat and 

nitrogen stresses, collectively called Pir proteins, were found 

to be overexpressed in the yeast cells under heat stress 

conferring them resistance to tobacco osmotin (an antifungal) 

(Yun, et. al., 1997). Late embryogenesis abundant (LEA) 

proteins can prevent aggregation and protect the citrate 

syntheses from desiccating conditions like heat- and droughtstress 

(Goyal, et. al., 2005). Using proteomics tool, Majoul, et. 

al., 2003 determined enhanced expressions of 25 LEA proteins 

in hexaploid wheat during grain filling. Geranium leaves 

exposed to drought and heat stress revealed expression of 

dehydrin proteins (25-60 kDa), which indicated a possible 

linkage between drought and heat-stress tolerance (Arora, et. 

al., 1998). Recently, three low-molecular-weight dehydrins 

have been identified in sugarcane leaves with increased 

expression in response to heat stress (Wahid and Close, 2007). 

Function of these proteins is apparently slated to protein 

degradation pathway, minimizing the adverse effects of 

dehydration and oxidative stress during heat stress (Schoffl, 

et. al., 1999). 

Conclusion and future prospects 

Plants exhibit a variety of responses to high 

temperatures, which are depicted by symptomatic and 

quantitative changes in growth and morphology. The ability 

of the plant to cope with or adjust to the heat stress varies 

across and within species as well as at different developmental 

stages. Although high temperatures affect plant growth at all 

developmental stages, later phenological stages, in particular 

anthesis and grain filling are generally more susceptible. Pollen 

viability, patterns of assimilates partitioning, and growth and 

development of seed/grain are highly adversely affected. 

Other notable heat stress effects include structural changes 

in tissues and cell organelles, disorganization of cell 

membranes, disturbance of leaf water relations, and impedance 

of photosynthesis via effects on photochemical and 

biochemical reactions and photosynthetic membranes. 

ACKNOWLEDGEMENTS 

Author acknowledges University Grants Commission, 

India for the award of Junior Research Fellowship (Non- NET) 

for carrying out his M.Phil work at University of Delhi. 


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Recieved on 20-09-2012 Accepted on ..................



Knowledge and Gaps for Herbal Zinc in Relation to Their Role in Regulation of 

Male Fertility : A Review 

MANISH MATHUR 

18E-564, C.H.B., Jodhpur, Rajasthan, 342 003 

email: ravi_mm2099@yahoo.com 

ABSTRACT 

Zinc is currently the trace minerals of greatest concern when 

considering the nutritional value of vegetarian. It plays several 

important roles in a biological system and considered as the 

closest thing to a nutritional aphrodisiac. In the present review 

various aspects related with zinc and male fertility were 

discussed. Zinc plays a key role in spermatogenesis from several 

perspectives and located primarily in the Lyding cells. It takes 

part in formation of sperm motility, influences directly on sperm 

morphology and plays an important part in capacitation. Various 

biological factors, heavy or regular use of alcohol, smoking 

and endocrine abnormalities are the major factors associated 

with de crea se level of seminal plas ma zinc leve l. Its 

bioavailability is highly influenced by phytate content of food 

material that makes non-vegetarian diet more preferable then 

vegetarian diet. Many medicinal plants as well as their products 

are now being use and marketed for correcting the Zinc 

deficiency. In present review it was realized that synergistic 

studies involving Zinc and phytate concentration and their 

impacts on sperm dynamics need to taken up. 

Dr. Manish Mathur has done M.Sc. in 

Botany and Ph.D. in Plant Ecology from 

Jai Narain Vyas University, Jo dhpur, 

Rajasthan. He has also done M.Sc. in 

Ecology and Environment from Indian 

Institute of Ecology and Environment, New 

Delhi. He d id his d octo ral work on 

validation of some aphrodisiac plant. He 

has 21 papers to his credit and got trained 

in Remote Sensing, Intellectual property 

rights, Agricultural statistics and Nanotechnology. 

globally the incidence of male fertility is about 13-18% (Mathur, 

2012). There has been a rapid increase in reports of declining 

sperm counts and infertility. Such rapid increase cannot be 

attributed to genetic factor alone. Environmental exposure to 

chemical, physical agents such as heat, lifestyle factors such 

as smoking and chewing tobacco, nutritional status and air 

Key words 

Zinc, Male infertility, Phytate, Medicinal Plants, Zn 

bioavailability 

Zinc is ubiquitously present throughout all biologic 

systems and has abundant and varied functions. It is the 

second most abundant trace element in the human body, 

totaling nearly 2 g. (Grahn, et al.,2001). Found in more than 

300 enzymes and a cofactor for multiple biologic processes 

including DNA, RNA, and protein synthesis. Zn plays a key 

role in spermatogenesis from several perspectives (Croxford, 

et al.,2010). Located primarily in the Lyding cells, the late type 

B spermatogonia, and the spermatids and essential for the 

production and secretion of testosterone with follicale 

stimulating hormone of spermatogenesis (Ruwanpura, et al., 

2010). As a result, Zn deficiency has been associated with 

reduced function of the luteinizing hormone receptor (Song, 

et al., 2010), reduce steroid synthesis (Prasad, 1985)), and 

Leyding cell damage (Hesketh, 1982) emanating from oxidative 

stress (Oteiza, et al., 1996). Zn is quite high in the developing 

spermatocytes due to the need for Zn during DNA 

condensation and Meosis (Kundu and Rao, 1996). 

Zinc and Male Fertility 

It has been reported that male pattern contributes in 

40% of the cases of infertility (Wroblewska, et al., 2011), and 

Fig. 1. (Adopted from Sorensen, et al., 1999). Electron 

micrograph of human sperm cells autometallographically 

developed for zinc ions (A) Zinc grain are associated to the 

acrosome (ac). The segmented column (s), the mitochondria 

(m) and outer dense fibers (odf). (B) Cross-section of sperm 

tail Zinc grains are found at the outer dense fibers (odf) and 

the plasma membrane (pm).

MATHUR, Knowledge and Gaps for Herbal Zinc in Relation to their Role in Regulation of Male Fertility 15 

pollutants have also been reported to affect sperm quality 

(Doshi, et al.,2008). Zinc is often described as the closest 

thing to a nutritional aphrodisiac. The highest concentration 

of zinc in the male body is in the prostrate and the prostate 

fluid, ranges from 78.9 to 274.6 mg/L. (Dissanayake, et al.,2010). 

Its believed that zinc not only stimulate the prostrate 

testosterone and increase sperm count, but also stimulate the 

prostate glands. Zinc levels have been reported to influences 

sperm count (Chia, et al.,2000), antioxidant status (Gavella, 

and Lipovac, 1998) and seminal viscosity (Elzanaty, et al., 

2004). It was noticed that seminal fluid with higher percentage 

of motile spermatozoa contains plasma with higher Zn 

concentration (Wong, et al., 2001). 

Male fertility is influenced by zinc in several different 

ways, it takes part in formation of sperm motility (Kumar, et 

al., 2006), influences directly on sperm morphology (Massanyi, 

et al.,2004). Seminal plasma zinc concentration has been 

significantly correlated with sperm density, possibly 

contributing a positive effect on spermatogenesis (Chia, et 

al., 2000; Fuse, et al.,1999). 

In addition to the beneficial effects of zinc on fertility, 

the relationship of zinc in prostate health must also be 

mentioned. A correlation exists between low prostate (tissue 

and fluid) zinc levels and prostatic carcinoma. The 

concentration of zinc in the prostate is higher than that in any 

other tissue in the body. Prostatic zinc content decreases 

incrementally from normal prostate to benign prostatic 

hyperplasia (BPH) to cancer. It was shown that the semen pH 

increase was caused by the decrease of Zn concentration in 

seminal fluid. 

Factors Affecting Zinc Levels in Male 

Solomons, 1986 described that host factors and dietary 

factors determine the absorption of zinc by an individual. The 

bio-availability of zinc in most common foods typically is in 

the range of 10-30%. Some non-digestible plant constituents 

such as phytates, dietary fibres and lignin can bind zinc in 

ways that inhibit its absorption thereby engendering dietary 

zinc deficiency. 

Pathozoospermia (Dissanayake, et al., 2010), 

inflammatory conditions (Kruse, et al., 2002), accumulation of 

toxic heavy metals in the testicular tissues (Akinloye, and 

Arowojolu, 2006), chronic prostatitis (Wong, et al., 2001), low 

Zn/Cd ration (Kruse, et al.,2002), mumps-related orchitis, 

diabetes mellitus, small testicles (


Table 2. Zinc contents in different medicinal plants (sources Ansari, et al., 2004; Alwakeel, 2008; Khant, et al., 2008; 

Lokhande, et al., 2010; Annan, et al., 2010 and Stef, et al., 2010) 

S. No Plant Name Zn (µ/g) S. No Plant Name Zn (µ/g) 

1 Aichornea cordifolia 46.5 41 Hemidesmus indicus 29.4 

2 Acorus calamus 39.2 42 Hypercium perforatum 16.9 

3 Aloe barbedensis 1.1 43 Jatropha gossypifolia 56 

4 Aloe vera 12.6 44 Lawsonia inermie 30.7 

5 Amomum subulatum 45.3 45 Lepidium sativum 77 

6 Anethum graveloens 7.3 46 Lippia multiflora 45 

7 Artemisia absinthium 12 47 Mentha arvensis 40.3 

8 Artemisia herba alboa 45.1 48 Mentha piperita 11.2 

9 Artemisia vulgaris 38.14 49 Mimosa pudica 49.8 

10 Artiemisia annua 43.4 50 Mucuna pruriens 32 

11 Asparagus adscendens 32.87 51 Myristica fragrans 28 

12 Astralagus sarcocolla 2.9 52 Nerium oleander 42.2 

13 Azadirachta indica 33.3 53 Nigella sativa 52.3 

14 Boerhavia diffusa 43.5 54 Occimum basilicum 101.5 

15 Bryophyllum pinnatum 47.5 55 Occimum gratissimum 100.5 

16 Caffea arabica 8.1 56 Occimum sanctum 122.5 

17 Calendula officinalis 14.8 57 Peganum harmala 20.5 

18 Calligonum comosum 12.4 58 Pergularia daemia 63.5 

19 Capcicum frutenscens 22.8 59 Phyllanthus amarus 80.5 

20 Capsella bursa pastoris 11.4 60 Poligonum aviculare 18.3 

21 Cassia fistula 66.3 61 Ranunculus mariculatus 28.6 

22 Cassia siamea 47 62 Rauwolfia serpentine 32.6 

23 Chenopodium foliosum 50 63 Rauwoliflia vomitoria 47.5 

24 Chlorophytum borivilianum 13 64 Salvia officinalis 96.2 

25 Cichorium intybus 89.64 65 Sizygium aromaticum 13.6 

26 Cinnamomum zeylanicum 18.7 66 Stevia rebaudiana 47.18 

27 Commiphora myrrah 2.9 67 Syzigium cumin 1.1 

28 Coriandrum sativum 51.6 68 Tecoma stans 43.2 

29 Cuminum cyminum 43 69 Turraea heterophylla 66.5 

30 Curcuma longa 18.3 70 Urtica dioica 28.8 

31 Cyamopsis tetragonoloba 17.34 71 Vernona ampygdaliana 58.5 

32 Desmodium adscendens 90 72 Vetveria zinzanoides 21.2 

33 Elettaria cordamomum 50.6 73 Vinca roseus 98.5 

34 Equisetum arvense 10.7 74 Voacanga africana 70.5 

35 Eucalyptus citriodora 32.3 75 Wathania coagulans 23.1 

36 Euphorbia heliscopia 45 76 Withania somnifera 43.01 

37 Foeniculum vulgare 37.5 77 Withania sonifera 39.5 

38 Glycyrrhiza glabra 12.7 78 Zanthoxylum xanthoxyloides 48.5 

39 Gymnea sylvestris 65.5 79 Zizgiber officinale 19.7 

40 Heliotropium indicum 76.5 80 Ziziphus vulgaris 7.5 

from low intake of bioavailable Zn. Zinc deficiency is one of 

the five leading risk factors contribute to the burden of disease 

in developing countries (WHO, 2002). Zn intake in North 

Africa, the Eastern Mediterranean United States, and Canada 

at ~ 10% and in Southeast Asia, the prevalence of Zn 

deficiency is ~33% (Croxford, et al., 2010). The International 

Zinc Nutrition Consultative Group estimates that 26% of the 

Indian population is at risk of inadequate zinc intake (Hotz, 

and Brown, 2004). 

Zinc is released from food as free ions during digestion. 

These liberated ions may then bind to endogenously secreted 

ligands or to exogenous material in the intestinal lumen before 

their transcellular uptake in the distal duodenum and proximal 

junction (Cousins, et al., 1996). Zinc transport into the 

enterocyte demonstrates saturable kinetics, suggesting 

involvement of a specific carrier mechanism. With high intakes, 

zinc is also absorbed through a passive, paracellular route. 

Unlike nutrients such as vitamin A and calcium, which 

are concentrated in a relatively small number of foods or 

specific food groups, zinc occurs in a wide range of plant 

based food items as well as animal sources of food (Table 1). 

However, the bioavailability of zinc from vegetarian diets is 

comparatively less than that of non-vegetarian diet. Plant food 

rich in zinc such as legumes, whole grain, nuts, and seeds are 

also high in phytic acid, an inhibitor of zinc bioavailability 

(Hunt, 2003). Despite high phytate content that lower the 

fraction of zinc absorbed from unrefined food, the higher 

content of these food may make these foods preferable to 

more refined products lower in zinc. For example, nearly 50% 

more zinc was absorbed from a serving of whole-wheat bread 

compared with a serving of the white bread (0.22 compared 

with 0.15 mg, respectively) because the zinc content of the

Table 3. 

MATHUR, Knowledge and Gaps for Herbal Zinc in Relation to their Role in Regulation of Male Fertility 17 

Herbal preparation containing Zinc 

Product Name Company name and address Composition 

Prostate First XENA BIO HERBALS PVT LTD, 3-6-294, 

HYDERGUDA, Hyderabad - 500029, Andhra 

Pradesh, India 

Saw Palmetto: 160mg,Soy Isoflavanes: 40%- 60mg, Stinging Nettle: 

125mg,Pumpkin Seed Extract: 175mg , Pygeum: 100mg, Zinc: 15mg,Copper: 

1mg, Lycopene: 10mg 

Ultra zinc 50 

New Roots Herbal Inc. 3405 FX Tessier 

Vaudreuil-Dorion, Quebec, Canada 

Zinc (from zinc citrate)50mg, Taurine350, Vitamin B1 (thiamine HCl), 6mg, 

Vitamin B2 (riboflavin and riboflavin 5'-phosphate), 15mg, Vitamin B6 

(pyridoxal HCl and pyridoxal 5-phosphate), 67mg, Molybdenum (from 

molybdenum citrate), 200mcg, Butternut squash (Cucurbita pepo var. 

moschata, seed extract 4:1), 15mg, Green Tea extract (Camellia sinensis), 50% 

total polyphenols 75mg 

BETAMEP Alna Biotech pvt ltd, SCF-320, 2nd Floor, M. Betacaotene 30% 10mg, zinc sulphate monohydrate 27. 50 mg, selenium 

Market, Manimajra, Chandigarh, Chandigarh - dioxide 75mcg, manganese sulphate 2mg, copper sulphate 1mg. 

134 109, India 

MOXAFAV Alna Biotech pvt ltd, SCF-320, 2nd Floor, M. Soya isoflavones 40% 40mg, alfacalcidol 0. 25mcg, calcium citrate 500mg, 

Market, Manimajra, Chandigarh, Chandigarh - magnesium oxide 40mg, copper sulphate 2mg, zinc oxide 40mg. 

134 109, India 

RESMEP Alna Biotech pvt ltd, SCF-320, 2nd Floor, M. Resveratrol 30% 5mg, grape seed extract 50mg, omega-3 fatty acids 150mg, 

Market, Manimajra, Chandigarh, Chandigarh - zinc 20mg, chromium 200mcg, selenium 200mcg. 

134 109, India 

R-Gin Granules Westcoast pharmaceuticals works L-Arginione, Zinc & Folic Acid Granules 

Solvazinc ® Effervescent 

Tablets 

Galen limited Seagoe Industrial Estate, 

Craigavon, Co Armagh, Northern Ireland, BT63 

5UA, Ireland 

Zinc sulphate monohydrate , Sorbitol (E420), mannitol (E421), sodium 

hydrogen carbonate, citric acid, saccharin sodium, povidone K25, sodium 

citrate and sodium carbonate anhydrous 

Zinc (in form of mg zinc citrate trihydrate) 10 mg, Aspartame 15mg, 

Sorbitol 655 mg, sodium 8.4 mmol 

Redoxon Double Action Bayer limited, The Atrium, Blackthorn Road, 

Dublin 18 

R-Gin Granules Westcoast pharmaceuticals works L-Arginione, Zinc & Folic Acid Granules 

whole- wheat bread more than compensated for less efficient 

absorption of zinc (16.6% compared with 38.2% respectively). 

Further with respect of male infertility it is evident that zinc – 

rich plants will enhance fertility in males (Honig, et al.,1994; 

Glenville, 2008 and Ogunlesi, 2009). There are numerous 

attempts have been made to quantify the zinc contents in 

medicinal plants (Table 2), however the bioavailability of zinc 

content from these medicinal plants and their phytic acid 

concentration are synergistically not available. Thus 

comparative analysis of zinc, phytic acid content, their ration 

as well as correlation between these plant derived zinc with 

seminal zinc are required. From present review it was emerged 

that various Occimum species like sanctum (122.5 µ/g), 

basilicum (101.5 µ/g) and gratissimum (100.5 µ/g) contains 

highest zinc concentration among 80 different plant species, 

it indicates its synergism with seminal zinc. However, Sethi, et 

al., 2010 reported (Occimum sanctum) as a male contraceptive 

that reduces the sperm counts in treated albino rabbits that 

indicates that lower bioavailability of Zn from this plant that 

ultimately not supportive for sperm count. In fruit of Tribulus 

terrestris they have (Mathur, and Sundaramoorthy, 2013) 

reported that zinc concentration ranges from 2.0-2.4µ/g and 

their biological assay indicates the increase in weight (mg/ 

100g body wt) of seminal vesicle (462.63 to 731.57) and ventral 

prostrate (109.09 to 126.37) with 80% sperm motility as 

compared with control animal. Thus generalized relationships 

between the zinc concentration in herbal plants and its role in 

sperm dynamics still required. 

Zinc an important trace element for various features of 

sperm dynamics. Its roles in spermatogenesis are well defined 

and it treated as an essential element for male reproductive 

function. However zinc from vegetarian diets are generally 

less bioavailable than from non-vegetarian diets because of 

reduced meat intake as well as the tendency to consume more 

phytic acid and other plant-based inhibitors of zinc absorption. 

Various biological, environmental and life style factors 

affecting the zinc concentration in male that ultimately played 

an inductive role for male infertility. Complementary and 

alternative medicines have provide a positive hope for the 

treatment of various zinc associated male disorders, 

consequently produces many herbal and ayuervedic products. 

However there is a gap between the concentration of plant Zn 

and their efficacy for parameters associated with sperm 

dynamics. 


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Insectivorous Plants and Their Trapping Mechanism: A Review 

MURALI, S. 1 AND NARASA REDDY, G. 

Department of Agricultural Entomology, UAS, GKVK, Bengaluru 65 

e-mail: dr.mmrl@rediffmail.com 1 

ABSTRACT 

Insectivorous and carnivorous plants are those plants that 

maintain their life by trapping and eating insects and other 

creatures and are known as ‘Hunter plants’. They are among 

the curiosities of nature, being specialized in their mode of 

acquiring nutrition. These plants are usually associated with 

nutrient poor soils and wetlands, where anaerobic condition 

exists and partial decomposition of organic matter occurs. 

Insectivorous plants are successful; because they supplement 

their photosynthetic food by trapping insect and digesting their 

nitrogen rich bodies. They have brilliant colours, nectarines 

glands, sweet secretions and other curious ways to lure their 

innocent victims. In India insectivorous plants are present in 

Kerala, Assam, Karnataka, Andhra Pradesh and Nilgris in Tamil 

Nadu. The Pitcher plant, Nepenthes spp. are found in Khasie 

and Jaintia hills of Meghalaya. They have multipurpose uses. 

The sundew, Drosera indica is used in the preparation of goldbhasma 

and D. peltata is used for dying silk. Nepenthes along 

with the debris of trapped insects inside, mixed with water is 

capable of curing cholera. Utricularia aurea and U. reticulate 

are suitable for aquaria and rockeries. The insects attracted by 

insectivorous plants include mosquitoes, housefly, small 

butterflies and beetles. They digest the insects through the 

secretion of acetic acid, propionic acid, butyric and vaerianic 

acids. Insectivorous plants are very sensitive to pollutants. Some 

of the insectivorous plants have been extinct from the natural 

fauna and are included in red data book. Further field level 

studies and research are to be developed in this area to utilize 

these novel plants as a component of integrated pest 

management. 

Key words 

Insectivorous plants, Nepenthes spp., Drosera indica, 

D. peltata, Utricularia aurea, U. reticulate 

An insectivorous plant, also called as carnivorous plant, 

captures prey items, such as insects, spiders, crustaceans, 

mites and protozoan’s, as a nitrogen source. Many 

insectivorous species live in freshwater bogs, where nitrogen 

is not present in available form, because the pH of the water is 

extremely acid. About 670 species of insectivorous plants are 

found and representing nine angiospermous families such as 

Bromeliaceae (Brocchinia reducta), Byblidaceae (Bylis 

gigantean), Cephalotacea (Cephalotus follicularis), 

Dioncophyllaceae (Triphyophyllum peltatum), Droseraceae 

(About 110 species; Dionaea, Drosera, Drosophyllum), 

Lentibulariaceae (Genlisea, Pinguicula, Utricularia), 

Nepenthace (About 70 species; Nepenthes), Sarraceniaceae 

(Darligtonia, Heliamphora, Sarracenia). Way back in 1875 

Charles Darwin drew the attention of scientific community to 

the world of insects eating plants in one of his essays. The 

insectivorous plants often have several attractions such as 

brilliant colours, sweet secretions and other curious ways to 

lure their innocent victims. The mode of action, insect trapping 

mechanisms and other new aspects related to carnivorous 

plants are given in this topic. The potential of these plants for 

human welfare is also discussed below. 

Why do these plants hunt? 

Murali, S. presently pursuing Ph.D. degree 

in the Department o f Agricultural 

Entomology at UAS, GKVK, Bangalore. 

He was doing research on prospecting 

antimicro bial peptides from aculeate 

Hymenop tera: In vivo sc reening of 

antimicrobial peptides from bees and 

wasps. He was doing research on 

management of bringal shoot and fruit borer 

at NBAII, Bangalore. 

Narasa Reddy, G. Currently pursuing 

Ph.D. d egree in the dep artment of 

Agricultural Entomology at UAS, GKVK, 

Bangalore. He was engaged on Studies on 

the interrelationship b etween s oil 

mesofauna and nematodes in organic 

farming system. 

All plants need nitrogen and phosphorus to build 

proteins and nucleic acids in their cells. Most of plants get 

these elements from the soil, where they are released by 

bacteria and fungi during the break down of dead matter. 

However, in some habitats, the growth of these microorganisms 

is suppressed and the soil becomes impoverished. These 

conditions are found in blanket peat bogs, where the growth 

of the moss sphagnum produces organic acids, which kills 

many of the microbes that usually recycle nutrients in the 

soil. These plants are usually associated with rain-washed, 

nutrient poor soils or wet and acidic areas that are ill drained. 

Such wetlands are acidic due to anaerobic conditions, which 

cause partial decomposition of organic material into the 

surroundings. As a result, most microorganisms necessary


for complete decomposition of organic matter, cannot survive 

in such poorly oxygenated conditions. Normal plants find 

difficult to survive on such areas. The hunter plants are 

successful in such places because they supplement their 

photosynthetic food production by trapping insects and 

digesting their nitrogen rich bodies. However, carnivorous 

plants have evolved to steal the nitrates and phosphates of 

animals, which have become their prey. 

In carnivorous plants, the leaf is not just used for 

photosynthesis; it is also used as a trap. The problem with 

this is that changing the leaf shape to make it a better trap 

makes it less efficient at photosynthesis. For example, pitchers 

have to be held upright, so that only their lids directly intercept 

light. Even worse, the plant has to spend extra energy on nonphotosynthetic 

structures like glands, hairs, glue and digestive 

enzymes. The energy source for these things is ATP, so the 

plant has to respire more of its biomass away to keep up with 

the demand for energy. Therefore, a carnivorous plant will 

have both decreased photosynthesis and increased 

respiration, making potential for its growth (Lloyd, 1992). 

Habitat of Carnivorous plants: 

Carnivorous plants live on every continent except 

Antarctica. In USA, they are prevalent in every state. In India 

they are found in Kerala, Assam, Tamil Nadu, Ooty and Nilgiris. 

Biggest Carnivorous plants 

It comes under the genus Nepenthes. Sarracenia 

purpurea, these large vines grow up to 10 meter long. Plants 

in this genus also have traps that have evolved to capture 

some of the largest prey including creatures such as frogs, 

Very rarely, captures of birds and rodents have been reported. 

Smallest Carnivorous plant 

It comes under the genus Drosera. Drosera ultima is 

the smallest insectivorous plant. It captures tiny insects. 

Uses of Insectivorous Plants 

They posses ethno-botanical and medicinal values. 

Drosera species are important economically. Due to the rich 

content of organic acids and enzymes, these plants are capable 

of curdling milk. In ethno-medicine the brushed leaf either or 

without common salt, are applied on blisters (Pinguicula, 

Drosera), Making rope (Nepenthes ampullaris, Indonesia), 

Container for steaming rice (Nepenthes sp., Philippines), A 

food sweetner (Byblis, Austrilia), Fly catchers (Drosophyllum, 

potugal), Kinky sex toy (Utricularia, USA, Ornamental use 

(Sarracenia leucophylla, San Francisco USA), Drosera 

indica is reported to be used by ayurvedic practitioners in 

the preparation of gold-bhasma considered an anti-syphilitic 

and tonic, Yellowish brown crystalline pigment from Drosera 

peltata is used for dyeing silk. Nepenthes, the pitchers along 

with the debris of trapped insects inside are rubbed into paste, 

mixed with water and given to cholera patients. The liquid 

inside the pitcher is consumed as a remedy for urinary troubles. 

It is also used as eye drops for treating redness and itching of 

eyes. Utricularia aurea and U. reticulate are ornamental 

especially suitable for aquaria and rockeries. U. sterllaris is 

useful against cough (Somons, 1995). 

Pest control 

Carnivorous plants attract beetles, butterflies and some 

of the attract caterpillars which feed on vegetables. 

Insectivorous plants can be grown well in green houses. Those 

temperate vegetables grown under this condition, we can have 

these plants to trap caterpillars. Some carnivorous plants 

especially Drosera attract flies. This can be used as a vector 

control agent (to control house flies). In Venus flytrap many 

time mosquitoes were attracted. We can use it in future as 

mosquitoes trap. House with carnivorous plants leads to 

pollution free mosquitoes control bioagent. Largest 

carnivorous plants like Nepenthes species sometimes rodents 

were captured. If we developed it on the bunds with 

carnivorous plants acts as rodent trap. No need for rodenticide 

and training of birds etc. Anything we want to do, we can do 

through tissue culture. By multiplying these plants through 

tissue culture we can augment these wonder plants and we 

can create the condition that suits to our environment. 

Everything is possible. The ideas of persons and the way of 

utilizing technology are important. In future truly this is going 

to become a challenge to scientists. 

Insect Trapping Mechanism 

It includes four genera of which Drosera and 

Aldrovanda occurs in India. Here some of the insectivorous 

plants of India and their insect capacity mechanisms are given 

below. 

Table 1. Insectivorus plants 

S.N. Common Name Scientific Name Family 

1 Sundew plant Drosera rotundifolia Droseraceae 

2 Waterwheel plant Aldrovanda vesiculosa Droseraceae 

3 Venusfly trap Dionaea musipula Droseraceae 

4 Pitcher plant Nepenthes sp. Nepenthaceae 

5 Fly catcher Drosophyllum lusitanicum Droseraceae 

6 Bladder Utricularia sp. Lantibularaceae 

7 Butterwort Pinguicula Lantibularaceae 

Fig. 1. Pitcher Plant 

Fig. 2. Venus fly trap

MURALI AND REDDY, Insectivorous Plants and their Trapping Mechanism: A Review 21 

Pitcher Plants : Nepenthes sp. Nepenthaceae 

It conforms to the pitfall type of trap. Here the opening 

of pitcher, or pitfall, there is often a spot of bright colour, 

usually purple or white. Drops of nectar are secreted around 

the rim of pitcher and there is often a nectar baited pathway, 

from the ground to the mouth of the pitcher, that attract 

wingless insects and leads them to pitfall .Inside of pitcher is 

slippery. Ones the insect enter into the pitcher, it is unable to 

climb up because of slipperiness. Inner wall secrets photolytic 

enzyme. This helps to digest the body of trapped insects. 

Digestive enzymes are secreted by most of the species. But 

putrificative bacteria are often found in the liquid and it is 

probable that they also involved on the digestion of snapped 

insects. 

Venusfly trap: Dionaea muscipula, Droseraceae 

If the insects touch any one of the 6 filaments to cause 

both the lobes to close. The stimulus must radiate in all 

directions. From the bifurcate leaves towards the margin a 

continuous zigzag line of vessels impulse pass through the 

whole circumparence of the leaf and the midrib so all the part 

of the leaf have some communication. The central 

paranchymate cells are larger, loosely attached together and 

have delicate walls. The surface of Plants unlike animals do 

not have central nervous system. In case of Venus flytrap 

they can detect the presence of disturbance on their leaves 

by means of trigger hairs on leaf they are stimulated an 

electrical signal is generated which causes change in water 

pressure in different parts of leaf. 

As soon as the Insect is captured interlocking spines 

along the margin prevent the escape of the insect and the trap 

is more effective. It secrets digestive fluid. The leaf remains 

closed until soft tissues of insect body is completely 

reabsorbed. It will take one week. The leaf then opens and is 

soon ready to entrap other insect (Drosera within a second 

open tentacles) will open of filament by a touch followed by 

rapid closure of leaves. It captures more beetles from fast 

flying one like flies which struck into the sticky fluid. The 

most striking thing in Dionaea is in the lobes of a bit if meat 

was placed. The glands over the whole leaf surface secrete 

copiously. Glands on both sides are pressed against the meat; 

the secretion from the first is twice (Jentsch, 1970). Two lobes 

comes into close contact, the secretion, containing dissolved 

animal matter , spread by capillary attraction, causing fresh 

Glenda on both sides to begin secreting in a continually 

widening circle, and they are capable of digesting 

Reproduction of carnivorous plants 

Carnivorous plants reproduce either by sexually or 

asexually. In sexual reproduction those plants which produce 

flowers they make seeds and they reproduce sexually. In 

asexual reproduction, true plants often divide asexually. Side 

shoots develops and soon a single rosette becomes two, 

occasionally some carnivorous plants produce plantlets on 

their flower stalk. This strange behaviour is known as “false 

vivipary”. 

Cultivation of insectivorous plants 

Cultivation in home 

Majority of them are grown in indoors. Because it is 

more effective in control of house fly. Pot up the plant in 

sphagnum moss / sand then plant them up to 12 to 25 mm. (1/ 

2-1 inch). Hygiene is important. Remove all dead and diseased 

leaves. Since majority of them sun lovers best to grown in 

south facing, windows, close to the glass. Insufficient light 

leads to weakness, which unattractive leads to death. 

Artificial light 

If we wish to grow in indoors, but in places where 

insufficient light is we can go under florescent lighting. Some 

lights specially designed available in market. It consists of 

light in the red and blue wavelength, so use Gro-lux. Switch 

on light for 16 days in summer, 9hours in winter, in spring 16 

hrs photoperiod. 

Green House of Carnivorous Plants 

It is in the green house the majority of them have 

advantage. Because we can give the individual requirements. 

Different types of green houses are available. It includes cold 

frame, warm house and stove house. 

Tissue culture 

Tissue culture is the greatest utility in rapidly 

propagating rate species new cultivars. In ICPS they have the 

project on tissue culture of carnivorous plant. First it was 

started with the seeds, which were sterilized while keeping 

the seeds alive. There is an interesting phenomena associated 

with plants they have recently been planted from tissue culture 

to regular planting media. Because of hormonal imbalance it 

took years. Plants from tissue culture i.e. Sarracenia and 

Dionaea may grow in a dense clumped way. 

Chemical ecology of Carnivorous plants 

Carnivorous plants have flowers, which has some 

odours attract the pests. Simons, 1981 studied potentially 

carnivorous plant ecology. Most of the species in assemblage 

chemically attract their prey. Attraction of prey is an important 

one. In many familiar carnivorous plants chemical mimicry 

probably aids in the capture of insects Slack, 1979. Nectar 

glands help position for capture in many carnivorous plants 

including Dionaea, and the Pitcher plants, three families 

(Sarraceniaceae, Nepenthaceae, and Cephalotaceae). Various 

observations suggests that many species also attack prey 

from a distance with odours (and visual cues). Sweet nectar 

like scents is produced by many pitcher plant species.


Noxious Insectivorous Plants 

Some of the carnivorous plants Ibicella lutea is one of 

the plant, which is noxious because it has long seedpods that 

gets snagged in the feet and coats of livestock. Utricularia 

inflata will grow very faster in water become denser rapidly in 

the lake, possible displacing native species. It is noxious 

because of dense mats, which can impede boaters and others 

who would prefer open water. 

Future thrust 

Carnivorous plants are one of the nature’s gifts which 

are having lot of medicinal values cure many diseases. We 

can utilize it as a curative agent in the agenda of medicine. 

These plants have multipurpose value. It has to be 

conserved. They are threatened a lot. Some species are no 

verge of extinction he world over. In India species like Drosera, 

Nepenthes include in red data book. Particularly among the 

gardeners because of its medicinal value is the major cause 

for decline of these species. They are very sensitive to external 

environment. Pollution includes fertilizers, inorganic 

pesticides, and effluents from industries again cause decline. 

Hence it has to be conserved. Awareness has to be created 

among the students and public for its conservation. 

LIRETERTURE CITED 

Charles darwin, 1888, Insectivorous plants of the world. Dover publs. 

New York, pp.197. 

Jentsch, E. 1970. A cytochemical study of the leafy gland enzymes of 

the genus, Dionaea sp. Preceedings of Am. Phil. Soc., 57: 112- 

129. 

Lloyd, F. E. 1992. The carnivorous plants, Dover publ. New York, 

pp.144. 

Somons, H., 1995. What’s new in insectivorous plant physiology and 

Mucilaginous seed pellicles? Carnivorous pl. newsl., 8(6): 1-5. 

Slack, F. Carnivorous plant odours. Chronica botanica, Watham Mass, 

pp.112. 

Recieved on 12-01-2013 Accepted on 15-02-2013


Expression and Immunogenicity of E2 Glycoprotein Gene of Classical Swine Fever 

Virus Cloned in Eukaryotic Expression Vector 

NITIN SHARMA 1 , PURSHOTAM KAUSHIK 2 AND ANANT RAI 3 

1,2 

Department of Botany & Microbiology, Gurukul Kangri Vishwavidyalaya, Haridwar 249 404, UK, India 

3 

Institute of Biotechnology & Information Technology, 197, Mudia Ahamadnagar, Pilibhit Road, 

Bareilly 243 122, UP, India 

1 

e-mail: nitinsharma.com@gmail.com 

ABSTRACT 

The E2 glycoprotein of classical swine fever virus (CSFV) is 

immunogenic and induces neutralizing antibodies against 

CSFV. In the present study the recombinant plasmid pSin.csfv.E2 

transfected HeLa cell expressed E2 protein which was confirmed 

by immunoperoxidase test. The in vivo immunological response 

was studied in pigs immunized through IM route. The immune 

response on 28 th day post vaccination induced by 10 µg dose of 

the rplasmid revealed protective serum neutralizing antibody 

titer 60, while control group showed lower 1:5 non protective 

SN titer. Stimulation Index (SI) was found higher in vaccinated 

group as compared to healthy control animals. The T-cell 

response in the blood of vaccinated pigs suggested the role of 

cell mediated immunity in protection apart from neutralizing 

antibodies in serum. The challenge test revealed 100% 

protection of vaccinated pigs while all control pigs died of csfv 

infection. Therefore, this study demonstrates that DNA 

vaccination with 10 ìg dose of pSin.csfv.E2 can provide protective 

immunity 28 days post vaccination. 

Key words 

Classical Swine Fever Virus, DNA vaccine, E2 gene, 

immunity 

Classical swine fever (CSF), also known as hog cholera, 

is a highly contagious, multisystemic and hemorrhagic viral 

disease, included in the list of diseases notifiable to the OIE, 

distributed almost world-wide and is considered the most 

economically important but vaccine preventable disease of 

swine in areas of intensive pig farming. Natural host of classical 

swine fever virus are members of the family Suidae, which 

include domestic pigs and wild boars (Depner, et al., 1995; 

Laddomada, 2000). A considerable problem is the survival of 

CSF virus in wild pig population, which is considered to be 

the potential source of the infection. 

Genomic virus RNA possessed single ORF. Initially a 

polyprotein is formed which is further cleaved cotranslationally 

and post-tanslationally by the host and viral genome encoded 

proteases to yield four structural (C protein, Erns, E1 and E2) 

and seven non structural proteins. Erns, E1 and E2 are the 

viral envelope glycoprotein. After infection with CSFV, 

antibodies are produced against the structural glycoproteins 

E2 and Erns and non structural protein NS3 (Paton, et al., 

1991; Terpstra and Wensvoort, 1988). Antibodies against Erns 

and NS3 have only low neutralizing capacity or none at all 

(Rumenapf, et al., 1991; Konig et al., 1995). E2 glycoprotein 

contains most of the known humoral and cell mediated 

protective determinants of CSFV Ceppi, et al., 2005, highly 

immunogenic against which most of the neutralizing antibodies 

are induced and the only one capable of conferring protection 

against CSFV challenge (Rumenapf, et al., 1991; Hulst, et al., 

1993). 

Currently in India, attenuated lapinised vaccine is being 

used for conferring protection against CSFV. Although it 

ensures protection against CSFV, it however requires cold 

chain similar to any other live attenuated vaccines (Bouma, et 

al., 1999). DNA vaccines have several advantages including 

heat tolerance, safety, etc. (Patel, et al., 2007). In addition, 

they are also being used as marker vaccines to identify 

vaccinated infected and naturally infected animals (Oshop, et 

al., 2003). 

Considering the advantages of DNA vaccines, in the 

present study, the E2 gene of CSFV already cloned into pSin 

vector was undertaken for evaluating the immune response 

against the recombinant DNA construct (pSin.csfv.E2), for 

use as DNA vaccine. 

MATERIALS AND METHODS 

Recombinant plasmid 

The recombinant plasmid pSin.csfv.E2 was available in 

the Microbiology Laboratory, IBIT, Bareilly. pSin vector 

contains replicase gene which produces thousand copies of 

gene insert mRNA. 

Cell Line 

PK-15 and HeLa cell line was obtained from National 

Centre for Cell Science (NCCS, Pune). PK-15 cell line was 

maintained in EMEM (Sigma, USA), supplemented with 50 ìg/ 

ml gentamicin (Amresco) and 10 % new born calf serum (Gibco, 

NY) and was used for peroxidase based serum neutralization 

test. HeLa cell line was used in the study for in vitro expression 

analysis of recombinant plasmids (pSin.csfv.E2) and was 

maintained in GMEM (Micro lab), supplemented with 10% 

new born calf serum (Gibco, NY), penicillin 100 units/ml and 

streptomycin 100 µg /ml. 

Virus 

Virulent classical swine fever virus as well as PK-15 cell 

culture adapted CSFV were available in the Laboratory.


Conjugates and Hyperimmune serum 

Direct polyclonal antibody conjugate rabbit anti-mouse 

HRPO obtained from Bangalore Genei, Bangalore, was used 

in expression analysis of pSin.csfv.E2 and peroxidase based 

serum neutralization test and anti-CSFV E2 hyperimmune 

serum was raised in mice using recombinant plasmid 

(pSin.csfv.E2). The recombinant plasmid (50µg/mouse) was 

administered intramuscularly in thigh four times at interval of 

one week, to six mice. The blood was collected one week later 

from inner canthus of eye of the mouse and serum was 

harvested. 

In vitro expression analysis 

Transfection of HeLa cells 

Cells were trypsinised using trypsin-versenate solution 

(TVS) and then 4 ml of GMEM (Glasgow modified minimum 

essential medium) containing 10% FCS and penicillin and 

streptomycin (50 µl/ml) was added, to make cell suspension 

of 1X10 5 cells/ml. Harvested exponentially growing cells by 

trypsinization and prepared cell suspension in growth medium. 

Prepared the calcium phosphate-DNA coprecipitate as follows: 

combined 50 µl of 2.5 M CaCl 2 

with 10 µl of plasmid DNA in a 

sterile microfuge tube, Added 40 ìl DW, kept at room 

temperature. 

Immediately transfered the calcium phosphate-DNA 

suspension using 20 µl suspension for each wells of microtitre 

plate. Added 100 µl of cell culture suspension in each wells of 

96 wells microtitter plate. Rocked the plate gently to mix the 

medium, which will become yellow-orange and turbid. Carried 

out this step as quickly as possible because the efficiency of 

transfection declines rapidly once the DNA precipitate is 

formed. Kept control wells without transfection. Incubated at 

37 0 C in a humidified incubator with an atmosphere of 5% CO 2 

for 72 hours. Examined for gene expression by IPT. 

Raising primary antibody against pSin.csfv.E2 in mice 

Primary polyclonal antibody against the CSFV E2 gene 

was raised in mouse by hyper immunization of 6 mice with 

pSin.csfv.E2 plasmid. 50 µl of plasmid DNA was injected 

intramuscularly in tibialis anterior region of hind leg (lateral 

region of thigh muscles) of each mouse once a week and 

repeated for 4 weeks consecutively. One week later mice were 

bled through inner canthus of eyes with a capillary and serum 

was prepared. 

Immunoperoxidase test (IPT) 

After 72 hours, transfected HeLa cells were washed with 

1XPBS twice and fixed with 80% chilled acetone at 4 0 C for 10 

min and air-dried. Put a few drops of CSFV E2 hyperimmune 

serum and incubated at 37 0 C for 1 hr and again washed with 

PBS, added a few drops of horseradish peroxidase (HRPO) 

conjugated rabbit anti-mouse antibody to wells and incubated 

at 37 0 C for 1 hr in a humid chamber. The cells were again 

washed with PBS thrice and incubated with 2-3 drops of Nadi 

reagent for 5 min. After the development of color, cells were 

washed with PBS, dried in air and observed under microscope 

and photographed, protocol was carried out as per (Nakane 

and Kawaoi, 1974). 

Immune response studies 

Immunization of piglets 

The in vivo experimental setup was designed with eight 

weeks old piglets. One week before the start of immunization 

trial, all piglets were treated with antihelminthics 

(Albendazole). These piglets were then tested for 

seronegativity against classical swine fever. Piglets of group 

1 were injected intramuscularly with 10 µg recombinant plasmid 

(pSin.csfv.E2) in nuclease free water, and group 2 were injected 

intramuscularly 10 µg vector alone (pSin) in nuclease free 

water, while group 3 were kept as healthy control. 

Collection of blood and serum samples 

Blood samples were collected from each piglet with a 20 

gauge sterile needle of 1.5” length and 5 ml syringes at 0 day 

and 28 days of vaccination, from anterior vena cava, which is 

the most satisfactory location for obtaining the large blood 

samples. For processing under uncoagulated condition, blood 

was collected in EDTA coated vials (Unique Lab Aids), while 

for serum collection blood was collected in sterilized vials 

without any anticoagulant, kept slanted overnight at 4 0 C and 

centrifuged at 2,500 rpm for 10 min, serum was collected 

without disturbing the clot. 

Serum neutralization test (SNT) using NPLA 

The neutralizing antibodies in the blood were measured 

through neutralizing peroxidase- linked assay (NPLA) 

(Terpstra, et al., 1984). 

Lymphocyte proliferation assay 

Whole blood was collected as per the method described 

earlier in EDTA coated vial. 4 ml of lymphocyte separation 

medium (LSM 1077, PAA) was taken in a 15 ml conical tube. 

Equal volume of whole blood sample was carefully layered 

over the LSM. The tubes were centrifuged at 1800 rpm for 30 

min at 4°C. Lymphocytes were collected from the plasma-LSM 

interface and washed twice with PBS at 1200 rpm for 10 min 

and finally the cell pellet was resuspended in 2 ml of RPMI- 

1640 growth medium (Sigma) without having phenol red and 

supplemented with 10% FCS. The cell count and viability was 

determined by Trypan-blue dye exclusion method (Freshney, 

2006).The blastogenic response of lymphocytes was assessed 

by MTT colorimetry method (Mosmann, 1983). 

CSFV challenge 

All the pigs were challenged subcutaneously with 1 ml

SHARMA, et al., Expression and Immunogenicity of E2 Glycoprotein Gene of Classical Swine Fever Virus Cloned 25 

of 10 5 ID 50 

of challenge CSFV after collecting blood and sera 

on 28 th day post vaccination. Body temperatures were 

monitored daily and clinical symptoms of piglets were 

assessed by high temperature, constipation followed by 

diarrhea, staggering gait, flushing of skin upto 11 th day after 

challenge. 

RESULTS AND DISCUSSION 

Classical swine fever is a highly contagious and 

economically significant disease of domestic pigs and wild 

boars. The E2 gene engineered into an expression plasmid 

has been evaluated as DNA vaccine against CSF (Ganges, et 

al., 2005; Ratta, et al., 2010). E2 is one of the three envelope 

glycoprotein of the CSFV that is involved in virus attachment, 

entry to target cells, virulence and immunogenicity (Risatti, et 

al., 2007). 

Expression of recombinant plasmid 

The expression ability of recombinant plasmids was 

checked by immunoperoxidase test (IPT) in HeLa cell line and 

cells were found to express the protein by development of 

purple color. Intense purple coloration of cells was observed 

in which HeLa cells were transfected with pSin.csfv.E2 (Fig.1), 

while healthy cell control did not show any color change. 

Table 1. Antibody titer of piglets after immunization 

Sl. 

No. 

stimulation index (SI) of 1.11 at 28 th days of immunization with 

cell culture adapted CSFV. Unstimulated samples from each 

group maintained maximum absorbance (A 550 

) near about 0.7. 

The SI (with virus) showed an increasing trend from control 

groups to vaccinated groups. The results obtained are 

presented in the (Table 2) and (Fig.2). 

Table 2. 

Groups 

Sl. Groups 

No. 

Antigen specific response of lymphocytes at 28 th 

day post immunization as assessed by MTT assay 

* 

Mean values with a common superscript do not differ significantly at 

P


showed congestion in their mesenteric lymph nodes. Out of 

the 2 remaining animals of vector control group, 1 exhibited 

weaving gait and one pig showed erythematous lesion on its 

limbs. All the 2 pigs of vector control group showed infarction 

at the margin of spleen and hemorrhagic lymph nodes typical 

of classical swine fever in necropsy (Table 3). 

Table 3. 

Sl. 

No 

Groups 

Protection test of pigs challenged with 10 5 ID 50 

of 

CSFV 

Pig 

No. 

1. Vaccinated pSin.csfv.E2 1 

(10 µg) 

2 

3 

2. Vector control pSin (10 4 

µg) 5 

3. Healthy control 

6 

7 

No. 

of 

pigs 

No. of pigs 

showing 

symptoms 

Protection 

% 

3 0 100 

2 2 0 

2 2 0 

pSin is an alpha virus (Sindbis virus) based plasmid 

vector. After entering into the nucleus it is transcribed by 

host RNA polymerase enzymes from CMV promoter into a full 

length RNA transcript. This full length transcript then acts as 

positive sense alpha virus which in turn is translated in 

cytoplasm to form replicase protein. This protein serves as 

RNA dependent RNA polymerase enzyme and forms negative 

sense RNA from positive sense transcript. From this negative 

sense strand full length as well as smaller fragments from 

subgenomic promoters are transcribed which in-turn is 

translated into proteins. Since the cloned insert is downstream 

to the subgenomic promoter, the translated proteins represents 

our target proteins. The subgenomic promoter of alpha virus 

is very strong so that it makes large number of target mRNA 

from the sequence downstream to it. Replicase based vectors 

are superior over other conventional vectors in terms of its 

lower requirement of dose of immunization (Xiong, et al., 1989; 

Hariharan, et al., 1998; Berglund, et al., 1998; Leitner, et al., 

2000). 

The DNA vaccines induce antigen specific immune 

responses (Yokoyama, et al., 1995). Direct injection of DNA 

into skeletal muscle results in synthesis of proteins that can 

trigger the host immune system (Michel, et al., 1995). In animal 

models, DNA vaccines have been shown to elicit a broad 

range of immune responses similar to live vaccines (Yasutomi, 

et al., 1996). Therefore, the E2 glycoprotein of CSFV is also 

being targeted for the specific diagnosis or immunoprophylaxis 

of CSF (Beer, et al., 2007; Ganges, et al., 2005). Although the 

work on DNA vaccines against various strains of CSFV has 

been well focused in many other countries, however, so far no 

such reports have been documented against Indian strains 

from India. The results of the present study would be useful 

for the development of DNA vaccine against CSF disease. 

Studies are required to test pSin.CSFV.E2 developed in 

inducing protective immune response in domesticated pigs. 

The 10 µg dose of pSin.csfv.E2 with regard to their immune 

response as assessed on 28 t h day post vaccination by 

lymphoproliferative test and stimulation index, SN antibody 

titer and was found to confer total protection 28 th days post 

vaccination through intramuscular route. Thus, study 

confirmed that 10 µg recombinant plasmid was optimum for 

induction of immunity and protection in pigs. 

The 10 µg dose of pSin.csfv.E2 with regard to their 

immune response as assessed on 28th day of post vaccination 

by lymphoproliferative test and stimulation index, SN antibody 

titer was found to confer total protection on 28th days post 

vaccination through intramuscular route. 

ACKNOWLEDGEMENT 

Authors wish to thank the Director, Institute of 

Biotechnology & Information Technology, 197, Mudia 

Ahamadnagar, Pilibhit Road, Bareilly 243 122, UP, India for 

providing necessary facilities to carry out this work. 


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Morphological and Anatomical Aspects of Callogenesis and Somatic Embryogenesis 

of Date Palm (Phoenix dactylifera L.) 

AKSHAYA BHATI 1 AND ATUL CHANDRA 2 

1 

SK Rajasthan Agricultural University, Bikaner 

2 

KVK, SK Rajasthan Agricultural University, Bikaner 

email: akshaya.horti@gmail.com 1 

ABSTRACT 

This study was carried out to investigate the morphological 

and ana tomical aspe cts of c allo gene sis and soma tic 

embryogenesis in date palm (Phoenix dactylifera L.). Shoot tip 

explants excised from young offshoots were cultured on MS 

medium with 2,4-D, BA and 2-ip. Somatic embryogenesis was 

produced by transferring the calli produced on BA medium. 

Sectioning of specimens was carried out using the Somatic 

embryos, callus tissue and its differentiating structures. The 

sections were stained with safranin and fast green. Histological 

examination of swollen explants, after eight weeks (60 days) 

showed formation of new parenchymatous cells resulting into 

swelling of explants. The new cells were small, packed densely 

and appeared in concentric layers. Around 130-140 days, when 

very small amount of callus was seen first, the sections revealed 

formation of periclinal concentric layers of cells from the 

peripheral region of explants. At this stage, cells surrounding 

the callus cells looked empty. Inner region of callus exhibited 

a core region composed of densely packed meristematic cells 

with cortical zone surrounded by layer cells 1-2 vacuoles. This 

region was surrounded by loosely arranged parenchymatous 

cells, which constituted the visible callus. Small individualized 

embryogenic masses and globular embryos were observed 

embedded in degenerating parenchyma, soon they matured 

into individual globular embryos surrounded by an epidermal 

cell layer. Longitudinal section of further developed embryos 

displayed a well differentiated vascular system along the 

cotyledon and a fully organized shoot apex. 

Key words 

Somatic Embryogenesis, Callogenesis, date palm 

Date palm (Phoenix dactylifera L.) is one of the most 

important horticultural crop cultivated in arid regions. Date 

palm is usually propagated by offshoots, which are mainly 

produced during the early life of the palm in limited numbers, 

depending on variety and other factors. Propagation by seeds 

is undesirable and impractical due to their dioecious nature. 

Plant tissue culture techniques have been used to propagate 

date palm for rapid and mass multiplication. In spite of great 

potential of tissue culture techniques for palms, there are some 

difficulties in the propagation of date palm. An extensive effort 

has been carried out over the last two decades to establish 

effective micropropagation systems. Somatic embryogenesis 

has been accomplished in several palms, including date palm, 

but only a small percentage of the somatic embryos produce 

vigorous plantlets. Increasing the conversion rate of the 

embryos to plantlets are very important factors in improving 

somatic embryo efficiency. 

Studying the morphological and anatomical aspects of 

embryogenesis at different stages in tissue culture can be 

guide for increasing the effectiveness of the system, in 

addition to increasing our knowledge about in vitro associated 

phenomena. In few histological studies on date palm somatic 

embryogenesis has been done (Davoodi, et al., 2002; Sane, et 

al., 2006), although the different investigators have not 

reached to the same conclusion about the developmental 

pattern of somatic embryos in date palm. The potential for 

obtaining somatic embryos from callus tissue produced from 

shoot tips and primordial leaves has been reported. This study 

has been carried out to investigate the morphological and 

anatomical aspects of somatic embryogenesis in cultivar 

Halawy and Medjool. 


Tissue Culture 

Offshoots of date palm cv. Medjool, Shamran and 

Halawy separated from adult trees were used as a plant 

material. The outer leaves were gradually removed till the 

tender portion was reached. The primordial leaf and apical 

meristem with sub-apical tissues were excised and kept in 

anti-oxidant solution (150 mg/l citric acid + 100 mg/l ascorbic 

acid) for 30 minutes. The explants were surface sterilized with 

0.1% HgCl 2 

containing few drops of Tween-20 with continuous 

stirring for 10-12 minutes followed by washing with autoclaved 

water for 6-8 times. Shoot tips of 1.5 cm and pieces of internal 

leaves were cultured on MS medium supplemented with 10 

mg/l 2,4-D + 3 mg/l BA + 1 mg/l 2-ip, additives (100 mg/l inositol, 

40 mg/l adenine sulphate, 170 mg/l NaH 2 

PO 4 

, 200 mg/l 

glutamine and 5 mg/l thiamine), 1.5 gm/l activated charcoal. 

The cultures were incubated in dark and 27±2ºC. At 15 days 

intervals, they were sub-cultured in order to obtain callus 

with granular appearance. For inducing morphogenesis, the 

MS medium with 0.1 mg/l BA supplemented with additives 

(100 mg/l inositol, 40 mg/l adenine sulphate, 170 mg/l NaH 2 

PO 4 

, 

200 mg/l glutamine and 5 mg/l thiamine) was used. Cultures 

were incubated under constant light (1000 lux) at 27±2ºC 

temperature. 

All culture media were solidified with 0.7% agar and 

adjusted to pH 5.8 before autoclaving at 1.5 kg/cm 2 

for 20 

minutes.

BHATI & CHANDRA, Morphological and Anatomical Aspects of Callogenesis and Somatic Embryogenesis 29 

Histological studies 

The callus tissue and differentiated structure were 

sampled at different stages (initial, secondary and differntiated 

calli) fixed in FAA (Formalin: Acetic acid: Alochol 90: 5: 5) for 

24 hours then dehydrated using 50, 70, 80, 90% and absolute 

concentrations of ethanol (Davoodi, et al., 2002). After 

dehydration, the specimen were embedded in parrafin and cut 

into 5-10 µm sections. The sections were double stained with 

safranin and fast green. 


The callus induction and its proliferation took a minimum 

of 159 days. Histological examination of swollen explants, after 

eight weeks (60 days) showed formation of new 

parenchymatous cells resulting into swelling of explants. Callus 

formation was not, however, evident at this stage. The new 

cells were small, packed densely and appear in concentric 

layers. 

Around 130-140 days, when very small amount of callus 

was seen first, the explants were sectioned again. The sections 

revealed formation of periclinal concentric layers of cells from 

the peripheral region of explants. At this stage, cells 

surrounding the callus cells looked empty (Fig. 1a). Inner 

region of callus exhibited a core region composed of densely 

packed meristematic cells with cortical zone, surrounded by 

layer cells with 1-2 vacuoles. This region was surrounded by 

loosely arranged parenchymatous cells, which constituted 

the visible callus (Fig. 1a). 

After about 20-25 days of transfer of primary callus on 

induction and maturation medium (MS + additives), the calli 

were examined for embryonic masses. The examination revealed 

that calli have further proliferated into microcalli comprised of 

two zones of cells. In the inner part of microcalli, the cells were 

meristematic and small (Fig.1c) with dense cytoplasm without 

any visible vacuole. This region was surrounded by loosely 

arranged parenchymatous cells, which constituted the bulk 

of visible callus (Fig 1a.). 

The calli were examined for embryonic masses. The 

examination revealed that calli have further proliferated into 

microcalli. The cells were meristematic and small (Fig. 1b) with 

dense cytoplasm without any visible vacuole. In the outer 

part, the cells were embryogenic with less dense cytoplasm 

and appeared to be rich in starch. Fragmentation zones were 

also visible. 

Small individualized embryogenic masses and globular 

embryos were observed embedded in degenerating 

parenchyma (Fig.2e), soon they matured into individual 

globular embryos surrounded by an epidermal cell layer 

(Fig.1f). longitudinal section of further developed embryos 

displayed a well differentiated vascular system along the 

cotyledon and a fully organized shoot apex (Fig.2h). 

The shoot apex consisted of a meristematic dome 

surrounded by a leaf primordium. The most meristem area, 

however was less organized and diffused. This probably led 

to precocious germination of abnormal embryos (Sane, et al., 

2006). 

Before becoming committed into a new morphogenesis 

in vitro programme, somatic cells undergo several successive 

physiological processes viz., Organgenetic competence 

acquisition, determination for a particular organ formation and 

ultimately the organ formation. Reorientation is possible before 

the determination step under the influence of PGR’s and the 

Fig. 1. 

Histological features of shoot morphogenesis in date 

palm. (a) Compact Callus (b) Meristematic Zones 

(c) Mic rocalli showing proembryonic mas ses 

(d) Globular somatic embryos 

Fig. 2. 

Histological features of shoot morphogenesis in date 

palm. (e) Embryo embed ded in to the callus 

(f) Epidermal layer of somatic embryo (g) Shoot apex 

from somatic embryo (h) Fully organized shoot apex


totipotency of plant cells confer flexibility to their 

organogenetic programs. Observed that cells have to either 

die (apotosis) or to differentiate and divide to undergo 

morphogenesis patterns such as embryogenesis. It is now 

assumed that the plant developmental program can give rise 

to alternative pathways have detected the concept of 

totipotent cells versus pluripotent cells. 

In date palm callus cultures, three phases of 

embryogenesis can be determined (i) the acquisition phase, 

which is mostly under the influence of a strong auxin like 2,4- 

D (100 mg/l), mostly in high concentration (Kackar, et al., 

1989; Yadav, et al., 2001). But also there are reports where low 

level of 2,4-D (2 mg/l) were found to be equally effective 

(Bhargava, et al., 2003; Zouine, and El-Hadrami, 2007; Taha, 

et al., 2007). In majority of these reports BAP / Kn was one of 

the cytokinin; (ii) the determination phase for embryogenic 

pathway mostly depending on an auxin like NAA / IAA with 

or without a cytokinin (2.5-10 mg/l, Bhargava, et al., 2003; 

Zouine and El-Hadrami, 2007; Taha, et al., 2007), (iii) 

differentiating phase where maturation and germination occurs 

under the influence of a cytokinin/ auxin or only ½MS medium. 

In present investigation, it looks not less than 15 days 

when primary callus was produced under the influence of 2,4- 

D + BA at places it was composed of meristematic zones 

surrounded by parenchymatous cortex. However, uptill this 

stage it was difficult to ascertain that whether these 

meristematic structures would develop into shoots or 

embryos. However, the presence of meristematic zones in 

primary callus indicated the acquisition of competence, the 

first stage of embryogenesis. Although in the callus induction 

medium BAP (3 mg/l) was present but dominating role of 2,4- 

D (10-100 mg/l) was evident. Auxins have many effects and 

can modulated diverse processes and tropic responses. 

Exogenous auxins are assumed to orient developmental 

pathways and to favour either callogenesis or rhizogenesis. 

In our experiments when primary competent callus was 

transferred on to maturation medium, globular embryogenic 

masses and embryos were observed within 20-25 days (Fig. 

2d) due to hard well developed vascular supply. Absence of 

phytohormones has been reported to favour embryo 

maturation in various date palm tissue culture studies (Kackar, 

et al., 1989 and Yadav, et al., 2001). 

It is well established that 2,4-D plays a role in the 

induction of somatic embryogenesis which is presumably 

mediated by a signal cascade triggered by this exogenous 

auxin (Zouine and El Hadrami, 2007). 

Increasing the proportion of cytokinin in the hormonal 

balance is thought to promote the expression of somatic 

embryogenesis and later development of embryos. In present 

investigation low level of cytokinin (BA, 3 mg/l) was more 

effective in maturation of somatic embryos. Bhargava, et al., 

2003 have observed that both the level of 2,4-D in the callus 

induction medium and level of cytokinin (0.25 mg/l) was very 

important for normal somatic embryos induction. More high 

was the level of these hormones, more was the number of 

abnormal embryos. 

Sane, et al., 2006 have also observed that application of 

cytokinin (BA) promoted the appearance of meristematic 

territories in the tissues whose successive divisions led to 

polarization of the pro-embryo within 3-4 weeks of culture. 

They undertook a detailed study of histological events during 

somatic embryogenesis in date palm and has described 

different stages: the division of perivascular cells on primary 

explants, the secondary caulogenesis in primary, the 

unicellular or pluricellular origin of the embryos and the 

development of embryos into complete explants. All these 

stages were observed in present study as well. Abnormal 

embryos and precocious germination of embryos was also 

observed in present study. This could be due to precocious 

cell reactivation as suggested by Gueye, et al., 2009. As for 

many monocotyledons, callogenesis is a prerequisite for the 

initiation of somatic embryos and requires the presence of an 

auxin in the medium (Davoodi, et al., 2002). 


Bhargava, S.C., Saxena, S.N. and Sharma, R. 2003. In vitro 

multiplication of Phoenix dactylifera. J. Plant Biochemistry and 

Biotechnology. 12: 43-47. 

Davoodi, D., Majidi, E. and Khoshkam, S. 2002. Some morphological 

and anatomical aspects of date palm (Phoenix dactylifera L.) somatic 

embryogenesis in tissue culture. J. Agric. Sci. Technol., 4: 63-71. 

Gueye, B., Said Ahmed, H., Morcillo, F., Borgel, A., Sane, D., Hilbert, 

J.L., Verdeil, J.L. and Blervacq, A.S. 2009. Callogenesis and 

rhizogenesis in date palm leaf segments are there similarities between 

the two auxin induced pathways. Plant Cell Tissue Organ Culture. 

98: 47-58. 

Kackar, N.C. ; Solanki, K.R. and Joshi, S.P. 1989. Micropropagation of 

Date palm (Phoenix dactylifera L.) cv. Khadrawy using tissue culture 

technique. Ann. of Arid Zone, 28: 137-141. 

Sane, D., Aberlenc-Bertossi, F., Gassama-Dia, K., Sagna, M., Trouslot, F., 

Duval, Y. and Borgel, A. 2006. Histocytological analysis of callogenesis 

and somatic embryogenesisfrom cell suspensions of date palm 

(Phoenix dactylifera L.). Annals of Botany. 98: 301-308. 

Taha, H.S., Hassan, N.M. and Al-Bahr. 2007. Micropropagation of 

some Egyptian date palm dry cultivars 1- maturation of somatic 

embryos. Arab J. Biotech. 10(2): 333-340. 

Yadav, N., Yadav, R.C., Chowdhury, V.K. and Chowdhury, J.B. 2001. 

Explant and cultivar response to in vitro clonal propagation of 

female date palm. Paper presented in II nd International Conference 

on Date palm (Al- Ain, UAE, March 25-27). 

Zouine, J. and El Hadrami, I. 2007. Effect of 2,4-D, glutamine and 

BAP on embryogenic suspension culture of date palm (Phoenix 

dactylifera L.). Scientia Horticultrae, 112: 221-226. 



Successful Therapeutic Management of Snake Bite in A Dog 

SONAL SHRIVASTAVA 1 , DEBOSRI BHOWMICK 2 AND P.C. SHUKLA 3 

1,2 

Department of Veterinary Surgery & Radiology, 3 Department of Veterinary Medicine College of Veterinary 

Science & A.H., N.D.V.S.U., Jabalpur 

ABSTRACT 

A dog was brought to the hospital with the history of snake bite 

in the forelimb. The snake was caught and identified by the 

snake expert, to be Russell’s viper. Clinical examination 

revealed swelling of the bitten limb with oozing of blood from 

the bitesite. Dullness, depression, vomition and frothy 

salivation were also observed. The dog was treated with the 

Snake Venom Antiserum. Supportive therapy icluding 

intravenous fluid (5% DNS), Paracetamol, Atropine Sulphate, 

Ranitidine Hydrochloride, Dexamethasone, Botropase, 

Ceftriaxone and Tetanus Toxoid was also given. The dog 

recovered completely in 5 days. 

Key words 

Dog, Russell’s Viper, snake bite, snake venom 

antiserum 

Snake bite, with envenomation, is a true emergency with 

significant morbidity and mortality in canines. In comparison 

to other domestic animals, fatal snakebites are more common 

in dogs due to their relatively small size, in proportion to the 

amount of venom injected at the bitesite. In India, the Big 

Four are the four venomous snake species responsible for 

causing the most snake bite cases. According to Whitaker, 

1990, the big four includes, Indian cobra (Naja naja), Common 

krait (Bungarus caeruleus), Russell’s viper (Daboia russelii), 

and Saw-scaled viper (Echis carinatus). The present paper 

describes snake bite in a dog and its successful therapeutic 

management. 


Case History and Clinical Examination 

A male German Shepherd dog aged 4 years was brought 

to the hospital with the history of snake bite in the forelimb. 

According to the owner the snake was caught and identified 

by the snake expert, to be Russell’s viper. Clinical examination 

revealed swelling of the bitten limb with oozing of blood from 

the bitesite (Fig.1). Fang marks were noticed at the site (Fig.2). 

The dog was dull and depressed; vomiting and frothy 

salivation were also observed. Haemato-biochemical 

parameters did not show any significant alterations. 

Therapeutic Management 

The dog was treated with the Snake Venom Antiserum 

(Lyophilised, Polyvalent, Enzyme refined, Equine 

Immunoglobulins). Freeze dried powder was reconstituted with 

10 ml of sterile water for injection supplied with the vial. 

Reconstituted antivenom was administered by slow 

intravenous injection (2 ml/minute). Intravenous fluid therapy 

(500 ml of 5% DNS), injection Paracetamol @ 15mg/kg b.wt. 

IM, injection Atropine Sulphate @ 0.04 mg/kg b.wt. IV, injection 

Ranitidine Hydrochloride @ 2 mg/kg b.wt. IV, injection 

Dexamethasone @ 0.15 mg/kg b.wt. IV, injection Botropase 

1ml IV, and Tetanus Toxoid 2 ml IM were also administered. In 

addition, injection Ceftriaxone 500 mg IV was administered for 

5 days. Betadine solution was topically applied on the bitesite. 

Fig. 1. 

The bitesite on the forelimb. Note the swelling and 

oozing of blood. 

Fig. 2. 

Fang marks at the bitesite


Fig. 3. Animal recovered completely after 5 days 

The supportive therapy was continued for 5 days. The animal 

completely recovered after 5 days of treatment (Fig.3). 


Snake venom is highly modified saliva containing 

zootoxins. According to Halliday and Tim, 2002 venoms 

contain more than 20 different compounds, mostly proteins 

and polypeptides. A complex mixture of proteins, enzymes, 

and various other substances with toxic and lethal properties 

are responsible for the biological effects (Bauchot, 1994). In 

the case of Russell’s viper the usual manifestations comprise 

persistent pain and swelling of the bitten limb with oozing of 

blood from the bitesite. It may be followed by generalized 

vascular injury with severe external and internal haemorrhage. 

Death may result from cardio vascular shock or renal failure. 

Antivenom should be administered as soon as possible after 

the diagnosis of envenomation. Antivenin is highly beneficial 

because its action is the only direct and specific mechanism 

for neutralizing snake venom. Moderate to severe pain caused 

by snake bite responds well to paracetamol. Envenomation 

requires intensive treatment, starting with IV fluids to combat 

hypotension. Administration of corticosteroids helps to 

control shock, protect against tissue damage caused due to 

envenomation, and reduces the possible allergic reactions to 

antivenin. Tetanus antitoxin and antibacterial agents are 

administered to prevent tetanus and bacterial infections 

respectively, as the fangs of snake are likely to contain 

bacteria. Other supportive treatment (eg. antacids, haemostats 

etc.) is given as and when needed. 


Bauchot, R. 1994. Snakes: A Natural History. New York City, NY, USA: 

Sterling Publishing Co., Inc. pp. 194-209. 

Halliday, A. and Tim, Kraig 2002. Firefly Encyclopedia of Reptiles and 

Amphibians. Toronto, Canada: Firefly Books Ltd. pp. 202-203 

Whitaker, Z. 1990. Snakeman. Penguin Books Ltd. pp. 192. 

Recieved on 08-11-2012 Accepted on 23-01-2013


Biology and Feeding Potential of Cheilomenus sexmaculata (Fab.) on Bean Aphid 

(Aphis craccivora Koch) and Mustard Aphid (Liphaphis erysimi L.) 

AKSHAY KUMAR, C.S. PRASAD AND G.N. TIWARI 

Department of Entomology, Sardar Vallabhbhai Patel Uni. of Agric. and Tech., Meerut (U.P.) India 

email: akshaypanwar1259@gmail.com 

ABSTRACT 

An experiment “biology and feeding potential of Cheilomenus 

sexmaculata on bean aphid (Aphis craccivora Koch) and mustard 

aphid (Liphaphis erysimi L.)” was carried out at Biocontrol 

Laboratory, S.V.P.U.A. & T., Meerut (U.P.) India. The ladybird 

beetle C. sexmaculata completed their life cycle in 18-27 days. C. 

sexmaculata laid eggs ranged 150-210 eggs during her life period. 

The 1 st , 2 nd , 3 rd and 4 th instar larval period lasted for 2.67±0.33 

days (ranged 2-3 days), 2.00±0.58 (ranged 1-3 days), 2.67±0.33 

(ranged 2-3 days) and 4.00±0.58 (ranged 3-5 days), respectively. 

Fourth and final instar grubs were deep black in colour, when 

freshly moulted but changed to dull black in colour before prepupation. 

The pre-pupal period was lost in 2.67±0.33 (ranged 2- 

3 days) while the pupal period was lost in 4.67±0.33 (ranged 4-5 

days). Male and female beetles survived for 29-33 and 33-38 days 

with a mean survival period of 30.67±0.88 and 35.33±1.45 days, 

respectively. The total number of aphid consumed during whole 

larval duration of C. sexmaculata varied from 70-95 Aphis 

craccivora and 63-85 Lipaphys erysimi, respectively. A male beetle 

consumed 510.33±9.82 A. craccivora (ranged 500-521 aphids) and 

410.33±4.26 L. erysimi (ranged 400-416 aphids), with an average 

of 28.72 A. craccivora and 32.92 L. erysimi per day, respectively. 

While, a female beetle consumed 718.00±17.21 A. craccivora 

(ranged 710-725 aphids) and 555.67±27.63 L. erysimi (ranged 547- 

571 aphids), respectively with an average of 35.93 A. craccivora 

and 27.90 L. erysimi per day, respectively. 

Key words 

Biology, Feeding potential, Cheilomenus sexmaculata, 

aphid 

In balanced ecosystem, insect pest are kept in check by 

their natural enemies (predators and parasitoids) these agents 

are beneficial in agricultural systems. Cheilomenus 

sexmaculata is an important predator of aphid, coccids, 

psyllids and many ecosystems of South East Asia (Parker, et 

al., 1976; Gupta and Yadava, 1986; Lokhanda and Mohan, 

1990; Hussein, et al., 1991; Maisini, et al., 1994). It is an 

effective predator to be used as bio-control agent but the 

major challenge is its mass rearing and augmentation. Hence 

the present study was therefore, conducted biology and 

feeding potential of C. sexmaculata (Fab.) on bean aphid 

(Aphis craccivora Koch) and mustard aphid (Liphaphis 

erysimi L.). 


Biology of Cheilomenus sexmaculata 

A pair of beetles was placed in raring jar with sufficient 

amount of aphids as food and substrate for egg laying. The 

eggs laid by female on the surface of glass jars and given 

substrates were removed with the help of a moist camel hair 

brush, while those deposited on the substrate were removed 

along with the substrate. Care was taken so that the eggs 

were not damaged. 

Study of incubation period 

Ten eggs of each coccinellid beetles were kept in a 

plastic vial. To maintain the moisture, the bottom of the vial 

was covered with moistened filter papers. These eggs were 

checked twice per day i.e. 10 a.m. and 5:30 p.m. to observe the 

incubation period and number of eggs hatched. 

Larval period 

The larval period was recorded from the day of hatching. 

Neonates which hatched from the eggs kept for incubation 

were used for the study. Rearing of the larvae were carried out 

individually in plastic vials (14X1.5) and aphid of almost 

different size were provided as food. Fresh aphids were 

provided to the growing larvae every morning. To observe 

the moulting of the developing larvae, the exuviae were 

checked twice daily at morning and evening hours. The 

duration of all larval (grubs) instars were recorded 

continuously from 1 st to 4 th instar. 

Pre-pupal to pupal period 

The pre-pupal period i.e., the period when the 4 th instar 

grub stopped feeding up to the period when it was completely 

transformed in to pupa, was also observed. 

Emergence of adults from pupae 

For studying the pupal period, freshly formed pupae 

from the batch of the grub used for grub period studies were 

kept in rearing plastic vials and observations on the emergence 

of coccinellids beetles from the pupae were recorded twice 

daily. 

Oviposition period and adult longevity 

Newly emerged adults were released in pairs in rearing 

glass jar to observe the duration of mating and ovipositon 

period. The adults after emergence were sexed on the basis of 

shape and size as well as morphological character. It was 

observed that the males were smaller than the females in 

general. After pairing, their mating behaviour was also studied 

by daily visual observations. To study adult longevity, newly


emerged adults were kept in rearing jar and were provided 

with aphids as food materials. Fresh food was provided every 

day and observations on the mortality, if any, were recorded. 

Feeding potential of C. sexmaculata 

Fifty or sixty freshly collected aphids for adult 

coccinellids and 10 for grubs viz., A. craccivora and Lipaphis 

erysimi were placed on fresh and tender mustard and bean 

leaves/twigs, respectively. The aphids were then allowed to 

settle. Different instars of grubs and adults of coccinellids 

starved for 2 hours except first instar grub were released singly 

inside each petridish with the help of a fine hair brush. The 

daily consumption by instars all of larvae and adult of 

coccinellids was calculated. Thereafter, the number of 

unconsumed aphids left over in the petridishes were collected 

daily and calculated for consumed ones. The actual number 

of aphids consumed by grubs/adult was obtained by 

subtracting the number of aphids left over from the total number 

of aphids supplied every day. 


Description of different stages of C. sexmaculata 

Egg 

Freshly laid eggs were cigar shaped, yellow in colour 

with smooth chorion and without any reticulations. The eggs 

turned blackish with advancement of age and became 

completely black before hatching. The eggs were usually laid 

in clusters of 4 to 10 and each cluster consisted of 5 to 25 

eggs. Number of eggs laid by an individual female of C. 

sexmaculata ranged from 150-210 eggs during her life period. 

These findings are supported by the findings of Zala, 1995 

and Tank, et al., 2007. The incubation period varied from 2-4 

days with an average of 3.00±0.58 days (Table 1), which is in 

accordance with the findings of Tank, et al., 2007. 

Grub 

During the present study, it was observed that the larvae 

of C. sexmaculata moulted thrice and thus there were four 

instars. Freshly hatched larvae were dark grey in colour with 

shining dark head capsule and legs. The first instar larval 

period lasted for 2.67±0.33 days (ranged 2-3 days) (Table 1). 

Second instar larvae were glistening black in colour with 

yellow coloured head capsule and black legs. Development 

of white coloured patches on meso and metathorax and also 

on fourth and sixth abdominal segments was observed. The 

second instar larval period lasted in 2.00±0.58 (ranged 1-3 

days) (Table 1). 

The freshly moulted third instar larvae were dull black in 

colour with yellow head capsule. The colour pattern was more 

intensified with additional development of white spots on 

mid-dorsal line of other segments except prothorax. The third 

instar larval period was lost in 2.67±0.33 (ranged 2-3 days). 

Table 1. Developmental period of different stages of C. 

sexmaculata 

S. No. Stage 

Duration (days) 

Range Mean* 

1 Egg (incubation period) 2-4 3.00±0.58 

2 Grub 

First instar 

Second instar 

Third instar 

Fourth instar 

2-3 

1-3 

2-3 

3-5 

2.67±0.33 

2.00±0.58 

2.67±0.33 

4.00±0.58 

3. Pre-pupa 

Pupa 

2-3 

4-5 

2.67±0.33 

4.67±0.33 

4 Adult longevity 

(a) Female 

(b) Male 

33-38 

29-33 

35.33±1.45 

30.67±0.88 

5 Total period taken from egg 

laying to adult emergence 

16-26 

*Mean of three replications with 10 pairs of adult male and females 

Fourth and final instar grubs were deep black in colour, when 

freshly moulted but changed to dull black in colour before 

pre-pupation. It developed additional rectangular dark grey 

spots in a continuous series mid-dorsally on abdominal 

segments, whereas the spots on the fourth abdominal segment 

were white. The fourth instar larval period was lost in 4.00±0.58 

(ranged 3-5 days) (Table 1). 

The present investigation revealed that the mean 

duration of first, second, third and fourth instars larvae of C. 

sexmaculata were 2.67±0.33, 2.00±0.58, 2.67±0.33 and 4.00±0.58 

days respectively and the total larval period lasted for 8-14 

days. Whereas Tank and Korat, 2007 observed that the mean 

duration of first, second, third and fourth larval instars were 

1.80±0.50, 1.72±0.46, 1.88±0.53 and 1.96±0.73 respectively and 

the total larval period ranged from 5-10 days. More or less 

nearest values of duration of different larval instars had been 

recorded by Rai, et al., 2003. 

Pre-pupa and Pupa 

When larva was about to pupate, it turned wood brown 

in colour and assumed curved shape and attached itself 

posteriorly to the substrate. The pre-pupal period was lost in 

2.67±0.33 (ranged 2-3 days) while the pupal period was lost in 

4.67±0.33 (ranged 4-5 days) (Table 1). Freshly formed pupae 

appered shinning yellow in colour which later on became pale 

appering orange yellow in colour. There were symmetrically 

orange black spots on each segment. This is in accordance 

with the report of Sureja, 1991 and Tank and Korat, 2007 

observed pre-pupal and pupal period was 1.72±0.45 and 

3.36±0.70 days respectively. 

Adult 

Freshly emerged adults were soft bodied, yellowish in 

colour without any markings which turned shining yellow or 

warm buff with black spots which developed gradually. The 

adults were small, oval, convex dorsally and flat ventrally. 

Abdomen and eyes were light yellow in colour whereas elytra 

and pronotum were marked with zigzag markings. Similar

KUMAR, et al., Biology and feeding potential of Cheilomenus sexmaculata (Fab.) on bean aphid (Aphis craccivora Koch) 35 

description had also been narrated by Sureja, 1991. Male and 

female beetles survived for 29-33 and 33-38 days with a mean 

survival period of 30.67±0.88 and 35.33±1.45 days, respectively 

(Table 1). Tank, et al., 2007 reported that the male and female 

beetles survived from 10-21 and 15-26 days respectively and 

also observed that the longevity of male without food (prey) 

was 2 to 4 (2.5±0.68) days while for female it was 3 to 5 

(3.35±0.58) days, indicating females survived slightly longer 

than the males in absence of food supporting the view that 

females survived longer than the males. Tank, et al., 2007 

reported that the average duration of entire life- span for male 

and female was 29.72±2.20 and 34.15±2.54 days, respectively. 

Feeding potential of C. sexmaculata 

For studying feeding potential of C. sexmaculata, the 

observations were recorded at temperature varying between 

21.0 and 25.0 0 C with a relative humidity of 61 to 64 per cent. 

The data presented in Table 2 indicates that the rate of feeding 

among different larval instars and adult varies greatly. 

Larval feeding potential 

First instar grub consumed an average of 10.67±0.882 A. 

craccivora (ranged 8-12 aphids) and 6.33±0.33 L. erysimi 

(ranged 5-8 aphids), respectively (Table 2). The second instar 

grub consumed an average 24.33±3.67 A. craccivora (ranged 

20-28 aphids) and 20.00±3.66 L. erysimi (ranged 16-25 aphids). 

The third instar grub consumed an average of 24.67±0.88 A. 

craccivora (ranged 21-26 aphids) and 21.67±0.88 L. erysimi 

(ranged 18-24 aphids), respectively. The fourth and final instar 

grub consumed an average of 26.33±8.19 A. craccivora (ranged 

21-29 aphids) and 26.00±0.58 L. erysimi (ranged 24-28 aphids), 

respectively. The total number of aphid consumed during 

whole larval duration varied from 70-95 A. craccivora and 63- 

85 L. erysimi, respectively. Chowdhury, et al., 2008 recorded 

that the first, second, third and fourth instar grubs of C. 

sexmaculata were consumed 21.70±5.81, 27.10±7.42, 

72.60±5.32 and 186.30±24.59 aphid, respectively, during their 

developmental period. However, Pandey and Khan, 2002 

reported that generally grubs consumed more aphids than 

adults. 

Adult 

The observed data indicates that, a male beetle 

Table 2. 

Feeding potential of predatory coccinellid beetle, 

C. sexmaculata 

Insect stages Consumed A. craccivora 

(No.) 

Consumed L. erysimi 

(No.) 

1 st instar 10.67±0.882 6.33±0.33 

Grub 

2 nd instar 24.33±3.67 20.00±3.66 

3 rd instar 24.67±0.88 21.67±0.88 

4 th instar 26.33±8.19 26.00±0.58 

Male 510.33±9.82 410.33±4.26 

Female 718.00±17.21 555.67±27.63 

Total consumed 1314.33±23.38 1040.00±30.24 

CD > 0.05% 

SE (m) 

36.999 

12.081 

*mean of 10 pairs of adult male and females 

47.869 

15.630 

consumed 510.33±9.82 A. craccivora (ranged 500-521 aphids) 

and 410.33±4.26 L. erysimi (ranged 400-416 aphids), with an 

average of 28.72 A. craccivora and 32.92 L. erysimi per day, 

respectively (Table 2). While, a female beetle consumed 

718.00±17.21 A. craccivora (ranged 710-725 aphids) and 

555.67±27.63 L. erysimi (ranged 547-571 aphids), respectively 

with an average of 35.93 A. craccivora and 27.90 L. erysimi 

per day, respectively. Chowdhury, et al., 2008 recorded that 

adult male and female Cheilomenes sexmaculata consumed 

825.90±64.11 and 1177.80±66.57 aphid, respectively, during 

their life span. 

The total consumption of A. craccivora and L. erysimi 

by the larva and adult of C. sexmaculata was 1314.33±23.38 

(1252-1355 aphids) and 1040.00±30.24 (ranged 995-1115 

aphids), respectively. Chowdhury, et al., 2008 recorded that 

the adult male and female Cheilomenes sexmaculata 

consumed 825.90±64.11 and 1177.80±66.57 aphid, respectively, 

during their life span 


Authors are very grateful to ICAR funded project “Niche 

Area of Excellence Programme” for their financial support 

during experimental work. 


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in regulating the aphid (Myzus persicae) Sulzer Population on 

Common in field. Indian J. Entomol., 51: 24-8. 

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requirement and predation efficiency of Menochilus sexmaculata 

Fab. Coleoptera Coccinellidae. Proc.2 nd Intl. Conf. Plant protection 

in Tropics, March 17-20, Genting High land Malaysia. pp.380-9. 

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craccivora Koch. By ladybird beetle, Menochilus sexmaculata F. in 

Chillies. Adv. Plant Sci., 3: 281-6. 

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patterns of predators on chilli plant. Proc. 4 th Intl. Conf. on Plant 

Protection in the Tropics, March 28-31, Kualalumpur. pp.96. 

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septempunctata L. (Coccinellidae: Coleoptera) on mustard aphid, 

Myzus persicae Sulzer. Pestology, 26(1): 23-26. 

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Maslation on fecundity, Longivity and Geotropism of Menochilus 

sexmaculata. Environ. Entomol., 5: 575-9. 

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biology of the coccinellid predator, Cheilomenes sexmaculata (Fab) 

on Aphis craccivora. Annals of Plant Protection Science, 11: 7-10. 

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for the management of aphid. Ph. D. thesis, Rajasthan Agricultural 

University, Udaipur, India. 

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Species of Predatory Coccinellid in Anand Region of Gujarat. Karnataka 

Journal of Agricultural Sciences, 20(3): 637-638. 



Survey, Vector Relationships and Host Range Studies of Tomato Leaf Curl Karnataka 

Virus Causing Sunflower Leaf Curl Disease 

VANITHA, L.S. 1 , RANGASWAMY, K.T. 1 , GOVINDAPPA, M.R. 2 , MANJUNATHA, L. 1 , SHIVAKUMAR S. 

CHINCHURE 2 AND GOVARDHANA, M. 1 

1 

Department of Plant Pathology, University of Agricultural Sciences, GKVK, Bengaluru 560 065, Karnataka, 

India 

2 

Main Research Station, University of Agricultural Sciences, Raichur 584 102, Karnataka, India 

1 

e-mail: vanithalakshmanaih@gmail.com 

ABSTACT 

Sunflower leaf curl disease (SuLCD) is a recently reported 

disease in India. The survey revealed the occurrence of the 

disease in the range between 8.00-58.20 per cent. Twenty 

viruliferous whiteflies were required for 100% transmission 

of SuLCV from Sunflower to Sunflower. Among twelve plant 

species tested for infectivity by sunflower leaf curl virus, 

Nicotiana benthamiana, Lycopersicon esculentum, Zinnia elegans, 

Acanthospermum hispidum and Parthenium hysterophorus took 

infection and developed symptoms. 

Key words 

Sunflower, leaf cu rl, whitefly, b egomovirus, 

viruliferous 

Sunflower (Helianthus annuus L.) is one of the three 

important edible oilseed crops grown in the world, after 

soybean and groundnut. The major sunflower producing 

states are Karnataka, Andhra Pradesh and Maharashtra. The 

crop has been found suffering from few viral diseases of which 

sunflower leaf curl disease (SuLCD) caused by a begomovirus 

is the recent one. Leaf curl disease on sunflower was first 

reported on sunflower hybrids in Main Research Station, 

Raichur District, Karnataka, India with disease incidence up 

to 40% during 2009 (Govindappa, et al., 2011). 

Viruses of genus Begomovirus, family Geminiviridae 

are circular single-stranded DNA plant pathogens which cause 

damage to many crop plants in many continents. Among these 

viruses, whitefly [Bemisia tabaci (Gennadius)]-transmitted 

begomoviruses are considered to be one of the largest and 

most important groups of plant viruses infecting a wide range 

of crops, particularly in tropical and subtropical regions. They 

can be monopartite or bipartite depending upon the presence 

of one (DNA-A) or two (DNA-A and DNA-B) genomic 

components, each of approximately 2.5–2.8 kb size (Stanley, 

1983). 


Survey for Sunflower Leaf Curl Disease 

Surveys were conducted from 2011 to 2012 to determine 

the incidence of SuLCD in major sunflower growing areas of 

Raichur, Koppal, Tumkur, Chitradurga, Kolar, Chikkaballapur 

and Bengaluru in Karnataka. Disease incidence was recorded 

based on the visual observation of the affected plants. 

Collection and maintenance of virus and whitefly vector 

Sunflower leaves with characteristic leaf curl symptoms 

on hybrid (Sun breed 275) were collected from the field of 

Main Research Station, Raichur. A virus free colony of B. 

tabaci was established on cotton plants. Whiteflies were 

allowed 24-h acquisition access period (AAP) on source plants 

and transferred to susceptible sunflower hybrid (Sun breed- 

275) for 24 h inoculation access period (IAP). Inoculated plants 

were maintained in the greenhouse and observed for symptom 

development. Plants that developed leaf curl and other 

begomovirus specific symptoms were transferred to a separate 

room and used as source plants for studies reported here. 

Virus-vector transmission relationships 

The SuLCV isolate collected from naturally infected 

plants at MRS, Raichur genotype KBSH 41 was maintained 

on sunflower. The whitefly (Bemisia tabaci) colonies was 

maintained on cotton and cucumber plants were used for virus 

transmission studies to determine the number of Bemisia 

tabaci required for the successful transmission of SuLCV, 

non-viruliferous B. tabaci were given an AAP of 24 h on 

SuLCV-infected susceptible sunflower hybrid. Viruliferous 

whiteflies were then transferred to 8-10 days old young healthy 

sunflower seedlings at the rate of 1, 3, 5, 10, 15 and 20 insects 

per seedling separately and 10 plants were inoculated in each 

treatment. After an IAP of 24-h, whiteflies were killed by 

spraying 0.03 % Imidachloprid. The plants were kept in insectproof 

cages in glasshouse for symptom expression and per 

cent transmission was recorded. 

Screening of different hosts against Sunflower leaf curl 

virus 

Eight crop plant species and four weeds were tested for 

their susceptibility to SuLCV. Healthy seedlings of different 

species of cultivated plants and weeds were raised in small 

plastic bags (5-10 cm) containing soil and compost in insect 

proof glasshouse and seedlings were inoculated with 10 

whiteflies after 24 h acquisition access feeding on SuLCV 

infected plant. The viruliferous whiteflies were fed on healthy

VANITHA et al., Survey, vector relationships and host range studies of tomato leaf curl Karnataka virus 37 

test seedlings for 24 h in insect proof inoculation tubes, after 

inoculation viruliferous whiteflies were killed by using 0.05% 

Confidor (systemic insecticide) and maintained in an insect 

proof cage for 2 months for symptom development. 


Disease incidence, virus transmission characters and host 

range studies of sunflower leaf curl disease 

Sunflower leaf curl disease incidence was more severe 

at Main Agricultural Research Station trails, Raichur with 

incidence of 58.20%. The disease incidence ranged from 0.00% 

to 58.20% and varied with hybrids and varieties. The disease 

incidence varied from mild (10%) at Murdi village to moderate 

level (25%) in Narasapur village of Kustagi Taluk and to the 

tune of 12.50% in Kanakagiri village and 16.00% in 

Somanakoppa village of Gangavathi Taluk. In Yelburga Taluk, 

Mangaluru village, the disease was recorded to extent of 

25.00% while it was up to 15.50% in Beur village. In Koppal 

Taluk, the disease incidence of 8.00% was noticed in 

Gundlanur village, Betagere village (10.50%) and 20.00% in 

Katarki village. The disease was not noticed in Tumkur, 

Chitradurga, Kolar, Chikkaballapur and Bengaluru districts of 

Karnataka, India (Table 1). 

The differences in the incidence of disease in areas 

surveyed might be due to the variation in the source of 

inoculum, vector population, climatic conditions and the area 

under the crop. In recent years there has been wide spread 

occurrence of begomoviruses on many crop plants 

(Rajeshwari, et al., 2005). The B biotype B. tabaci reported 

during 2000 appears to have spread to other parts of the state. 

To ascertain the minimum number of B. tabaci required 

for efficient transmission, different groups of whiteflies (e.g., 

1, 3, 5, 10, 15, and 20) per plant were used for virus inoculation. 

Viruliferous whiteflies were enclosed on test plants with AAP 

and IAP of 24-h each. Single whitefly was able to transmit 

SuLCV with 10.00 per cent efficiency. The transmission 

efficiency increased to 30% when three whiteflies were caged 

on healthy sunflower seedlings. Transmission efficiency was 

Fig. 1. 

Determination of number of whiteflies (B. tabaci) 

required for transmission of Sunflower leaf curl virus. 

Table 1. 

Sl. 

No. 

Incidence of Sunflower leaf curl disease in Karnataka 

District Taluk Location Hybrid 

No. of plants 

infected/number of 

plants examined 

Disease incidence 

(%) (Average) 

Symptoms 

1. Raichur 

Vt- Vein thickening, En- 

Raichur MARS, Raichur KBSH 41 71/122 58.20 Enation, UCu- Upward 

curling, St-stunting 

Lingasugur Pamanakallur KBSH 41 11/110 10.00 ” 

Gangavathi 

Kanakagiri KBSH 41 6/49 12.50 ” 

Somanakoppa KBSH 44 15/108 16.00 ” 

Kustagi 

Narasapur KBSH 41 31/124 25.00 ” 

Murdi KBSH 41 9/90 10.00 ” 

2. Koppal 

Mangaluru KBSH 44 27/108 25.00 ” 

Yellaburga 

Beur KBSH 41 13/84 15.50 ” 

Gundlanur KBSH 41 8/100 8.00 ” 

Koppal Katarki KBSH 41 12/60 20.00 ” 

Betagere KBSH 41 13/120 10.50 ” 

3. Tumkur Sira 

Thagarigunte KBSH 44 0/123 0.00 No symptoms 

Malangi KBSH 44 0/131 0.00 ” 

4. Chitradurga Hiriyur Hiriyur KBSH 44 0/86 0.00 

5. Kolar Srinivaspur Srinivaspur KBSH 44 0/120 0.00 

6. Chikkaballapur Chikkaballapur Bagepalli KBSH 44 0/98 0.00 

Vishwanathapura KBSH 44 0/114 0.00 ” 

7. Bengaluru rural Devanahalli 

Budigere KBSH 44 0/98 0.00 ” 

Chikkahalli KBSH 41 0/109 0.00 ” 

Battaramarenahalli KBSH 41 0/139 0.00 ” 

8. Bengaluru urban Bengaluru urban UAS, GKVK. KBSH 41 0/142 0.00 ” 

Note: Vt-vein thickening, UCu-upward curling, En- enation on lower surface of leaves, RL-reduction in leaf size, Ab-abscission of flower, St-stunting 

of plant


Tabl e 2. Screeni ng of hosts of Begomovirus against Sunflower leaf curl virus 

Sl. 

No. 

Name of the plant species 

Family 

No. of plants 

infected / 

No. of plants 

inoculated 

Transmission 

(%) 

Incubation 

period 

(days) 

Symptoms 

observed 

1 Tobacco (Nicotiana benthamiana) Solanaceae 6/10 60.00 22 DCu, RL, St 

2 Tomato (Lycopersicon esculentum) Solanaceae 7/10 70.00 21 DCu, Bl, RL, St 

3 Zinnia (Zinnia elegans, L.) Asteraceae 6/10 60.00 16 UCu, RL, St 

4 Acanthospermum hispidum Asteraceae 5/10 50.00 23 DCu, RL 

5 Parthenium (Parthenium hysterophorus) Asteraceae 2/10 20.00 28 DCu, RL, St 

6 Euphorbia (Euphorbia geniculata L.) Euphorbiaceae 0/10 0.00 ____ ” 

7 Cotton (Gossypium hirusutum.) Malvaceae 0/10 0.00 ____ ” 

8 Cucumber (Cucumis sativus) Cucurbitaceae 0/10 0.00 ____ ” 

9 Cassia (Cassia sp.) Fabaceae 0/10 0.00 ____ ” 

10 Bhendi (Abelmoschus esculentus) Malvaceae 0/10 0.00 ____ ” 

11 Pumpkin (Cucurbita sp.) Cucurbitaceae 0/10 0.00 ____ ” 

12 Beans (Phaseolus vulgaris) Fabaceae 0/10 0.00 ____ ” 

Note: Bl: blistering; UCu: upward curling; DCu: downward curling; Vt: Vein thickening; RL: reduction of leaf size; SI: shortening of internodes; 

St: stunting 

100% with 20 whiteflies per plant. This indicates that the 

number of insects and the transmission efficiency was 

positively correlated (Fig.1). Low transmissibility of Indian 

Cassava Mosaic Virus (ICMV) associated with woody plant 

species like cassava was reported to be only 18.7% with 50 

whiteflies per plant respectively (Mathew, 1988). 

In order to identify the natural hosts and weeds, the 

virus was inoculated to different plant species through 

whitefly (Table 2). Of the 12 plant species inoculated through 

whiteflies, successful transmission of virus was obtained only 

with Nicotiana benthamiana (Fig. 2a), Lycopersicon 

esculentum (Fig. 2b), Zinnia elegans (Fig. 2c), 

Acanthospermum hispidum (Fig. 2d), and Parthenium 

hysterophorus (Fig. 2e), The common symptoms observed 

on host plants are leaf curling, reduction in the leaf size and 

stunting of plants. Euphorbia geniculata, Gossypium 

barbadense, Cassia sp., Cucumis sativus, Abelmoschus 

esculentus, Cucurbita moschata and Phaseolus vulgaris did 

not show any visible symptoms. 

Host range studies revealed that the virus was limited 

to the only few plant species belonging to Solanaceae and 

Asteraceae. Out of 12 plant species inoculated, two plant 

species belonging to Solanaceae viz., N. tabacum, L. 

esculentum were infected with SuLCV. Three plant species, 

Zinnia elegans, Acanthospermum hispidum and Parthenium 

hysterophorus belonging to Asteraceae were infected with 

SuLCV through whiteflies and expressed the systemic 

symptoms within 30 days. Begomovirus specific primers 

(Deng, et al., 1994) were used to confirm infection by SuLCV 

in Nicotiana benthamiana, Lycopersicon esculentum, Zinnia 

elegans, Acanthospermum hispidum and Parthenium 

hysterophorus by amplifying ~530 bp of core region of CP 

gene. These primers used elsewhere, successfully detected 

Croton leaf curl virus (CrLCuV) (Mahesh, et al., 2010), Cotton 

leaf curl Multan virus-Hibiscus [Bangalore] 

(CLCuMVHib[Ban]) (Rajeshwari, et al., 2005), ( Zinnia leaf 

curl virus (ZLCV) (Shivakumar, 2010) and Jatropha Mosaic 

Virus (Rangaswamy, et al., 2005; Aswathanarayana, et al., 

2007). 

Fig. 2. (a-e). Screening of hosts of Begomovirus against Sunflower leaf 

curl virus

VANITHA et al., Survey, vector relationships and host range studies of tomato leaf curl Karnataka virus 39 


Aswathanarayana, D.S., Rangaswamy, K.T., Shankarappa, K.S., Maruthi, 

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Murthy, K.V. 2007. Distinct begomoviruses closely related to cassava 

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Losses Caused Due to Alternaria Blight Disease in the Grain Yield of Pigeonpea 

LAXMAN PRASAD BALAI 1 AND R.B. SINGH 

Department of Mycology and Plant Pathology, Institute of Agricultural Sciences, Banaras Hindu University 

(BHU), Varanasi 221 005, India 

e-mail: laxmanbhu08@gmail.com 1 

ABSTRACT 

A field trial was conducted during 2010-11 and 2011-12 to 

ascertain the losses caused by alternaria blight disease in the 

grain yield of pigeonpea. A susceptible variety ‘Bahar’ was sown 

in the research field. The artificial inoculations of mass culture 

of Alternaria tenuissima (Kunze ex pers.) Wiltshire was done in 

the inoculated plots at 60 DAS to create the desirable level of 

disease pressure of alternaria blight. The uninoculated plots 

(control) were sprayed with Mancozeb 75 WP (0.25%) to check 

the alternaria blight disease. It revealed that inoculated plots 

showed maximum disease intensity (44.19%) was recorded in 

plots which did not receive any spray of Mancozeb. In plots 

with one, two three and four sprays of Mancozeb, disease 

intensity was recorded as 31.01, 20.57, 13.75 and 11.88% and 

the mean % disease control was 29.88, 53.45, 68.88 and 73.09% 

respectively. Almost similar trend of disease intensity, crop 

yield and number of 100 grain seed weight was recorded in 

different treatments with Mancozeb. 

Key word 

Pigeonpea, alternaria blight, losses and disease 

intensity 

Pigeonpea [Cajanus cajan (L.) Millsp.] is one of the 

important food legume crops and rank second after chickpea 

in area, production and productivity of the tropics and subtropics. 

The crop suffers from many fungal diseases, of which 

foliar diseases take a heavy toll. Assessment of the effect of 

any disease on yield of a crop is pre-requisite for preparing 

rational disease management programme. Among the diseases, 

alternaria blight disease has been reported to cause yield 

losses between 20-80% (Sharma, et al., 2012, Kushwaha, et 

al., 2010). Alternaria blight (Alternaria tenuissima (Kunze ex 

pers.) Wiltshire) was reported for the first time from Varanasi, 

India by Pavgi and Singh, 1971. Later, Kannainyan and Nene, 

1977 reported its occurrences from Hyderabad as a disease of 

minor importance but recently alternaria blight has become 

one of the serious fungal diseases of pigeonpea, especially in 

September sown crop. This disease has thus become the most 

limiting factor for rabi cultivation of pigeonpea in eastern 

Uttar Pradesh, Bihar and Bengal. The disease symptoms start 

appearing as circular, chlorotic, dark brown minute spots with 

yellow halo on the upper surface of leaf lets, followed by 

development of black spots which increase in size showing 

purple margin around the black necrotic spots. The spots 

enlarge and coalesce with each other forming big lesions. 

These symptoms appeared in the second or third weeks of 

November and attain maximum disease severity at the second 

or third weeks of January onward. Reliable estimates of loss 

facilitate the objective identification of the relative importance 

of biotic pests. Consequently, limited resources can be 

assigned on a priority basis and optimize returns from a given 

effort. Accurate information concerning losses is also needed 

by growers and plant protection specialists to develop 

decision thresholds for determining, when cost effective 

control measures should be deployed (Nutter, 1993). The need 

for reliable crop loss assessment methodology (to develop 

reliable decision aids) assumes added importance given, the 

current worldwide concern about improving or maintaining 

environmental quality by reducing the use of pesticides (Stern, 

et al., 1959). Keeping in view the above facts, the present 

investigation was undertaken the estimation of yield losses 

in pigeonpea production under agro-climatic conditions. 


An experiment was conducted during the years 2010-11 

and 2011-12 at the Entomological Research Trials Field, 

Banaras Hindu University (BHU),Varanasi in the crop season 

under artificial inoculation condition to find out the extent of 

grain yield loss due to Alternaria blight. Susceptible cultivar 

(Bahar) was used in randomized block design with three 

replications. The plot size was 4.50 3.0 m with row spacing 

of 45 cm and plant to plant distance of 20 cm. Seeds were 

sown in second week of September. Weeds were controlled 

by using Glyphosate @ 3 liter/ ha pre-emergence application 

15 days before sowing. Optimum moisture in the soil for proper 

seed germination was maintained by rolling the field with a 

roller. Endosulphan one liter/ha was sprayed twice to prevent 

crop damage by different insects. After attaining the age of 

two months, the plants were artificial inoculated with spore 

cum-mycelial suspension of the pathogen and the plots were 

irrigated from time to time to maintain proper moisture. One, 

two, three and four sprays of Mancozeb 75 WP (0.25%) were 

applied separately in each plot. The first spray of fungicide in 

recommended dose was given 48 hrs after inoculation and 

subsequent sprays were done at 10 days interval. Check plot 

was sprayed with sterilized water only. Observations on 

disease intensity were recorded on the basis of percent of leaf 

area affected after 15 days of last spray and disease intensity 

recorded using 0-5 scale (Mayee and Datar, 1986) following 

Table 1. 

Data on the per cent infection of the disease were also 

converted into angular values and analyzed statistically. Grain 

yield (kg/ha), 100 seed grain weight (g) per plot and per cent 

disease control was recorded after harvest of the crop. Finally

Table 1. 

BALAI AND SINGH, Losses Caused due to Alternaria Blight Disease in the Grain Yield of Pigeonpea 41 

Disease intensity recorded using 0-5 scale 

Scale Description Degree of resistant 

0 No Disease High resistant 

1 0.1-5.0% Resistant 

2 5.1-10% Moderent resistant 

3 10.1-25.0% Moderent susceptible 

4 25.1-50.0% Susceptible 

5 50 % and above High susceptible 

Table 2. 

grain yield loss was calculated using following formula. 

Yieldin treatment Yieldin check 

Per cent yield loss = 100 

Yield in treatment 

a) Per cent disease intensity (PDI): Per cent disease 

intensity was calculated by using following formula 

(Mayee and Datar, 1986). 

Sum of individual rating scale 

Per cent disease intensity = —––––––——————— × 100 

No. of disease plant observation × Maxi. Dis. Rating 

b) Yield: Crop was harvested at maturing stage and yield 

of net plot was recorded (kg/ha) and expressed in q/ha. 

c) Per cent reduction over control 

Disease severity in control – Disease severity in treatment 

Disease reduction (%) = ––––––––––––––––––––––– × 100 

Disease severity in control 

Yield in treatment plot - Yield in control plot 

Yield increase (%) = –––––––––––––––––––––––––– × 100 

Table 3. 

Spray schedule. 

Sl. No. Treatment Spray schedule 

1 T 1 One spray at the onset of the disease 

2 T 2 Two sprays, at T 1 + 10 days after T 1 

3 T 3 Three sprays, at T 1 + T 2 +10 days after T 2 

4 T 4 Four sprays, at T 1+T 2+T 3+10 days after T 3 

5 T 5 No spray (control) 

Yield in control plot 

Estimation of yield losses in Alternaria blight affected pigeonpea 


The data presented in the Table 3 clearly shows that 

increased in number of sprays of Mancozeb (1-4) significantly 

decreased the disease intensity of alternaria blight. Disease 

intensity recorded in 2010-11 crops season was 30.01, 20.02, 

14.01 and 12.44% at 1, 2, 3 and 4 spray of Mancozeb, 

respectively. Whereas, in crop season 2011-12 disease 

intensity was recorded 32.01, 21.12, 13.50 and 11.32%. It 

revealed that inoculated plots had maximum disease intensity 

43.88 and 44.50% were recorded in 2010-11 and 2011-12, 

respectively in control plots which did not receive any spray 

of Mancozeb. Foliar sprays of Mancozeb significantly reduced 

the disease intensity over control the maximum reduction 

being plots with 4 sprays. Number of spraying significantly 

increased the grain yield (gram/plot). 

Minimum yield (1503.60 and 1496.40g/plot) was recorded 

in control plots which did not receive any spray of Mancozeb. 

One, two, three and four sprays of Mancozeb yield recorded 

(1677.00, 1776.60, 1869.00 and 19.50g/plots) 2010-11, whereas 

crop season 2011-12 (1667.90, 1776.60, 1855.00 and 1945.90g/ 

plots) were recorded and disease control of 31.51, 54.31, 68.03 

and 71.61% respectively, during the year 2010-11.Whereas, 

disease control recorded in 2011-12 was 28.07, 52.54, 69.66 

and 74.56%. Over all maximum and minimum disease intensity 

(44.19 and 11.88%), grain yield (1948.08 and 1500.00g/plot) 

and disease control (73.09 and 29.79%) were recorded during 

this crop season. 

Four sprays of Mancozeb consistently gave maximum 

disease control in both the year crop seasons. Hence, it could 

be concluded that crop protection with fungicide is essential 

in pigeonpea to minimize the reduction in the yields caused 

due to alternaria blight disease. This corroborates the findings 

of Carson, 1985 that alternaria blight caused yield losses up 

to 51 to 60%, when seed yields were compared to those of 

check plots sprayed with fungicide. The 100-seed weight was 

also significantly reduced in treatments. Yield losses and 

Treatment 

Disease intensity Grain yield/plot (g) Test wt (100 seed) Per cent disease control 

2010-11 2011-12 Mean 2010-11 2011-12 Mean 2010-11 2011-12 Mean 2010-11 2011-12 Mean 

T 1 43.88 44.50 44.19 1503.60 1496.40 1500.00 8.92 8.88 8.9 0 0 0 

(41.50)* (41.86) (41.68) 

T 2 30.01 32.01 31.01 1677.00 1667.90 1672.45 9.30 9.26 9.28 31.51 28.07 29.79 

(33.21) (34.44) (33.83) 

T 3 20.02 21.12 20.57 1776.60 1776.60 1776.60 9.80 9.76 9.78 54.31 52.54 53.43 

(26.71) (27.36) (27.04) 

T 4 14.01 13.50 13.75 1869.00 1855.00 1862.00 9.96 9.98 9.97 68.03 69.66 68.85 

(21.97) (21.57) (21.77) 

T 5 12.44 11.32 11.88 1950.25 1945.90 1948.08 10.50 10.60 10.55 71.61 74.56 73.09 

(20.64) (19.63) (20.14) 

Sem± 0.50 0.34 16.22 15.34 0.10 0.11 - - - 

C.D. (0.05) 1.46 1.00 47.47 44.87 0.31 0.32 - - - 

*Values in parentheses are angular transformed values, where as, T 1 

= Control, T 2 

= One spray Indofil M-45 (mancozeb), T 3 

= Two sprays Indofil 

M-45 (mancozeb), T 4 

= Three sprays Indofil M-45 (mancozeb), T 5 

= Four sprays Indofil M-45 (mancozeb)


losses in 100-seed weight were consistently greater, indicating 

that alternaria could be a greater problem in seed production 

fields. Shivankar, et al., 2000 reported that inoculated plots 

showed highest intensity of leaf blight disease within the 

range of 58.42 to 64.91% and gave significant reduction in the 

yields in the range of 19.95 to 39.32% along with also reduction 

in 100 grain weight i.e., 6.80 to 17.08% as compared to uninoculated 

plots. Assessment of the effect of any disease on 

yield of a crop is pre-requisite for preparing rational disease 

management programme. Among the diseases, alternaria blight 

disease caused the yield losses up 20-80% (Sharma, et al., 

2012). The crop suffers greatly from alternaria blight and the 

pathogen may cause 40-50% reduction in yield, in most 

pigeonpea growing states of India (Kushwaha, et al., 2010). 


Authors are grateful to Head, Department of Mycology 

and Plant Pathology, Institute of Agricultural Sciences, 

Banaras Hindu University (BHU), Varanasi for providing 

necessary facilities during the course of investigation. 


Carson, M. L. 1985. Epidemiology and yield losses associated with 

alternaria blight of sunflower. Phytopathology. 75:1151-1156. 

Kannaiyan, J. and Nene, Y. L. 1977. Alternaria leaf spot of pigeonpea. 

Tropical Grain Legume Bulletin. 9: 34. 

Kushwaha, A., Srivastava, A., Nigam, R. and Srivastava, N. 2010. 

Management of alternaria blight of pigeonpea crop through 

chemicals. International Journal of Plant Protection. 3:313-315. 

Mayee, C.D. and Datar, V.V. 1986. Phytopathometry. Department of 

Plant Pathology, Marathwada agricultural University, Parbhani, 

Technical bulletin No. 3, pp. 110. 

Nutter, F.W. 1993. Terms and concepts for yield, crop loss and disease 

thresholds. Plant Disease, 77: 211-215. 

Pavgi, M.S. and Singh, R.A. 1971. Parasitic fungi from north India, 

VIII. Mycopath. et Myco. Appl. 43:117-125. 

Sharma, M., Ghosh, R., Mangla, N., Saxena, K. B. and Pande, S. 2012. 

Alternaria tenuissima causing alternaria blight on Pigeonpea 

[Cajanus cajan (L.) Millsp.] in India. Plant Disease, 96: 907.2- 

907.296. 

Shivankar, S.K., Shivankar, R.S. and Nagone, A.H. 2000. Losses caused 

due to leaf blight disease in the grain yield of irrigated Wheat. 

Agricultural Science Digest, 20: 203-204. 

Stern, V.M., Smith, R.F., Van den bosch, R. and Hagan, K. S. 1959. The 

integrated control concepts. Hilgardia, 9: 81-101. 



Relative Performance of Resource Conservation Technologies in Maize Based 

Cropping System Under Temperate Kashmir 

FAYAZ AHMED BAHAR 

Department of Agronomy, FOA & RRS, Wadura, Sher-e-Kashmir University of Agricultural Sciences and 

Technology Kashmir 193 201 

e-mail: bharfayaz@redifmail.com 

ABSTRACT 

A field experiment was conducted during kharif, 2008-09 and 

2009-10 at the Experimental Farm of the Regional Research 

Station and Faculty of Agriculture, Sher-e-Kashmir University 

of Agricultural Sciences and Technology of Kashmir to study 

the relative performance of resource conservation technologies 

in maize based cropping system. Four different tillage methods 

viz; conventional tillage, minimum tillage, zero tillage and 

bed planting were used in the main plots alongwith two sub 

plot treatments viz; no mulch and straw mulch. Based on the 

findings of experimentations, higher yield attributes and grain 

yield of maize were obtained in conventional tillage which was 

at par to bed planting. Similar trend was observed in the 

succeeding crops of maize based cropping system during both 

the years. The yield under conventional tillage was about 19.6% 

higher than zero tillage. Zero tillage and minimum tillage 

were found at par. Mulching with paddy straw significantly 

increased the grain yield of maize by about 14.4%. Significantly 

lowest total weed density and dry matter accumulation were 

recorded in conventional tillage than minimum and zero tillage. 

Conventional tillage recorded the lowest depletion of nutrients 

in weeds and minimum B:C ratio among tillage practices. Straw 

mulch recorded the significantly highest net returns and B:C 

ratio than the no mulch treatment. 

Key words 

Cropping system, maize, tillage, conservation 

technologies 

In Jammu and Kashmir State, the total area under maize 

crop is 0.33 m ha with a production of 0.525 m t (Sharma, 2004) 

and is cultivated mostly by the tribal people on rainfed areas 

and karewas. The production of this crop is low in Jammu 

and Kashmir State as compared to the rice and wheat. Maize 

crop has a good potential under Kashmir conditions but is 

facing some of the major constraints. Inadequate land 

preparation coupled with over exploitation of resources often 

leads to soil deterioration and thus poor crop stand and 

productivity. Therefore, it becomes imperative to sustain the 

productivity of maize based cropping system through 

alternative and efficient technologies which are less labour 

intensive, makes best utilization of the residual moisture and 

enable the farmers to produce more with less cost of 

cultivation. This can be accomplished by making the 

comparative studies on different tillage methods. Beneficial 

effects of tillage and mulching on crop productivity are 

reported (Sharma, et al; 2009) but the information on these 

aspects under rainfed valley conditions is meager. 

Management of weeds through integration of tillage methods 

and mulching can increase the uptake of nutrients and 

productivity of the crop by decreasing the biomass and 

nutrient removal by the weeds. Unchecked weed growth in 

this crop may result in grain yield loss to the extent of 100% 

(Sharma, 2005). Keeping in view the above facts the present 

investigation was undertaken to study the relative performance 

of resource conservation technologies in maize based cropping 

system. 


A field experiment was conducted during 2008-09 and 

2009-10 at Experimental Farm of the Regional Research Station 

and Faculty of Agriculture, Sher-e-Kashmir University of 

Agricultural Sciences and Technology of Kashmir to study 

the relative performance of resource conservation 

technologies in maize based cropping system. The 

experimental soil was well-drained, silty clay-loam in texture 

and had pH of 7.4, organic carbon 0.75%, available N 292 kg/ 

ha, available P 22.4 kg/ha and available K 107.7 kg/ha. The 

experiment consisted of four different tillage methods viz; 

conventional tillage (CT), minimum tillage (MT), zero tillage 

(ZT) and bed planting (BP) in the main plots and two sub plot 

treatments viz; no mulch (NM) and straw mulch (SM) laid out 

in split plot design (SPD) with three replications. Spacing of 

60cm x 20cm was maintained in maize by thinning and gap 

filling and the subsequent crops were grown as per the 

recommended package of practices. Weed count (No./m 2 ), 

weed dry matter accumulation (g/m 2 ) and nutrient uptake of 

weeds and crop were recorded at 60 days stage. Data on weed 

count and weed dry matter accumulation was subjected to 

square root transformation. The nutrient uptake by weeds 

and crop was calculated by multiplying the content of each 

nutrient with dry matter of weeds and crop respectively. 


Grain yield 

Significantly taller plants were observed in conventional 

tillage and bed planting than minimum tillage and zero tillage 

methods among tillage practices whereas straw mulch recorded 

significantly taller plants than no-mulch treatment. 

Conventional tillage and bed planting, being statistically at 

par with each other produced significantly higher yield 

contributing characters (viz; cob length, cob girth, number of


cobs/plant, grains/cob and 1000 grain weight) and grain yield 

of maize than minimum tillage and zero tillage. Grain yield with 

minimum tillage was at par with zero tillage. Conventional 

tillage recorded the highest maize grain yield of 34.01q/ha 

whereas it was lowest with zero tillage (28.42 q/ha) thereby 

indicating an increased grain yield of 19.6% over zero tillage. 

Similar trend was observed in the succeeding crops of maize 

based cropping system (Table 1). This was due to the improved 

physico-chemical condition of the soil which favourably 

increased the uptake of nutrients by the crop and also reduced 

the state of crop weed competition by lowering the total weed 

population and weed dry matter accumulation, leading to 

increase in values of yield contributing characters. Similar 

findings have been reported by Chopra and Angiras, 2008. 

Zero tillage and minimum tillage were at par with respect to 

the average grain yield of crops in maize based cropping 

system. Mulching with paddy straw significantly increased 

the grain yield of maize by about 14.4%. The residual effect of 

the kharif tillages and mulching was nonsignificant on the 

grain yield of succeeding rabi crops. However the highest 

grain yield of the succeeding crops was recorded with 

conventional tillage which was statistically at par with bed 

planting followed by minimum tillage and zero tillage 

respectively. These findings are in lines with the findings of 

Sharma, et al., 2009. Mulching application significantly 

increased the yield of crops. This was due to decrease in the 

evaporation and improved physico-chemical properties of soil, 

availability of adequate soil moisture for longer period and 

effective weed control than un-mulched treatment besides 

shifting of nutrients towards the crop. Uncontrolled water 

supply through the conservation of moisture and regulation 

of soil temperature which in turn increased maize yield and 

yield of crops in maize based cropping system (Ondal, et al., 

2008). Inadequate moisture supply under no mulching resulted 

in low grain yield which was due to deleterious effect on most 

of the physiological process of the crop. 

Weed growth 

The predominant weeds which have been observed in 

the experimental field were grouped in to (i) broad leaved 

weeds viz; Amaranthus spp. (32.08%), Chenopodium album 

(20.37%), Convolvulus arvensis (11.84%) (ii) narrow leaved 

weeds viz; Sorghum halepense (10.54%), Poa spp. (5.6%), 

Cynodon dactylon (6.4%), Lolium spp.(5.5%) and (iii) Cyperus 

Table 1. 

Table 2. 

Effect of tillage and mulching on grain yield of maize, wheat, lentil and brown sarson (mean yield of two years). 

Treatment 

Maize yield (qha -1 ) Wheat yield (qha -1 ) Lentil yield (qha -1 ) Brown sarson (qha -1 ) 

2008 2009 Mean 08-09 09-010 Mean 08-09 09-010 Mean 08-09 09-010 Mean 

Tillage 

Conventional tillage 33.50 33.6 34.01 30.00 29.25 29.62 6.90 6.73 6.81 12.25 11.88 12.06 

Minimum tillage 29.80 29.95 29.87 26.35 26.69 26.52 6.05 5.90 5.97 9.80 9.51 9.65 

Zero tillage 28.35 28.50 28.42 24.75 24.13 24.44 5.69 5.55 5.62 9.61 9.33 9.47 

Bed planting 32.85 32.95 32.9 32.75 31.93 32.34 7.40 7.22 7.31 13.00 12.60 12.80 

C. D.(P=0.05) 2.69 2.71 2.7 2.39 2.33 02.36 0.56 0.55 0.55 0.95 0.92 0.93 

Mulching 

No mulch 29.08 29.15 29.11 27.78 27.09 27.43 5.92 5.77 5.84 10.70 10.38 10.54 

Straw mulch 33.18 33.35 33.26 29.15 28.42 28.78 7.10 6.93 7.01 11.63 11.28 11.45 

C. D.(P=0.05) 3.41 3.42 3.41 NS NS NS 0.72 0.70 0.71 NS NS NS 

Effect of tillage and mulching on yield attributing characters, weed growth and nutrient uptake (kg/ha) by weeds in 

maize (mean of two years) 

Treatment Plant height Cob length Cob girth No. of cobs Grains 1000 grain TWD* TWDMA # Nutrient uptake 

(cm) (cm) (cm) plant -1 cob -1 weight (No./m 2 ) (g/m 2 ) N P K 

Tillage 

Conventional tillage 152.9 15.8 4.3 1.15 330 265.7 14.7 9.8 17.76 3.98 19.10 

**(238.0) (107.3) 

Minimum tillage 139.8 12.5 3.9 1.14 307 224.4 16.1 11.1 22.7 4.98 24.3 

(270.0) (131.3) 

Zero tillage 136.0 12.3 3.8 1.10 299 238.6 16.5 11.4 24.81 5.72 26.9 

(283.3) (139.4) 

Bed planting 150.6 15.7 4.1 1.14 322 264.3 15.3 10.2 22.1 4.83 23.99 

(241.0) (113.4) 

C. D.(P=0.05) 9.5 2.9 NS 0.06 5.6 2.4 0.5 0.3 1.92 0.40 1.32 

Mulching 

No mulch 113.4 11.2 3.4 1.10 313 218.3 16.2 10.8 40.63 11.35 45.45 

(260.0) (115.8) 

Straw mulch 141.9 14.4 3.6 1.14 327 224.4 13.1 7.8 15.09 2.97 11.98 

(171.1) (59.0) 

C. D.(P=0.05) 10.3 2.0 NS NS 4.6 2.6 0.5 0.3 1.30 0.60 2.25 

*TWD:- Total weed density; # TWDMA:- Total weed dry matter accumulation; **Values in parantheses are means of original values

Table 3. 

BAHAR, Relative Performance of Resource Conservation Technologies in Maize based Cropping System 45 

Effect of tillage and mulching on yield attributing characters, weed growth and nutrient uptake (kg/ha) by weeds in 

maize (mean of two years) 

Tillage / Mulching 

Net returns 

Benefit : Cost ratio 

2008 2009 2008 2009 

CT MT ZT BP Mean CT MT ZT BP Mean CT MT ZT BP Mean CT MT ZT BP Mean 

No mulch 5727 6667 5934 6161 6122 6356 6678 5637 5253 5981 0.64 0.94 0.84 0.80 0.81 0.70 0.92 0.8 0.7 0.78 

Straw mulch 7946 9839 9405 8491 8920 9893 10291 10528 10929 10410 0.8 1.16 1.21 1.02 1.05 1.02 1.26 1.28 1.23 1.19 

Mean 6837 8253 7670 7326 8125 8485 8083 8091 0.72 1.05 1.02 0.91 0.86 1.09 1.04 0.98 

C.D. T M TxM T M TxM T M TxM T M TxM 

(0.05) 350 280 407 548 635 0.08 0.12 0.12 0.06 0.08 0.11 

spp. (6.5%). Predominance of these weed species in maize 

crop had also been reported by Sharma, 2005. 

Significantly lowest total weed density and weed dry 

matter accumulation were recorded in the conventional tillage 

(Table 2) and bed planting. This was owing to delay in 

emergence of weeds in more number due to the presence of 

less moisture content in the top layer under these two tillage 

methods compared with zero tillage and minimum tillage 

practices. Zero tillage and minimum tillage were at par with 

respect to reduction in weed density and weed dry matter. 

These findings are in conformity with the findings of Chopra 

and Angiras, 2008. Mulching with straw recorded significant 

reduction in weed growth and dry matter accumulation than 

un-mulched treatment (Table 2). It can be attributed to vigorous 

crop growth and less crop weed competition in mulched plots 

and very low degree of crop weed competition in un-mulched 

plots thus creating smothering effect on weed growth. 

Nutrient uptake and Economics 

Nutrient uptake (kg/ha) is a product of nutrient content 

in per cent and dry matter accumulation. Conventional tillage 

reduced the N, P and K depletion by weeds followed by bed 

planting compared with zero tillage and minimum tillage (Table 

2). The weeds in the zero tillage and minimum tillage on an 

average removed 24.81, 5.72, 26.9 and 22.7, 4.98, 24.3 kg/ha N, 

P and K respectively. Straw mulch significantly lowered the 

uptake of N, P and K by weeds as compared to un-mulched 

plots. It can be attributed to the effective control of weeds by 

straw mulch and provided a competition free environment 

which led to increased growth of the crop and thereby increase 

in nutrient uptake by increasing the grain yield of maize. 

Minimum tillage fetched the significantly highest mean net 

returns of Rs. 8369 followed by zero tillage (Rs. 7877), bed 

planting (Rs. 7709) and conventional tillage (Rs. 7481) in 

decreasing order (Table 3). Similar results have been reported 

by Sharma, et al., 2009. The highest benefit: cost ratio of 1.05 

and 1.09 was observed in minimum tillage and lowest with 

conventional tillage (0.72 and 0.98) respectively. Straw mulch 

recorded the highest mean net returns (Rs. 9665) and benefit: 

cost ratio (1.05 and 1.19) whereas no-mulch provided the 

lowest net returns (Rs. 6052) and benefit: cost ratio (0.81 and 

0.78) respectively. 

Hence it can be concluded that maximum yield attributing 

characters and grain yield was recorded in conventional tillage 

alongwith reduced weed growth and nutrient uptake among 

tillage practices. Straw mulch recorded the significantly highest 

net returns and B: C ratio than the no mulch treatment. 

LITERATUREC CITED 

Chopra, P. and Angiras, N.N. 2008. Effect of tillage and weed 

management on productivity and nutrient uptake of maize (Zea 

mays). Indian Journal of Agronomy, 53(1): 66-69. 

Ondal, N.A.M., Ossain, S.M.A.H., Huiya, S.U.B. and Ahiruddin, M.J. 

2008. Productivity of rainfed mustard in relation to tillage and 

mulching. Bangladesh Journal of Agricultural Research. 33: 597- 

606. 

Sharma, P., Abrol, V., Sankar, G.R.M. and Singh, B. 2009. Influence of 

tillage practices and mulching options on productivity, economics 

and soil physical properties of maize (Zea mays L.) – wheat (Triticum 

aestivum) system. Indian Journal of Agricultural Sciences, 79(11): 

865-70. 

Sharma, A.R. 2004. Fertilizer management in maize wheat cropping 

system in the western Himalayan region. Fertilizer News, 49(2): 

13-32. 

Sharma, R. 2005. Integrated weed management in kharif maize. Intensive 

Agriculture May-June pp. 6-9. 



Influence of Deltamethrin and Achook ® on Activities of Phosphatases in the Nervous 

Tissue of Zebrafish, Danio rerio 

DILIP KUMAR SHARMA AND BADRE ALAM ANSARI 1 

Zebrafish Laboratory, Department of Zoology, D.D.U. Gorakhpur University, Gorakhpur 273 009, U.P. India 

e-mail: ba.ansari@rediffmail.com 1 

ABSTRACT 

number of medicinal properties. Azadirachtin (a 

A comparative study of synthetic pyrethroid, Deltamethrin and 

a neem based pesticide, Achook ® was performed to evaluate the 

effect on the activity of phosphatases in the nervous tissue 

(brain) of Danio rerio at different concentrations (96 h LC 5 

, 

LC 10 

and LC 20 

) and exposure periods (4, 8, 12 and 16 days). 

Exposure of fish to each concentration of each pesticide resulted 

in concentration-dependent inhibition in phosphatases 

activities in comparison with control group. The activity of acid 

phosphatase (ACP) was reduced to 44% for Deltamethrin 

whereas 48% of controls (100%) for Achook ® treated fishes 

after 16 days exposure to LC 20 

. After 12 days the alkaline 

phosphatase (ALP) activities of the LC 20 

of Deltamethrinexposed 

fish were more strongly inhibited (31%) than that of 

the fish exposed to Achook ® (37%). With the present study it is 

attributed that the natural pesticides are also deadly toxic to 

fishes and hence these pesticides should be used with great 

caution and in sustainable way to minimize the hazards in 

aquatic environment and to human beings. 

Key words 

Danio rerio, Deltamethrin, Achook ® , Phosphatases, 

Nervous tissue 

The main sources of water pollution are industrial waste, 

domestic sewage, drainage and pesticides used for food 

production. Aquatic contamination of the pesticides causes 

acute and chronic poisoning of fish and other organisms. 

Because of their rapid knock out properties, synthetic 

pyrethroids have attracted farmers and health departments to 

use them in pest control. Type-II pyrethroids including 

deltamethrin are potentially toxic to fish and least toxic to 

mammals (W.H.O., 1990). They are more effective than the 

organophosphate pesticides, replacing them in many 

agricultural, commercial and residential applications. All 

pyrethroids are potent neurotoxicant (Oros, et al., 2005) and 

interfere with nerve cell function by interacting with voltagedependent 

sodium channels as well as other ion channels, 

resulting in repetitive firing of neurons and eventually causing 

paralysis (Shafer and Meyer, 2004). Fish sensitivity to 

pyrethroids may be explained by their relatively slow 

metabolism and elimination of these compounds (David, et 

al., 2003). They are reported to alter the various biochemical 

constituents in various tissues of fish (Anita Susan, et al., 

2010; Sharma and Ansari, 2011; Sharma, et al., 2012). 

The neem (Azadirachta indica A. Juss) is a tropical 

evergreen tree, native to Indian sub-continent which is a 

natural source of insecticides and agrochemicals along with a 

tetranotriterpenoid) is one of a major component of neem, 

which have pesticidal property (N.R.C., 1992). However, the 

disturbance in the total protein level due to different neem 

extracts have been reported (Winkaler, et al., 2007). These 

pesticides may damage the vital organs (Joshi, et al., 2007) 

and found toxic to embryo, fingerlings and adult of zebrafish 

(Ansari and Sharma, 2009; Ansari and Ansari, 2012; Sharma, 

et al., 2012). 

Enzymes are biochemical molecules that control 

metabolic processes of organisms, thus a slight variation in 

enzyme activities would affect the organism. Hence, a need 

was felt to investigate the changes in the activities of acid 

phosphatase (ACP) and alkaline phosphatase (ALP) in the 

brain of zebrafish, Danio rerio after sub-lethal exposure of 

synthetic pyrethroid Deltamethrin and neem based formulation 

Achook ® . 


For the present experiment zebrafish were procured from 

our stock aquarium. Water temperature of the aquarium ranged 

between 25 ± 2 o C and the pH was maintained between 6.6 and 

8.5. In this study, commercial formulations of synthetic 

pyrethroid Deltamethrin available in 2.8% emulsifiable 

concentrate (E.C.) and neem pesticide Achook ® (0.15% E.C.) 

were used. Mature adult fishes were exposed to sublethal 

concentrations viz., 96-h LC 5 

, LC 10 

and LC 20 

of Deltamethrin 

and Achook ® for 16 days continuously as per the values 

calculated in earlier experiment (Sharma, et al., 2012). Fifty 

fishes for each concentration of the pesticides were used. In 

these aquaria water was replaced daily with fresh treatment of 

pesticides so as to maintain the constant concentration of the 

toxicant. Each experiment was accompanied by its respective 

controls. Zebrafish was used as the test species as per 

recommendation of the International Organization for 

Standardization and the Organization for Economic Cooperation 

and Development (O.E.C.D., 1992). 

After the expiry of the exposure periods (4, 8, 12 and 16 

days), required number of exposed fishes were taken out from 

experimental and control groups. Activities of ACP and ALP 

in the brain of zebrafish were estimated according to the method 

originally proposed by Andersch and Scyzpinski, 1947 later 

modified by Bergmeyer, 1967 using p-nitrophenylphosphate 

as substrate. The activities of phosphatases have been 

expressed as micro mole (µM) substrate hydrolyzed/30

SHARMA AND ANSARI, Influence of Deltamethrin and Achook® on Activities of Phosphatases in the Nervous Tissue 47 

minutes/mg protein. Analysis of variance (ANOVA) was 

employed to test the significance of the data using StatPlus ® 

version 2009 computer software purchased from Analystsoft 

Vancouver, Canada. 


The phosphatases activity in the brain tissues of control 

fish and the fish exposed to sublethal concentrations of the 

pesticides for 16 days are presented in Tables 1-4. The activity 

of ACP was reduced to 88% of controls (100%) in brain for 

Deltamethrin whereas 91% for Achook ® treated fishes at LC 5 

exposure for 4 days. After 8 days exposure to LC 10 

the ACP 

activities reduced to 60 and 71% in brain tissues for 

Deltamethrin and Achook ® treatment, respectively. Greater 

inhibition of the ACP activity in brain tissues was recorded 

with the increasing concentration of the pesticide especially 

after 16 days of exposure (Tables 1 and 2). 

After 12 days the ALP activities of the LC 20 

of 

Deltamethrin-exposed fish were more strongly inhibited (31%) 

than that of the fish exposed to Achook ® (37%) (Tables 3 and 

4). The ALP activity was reduced to 30% of controls (100%) in 

brain after 16 days exposure to LC 20 

of Deltamethrin. The 

reduction in ALP activity from the control was 35% in brain 

due to Achook ® at the same concentration and exposure period 

as that of Deltamethrin. There was a concentration as well as 

time dependent inhibition in the activities of phosphatases. 

All the data were statistically significant (P


Table 2. 

Effect of Achook on Brain ACP (µM substrate hydrolyzed/30 minutes/mg protein) in Danio rerio 

Treatment 

period 

(days) 

4 

8 

12 

16 

Control 

(0.00) 

9.05±0.41 

(100) 

9.10±0.69 

(100) 

8.82±0.39 

(100) 

9.02±0.69 

(100) 

Values represent mean ± SD (n=6); data were significant at P

Table 4. 

SHARMA AND ANSARI, Influence of Deltamethrin and Achook® on Activities of Phosphatases in the Nervous Tissue 49 

Effect of Achook on Brain ALP (µM substrate hydrolyzed/30 minutes/mg protein) in Danio rerio 

Treatment 

period 

(days) 

4 

8 

12 

16 

Control 

(0.00) 

5.29±0.44 

(100) 

5.02±0.33 

(100) 

5.15±0.60 

(100) 

4.93±0.63 

(100) 

Concentration of Achook (µg/L) 

LC 5 

LC 10 

(0.025) 

(0.17) 

4.56±0.61 

3.61±0.30 

(86) 

(68) 

4.23±0.08 

3.11±0.08 

(84) 

(62) 

3.90±0.07 

2.67±0.09 

(76) 

(52) 

2.88±0.09 

2.04±0.03 

(58) 

(41) 

Values represent mean ± SD (n=6); data were significant at P


Herbal Rumenotoric Drugs and Their Effect on Digestion and Growth Performance 

of Crossbred Calves 

R.D. GAUTAM 1 , D.P. SINGH, RAM NIWAS AND ABED M. ALBIAL 

Department of Animal Husbandry and Dairying, Institute of Agricultural Sciences, Banaras Hindu University, 

Varanasi 221 005, India 

e-mail: rajdeepak.gautam@gmail.com 1 

ABSTRACT 

The aim of present study to evaluate the effect of market drug 

(Rumizyme) and self formulated rumenotoric drug on the growth 

performance, dry matter intake (DMI), digestibility coefficient 

and rumen pH of crossbred calves. For said purpose the 

investigation was carried out in the Department of Animal 

Husbandry and Dairying, Institute of Agricultural Sciences, 

Banaras Hindu University, Varanasi, India with Twenty seven 

crossbred calves weighing about 77 to 80 kg (6 to 12 months of 

age) were selected and divided randomly into three similar 

group (3 animals in each) for three different trials (1 st Trial, 2 nd 

Trial and 3 rd Trial) and each trial was conducted for 60 days 

excluding pre experimental period. All the calves were fed on 

seasonally available feeds and fodders to meet out the nutritional 

requirement as per NRC, 1989 recommendation. Group T 2 

and 

T 3 

were supplemented with marketed drug (one bolus/day) and 

self formulated rumenotoric drug (40gm/day) respectively and 

group T 1 

(control) was fed without any drug. During each trial 

at 15, 30, 45 & 60 days of interval the body weight, heart girth 

circumferences, DMI, digestibility coefficient and rumen pH 

were analyzed. Herbs used in the rumenotoric drugs not only 

improve the appetite, digestion process but also it stimulates 

the growth parameters and improves the immune system. There 

was significant increase (P

GAUTAM et al., Herbal Rumenotoric Drugs and their Effect on Digestion and Growth Performance of Crossbred Calves 51 

standard farm ration comprising green fodder (Bajara, Maize 

and Berseem etc. depending on seasonal availability) and 

wheat busha as the dry roughage along with a balanced 

concentrate mixture and mineral to meet their nutritional 

requirements. During the entire period possible scientific care 

was exercised to maintain hygienic conditions and to avoid 

infectious diseases in the experimental animals. These animals 

were dewormed using bolus of Bandy Kind Plus before 

initiating the experiments. During each trial at 15, 30, 45 and 60 

days interval the body weight gain, heart girth circumference, 

dry matter intake (DMI), digestibility coefficient (DC) and rumen 

pH were measured. 

In digestibility experiments attempts were made to find 

out the digestibility coefficient of the food as a whole or some 

constituents of the food. The usual calculation of the digestion 

coefficient (DC) by following equation: 

DMI (kg/head/day) = DM in feed offered – DM in feed residue 

DM intake (kg) - DM in faeces (kg) 

Digestibility Coefficient of DM (Per cent) = ———— × 100 

DM intake (kg) 

Statistical Analyses: The data were statistically analyzed 

using GLM procedure of SAS 1992. Duncan’s test 1955 was 

applied in experiment whenever to test differences. The 

following model was used: 

Where: 

Y = µ + Ti + P n 

+ TP in 

+ e ins 

Y = observed trait, ì = overall mean 

Ti = effect of i th treatments (i th = T 1 

, T 2 

, T 3 

) 

e = random error 

P n 

= effect of n th periods (n th = 0, 30, 60) TP in 

= interaction 

between T i 

and P n 


During each trial heart girth circumference was measured 

in cm, while body weight and dry matter intake (DMI) in kg 

and digestibility coefficient (DC) in per cent and rumen pH at 

15, 30, 45 and 60 days of interval. The heart girth was found 

best in T 3 

followed by T2 and lowest in T1 (Control) groups in 

all three trials; showed a significant difference (P

Table 2. 

Group 

Control Group (T 1) 

Market herbal drug 

(T 2) 

Self Compounded 

Herbal drug (T 3) 

Physical parameters, Dry matter intake, Digestibility coefficient and Rumen pH in different groups of crossbred calves at different intervals 

Parameters 

Hearth girth 

(cm) 

Body weight 

(kg) 

DM 

(kg) 

Intake 

Digestibility 

(%) 

1 st Trial 2 nd Trial 3 rd Trial 

15 Days 30 Days 45 Days 60 Days 15 Days 30 Days 45 Days 60 Days 15 Days 30 Days 45 Days 60 Days 

104.66 defg ±1.20 109.00 cdef ±0.57 113.00 abcd ±0.57 118.00 ab ±0.57 123.33 cd ±2.40 128.00 bc ±2.52 135.67 ab ±3.84 140.67 a ±3.33 142.33 c ±2.91 144.00 bc ±3.46 145.33 bc ±3.18 147.33 abc ±3.18 

78.00 a ±1.15 82.00 a ±1.15 86.66 a ±1.20 91.00 a ±0.57 97.67 ab ±8.11 100.67 ab ±8.11 104.00 ab ±8.39 107.33 ab ±8.37 113.33 a ±2.96 115.67 a ±2.91 118.33 a ±2.40 123.33 a ±3.84 

1.63 e ±0.03 1.70 e ±0.08 1.94 c ±0.03 1.70 e ±0.04 2.70 ab ±0.12 2.78 ab ±0.12 2.88 a ±0.12 2.98 a ±0.11 3.09 bcd ±0.11 3.21 abc ±0.05 3.53 ab ±0.02 3.54 ab ±0.17 

45.02 abcd ±0.96 43.34 d ±0.78 44.04 cd ±0.30 46.66 a ±0.08 46.35 b ±0.36 46.30 b ±0.61 45.89 bc ±0.79 46.70 b ±0.49 47.45 bc ±0.93 47.23 c ±1.32 47.74 abc ±1.15 47.47 bc ±1.69 

Rumen pH 7.38 ab ±0.15 7.22 abcd ±0.05 7.43 a ±0.07 7.34 abc ±0.05 7.11 ab ±0.04 7.20 ab ±0.03 7.10 ab ±0.01 7.12 ab ±0.04 6.15 b ±0.03 6.58 b ±0.35 6.51 b ±0.20 6.35 b ±0.11 

Hearth girth 

(cm) 

Body weight 

(kg) 

DM 

(kg) 

Intake 

Digestibility 

(%) 

100.00 g ±3.46 103.66 efg ±3.38 107.66 cdefg ±3.38 112.00 bcde ±3.00 121.33 cd ±0.88 127.67 bc ±1.20 135.33 ab ±1.45 142.67 a ±1.76 142.67 c ±3.38 145.33 bc ±2.91 148.00 abc ±3.21 150.33 abc ±2.91 

76.66 a ±3.33 82.00 a ±4.00 83.33 a ±3.38 89.00 a ±3.05 94.67 ab ±0.33 99.00 ab ±0.58 104.33 ab ±0.88 109.00 ab ±1.00 109.67 a ±8.67 113.67 a ±7.84 116.67 a ±7.80 119.00 a ±7.81 

1.88 cd ±0.03 1.70 e ±0.03 1.91 cd ±0.01 2.12 b ±0.02 1.79 d ±0.37 1.93 cd ±0.36 2.08 bcd ±0.36 2.27 abcd ±0.34 3.10 bcd ±0.13 3.50 ab ±0.04 3.66 ab ±0.14 3.91 a ±0.14 

44.22 bcd ±1.17 44.69 abcd ±0.88 44.50 abcd ±0.47 44.02 cd ±0.68 46.67 b ±0.49 48.20 ab ±0.66 47.84 ab ±0.32 49.38 a ±0.26 46.68 c ±0.49 46.99 c ±0.81 46.52 c ±0.93 47.12 c ±0.53 

Rumen pH 7.14 cd ±0.01 7.24 abcd ±0.00 7.25 abcd ±0.04 7.16 bcd ±0.01 7.34 a ±0.06 7.16 ab ±0.12 6.80 bc ±0.28 6.62 cd ±0.10 7.09 a ±0.03 7.13 a ±0.08 6.58 b ±0.03 6.56 b ±0.10 

Hearth girth 

(cm) 

102.00 fg ±4.04 107.33 cdefg ±2.90 114.00 abc ±2.30 121.00 a ±2.08 115.33 d ±3.18 123.00 cd ±2.65 132.00 b ±2.31 141.00 a ±3.21 144.67 bc ±1.45 147.67 abc ±1.45 152.00 ab ±1.15 155.67 a ±0.88 


Body weight 

(kg) 

77.33 a ±8.96 83.33 a ±9.13 89.00 a ±9.07 95.66 a ±8.95 92.33 b ±2.96 98.33 abab ±3.93 104.00 ab ±3.61 111.00 a ±3.61 111.00 a ±1.00 113.67 a ±0.88 117.33 a ±0.88 121.33 a ±0.88 

DM 

(kg) 

Intake 

1.69 e ±0.04 1.76 de ±0.05 2.27 ab ±0.10 2.34 a ±0.05 2.33 abcd ±0.02 2.53 abc ±0.02 2.77 ab ±0.04 2.98 a ±0.07 2.39 d ±0.37 2.70 cd ±0.41 3.22 abc ±0.29 3.87 a ±0.35 

Digestibility 

(%) 

45.85 abc ±0.60 46.09 abc ±0.39 45.35 abcd ±0.28 46.34 ab ±0.35 44.03 c ±0.69 46.16 bc ±0.91 47.21 ab ±1.32 47.45 ab ±0.94 49.38 abc ±0.26 50.55 a ±0.91 50.38 ab ±0.52 50.40 ab ±0.04 

Rumen pH 7.11 cd ±0.02 7.15 bcd ±0.03 7.80 de ±0.02 6.87 e ±0.14 7.17 ab ±0.01 6.65 cd ±0.23 6.30 de ±0.13 6.13 e ±0.03 6.61 b ±0.08 6.30 b ±0.02 6.32 b ±0.15 6.26 b ±0.10 

Means with different superscript in a row differ significantly (p

GAUTAM et al., Herbal Rumenotoric Drugs and their Effect on Digestion and Growth Performance of Crossbred Calves 53 


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Studies on Nutritional Quality of Idli Fortified with Whey Protein Concentrates 

ALKA YADAV 1 , RANU PRASAD 1 AND RAMESH CHANDRA 2 

1 

Department of Foods and Nutrition, Ethelind School of Home Science and 2 Warner School of Food and 

Dairy Technology, Sam Higginbottom Institute of Agriculture, Technology & Sciences (Deemed to be University), 

Allahabad 211 007, India 

email: alkayadav1122@gmail.com 1 

ABSTRACT 

Most of the developing countries today depends upon various 

fermented foods that are staples in the diet. Idli is most popular 

breakfast food in south India which is very delicious and 

nutritious. The fermentatio n proces s increa ses the 

bioavailability of proteins and enhances the vitamin B content 

of the food. As it is steamed food, fat content is low and it is 

easily digestible. Idli fortified with whey protein concentrate 

(WPC) @ 5%, 10%, 15% and 20% were evaluated for nutritional 

characteristics. The protein content of WPC fortified idli were 

significantly higher then the control (p < 0.05). As a result of 

increase in the protein content of the idli sample with the 

increase in the levels of WPC. Idli fortified with WPC is high 

in protein and low in fat content and which is helpful for people 

suffering from diabetes, liver disease and protein energy 

malnutrition (PEM). 

Key words 

Whey protein concentrate, batter, fermentation, 

fortification, idli 

All over the world, production of fermented foods is 

one of the oldest food processing technologies known to 

man. In many oriental countries, consumption of fermented 

foods is common and usually produced by traditional 

methods. The traditional fermented foods contain high 

nutritive value and developed a diversity of flavours, aroma 

and textures in food substrates. Idli is a traditional cereal/ 

legume based naturally fermented steamed product with a 

soft and spongy texture, which is highly popular and widely 

consumed as a breakfast food item in India. (Steinkrans, 1995). 

Whey protein concentrate (WPC) is the collection of 

globular proteins, isolated from whey, a by - product of cheese 

manufactured from cow’s milk. It is pure, all natural, high 

quality product that contains little to no fat, lactose or 

cholesterol and is a rich source of essential amino acids. In its 

purest form, whey protein concentrate (WPC), it provides 

benefits for men and women of all ages, including infants and 

toddlers. (Patel and Kilara, 1990). 

Whey protein concentrate (WPC) is a high quality, 

complete protein, with all the essential amino acids. WPC is 

also the richest known source of naturally occurring branched 

chain amino acids (leucine, isoleucine and valine). WPC is a 

soluble and very easy to digest protein. It quickly enters the 

body to provide the important essential amino acids needed 

to nourish muscles and other body tissues. (Renner and Salam, 

1991). 

Semolina is the coarse, purified wheat middlings of 

durum wheat used in making pasta, breakfast, cereals, 

puddings and couscous. Rava idli or rave idli is a variation 

of idli, made with Rava / sooji / semolina instead of the usual 

rice and urad daal. In India semolina is used to make breakfast 

dishes such as upma, sheera, rava idli, rava dosa, rava 

uttappam etc. It is also used to make snacks such as maddur 

vada etc. Semolina is used to make sweets such as rava ladoo, 

rava kheer, semolina puddings etc. It is used in making breads 

in small quantity to get a tasty crust. 


Preparation of idli 

Whey protein concentrate (WPC) was purchased from 

Mahan Dairy in New Delhi. Other ingredients such as 

Semolina, Eno powder and salt were collected from local market 

of Allahabad. 

Whey protein concentrate (WPC) was added to semolina 

at different ratio @ 5%, 10%, 15%, 20% and holding time for 

10, 15, 20 and 25 minute for fermentation and then addition of 

water to required consistency of batter. Then addition of small 

amount of Eno powder and salt in batter. After fermentation 

the batter was steam cooked for 10 minute to make idli. 

Chemical analysis 

Protein, fat, moisture, ash, carbohydrate, energy and 

calcium contents were determined using AOAC, 1980 method. 

Statistical analysis 

The results were analysed statistically for test of 

significance by using two way classification and critical 

difference technique (CD). (Imran and Coover, 1983) 


With the increase in the level of WPC in the semolina, 

there was an increase in protein content of experimental idli 

sample (Table 1). The protein content increased from 5.88% to 

8.09% with increasing the level of WPC from 5 % to 20%. The 

protein content in the WPC fortified idli was significantly 

higher then that of control (p < 0.05). The increase in the 

protein content of WPC fortified idli might be due to the 

appreciably higher protein content of WPC. The highest 

protein percentage was observe in idli sample T 4 

H 3 

and T 4 

H 4 

(8.09) then compared to other samples.

Table 1. 

YADAV et al., Studies on Nutritional Quality of Idli Fortified with whey Protein Concentrates 55 

Average value per 100 gm of idli fortified with whey protein concentrate 

Treatments 

Nutrients 

Protein (%) Fat (%) Moisture (%) Ash (%) CHO (%) Energy (kcal) Calcium (mg) 

T 0H 0 5.24 0.48 58.17 0.36 35.74 168.24 7.62 

T 1H 1 5.88 0.53 57.33 0.54 35.72 171.18 19.59 

T 1H 2 5.87 0.53 57.41 0.54 35.66 170.86 19.55 

T 1H 3 5.88 0.53 57.30 0.54 35.75 171.29 19.60 

T 1H 4 5.83 0.52 57.64 0.53 35.47 169.93 19.45 

T 2H 1 6.55 0.58 55.98 0.71 36.17 174.66 31.20 

T 2H 2 6.51 0.58 56.28 0.70 35.93 174.97 30.99 

T 2H 3 6.39 0.57 57.08 0.69 35.27 171.78 30.42 

T 2H 4 6.39 0.57 57.05 0.69 35.30 171.89 30.44 

T 3H 1 7.25 0.64 54.44 0.88 36.79 181.97 42.59 

T 3H 2 7.19 0.64 54.81 0.87 36.49 180.46 42.24 

T 3H 3 7.09 0.63 55.40 0.86 36.01 178.13 41.69 

T 3H 4 7.15 0.64 55.02 0.87 36.32 179.63 42.04 

T 4H 1 8.06 0.71 52.10 1.06 38.06 190.92 54.63 

T 4H 2 8.08 0.72 51.96 1.06 38.18 191.48 54.79 

T 4H 3 8.09 0.72 51.93 1.06 38.20 191.62 54.83 

T 4H 4 8.09 0.72 51.93 1.06 38.20 191.61 54.82 

The highest fat percentage was observe in idli sample 

T 4 

H 2, 

T 4 

H 3 

and T 4 

H 4 

(0.72) and lowest in T 0 

H 0 

(0.48). The 

moisture content was highest in idli sample T 0 

H 0 

(58.17) and 

lowest in T 4 

H 3 

and T 4 

H 4 

(51.93). The ash content was highest 

in idli sample T 4 

H 1, 

T 4 

H 2, 

T 4 

H 3 

and T 4 

H 4 

(1.06) and lowest in 

T 0 

H 0 

(0.36). The carbohydrate content was highest in idli 

sample T 4 

H 3 

and T 4 

H 4 


H 3 

(35.27). The 

highest energy value in idli sample T 4 

H 3 

(191.62) and lowest 

in T 0 

H 0 

(168.24). The calcium content was highest in idli 

sample T 4 

H 3 


H 0 

(7.62). 

From the findings of this study, it can be concluded that 

idli fortified with whey protein concentrate (WPC) can be 

successfully used. The addition of different levels of WPC in 

semolina/sooji and different holding times for fermentation 

can be satisfactory effect on the quality of idli. As a result of 

increase in the protein content of the idli sample with the 

increase in the levels of WPC. Findings revealed that idli 

fortified with whey protein concentrate (WPC) significantly 

improves the nutritional value. It is high in protein and low in 

fat. This product will be helpful from therapeutic point of view 

for people suffering from diabetes, liver disease and protein 

energy malnutrition (PEM). 


Imran, R.L. and Coover, W.B. 1983. A modern approach to statistics. 

New York: John Willy and Sons Inc, pp. 497. 

Patel, M.T. and Kilara, A. 1990. Studies on whey protein concentrate: 

Foaming and emulsifying properties and their relationships with 

physico–chemical properties. J. of Dairy Science. 73(10): 2731- 

2740. 

Renner, E. and Abd-EL-Salam. 1991. Application of ultrafiltration in 

dairy industry. Elsevier Applied Sci., London. 

Steinkraus, K.H. 1995. Handbook of indigenous fermented foods. New 

York Marcel Dekker, Ine. Wang, H.L. and G.W. Hasseltin, 1982. 

Prescott and Dunne’s Ind. Microbiology. (ed. Read J) AVI Publishing 

Co. Inc. Weastport, 492, pp.776. 



Evaluation and Assessment of Ground Water of Shivpuri Town in Madhya Pradesh 

CHANDANA JAIN 1 AND RITU MENDIRATTA 

Department of Life Science, Boston College for Professional Studies, Gwalior, M.P. 

email: cjainjain91@gmail.com 1 

ABSTRACT 

This paper aims to the assess physico-chemical properties of 

underground drinking water of Shivpuri town of Madhya 

Pradesh in India. Underground water samples were collected 

from the various places of Shivpuri town and nearby area. The 

physico-chemical properties such as color, pH, total dissolved 

solid, salinity, electrical conductivity, acidity, alkalinity, 

chloride, iron, total hardness, calcium and magnesium 

hardness were studied and analyzed. The results obtained were 

compared with permissible limits of the drinking water set by 

Bureau of Indian Standards. Some water samples were found 

within desirable limit. Water from these sources can be used 

for drinking purpose. However some sources of water have some 

parameters more than permissible level and water from these 

sources can’t be used for drinking purpose without special 

treatment. 

Key word 

Ground wa ter, p hysico-chemical properties, 

evaluation 

In India primary sources of drinking water are surface 

water and ground water. About 2.5% of earth’s water is fresh 

and suitable for human consumption. Approximately 13% of 

this fraction is ground water, used as a important source of 

drinking The ground water has been used for drinking for a 

long time. Safe drinking water is linked closely to the well 

being of human life therefore WHO refers to “control of water 

supplies to ensure that they are pure and wholesome” as one 

of the primary objectives of environmental sanitation. Surface 

water as well as ground water is contaminated by different 

physical impurities, agricultural and industrial wastes and 

underground chemicals and minerals. It is a cause of growing 

concern that a sizable population in India does not have 

access to safe drinking water. 

Ground water is an essential and vital component of our 

life supporting system. The ground water resources are being 

utilized for drinking, irrigation and industrial purpose. However 

due to rapid growth of population, urbanization, 

industrialization and agricultural activities, ground water 

resources are under stress. Hence deterioration of ground 

water quality is taking place due to geogenic and 

anthropogenic activities. Trend in public health system in 

India is the growing gap between drinking water supply system 

and sanitation service (Jaiprakash, et al., 2000, Kumar, et al., 

2004, Prajapati, et al., 2007, Kumar and King, 2004, Dutta, et 

al., 2006, Fatima Rani and Bieta, 2007). Shivpuri district is 

located in Madhya Pradesh, India. Shivpuri is located at 25.60 0 

N and 77.65 0 E. The average rainfall of Shivpuri district is 

about 875 mm most of which is received in monsoon from late 

June to early September. In the present study the quality of 

drinking water was assessed in terms of physiochemical 

parameters in the month of December 2011. 


Ground water samples were collected in the month of 

December 2011 from 17 underground bore wells and hand 

pumps at various locations of Shivpuri town which is a famous 

tourist destination of Madhya Pradesh. The location was 

chosen as study area because recently presence of Fluoride 

is reported in some of the towns of the district. The samples 

were collected in sterilized bottles and were stored at 4 0 C till 

further investigations. The physico-chemical parameter such 

as color, pH, salinity, electrical conductivity, alkalinity, acidity, 

TDS, total hardness, Ca and Mg hardness, chloride, total iron 

were analyzed according to standard methods of APHA, 2000. 

The results obtained were compared with drinking water 

quality standards set by the Bureau of Indian Standards (BIS). 

On the basis of classification, the nature of ground water has 

been categorized as desirable, permissible and unfit for human 

consumption. 


The values of different parameters of different samples 

are given in Table 1. It was observed that colour of all the 

samples was transparent except the sample collected from 

Jhiri in Shivpuri was light brown in colour. 

pH measures the activity of hydrogen ions (H + ) in the 

water. Slightly acidic water have pH value between 6.5 to7 

while slightly alkaline water have pH between7.2-8.04. 

Therefore, the pH value indicates the alkalinity and CO 2 

Fig. 1. pH of water samples

JAIN AND MENDIRATTA, Evaluation and Assessment of Ground Water of Shivpuri Town in Madhya Pradesh 57 

concentrations of the water. Results show that pH of all the 

samples under study varied from 6.96 to 8.04. Water from Jhiri 

and Tharra is slightly acidic in nature with pH value 6.96, 

while the water form Mansapuran is slightly basic with (8.04 

pH).The pH values of all the water samples are within 

acceptable limits (pH 8.0) (Figure 1) hence all the samples are 

permissible for drinking on the basis of pH. 

Total dissolved solid (TDS) consists of inorganic and 

organic substances. Inorganic substances which include clay, 

silt, minerals, metals, etc. mainly in the form of carbonates, 

bicarbonates, chlorides, sulphates, phosphates, nitrates, 

calcium, magnesium and sodium can create taste, odor, 

hardness, corrosion and scaling problem. TDS of the different 

water samples ranged from 246 to 1769 mg/l. Only one samples 

(sample No.2) showed high percentage of total dissolved 

solids, but it is in the permissible limit. 

calcium may contribute to many health problems. Magnesium 

is often associated with calcium in all kinds of water but its 

concentration remains generally lower than the calcium. 

Magnesium content in the samples under study ranged from 

9.52mg/L to 142.86 mg/L. It was above the acceptable limit in 

sample number 6,7and 14 (Figure 3). 

Fig. 3. Mg contents in water samples 

Fig. 2. TDS in water samples 

Electrical conductivity is directly linked to the 

concentration of the ionic impurities in the water. Conductivity 

measurements are influenced by pH levels and the temperature. 

Conductance of all the samples (except sample number 5 and 

14) were ranged between 238µs/cm to 678µs/cm. Conductivity 

of sample number 5 and 14 are 1711µs/cm and 949µs/c 

respectively, it shows these water samples have high ionic 

species. (According to BIS). 

Hardness of water is caused by the presence of 

multivalent metallic cations and is largely due to calcium and 

magnesium ions. Hardness is reported in terms of CaCO 3 

. It is 

not caused by single substance but by a variety of dissolved 

polyvalent metallic ions, predominantly calcium and 

magnesium cations. The low and high value of hardness has 

advantages and disadvantage. Hardness of all the water 

samples is found in the range of 238mg/L to 485.71mg/L. Hence 

all the samples fall in the category of hard and very hard. 

Calcium is a major constituent of various types of rock. 

It is one of the most common constituent presents in natural 

water ranging from zero to several hundred milligrams per 

liter. Calcium ranged from 190.47mg/L to 438.95mg/L. It shows 

all the samples have high concentration of calcium. Excessive 

Fig. 4. Calcium contents in water samples 

Alkalinity is the content level of carbonate (CO 3 

2-) 

, 

bicarbonate (HCO 3- 

) and hydroxide (OH - ) in water. Alkalinity 

is the main control factor for the aggressiveness of the water. 

Alkalinity of the samples varies from 12 mg/L to 1240 mg/L. 

Alkalinity of sample number 5 is found to be very high than 

permissible range (30-500mg/L, according to BIS). 

Chloride is present in all natural water mostly at low 

concentrations. It is highly soluble in water. There is no known 

evidence that chlorides constitute any human health hazard. 

Chlorides are generally limited to 250mg/L to1000mg/L 

(according to BIS). Chloride ion concentration of all the 

samples varied from 28.54 to 190.28mg/L and falls in 

permissible limits. 

Iron is common constituent in ground water. It is present 

in water either as soluble ferrous iron or insoluble ferric iron. 

Taste of iron is not usually noticeable at iron concentrations 

below 0.3mg/L. Although iron has got little concern as a health


Table 1. 

Sample 

No. 

Ground Water Quality Assessment of Shivpuri. 

Colour pH TDS 

mg/L 

Electrical 

conductivity 

µs/cm 

Acidity 

mg/L 

Total 

hardness 

mg/L 

1. Sakhya Sagar Lake 2. Bilava village 3. Jasrajpur village 4. Mansapuran 5. Chhatries 6. Sirsod village 7. Ramkhedi village 8. Sighniwas 9. Jhiri village 

10. Khurd village 11. Tharra village 12. Balaji Dham 13.Old Shivpuri 14. Bus Stand 15. Govt Hospital 16. Survaya Fort 17. Tourist Village. 

Ca 

mg/L 

concentration in ground water is less than 1.0mg/L as BIS 

standard. Total iron content in the samples lie between 0.1mg/ 

L to 1mg/L, hence fall in permissible range. 


Mg mg/L 

Alkalinity 

mg/L 

Cl mg/L 

1. Transparent 7.83 394.0 678.0 780.0 333.33 228.57 104.76 470.0 28.54


Spatial and Modular Variability in Phytosterol Composition in Tribulus terrestris 

MANISH MATHUR 

18E/564, Chopasni Housing Board, Jodhpur, 342 003, Rajasthan 

e-mail: ravi_mm2099@yahoo.com, eco5320@gmail.com 

ABSTRACT 

In the present investigation spatial and modular variability in 

phytosterol concentration were quantified in an important 

aphrodisiac plant Tribulus terrestris. The study revealed that 

variations in phytosterol concentration were brought by both 

spatial and modular factors as well as by their interaction. 

Among different modules leaf possessed highest concentration. 

Path analysis established various relationships of phytosterol 

with edhaphic, plant metabolites and biomass factors. Higher 

soil organic carbon and clay content were the most crucial 

factors for harvesting phytosterols in larger quantities. Habitat 

specific study indicates that older alluvial plains and piedmonts 

supports higher phytosterol concentration compared to younger 

alluvial plain and sandy plain habitats. 

Key words 

Phytosterol, Spatial and Modular Variations, Path 

Analysis, Steroidal Sapogenin, Clay 

Phytosterols are sterols that are synthesized only in 

plants and that are structurally similar to cholesterol but with 

the inclusion of an extra hydrophobic carbon chain at the C- 

24 position. Phytosterols and their esters reduce cholesterol 

level in the blood in spite of the fact that they are poorly 

absorbed into the blood stream. Sterols are the precursors of 

steroid hormones and bile acids in humans, brassinosteroidsphytohormones 

in plants and, as the recent identifications of 

sterol mutants have shown, they are involved in important 

growth and developmental processes in living organisms. 

Phytosterols have been isolated from a large number of species 

and according to numerous publications (Gordon and Miller, 

1997; Dutta and Normen, 1998; Piironen et al., 2000) they 

probably exist in all angiosperm and gymnosperm species. 

Although, there are more than 40 different phytosterols found 

in higher plants, sitosterol, campesterol and stigmasterol often 

predominate, while other phytosterols are usually typical only 

for certain plant family or even species. Brassicasterol is for 

example typical only for Brassicaceae family, and therefore it 

could be used for identification (Benveniste, 2002). 

Phytosterol production in Digitalis purpurea has been 

reported by Evans, 1973 while 67 different medicinal plants 

have also been screened for this natural product by Shirley, et 

al., 1984. Sarin and Bansal, 2011 have reported phytosterols 

in Adhatoda vasica and Ageratum conyzoides from semi arid 

Rajasthan. Very informative review on phytosterols 

concentration in tomato, apple, potato, carrot, cabbage, canola, 

Cucumis melo, Asparagus, Discorea spp., Solanum 

melongena, Trigonella foenum-graceum have been made by 

Moreau, et al., 2002. 

With above information’s a study was conducted to 

quantify the spatial and modular variation in phytosterol 

concentration in an important medicinal plant Tribulus 

terrestris. It is an annual plant cosmopolitan in nature. 

Decoction of whole plant as well as fruits is traditionally uses 

as aphrodisiac. The objectives includes (a) to find out the 

variation in phytosterol composition in different modules of 

Tribulus tereestris at various spatial conditions and (b) to 

find out relationship between phtyosterol compositions with 

bottom up (community dynamics like, Relative Importance 

Value {RIV} of Tribulus terrestris, Shannon and Weaver Index 

{H’}, evenness and richness), top down factors like soil 

parameters (organic carbon, nitrogen, moisture, phosphorus, 

soil pH, electric conductivity, soil textures) and primary and 

secondary plant metabolites. 


Plant materials were collected from five natural sites 

during rainy (Pulse events) seasons. The coordinates, habitat 

types and other attributes have been mentioned in Table 1. 

The phytosterol concentration in the ethanolic plant extract 

was done by sperctro-photometrically through Liebermin- 

Burchard method (Bloor, 1916). In this method the standard 

was prepared with 20 mg. cholesterol and Acetic-anhydride- 

H 2 

SO 4 

was the main reagent. The mechanism of this test 

includes that acetic anhydride break down the sterol into 

chleostra-3, 5 diene and H 2 

SO 4 

converted it into Bioscholestra 

-3, 5 diene monosulphonate, this produces green colour which 

measured spectro-photometrically at 680 nm (Okpuzor, et al., 

2009). This method was chosen for its simplicity and rapid 

analysis and it generally gave an approximate level of 

phytosterol. Further, HPLC protocol available only for some 

particular phytosterol composition of vegetable oil (Okpuzor 

et al., 2009 and Mathur, 2012a). 

Statistical Analysis 

Two way Analysis of Variance (ANOVA) in strip plot 

design were carried out accordingly Gomez and Gomez, 1984, 

where sites were considered as vertical factor and different 

modules as horizontal factor. 


The mean (µ g g -1 ) concentrations of phtytosterol in 

various modules at different spatial levels are presented in 

Table 2. Among modules root (295.91 µ g g -1 ) and leaves (636.19 

µ g g -1 ) from site 3 possessed maximum phytosterol


concentration whereas same modules from site 5 showed 

minimum concentration 8.07, 119.83 µ g g -1 respectively. 

However, stem (274.23 µ g g -1 ) and fruit (195.24 µ g g -1 ) parts 

from site 2 and site 4 showed maximum phytosterol 

concentration, respectively, while both these modules 

possessed their minimum concentration at site one (Table 2). 

Analysis of variance (strip plot design) revealed that variations 

in phytosterol concentration were brought by both spatial, 

modular as well as by their interactions at 99% probability 

levels (Table 3). 

Path analysis 

Regression analysis were carried out to find out the 

factors affecting the phytosterol concentration in various 

modules and for this different edaphic, plant primary and 

secondary metabolites and biomass parameters were 

regressed with phytosterol concentration in respective 

modules. Steroidal sapogenin showed quadratic relation with 

both root (root phytosterol = 1864.10+-23.86 root steroidal 

sapogenin+0.074 root steroidal sapogenin^2; P

MATHUR, Spatial and Modular Variability in Phytosterol Composition in Tribulus terrestris 61 

a 

b 

g 

h 

c 

d 

i 

j 

e 

f 

k 

L 

m 

n 

put; P


matter production is 2 to 3 times more effective than any direct 

effect on special production. Thus regression analysis of 

phytosterol with above and below ground biomass revealed 

the important information about the production ecology of 

this important secondory plant product. 

The present study provides useful information about 

spatial and modular variation in phytosterols compositions 

as well as factors affecting it. Such studies are useful for 

formulation of phytosterols based product. Study revealed 

that leaves and higher soil organic carbon and clay content 

are the most crucial factors for harvesting phytosterols in 

larger quantities. From above study habitat specific study 

indicates that older alluvial plains and piedmonts supports 

higher phytosterol concentration compared to younger alluvial 

plain and sandy plain habitats. Products like Lipton (by 

Uniliver company), Lecithin-phyosterol preparation (by Forbes 

medi-tech company), phytosteol-emulsifier complex (by Kraft 

company) and corn fiber oil (by Monsanto company) supports 

the present study (Moreau, et al., 2002). 


Ali, M.M., Humrawali, N. and Latif, M.T. 2009. Phytosterol compost 

ion in surface sediments of kuala Selangor, Selangor, Malaysia. 

European Journal of Scientific Research. 33(1): 187-194. 

Ash, M.M. 2010. Modulation of lipid metabolism by phytosterol sterates 

and black raspberry seed soils. Ph.D. Thesis, University of Nebraska, 

Lincoln, pp.114. 

Benveniste, P. 2002. Sterol metabolism. In: The Arabidopsis, (eds, CR 

Somerville, EM Meyerowitz) American Society of Plant Biologists, 

Rockville, MD. 

Bernath, J. 2002. Production ecology of secondary plant products. In: 

Herbs, Species, and Medicinal Plants, (eds.), LE. Craker and JE. 

Simon), CBS Publishers and Distributers, New Delhi, India. Vol. 1. 

pp.185-234 

Bloor, W.R. 1916. The determination of cholesterol in small amount 

of blood. Journal of Biological Chemistry, 24: 227-231. 

Dutta, C. P. and Normen, L. 1998. Capillary column gas-liquid 78 M. 

Kemal G.l and Samija Amar chromatographic separation of 5- 

unsaturated and saturated phytosterols. Journal of Chromatography 

A, 816: 177-184. 

Evans, F. J. 1973. Production of phytosterol by mature Digitalis 

purpurea L. plant. Planta, 3: 33-40. 

Goad, L.J. and Goeodwin, T.W. 1966. The biosynthesis of sterol in 

higher plants. Biochemistry Journal, 99: 735-747. 

Gomez, K. A. and Gomez, A. A. 1984. Statistical Procedures for 

Agricultural Research. 2, John Wiley and Sons, New York, USA. 

Gordon, M. H. and Miller, L.A.D. 1997. Development of steryl ester 

analysis for the detection of admixtures of vegetable oils. Journal 

of American Oil Chemists Society, 74: 505-510. 

Grusak, M.A. and Penna, D.D. 1999. Improving the nutrient 

composition of plants to enhance human nutrients and health. 

Annual Review of Plant Physiology and Plant Molecular Biology, 

501: 133-161. 

Helfman, E. 1971. Function of sterol in plants. Lipid, 6(2): 128-133. 

Itkin, M., Rogachev, I., Alkan, N., Rosenberg, T., Malitsky, S., Masini, 

L., Meir, S., Iijima, Y., Aoki, K., de Vos, R., Prusky,D., Burdman, S., 

Beekwilder, J. and Aharoni, A. 2011. Glycoalkaloid metabolism is 

required for steroidal alkaloid glycosylation and prevention of 

phytotoxicity in tomato. Plant Cell, 23(12): 4507-4525. 

Liu, H., Chou, G.X., Wang, J.M., Ji, L.L, and Wang, Z.T. 2011. Steroidal 

saponins from the rhizome of Dioscorea bulbifera and their 

cytotoxic activity. Plant Medica, 77(8): 845-848. 

Mathur, M. 2012a. Variations in phytosterol composition in Corchorus 

depressus and their relation with bottom-up, top-down and plant 

metabolites. Journal of Natural Products, 5: 179-187. 

Mathur, M. 2012b. Phytosterol composition in seeds of Blepharis 

sindica and its relation with bottom up, top down and plant 

metabolites factors. Medicinal Plants International Journal of 

phytomedicine and Related Industries, 4(3): 126-132. 

Mbatchou, V.C. and Kosoono, I. 2012. Aphrodisiac activity of oil from 

Anacardium occidentale L. seeds and seed shells. Phytopharmacol., 

2(1): 81-91. 

Moreau, R.A., Whitaker, B.D. and Hicks, K.B. 2002. Phytosterols, 

phytostanols and their conjugates in food: structural diversity, 

quantitative analysis and health promoting uses. Progress in Lipid 

Research, 41: 75-95. 

Okpuzor, J., Okochi, V.I., Ogbungafor, H. A., Ogbonnia, S., Fagbayi, T. 

and Obidiegwu, C. 2009. Estimation of cholesterol level in different 

brands of vegetable oils. Pakistan Journal of Nutrition, 8(1): 57- 

62. 

Pironen, V., Lindsay, D. G., Miettinen, T. A., Toivo, J. and Lampi A. M. 

2000 Review Plant sterols: Biosynthesis, biologicalfunction and 

their importance to human nutrition. Journal of the Science of 

Food and Agriculture, 80: 939-966. 

Sarin, R. and Bansal, N. 2011. Phytosterls from In Vivo and In Vitro 

clusters of two medicinal plants viz. Adhatoda vasica and Ageratum 

conzyoide. International Journal of in Ayurveda and Pharmacy, 

2(3): 927-930. 

Shirley, N., Hooper, R. and Frunk, C. 1984. Herbal remedies of the 

maritime Indians: phytosterol and triterpines of 67 plants. Journal 

of Ethnopharmacology, 10(2): 181-194. 



Performance Evaluation of Commercial Reapers for Finger Millet Harvesting 

H.G. ASHOKA 1 , G.M. PRASHANTHA 2 , E.G. ASHOK 3 , AND M.V. CHANNA BYREGOWDA 4 

1,2 

Division of Agricultural Engineering, 3,4 AICRP on Small Millets, University of Agricultural Sciences, G.K.V.K., 

Bangalore 560 065, Karnataka, India 

e-mail: agrilengineeruasb@gmail.com 1 

ABSTRACT 

The experiment was conducted during 2011-12 in finger millet 

plot for harvesting Fingermillet (Eleusine coracona gaertn) by 

using different commercial reapers viz. Shrachi, Fortune, 

KAMCO and Vinayaka make power tiller reaper attachment. 

The different reapers were used for testing the suitability for 

harvesting of Fingermillet crop and the different genotypes 

used were MR I, L 5, GPU 66, GPU 67 and GPU 28. Out of these 

genotypes, GPU 67 was considered good for mechanical reaper 

harvesting as the crop stand was medium with better inclined 

angle of 85 0 . The results indicated that out of four commercial 

reapers, Shrachi make is more suitable for harvesting finger 

millet crop grown under rainfed condition. The minimum and 

satisfactory harvesting stubble height of 8-9 cm with negligible 

shattering losses was observed by using Shrachi reaper 

compared to other reapers. 

Key words 

Finger millet, genotypes, Reaper, Harvesting, Field 

efficiency, Stubble height, Plant shoot, Shuttering loss 

Finger millet (Eleusine coracona gaertn) also known 

as Ragi is the most important small millet food and fodder 

crop. It is widely grown in Karnataka, Tamilnadu, Andhra 

Pradesh, Orissa, Bihar, Gujarat, Maharashtra and in hilly 

regions of UP, Sikkim and Himachal Pradesh. Karnataka is the 

major finger millet producing state in India covering about 

60% of the total area (Anonymous, 2010). This crop is grown 

both in dry land as well as in irrigated conditions where 

irrigation facilities are available. Rainfed finger millet 

constitutes major area of 95% and it is usually grown in Kharif 

and irrigated finger millet in Rabi or summer. 

Harvesting is one of the high energy consuming 

operations involving high cost of production for finger millet. 

It is estimated that harvesting consumes about one third of 

the total requirement of production system (Ohja and 

Devamani, 1987). During the peak harvesting season, getting 

the required labour is becoming a difficult task which is forcing 

to mechanise the operations. In view of the demand for 

mechanising peak seasonal operations, several commercial 

reaper harvesters are available for harvesting paddy, wheat 

and maize. These reapers cut the crops at ground level and 

lay the harvested crop in the form of windrows. However the 

reapers are not used to harvest the Finger millet crop due to 

the hardy stem of the crop. Thus require sturdier reaper to 

withstand field vibrations and cutting load than the one 

available for harvesting of paddy and wheat (Kumar, et al., 

2006). Keeping in view of the arising situation demanding for 

mechanizing the harvesting operation, a performance 

evaluation of four different popular reapers was carried out 

during 2011-12 at the UAS, GKVK, Bangalore. 


The field performance of the four popular commercial 

reapers viz., Shrachi, Fortune, KAMCO and Vinayaka make 

power tiller operated reaper attachment was evaluated for 

harvesting finger millet crop at the Division of Agricultural 

Engineering, UAS, GKVK, Bangalore. The five different 

popular genotypes of Finger millet grown as dry land crop 

were utilized for the studies. The different genotypes grown 

are MR I, L 5, GPU 66, GPU 67 and GPU 28. The reapers used 

for evaluation were kept in good working condition and 

operated by a skilled operator. The field of 474 m 2 areas for 

each variety was considered for the study. The parameters 

like height of cut, speed of operation, fuel consumption, field 

capacity, incomplete harvest/left out, shattering losses and 

break down for reaper blades were recorded. The crop 

parameters like plant height, angle of inclination, plant 

population, moisture content at the time of harvest, diameter 

of stem at cutting height were also recorded for each variety. 


Among the five different genotypes MR 1 was the most 

succulent genotypes and having the moisture content of 71% 

and GPU 66 had the lowest moisture content of 43%. Whereas 

the genotypes viz., GPU 67, L 5 and GPU 28 had the moisture 

content of 67.5, 57 and 53.4% respectively. The plant height 

was observed to be highest in MR 1 (116.4 cm) followed by 

GPU 66 (104.13cm), (Table 1). Except GPU 66, the remaining 

genotypes had much greener fodder at the time of harvest. 

The inclined angle of 85 0 was observed in GPU 67 and in the 

remaining genotypes, the crop stand was highly inclined and 

lodged. The plant population was uniform in all the genotypes 

and there was not much difference found for number of tillers/ 

ears per plants. Among the five genotypes GPU 67 had the 

greenish fodder due to its staggered nature and medium in 

height having the better inclined angle for mechanized 

harvesting (Table 1). 

The commercial reapers Vinayaka brand reaper was 

attached to 15 HP Shrachi power tiller (Table 2). The other 

three reapers are Shrachi (Petrol), Fortune (Diesel) and 

KAMCO (Petrol) engines attached to exclusive self propelled 

reapers having the power source of 4.6, 4.0 and 3.5 HP engines


Table 1. 

Table 2. 

Technical specification of commercial vertical conveyor reapers for Finger Millet 

Sl. 

No. 

Particulars Shrachi-petrol Engine 

operated 

Fortune-diesel Engine 

operated 

KAMCO, 

petrol Engine 

Vinayaka, petrol Engine 

Power tiller attached 

1. Power source, hp 4.6 4.0 3.5 15 

2 Power transmission Clutch Clutch Clutch Clutch 

3. No. of cutting blades 24 23 25 17 

4. Width of cutting, cm 90 110 126 132 

5. Windrowing Facility 

Fibre plastic Star wheel and 

metallic chain with finger 

respectively. The shrachi and fortune brands are the imported 

units such as; shrachi and fortune have higher operating speed 

of 3.5 and 4.0 kmph. The KAMCO was able to operate at the 

lowest speed of 3.5 kmph. Among the reapers, the shrachi 

reaper could harvest the crop very close to the ground (8- 

9cm) and the KAMCO could able to harvest at much higher 

height (15cm). Also the shrachi reaper harvested the crop to 

the extent 100% compared to other reapers. However, there 

was not much shattering loss observed while harvesting by 

any of the reapers. The harvesting loss was observed to be 

0.2% but it was 0.4% in the case of Fortune reaper. The effective 

field capacity was higher in Fortune (0.383 ha/hr) and Vinayaka 

(0.373 ha/hr) reapers. Higher effective field capacity of Fortune 

is attributed to higher working speed with higher fuel 

consumption though the width of operation was lower than 

KAMCO and Vinayaka. All the four reapers recorded higher 

field efficiency of more than 90% and were comparable to 

each other (Table 3). 

Among the five different genotypes GPU 67 was having 





the green fodder with medium height having the better inclined 

angle for mechanized harvesting. Among the four different 

reapers Shrachi reaper harvested the crop completely 

compareable to other reapers. Though the width of cut is 

90cm, the Shrachi with 4.6HP petrol engine recorded 0.2% 

shattering loss with 1.8 lit/hr fuel consumption. The better 

harvesting of finger millet was possible by using Shrachi reaper 

fitted with 24 cutting blades optimized with 90cm cutting width. 



non metallic belt with finger 

6. 

Speed, Forward/ 

Reverse 

2+1 2+1 2+1 2+1 

7. Break down Nil Nil Nil Nil 

Table 3. 

Crop parameters of Finger Millet genotypes at the harvesting stage 

Sl. No. Particulars MR I L 5 GPU 66 GPU 67 GPU 28 

1. 

Green fodder, dry, 

Green fodder, medium Green fodder, medium 

Appearance of crop at the time of Greenish and tall 

medium height, Lodged height, good for height, good for 

harvest 

(tied) 

tied 

mechanised harvest mechanised harvest 

2. 

Moisture content at the time of 

harvest, % wb 

71 57 43 65.7 53.4 

3. Plant height, cm 116.4 93.6 104.13 68.8 95.4 

4. Inclined angle of plant 

Lodged 

Lodged Lodged 

Lodged 

85 

(0-20) (0-22) (0-23) 

(0-24) 

5. Plant population, no/m² 47 48 45 44 46 

6. No. of tillers/ears per plant 1-2 1-2 2-3 2-3 1-2 

7. 

Diameter of stem at cutting 

height, cm 

0.9-1.0 0.8 0.8-1.0 0.8-1.0 1.0 

8. Yield of grain, q/ha 35 35 38 35 35 

Performance evaluation of commercial vertical conveyor reapers for Finger Millet 

Sl. No. Particulars Shrachi-petrol 

Engine 

Fortune-diesel 

operated 

KAMCOpetrol 

Engine 

Vinayaka–diesel engine 

Power tiller attachment 

1. Power source, hp 4.6 4.0 3.5 15 

2. Operating speed, kmph 3.5 4.0 2.5 3.2 

3. Effective field capacity, ha/hr 0.378 0.396 0.13 0.28 

4. Field efficiency, % 90 90 90 92 

5. Stubble height, cm 8-9 12.5-15.0 10.2 13.6 

6. Left out plant shoots/M 2 Nil 3 1 2 

6. Shattering losses, % 0.2 0.4 0.2 0.2 

7. Fuel consumption, lit/hr 1.8 2.0 1.5 1.7 

Anonymous. 2010. http://dacnet.nic.in/extension/document/chapter/ 

66.htm. 

Kumar, K.G., Chowdegowda, M. and Jayamala, G. B. 2006. Comparative 

performance of mechanical reapers for harvesting rainfed and 

irrigated Fingermillet. Mysore J.agric.Science., 40(3): 351-355. 

Ohja, T.P. and Devnami. 1987. Status of harvesting machinery in 

India-a country report ‘Regional workshop on Design and 

Development of harvesting and threshing equipment’ IARI, New 

Delhi, October 4-14. 



Induced Mutation Through Gamma Irradiation at Different Doses to Create Genetic 

Variability and Study the Improvement in Yield and Yield Attributes of Genotype 

HD 2867 

SHUBHRA SINGH 1 , RAM, M., S. MARKER, ABRAR YASIN, B. AND AKHILESH KUMAR 

Department of Genetics and Plant Breeding, Sam Higginbottom Institute of Agriculture Technology and 

Sciences, Deemed to be University, Allahabad 211 007 

email: shubhra_urmi@rediffmail.com 1 

ABSTRACT 

Different doses of gamma rays (20kR, 25kR and 30kR) were 

used to irradiate seeds of wheat genotype HD 2867. Treated 

seeds were sown along with control to study the induced 

variation and improvement in yield and yield contributing traits 

in M 2 

and M 3 

generations. The results revealed significant 

differences among the treatments. All three doses were quite 

effective in inducing genetic variability. The mean performance 

showed improvement in most of mutagenic treatments in M 3 

as compared to the corresponding treatments in M 2 

generation 

over untreated check. The most beneficial dose was 20kR. The 

impact of this dose was promising in days to flowering, number 

of tillers/plant, plant height, days taken from anthesis to 

maturity, days to maturity, test weight and yield/plant. However, 

high reduction in the mean value for all the characters were 

obtained in response to higher dose of gamma rays (30kR). It 

was concluded from this study that there was significant genetic 

variability induced through all the three mutagenic treatments. 

Significant enhancement in yield and yield contributing traits 

were observed at 20kR followed by 25kR. Under the influence 

of higher dose of gamma rays (30kR) significant reduction were 

observed in yield and yield attributes. It indicates that inducing 

genetic variability and improvement in quantitative traits would 

be possible through gamma rays. 

Key words 

Gamma rays, wheat cultivar, variability, yield and 

yield traits 

Mutation breeding is recognized as one of the driving 

force of evolution. Mutation breeding is relatively quicker 

method for improvement of various crop species. It is an 

important tool to create variability for quantitatively inherited 

traits in different plants and is considered as an alternative 

method to increase genetic variability in plant breeding 

(Camargo, et al., 2000). It is often used to correct defects in a 

cultivar, which has a set of good agronomic characteristics 

(Sigurbjornsson, 1977). Among various physical mutagens 

such as X-rays, fast neutrons, thermal neutrons, ultraviolet 

and beta radiation, gamma rays in particular are well known 

with their effect on the plant growth and development by 

inducing cytological, physiological and morphological 

changes in cell and tissues (Thapa, 2004). Gamma radiation is 

an important tool for inducing the genetic variability, enhancing 

yield and yield contributing traits. However, there is a need to 

predict the most beneficial dose of gamma rays for 

improvement of specific traits of crop plants because gamma 

radiation can induce useful as well as harmful effects. In India, 

NP 836 which is an awned mutant from the awnless wheat 

variety NP 799, Sharbati Sonara which is an amber grain 

colour mutant from the red grain colour wheat variety Sonora- 

64 (Singh, 2000) have been developed. Reddy and 

Viswanathan, 1999 induced rust resistance in wheat variety 

WH 147. Mackey, 1954 reported some beneficial radiation 

induced mutants in wheat with increase straw strength, 

resistance to stem rust and slightly early maturity. So far in 

the world 222 mutant varieties of wheat have been released 

(Phundhan Singh, 2010). The present investigation was 

undertaken with the objective to induce genetic variability, 

study the effect of various doses of gamma rays on yield and 

yield components of wheat cultivar HD 2867 and find out 

useful mutants in M 2 

and M 3 

generations under field 

conditions. 


The present investigation was carried out in Field 

Experimentation Centre, Department of Genetics and Plant 

Breeding, Sam Higginbottom Institute of Agriculture, 

Technology and Sciences, Deemed-to-be-University, 

Allahabad. Dry seeds of wheat genotype HD 2867 irradiated 

with 20, 25 and 30 kR doses of gamma rays from radioactive 

element Cobalt-60 ( 60 Co) source at National Botanical Research 

Institute (NBRI), Lucknow. As per the availability of literature 

regarding beneficial dose of gamma rays in wheat crop, 10, 15, 

20, 25 and 30 kR are considered to be much perfect dose for 

inducing genetic variability and producing desirable mutants 

(Arora, et al., 1989, Drozed, 1994, Shkvarnikov and Kulik, 1987). 

The effect of various doses of gamma rays was studied in M 2 

and M 3 

generations (M 1 

generation was already being raised 

during rabi 2007-08). Sowing of M 2 

generation was done during 

rabi season 2008-09 (25 th November) and M 3 

generation during 

rabi season 2009-10 (27 th November). The experiment was laid 

out in randomized block design with three replications. Each 

plot consisted of 4 rows, 2.5 mt. in length with row and plant 

to plant distance of 25 and 15 cm, respectively. In each plot 

about 65 treated seeds from 15 mutants that were already 

selected from each treatment (20, 25 and 30 kR) in M 1 

generation were dibbled along with non-irradiated seeds 

(control) to raise M 2 

and subsequently M 3 

generation. 

Selection was carried out in M 2 

generation and desirable plants 

from each treatment were harvested individually. The M 3


progeny was raised, from selected M 2 

plants and selection 

was further advanced on the basis of single plant selection 

method. The recommended cultural practices were followed 

during the crop growth period. The observations were recorded 

for days to flowering, number of tillers/plant, spike length, 

grains per spike, plant height, days taken from anthesis to 

maturity, days to maturity, 1000 grain weight and yield/plant. 

Data for no. of tillers/plant, spike length, no. of grains per 

spike and plant height were recorded on five randomly selected 

plants in each plot. Data on 1000 grain weight and yield per 

plant were recorded in gram. The data recorded for the above 

mentioned characters were averaged and subjected to 

statistical analysis as outlined by Steel and Torrie, 1980 and 

subsequently Duncans Multiple Range Test (Leclarg, et al., 

1963) was used to establish the differences among the different 

treatment means. 


The differences in the mean value of all the traits due to 

different radiation doses were highly significant in both M 2 

and M 3 

generation. The results correspond to those of Jamil 

and Khan, 2002, who irradiated wheat cultivar Bakhtawar 92 

by gamma rays at 5, 10, 15 and 25 kR, observed highly 

significant differences in the mean value due to different 

radiation doses. It is revealed from the Tables 1-2 that 

significant delay in flowering was recorded in cv. HD 2867 at 

different radiation doses, as the doses increased to higher 

level, a delay in days to flowering was noted. An increase of 

91 and 92 days was recorded by 30kR followed by 25kR dose 

(89 and 90 days) and 20kR (87 and 88 days) as compared to 

control (85 days) and also the extent of variability was recorded 

higher for this trait both in M 2 

and M 3 

generation. The present 

results are in conformity with the finding reported by Rahim, 

et al., 2003. It was observed (Tables 1 and 2) that number of 

tillers per plant varied significantly when radiated with various 

doses of gamma rays. 20kR dose increased number of tillers 

per plant in both M2 (14.16) and M3 (15.26) generations 

followed by 25kR dose. The highest dose of 30kR caused 

reduction in tillers per plant (8.40 in M2 generation and 9.31 in 

M3 generation) as compared to control. The results are in 

conformity with those of Din, et al., 2003; who found significant 

decrease in number of tillers per plant of different wheat 

varieties at higher intensity of gamma radiation. The data 

regarding spike length showed significant variability for wheat 

genotype HD 2867 due to different radiation doses. However, 

all the radiation doses showed reduction in spike length while 

comparing the mean values of gamma rays with one another. 

The minimum spike length (8.71 in M 2 

and 9.67cm in M 3 

generation) was recorded with 30kR dose and maximum spike 

length (12.21 in M 2 


generation) was recorded 

in control. The findings are in agreement with those already 

reported by Khan, et al., 2003. It was inferred from the tables 

1-2 that both in M2 and M3 generations there was significant 

variation observed for number of grains per spike. By 

comparing the mean values due to various radiation doses 

with one another it was observed that the number of grains 

per spike was reduced as the radiation dose increased. The 

maximum decrease in number of grains per spike was observed 

due to 30kR dose of gamma rays, 52.13 in M 2 

and 53.32 in M 3 

generation as compared to control. These results are in 

agreement with those of Khamankar, 1989. It was apparent 

from the results (Tables 1-2) that extent of variability in plant 

height increased in both the generations. 

The radiation dose of 30kR gamma rays reduced plant 

height, 84.48cm in M2 generation and 85.28cm in M3 generation 

over the control. It was noted that plants radiated with 20kR 

gamma rays showed significant increase in plant height, 

91.55cm in M 2 


generation over untreated 

Table 1. 

Mean performance of various characters of wheat genotype HD 2687 treated with different doses of gamma rays in 

M 2 

generation 

Radiation Days to No. of Spike No. of Plant Days taken from Days to 1000 Yield/plant 

dose (kR) flowering tillers/plant length grains per height anthesis to maturity grain (g) 

(cm) spike (cm) maturity wt (g) 

Control 85d 10.54c 12.21a 56.43a 90.39b 32c 116d 40.53c 10.39c 

20 87c 14.16a 10.87b 54.38b 91.55a 34a 121a 43.31a 12.16a 

25 89b 13.32b 9.68c 53.34c 88.71c 33b 120b 42.46b 11.36b 

30 91a 8.40d 8.71d 52.13d 84.48d 30d 118c 38.57d 9.43d 

Mean values sharing same letter does not differ significantly at 5% level of probability (P=0.05) 

Table 2. 

Mean performance of various characters of wheat genotype HD 2687 treated with different doses of gamma rays in 

M 3 

generation 

Radiation Days to No. of Spike No. of Plant Days taken from Days to 1000 Yield/plant 

dose (kR) flowering tillers/plant length grains per height anthesis to maturity grain (g) 

(cm) spike (cm) maturity wt (g) 

Control 85d 10.41c 12.29a 56.59a 90.41b 33c 117d 40.83c 10.68c 

20 88c 15.26a 11.68b 55.51b 92.34a 35a 122a 44.28a 13.16a 

25 90b 14.41b 10.54c 54.21c 89.53c 34b 121b 43.16b 12.21b 

30 92a 9.31d 9.67d 53.32d 85.28d 31d 119c 39.11d 10.22c 

Mean values sharing same letter does not differ significantly at 5% level of probability (P=0.05)

SINGH et al., Induced mutation through gamma irradiation at different doses to create genetic variability 67 

check. So, all the doses adversely affected the average plant 

height. Plant height was inversely proportional to the increase 

in the radiation intensity. The results are in conformity with 

Muhammad and Khalid, 2001. In response to various doses 

of gamma rays, significant differences in the mean values were 

observed for days taken from anthesis to maturity. By 

comparing the effect of various radiation doses, it was 

observed that all the doses (except 30kR) increased days taken 

from anthesis to maturity in both the generations, 34 days in 

M 2 

and 35 days in M 3 

generation by 20kR followed by 25kR 

over control. It was found from the present investigation that 

days to maturity significantly increased due to various doses 

of gamma radiation over untreated control. The extent of 

variability for this trait was higher in both M 2 

and M 3 

generations. The differences in the mean values for 1000 grain 

weight due to various doses of gamma rays varied significantly 

both in M 2 

and M 3 

generations. By comparing the mean values 

of various doses with one another it was found that 1000 

grain weight significantly decreased due to 30kR radiation 

dose, 38.57g in M 2 

and 39.11g in M 3 

generation as compared 

to control. The maximum increase in 1000 grain weight (gm) 

was observed in response to 20kR radiation dose, 43.31g in 

M 2 

and 44.28g in M 3 

generation followed by 25kR radiation 

dose over untreated check. In general, gradual decrease in 

1000 grain weight appeared due to increase in radiation intensity 

both in M 2 

and M 3 

generation. These findings are inline with 

Muhammad, et al., 2003. The differences in the mean values 

for grain yield per plant due to different doses of gamma rays 

were highly significant. The data from the Tables 1-2 revealed 

that there was significant increase in the grain yield per plant 

at 20kR radiation dose, 12.16g in M 2 

and 13.05g in M 3 

generation followed by 25kR radiation dose over control. The 

results are in agreement to those of Jamil and Khan, 2002 and 

Khan, et al., 2003. However, at 30kR radiation dose grain yield 

decreases significantly, 9.43g in M 2 

and 10.22g in M 3 

generation 

as compared to control. 

Change brought by mutation is permanent and heritable. 

If the changes would be brought by environment they are not 

fixable and heritable. For example, from present investigation 

it has been observed that in both M 2 

and M 3 

generation, there 

is continuous induction of genetic variability and all the 

treatments are showing their effect continuously. If it will be 

due to environmental fluctuation such permanent changes 

could not observed generation after generation. 

From the above foregoing results and discussion, it is 

concluded that different doses of gamma rays in wheat 

genotype HD 2867 provide enough scope by developing a 

wide range of variation in desirable plant attributes to select 

high yielding mutants. From the present study significant 

genetic variability was induced through all the three mutagenic 

treatment and also enhancement in yield and yield 

contributing traits were observed at 20kR followed by 25kR. 

Under the influence of higher dose of gamma rays (30kR) 

significant reduction were observed in yield and yield 

attributes. It indicates that inducing genetic variability and 

improvement in quantitative traits would be possible through 

gamma rays. Hence, gamma ray played a pivotal role in crop 

breeding through mutation and stability of genetic variability 

should be analyzed in succeeding generations and selection 

of desirable mutants could be performed for a successful 

breeding programme. 


Arora, R., Maherchandani, N., Uppal, S. 1989. Modulation of radiation 

effects in wheat by growth regulators. Ann. Biol., Ludhiana, 5: 

109-113. 

Camargo, C.E.D.O., Neto, A.T., Filho, A.W.P.F., Felico, J.C. 2000. 

Genetic control of aluminium tolerance in mutant lines of the 

wheat cultivar Anahuac. Euphy. 114: 47-53. 

Din, R., Khan, M.M., Qasim, M., Jehan, S., Khan, M.M.I. 2003. Induced 

mutability studies in three wheat (Triticum aestivum L.) varieties 

for some morphological and agronomical characteristics. Asian J. 

of Pl. Sci., 17(2): 1179-1182. 

Drozed, D. 1994. The effect of radiation on spring wheat properties. 

Int. Agrophys. 8: 209-213. 

Khamankar, Y.G. 1989. Gamma ray irradiation and selection for yield 

components in bread wheat. PKV Res. J., 13:1-5. 

Khan, M.M., Din, R., Qasim, M., Jehan, S., Iqbal, M.M. 2003. Induced 

mutability studies for yield and yield related characters in three 

wheat (Triticum aestivum L.) varieties for some morphological and 

agronomical characteristics. Asian J. of Pl. Sci., 2: 1183-1187. 

Leclarg, R.L., Leonard, W.H., Clark, A.G. 1962. Field plot technique. 

2 nd ed. Burgees publish. Co. South Minnesota. pp. 144-146. 

Madina Jamil, Umer Khan Q. 2002. Study of genetic variation in yield 

components of wheat cultivar buktawar-92 as induced by gamma 

radiation. Asian J. of Pl. Sci., 1(5): 579-580. 

Mackey, J. 1954. Neutron and x-ray experiments in wheat and revision 

of speltoid problem. Hereditas, 40:65-180. 

Muhammad Irfaq, Khalid Nawab, 2001. Effect of gamma irradiation on 

some morphological characteristics of three wheat (Triticum 

aestivum L.) cultivars. Online J. of Bio. Sci., 1(10): 935-937. 

Muhammad, Mohibullah, Khan, Rahim, Din, Muhammad, Qasim, Shah 

Jehan, Malik Muhammad Iqbal, 2003. Induced mutability studies 

for yield and yield related characters in three wheat (Triticum 

aestivum L.) varieties. Asian J. of Pl. Sci., 2: 17-24. 

Phundan, Singh. 2010. Essentials of Plant Breeding, Kalyani Publishers, 

New Delhi, pp. 219. 

Rahim, Din, Muhammad, Qasim, Khalil, Ahmad, Shah, Jehan. 2003. 

Study of days taken to earing initiation and earing completion in 

M 1 

generation of different wheat genotypes irradiated with different 

doses of gamma radiation. Asian J. of Pl. Sci,. 2(12): 894-896. 

Reddy, V.R.K., Viswanathan, P. 1999. Induced rust resistant mutants in 

hexaploid wheat “WH 147”, Crop Research (Hisar). pp. 443-445. 

Shkvarnikov, P.K., Kulik, M.I. 1987. Induction of mutation in wheat, 

Academy of Sciences, Ukrainian SSR, Kiev, USSR. 41: 204-217. 

Sigurbjörnsson, B. 1977. Introduction Mutations in Plant Breeding 

Programs. Manual on Mutation Breeding Second Edition Tech. 

Report Series. 119 IAEA,Vienna, pp. 1-6. 

Singh, B.D. 2000. Plant Breeding, Kalyani Publishers, New Delhi, pp. 

627-628. 

Steel, R.G.D., Torrie, J.H. 1980. Principles and procedures of statistics. 

McGraw Hill Book Comp. Inc., New York. 

Thapa, C.B. 2004. Effect of acute exposure of gamma rays on seed 

germination and seedling growth of Pinus kesiya gord and P. 

wallichiana A.B. jacks. Our Nature, 2: 13-17. 



Assesment of Genetic Divergence in Chickpea Genotypes (Cicer arietinum L.) 

SUDHANSHU JAIN 1 AND Y.M. INDAPURKAR 

Deptt. of Genetics and Plant Breeding, College of Agriculture, RVSKVV, Gwalior 

email: sudhanshujain07@yahoo.in 1 Table 2. Intra (diagonal) and inter-cluster average 

ABSTRACT 

A study was carried out on the thirty genotypes of chickpea to 

study the nature and magnitude of genetic divergence using 

Mahalanobis’s D 2 Statistics. The data were recorded on eight 

important quantitative traits from the genotypes grown in 

Randomized Block Design having three replication. The thirty 

chickpea genotypes were grouped into six clusters. The cluster 

I shows largest cluster with eight genotypes. Highest inter 

cluster distance was observed between cluster IV and cluster V, 

followed by cluster V and VI. The cluster VI was identified for 

pods per plant and seed yield per plant. Whereas genotypes 

included in cluster II were of early maturity with early flowering 

habit and average plant height. Genotypes included in these 

two clusters can be utilized for future crop improvement 

programme. 

Key word 

Chickpea, genetic divergence 

Assessment of genetic diversity of cultivated crop plants 

is very important to select proper genotypes for a hybridization 

program. Inclusion of genetically divergent parents in any 

breeding programme is essential to create new genetic stocks. 

Genetic diversity is the most important tool in the hands of 

the plant breeder in choosing the right type of parents for 

hybridization programme. The divergence can be studied by 

technique using D2 statistics developed by Mahalanobis, 

1936. It is based on multivariate analysis and grouped into 

various cluster. This is considered as the most effective method 

for qualifying the degree of genetic diversity among the 

genotypes included in the study. The present investigation 

aimed to estimate the magnitude of genetic divergence present 

in the genotypes and to identify the diverse genotypes for 

future. 


Thirty genotypes of chickpea were grown during rabi 

(winter) 2010-11 in a randomized block design with three 

replication at College of Agriculture, RVSKVV, Gwalior. 

Observations were recorded on five randomly tagged plants 

for Days to 50% flowering, Days to maturity, Plant height 

(cm), Number of branches per plant, Number of pods per plant, 

Number of seeds per pod, 100 seed weight (g) and Yield per 

plant (g). Wilks criteria was used to test the significance of 

differences in mean values of all the 10 characters. Genetic 

diversity was studied using Mahalanobis’s D 2 and clustering 

of genotypes was done according to Toucher’s method. Intra 

and inter-cluster distances and mean performance of the 

clusters for the characters were also computed. 


The analysis of variance for the experimental design 

showed highly significant differences among genotypes for 

all the characters studied. 30 genotypes were grouped into VI 

clusters using Mahalanobis D 2 statistics and Tocher’s method 

(Table 1). The cluster I was the largest and consisted of 8 

genotypes followed by clusters IV which had 7 genotypes. 

The cluster III and VI had 5 genotypes, cluster II had 3 

genotypes and cluster V had 2 genotypes. This suggested 

the presence of high degree of divergence in the material. 

Highest inter cluster distance was observed between cluster 

IV and cluster V followed by I and VI and V and VI respectively 

(Table 2). 

Average intra and inter cluster D 2 values among 30 

genotypes revealed that the cluster IV showed maximum intra 

cluster D 2 value (4.036) followed by cluster VI (3.818) and 

cluster I (3.530) indicating presence of diversity in these 

clusters. The inter cluster D 2 values ranged from 17.430 to 

6.105. Maximum inter cluster D 2 value was observed between 

cluster IV and V (17.430) which indicates that the genotypes 

Table 1. 

Distribution of 30 genotypes of chickpea into 

different clusters 

Cluster Number of 

Genotypes 

number genotypes 

I 8 C505,C513,C516,C532,C533,C534,C535,C537 

II 3 C502,C510,C523 

III 5 C504,C507,C514,C530,JG16, 

IV 7 C508,C509,C511,C517,C519,C522,C528 

V 2 C501,C525 

VI 5 C520,C521,C526,C529, C531 

distances (D 2 ) for 10 characters in 30 genotypes 

of chickpea 

Cluster number I II III IV V VI 

I 1.879 3.008 2.471 3.221 3.117 4.142 

(3.530) (9.084) (6.105) (10.374) (9.715) (7.156) 

II 1.547 2.927 3.065 3.292 2.857 

(2.393) (8.567) (9.394) (10.387) (8.265) 

III 1.722 3.912 3.441 2.716 

(2.965) (15.303) (11.840) (7.376) 

IV 2.009 

(4.036) 

4.175 

(17.430) 

2.790 

(7.784) 

V 1.516 

(2.298) 

3.713 

(13.786) 

VI 1.954 

(3.818)

JAIN AND INDAPURKAR, Assesment of Genetic Divergence in Chickpea Cultivars (Cicer arietinum L.) 69 

Table 3. 

Cluster mean values of 6 clusters for 8 characters 

in 30 genotypes of chickpea. 

S. Cluster No./ 

I II III IV V VI 

No. Character 

1 Days to 50% 72.42 62.98 72.00 78.00 68.50 72.20 

flowering 

4 Days to maturity 128.83 128.11 129.40 128.57 131.57 129.00 

3 Plant height (cm) 38.91 40.76 34.11 44.69 37.13 46.32 

4 Number of branches 3.39 4.02 4.33 4.19 3.83 3.89 

per plant 

5 Number of pods per 37.75 47.00 47.13 52.62 50.83 62.00 

plant 

6 Number of seeds per 1.10 1.40 1.08 1.44 1.13 1.43 

pod 

7 100 seed weight (g) 27.00 26.96 22.55 23.19 37.73 27.41 

8 Yield per plant (g) 12.67 16.98 13.24 13.54 15.97 22.61 

included in these different clusters may give high heterotic 

response in crop improvement programmes. Because crosses 

made between the genotypes of clusters separated by large 

inter-cluster distances (Sandhyakishore, et al., 2007; Chandra, 

et al., 2007) show high heterosis. Similar findings have been 

reported by Lal, et al., 2001. Minimum inter cluster D 2 value 

was observed between cluster I and III (6.105) indicating the 

close relationship among the genotypes included in these 

two clusters. The average cluster means for 10 characters 

(Table 3) revealed that genotypes included in cluster II were 

of early maturity with early flowering habit and average plant 

height. Selection of genotypes from this cluster for these 

characters may yield desirable results. Cluster III showed 

highest mean for number of branches per plant. Cluster IV 

showed higher mean performance for number of seeds per 

pod, where as cluster V showed maximum mean performance 

for 100 seed weight. Similar findings have been reported by 

Sable, et al., 2000. The cluster VI showed highest mean for 

pods per plant and seed yield per plant. The genotypes 

included in the cluster I showed minimum mean value for 

number of branches per plant, number of pods per plant and 

yield per plant. Diverse clusters namely, VI and II hold good 

promise as parents for obtaining potential hybrids for desirable 

characters and thereby creating greater variability to improve 

the yield. Similar result has been reported by Kumar, et al., 

1997. 


Chandra, B.S., Reddy, T.D. and Ansari, N.A. 2007.Genetic divergence 

in rice (Oryza sativa L.). Res. Crops, 8: 600-603. 

Kumar, N. 1997. Genetic diversity among chickpea accession. Ind. J. 

Genet. Plant Breed., 57(1): 12-15. 

Lal, D., Ram Krishna and S. Gurpreet, 2001. Genetic divergence in 

chickpea. Ind. J. Pulses Res., 14(1): 63-64. 

Mahalanobis, P.C., 1936. On the generalized distance in statistics. Proc. 

Natl. Inst. Sci. India. 2: 49-55. 

Sable, N.H., P.W., Khorgade and M.N., Narkhede, 2000. Genetic 

parameters and formulation of selection indices in chickpea. Annuals 

of Plant Physiology, Pub., 14(1); 83-87. 

Sandhyakishore, N., Babu, V.R., Ansari, N.A. and Chandran, R. 2007. 

Genetic divergence analysis using yield and quality traits in rice 

(Oryza sativa L.). Crop Improv., 34:12-15. 



Phytotoxic Effect of Asclepias curasavica Linn on Metabolism of Parthenium 

hysterophorus L 

T.G. NAGARAJA 1 AND A.H. PUDALE 2 

1 

Department of Botany, The New College, Kolhapur 416 012, Maharashtra State, 2 Department of Agrochemicals 

and Pest Management, Shivaji University, Kolhapur 416 004, Maharashtra State 

email: tgnagaraja2010@gmail.com 1 

ABSTRACT 

The present paper deals with the powdered leaves, stem and 

roots of Asclepias curasavica Linn adversely affected the 

metabolism of Parthenium hysterophorus L upto 30 DAS. It 

influenced plant height and number of leaves per plant. The 

biochemical constituents such as total chlorophylls, polyphenols 

and carbohydrate (sugar) contents were decreased in leaves, 

stem and roots of Parthenium hysterophorus L supplied with 

Asclepias curasavica L residues. The total lipid and protein 

contents were greatly reduced as compared to control. 

Therefore, residues of Asclepias curasavica L may be used as a 

potential bioherbicide. 

Key words 

Asclepias curasavica, Parthenium hysterophorus, 

sugars, lipids, chlorophylls and Proteins. 

Parthenium hysterophorus L is an annual herbaceous 

weed, aggressively colonizes in disturbed sites, native to 

Mexico, Central and South America. In India it is wide spread 

in grassland, fruit tree, orchards and arable land in natural and 

acidic soils. In India the weed is considered to be a major 

problem (Gupta and Sharma, 1977), because, it has high rate 

of fecundity, efficient seed dispersal and adaptability to grow 

under adverse environmental conditions. Because of high 

prolific rate of reproductive ability, this exotic weed dominated 

in arable and non-arable land. In some areas it has become an 

extremely serious in agricultural and rangeland habitats. 

Parthenium hysterophorus L is also know to be allergenic to 

some people and live stock, it creates allergic skin diseases, 

asthma in human beings, due to presence of allelopathic effect 

(Das and Das, 1995). The allelopathic potential of Parthenium 

hysterophorus L weed results from release of phytotoxic 

substances such as ferulic, chlorogenic, P-coumaric and P- 

hydrobenzoic acids, parthenin, ambrasin and coronopilines 

etc, which inhibits the germination growth of several crops 

and multipurpose trees (Basak, 1984) and also cause allergic 

dermatitis and asthma in human beings (Auld and Medd. 1987). 

Hence, it is a problematic weed in Indian agricultural system. 

At present it is controlled by mechanical and use of other 

herbicides. The used of herbicides created hazards on our 

ecosystem. Therefore, an interest created to study an 

alternative ecofriendly approach to control Parthenium. Thus 

the concept of allele chemicals utilized to control Parthenium 

hysterophorus L. 

Asclepias curasavica Linn commonly called milk weed 

or silk weed or butterfly weed, blooms almost continuously in 

uncultivated waste lands, sometimes cultivated, even in 

garden, as the blooms are quite attractive to butterflies, 

especially Monarch butterflies. Because of its popularly as 

an ornamental plant, it has become natural weed in tropical 

and sub tropical pastures, fields and disturbed areas 

throughout the world. The plant is toxic to animals, a resinoid 

(galitoxin) is the toxic principle found in the milk latex of the 

plant stem. The plant also possess other phytochemicals 

such as cardiac glycosides, acetyl-beta-glucosominidase, 

asclepin and beta fucosiodase. Even in ayurveda, whole plant 

is considered as emetic, styptic, and purgative. Powder of 

root mixed with equal quantity of Acorus root is given in 

chronic ulcers, sometimes root decoction is used against 

cancer (Joshi, 2000). As the plant Asclepias curasavica L 

possess several phytochemicals, therefore, an attempt has 

been made to study the effect of phytochemicals 

(Allelochemicals) on growth and metabolism of 

P.hysterophorus Lin to identify its suitability as a botanical 

herbicide. 


Freshly harvested leaves, stem and roots of Asclepias 

curasavica Linn from Ambai Defence colony, Kolhapur during 

the months of July-Nov 2010, for experimental study were 

collected. The collected sample were washed properly through 

distilled water and air dried and kept for oven at 70ºC for 

consecutive three days. The dried material of leaves, stem 

and roots were finely powdered in domestic grinder separately. 

Pot studies were carried out during September to December 

2010 in the Botanical Garden of Department of Agro Chemicals 

and Pest Management, Shivaji University, Kolhapur. 

The fire clay pots of 9 cms deep and 21 cm diameter were 

filled with 10 kg of loam soil. Finely powdered dried samples 

of leaves, stem and roots of Asclepias curasavica L were 

mixed at the rate of 100 gm per pot separately. In each fire clay 

pot five viable seeds of Parthenium hysterophorus L were 

sown at equal distance, one pot used as control. Uniform 

watering was done i.e., 100 ml per pot continuously up to 30 

days (Days after sowing). After 30 days treated plant parts 

were used for biochemical test and growth parameters. 

The treated (residues of A.curasavica L) leaves, stem 

and roots on Parthenium hysterophorus L were used for 

experiment which was laid down in randomized block design

NAGARAJA AND PUDALE, Phytotoxic Effect of Asclepias Curasavica Linn on Metabolism 71 

with two replicates. The Chlorophyll content from leaves and 

stem of treated samples were extracted with 80% acetone along 

with control samples and estimated by the method of Arnon, 

1949. The polyphenol content were determined by the method 

of Folin and Denis, 1915. The total carbohydrate and starch 

measured by the Anthrone method presented by Hodge and 

Hofreiter, 1962 and Thayumanavan and Sadasivan, 1984. The 

soluble proteins were estimated by the method of Lowery, et 

al., 1951. The reducing sugars were calculated by 

dinitrosalicylic acid method (Miller, 1972) and fatty acids were 

calculated by Cox and Pearson, 1962 method. 


Ground plant parts (leaves, stem and roots) of Asclepias 

curasavica Linn significantly inhibited the growth parameters 

like height and number of production of leaves on Parthenium 

hysterophorus L. 

The germination and growth of Parthenium 

hysterophorus L was hindered, 69.23% and 53.84% was 

reduced in leaves and stem extracts. While 38.46% was reduced 

in root treatment. A significant 35.29 and 29.41% of reduction 

was noticed after 30 DAS (Table 1) with least per cent growth 

(17.64%) in root extract was recorded. 

A parallel result were recorded on production of leaves 

in P.hysterophorus L due to application of residues of A. 

curasavica Linn (Table 1). The numbers of leaves were reduced 

as compared to control, after 30 DAS treatment. A maximum 

of 35.29 per cent of reduction was recorded in leaf extract 

followed by stem and root as compared to control. A similar 

investigation was carried out Nagaraja and Deshmukh, 2009 

in residues of Andrographis paniculata. Thus totally plant 

height and number of production of leaves hindered 

significantly may be due to phyto toxicity effect of Asclepias 

curasavica L. Hence, allelochemicals may have a great 

influence on physiology and metabolism process. Therefore, 

growth and inhibition of Parthenium hysterophorus L may 

be due to allele chemicals released from leaf, stem and roots of 

A. curasavica L (Thaper and Singh, 2006). 

Meanwhile application of leaf, stem and root biomass 

(residues) of A.curasavica L on Pathenium hysterophorus L 

shows (Table 2) reduced its phytochemicals. The total 

chlorophylls get reduced as compared to control. 11.70 mg of 

chlorophyll per 100 g of fresh tissue get reduced to 10.91 g in 

leaves, 6.10 mg in stem and 2.60 mg in root after 30 DAS 

treatment. A parallel findings were reported by Nagaraja and 

Deshmmukh, 2009 in Andrographis paniculata. The 

polyphenol content were responsible for resistant has been 

extensively worked out Deshpande, 1993, get reduced in 

treated parts of Parthenium hysterophorus L (Table 2) after 

30 DAS of treatment with residues. The total carbohydrate 

content in all treated plant parts such as leaves, stem and 

roots (P. hysterophous) get reduced, similarly starch and 

reducing sugar also reduced after 30 DAS of treatment (Table 

2) Hence, allele chemicals interferes with photosynthesis, in 

relation to water (Colton and Einhelling, 1980) and nutrient up 

take (Craig and Einhelling, 1980), which were required for the 

metabolism of number of organic compounds. 

The lowered synthesis of carbohydrates due to 

Table 1. Phytotoxic effect of Asclepias curasavica Linn on growth and development of Parthenium hysterophorus L 

Treatment 

by residues 

Number of leaves per plant 

after 15 days 

Percentage of reduction in 

production of leaves 

Plant height in 

cm. after 15 days 

Percentage of reduction of 

growth in height 

Control 6-7 -- 3-8 -- 

Leaf residue 4-5 69.23 2-9 76.31 

Stem residue 3-4 53.84 2-7 71.05 

Root residue 2-3 38.46 1-6 42.10 

AFTER 30 DAS 

Control 8-9 --- 5-7 ------ 

Leaf residue 2-4 35.29 2-8 49.12 

Stem residue 2-3 29.41 2-5 43.85 

Root residue 1-2 17.64 1-2 19.29 

Table 2. Effect of residues of leaves, stem and root of Asclepias curasavica L on Parthenium hysterophorus L-regarding 

Biochemical constituents after 30 DAS 

Sr.No Constituents Control Leaves Stem Root 

1. Chlorophyll a* 13.30 5.15 7.20 3.30 

Chlorophyll b* 13.40 5.70 6.40 1.80 

Total chlorophylls( a + b )* 11.70 10.91 6.10 2.40 

2. Polyphenols** 714 285 221 210 

3. Total carbohydrates** 5620 4002 3187 2400 

4. Starch* 60.61 12.38 11.94 12.05 

5. Reducing sugars** 253.6 158.26 141.20 039.12 

6. Proteins** 417 360 214 166 

7. Lipids* 2.244 0.033 0.016 0.011 

*g -100 -1 g fresh tissue. **mg 100 -1 g fresh tissue.


Graph 1. Growth parameters. 

application of residues (biomass) of A. curasavica Linn (Table 

2) on P. hysterophorus L reflects on biosynthesis of proteins. 

The protein content may also get reduced in leaves, stem and 

roots. Besides, proteins may be used as respiratory substrate, 

when carbohydrates supply was in adequate, due to effect of 

allelo chemicals. The protein content get declined may be due 

to degradation or inhibition of protein synthesis under the 

influence of allelo chemicals (Table 2). 

The lipid metabolism was highly modified by the 

application of residues of A. curasavica Linn on Parthenium 

hysterophorus L (Table 2). After 30 DAS treatment, the lipid 

content was highly reduced in leaves, stem and roots. This 

may be due degradation or lipid peroxidation or inhibited 

synthesis under the influence of allelo chemicals. Thus in all 

residues (allelochemicals) influence the growth and 

metabolism of Parthenium hysterophorus L, therefore, the 

biomass may be used as herbal herbicides for management of 

weeds in agriculture. 


The authors are thankful to Co-ordinator, Department 

of Agro chemicals and Pest Management, Shivaji University, 

Kolhapur for providing laboratory facilities. 


Arnon, D. I. 1949. Copper enzyme in isolated chloroplast, polyphenol 

oxidase in Beta Vulgaris., Plant Physiology., 24: 1-15. 

Auld, B.A. and Medd, R.W. 1987. Weeds on illustrated botanical guide 

to the weeds of Australia: Lnkata Melbourne- Sydney, pp. 255. 

Basak, S.L. 1984. Parthenium a big treat to agriculture and health; in 

1980’s. Indian Agriculturist., 28: 137-139. 

Colton, C.E and Einhelling, F.A. 1980. Allelopathic mechanism of 

Velvet leaf (Abutilon theophrasii) on Soya bean. American journal 

of Botany, 67: 1407-1413. 

Graph 2. Biochemical constituents 

Cox, H.E and Pearson, D. 1962. The chemical Analysis of Foods, 

Chemical Publishing Co. Inc; New York, pp. 426. 

Craig, C.E and Einhelling, F.A. 1980. Allelopathic mechanism of Velvet 

leaf on Soya Bean., American Journal of Botany., 67: 1407-142- 

13. 

Deshpande, A.V. 1993. Studies in phylloidy disease in Parthenium with 

respect to Morphology, Plant phenolics and Sesui terpene lactone, 

M.phil thesis Shivaji University, Kolhapur. pp. 1-99. 

Das, B. and Das, R. 1995. Chemical investigation in Parthenium 

hysterophorus L: an Allelopathi plant. Allelopathy. Jounal, 2: 99- 

104. 

Folin, O. and Denis, W. 1915. A Calorimetric method for determination 

of phenols and Phenol derivatives in Urine, Journal of Biological 

Chemistry., 22:305-308. 

Joshi, S.G. 2000. Medicinal plants Oxford & IBH Publishing Co Pvt. 

Ltd., pp. 62. 

Gupta, O.P. and Sharma, J.J. 1977. Elpeligro del parthenium enla India 

possible Medidas de control del : N Boletin Fistosanitario FAO, 25: 

112-117. 

Hodge, J.E. and Hofreiter, B.T. 1962. In carbohydrate chemistry 17 

(eds. Whistler, R.L. and Be Miller, T.N.). Academic Press, New 

Yor k. 

Lowry, A.H., Rosenbrough, N.J., Fan, A.L. and Randal, R.J. 1951. Protein 

Measurement with folin phenol reagent., Journal of Biological 

Chemistry, 193: 265-275. 

Miller, G.L. 1972. Anal. Chem., 31: 426. 

Nagaraja, T.G. and S.M. Deshmukh. 2009. Phytotoxic effect of 

Andrographis paniculata Nees on metabolism of Parthenium 

hysterophorus L., Journal of Biopesticides, 2(2): 165-167. 

Thapar, R. and Singh, N.B. 2006. Phytotoxic effects of Cassia tora on 

growth and Metabolism of Parthenium hysterophorus L., Allelopathy 

Journal, 17(2): 235-246. 

Thayumanvan, B. and Sadasivam, S. 1984. Qual Plant Foods Hum 

Nutr, 34: 253. 



Serological Detection and Host Range Studies of Peanut Bud Necrosis Virus Infecting 

Tomato 

MALLU GOVARDHANA 1 , GOPINATH, K. 2 , VANITHA, L.S. 3 , ESWAR REDDY MADDI 2 AND 

MANJUNATHA, L. 3 

1 

Department of Biotechnology, Indian Institute of Horticultural Research, Bengaluru 560 089, Karnataka, 

India, 2 Department of Plant Science, University of Hyderabad, Hyderabad, 500 046, Andhra Pradesh, India 

3 

Department of Plant Pathology, University of Agricultural Sciences, GKVK, Bengaluru 560 065, Karnataka, 

India 

e-mail: mrgyadav@gmail.com 1 

ABSTRACT 

Peanut bud necrosis virus (PBNV), a tospovirus has become 

serious disease in major tomato (Lycopersicon esculentum) 

growing areas of the world. The virus was obtained in purest 

form from naturally infected tomato sample. The virus was 

detected by DAC-ELISA using polyclonal antibodies against 

PBNV and TSV. PBNV was succesfully transmitted to Vigna 

unguiculata, Nicotiana rustica and Nicotiana benthamiana. 

Key words 

Lycopersicon esculentum, PBNV, tospovirus, TSV, 

ELISA 

Tomato (Lycopersicon esculentum) is an important 

vegetable crop next to potato and sweet potato in the world. 

In India the largest grower of tomato is Andhra Pradesh, 

Karnataka, Bihar, Maharashtra and Orissa. Among various 

diseases on tomato, viral diseases are considered one of the 

most severe, affecting tomato production in many countries. 

There are about 75 viruses which infect tomato causing 

significant losses in the sustainable production of tomato 

viz., Tobacco mosaic virus (TMV), Potato virus X (PVX), 

Tomato yellow leaf curl begomovirus (TYLV), Potato virus Y 

(PVY) Tomato leaf curl virus (ToLCV) and tospoviruses 

(Reddy, et al., 1968). 

Peanut bud necrosis disease (PBND) has been described 

in India since 1962 by different names: groundnut mosaic, 

groundnut rosette, bunchy top, chlorosis, ring mottle, bud 

blight and ring mosaic (Reddy, et al., 1991). PBNV virion is 

spherical single strand RNA molecule as genetic material, 80- 

120 nm in diameter and display surface glycoprotein 

projections of 5 to 10 nm, which are embedded in lipid bilayer 

of 5-7 nm thickness and derived from host membrane. The 

virion has three genome segments, large (L), medium (M), and 

small (S). All three genome segments of virus have the same 

complementary nucleotides at their 3’ and 5’ termini. The proper 

identification and studies on transmission of the virus is a 

prerequisite for development of suitable management 

strategies in this point of view the present work was undertaken 

to investigate on serological detection of PBNV through DAC 

ELISA and transmission studies. 


Detection of PBNV through DAC ELISA 

Serological properties of the virus present in the samples 

of naturally infected tomato were determined using DAC- 

ELISA with polyclonal antibodies against different viruses 

like Tobacco streak virus (TSV), Cucumber mosaic virus (CMV) 

and Peanut bud necrosis (PBNV). 

DAC-ELISA was used to screen the virus infected 

tomato fruit sample by directly coating the antigen (plant sap.) 

on the inner walls of the microtitre plate well (Hobbs, et al., 

1987; Mowat and Dawson, 1987). 200 mg of infected fruit 

sample was macerated with 500 µl of carbonate buffer 200 µl 

of extract was loaded to respective wells of microtiter plate 

and kept for 2 hours incubation at room temperature. The 

plate was washed with PBS-T buffer with three times at 10 

minute interval. 200 µl of respective antisera for positive control 

CMV (1:5000), TSV (1:10000) and PBNV (1:10000) was added 

and incubated for 2 hours in rocker at room temperature. Plate 

was emptied and washed with PBS-T three times at 10 minute 

interval and 200 µl of secondary antibody-alkaline 

phasphatase conjugate was added to the well and incubated 

for 2 hours in rocker at room temperature, plate was washed 

with PBS-T three times at 10 minute intervals. Then 200 µl of 

the freshly prepared substrate p-nitro phenol phosphate 

(PNPP) was added and kept for 1 hour in dark. The reaction 

was terminated by adding 50 µl of 3 M NaOH solution in each 

well. The positive samples were screened by measuring 

absorbance at 405 nm wavelength. 

Virus Source 

The infected samples of tomato fruits showing chlorotic 

ring like symptoms was obtained from field of Andhra Pradesh 

Province (Figure 1) found positive for PBNV, CMV and TSV 

through DAC-ELISA. The infected tomato samples was 

maintained under -20 o C condition for further experimentation. 

About 500mg of pulp was taken as a source of virus and 

macerated in 1ml of ice cold 20mM phosphate buffer pH 8.0 

containing Sodium thioglycolate and beta mercaptoethanol


under chilled condition. 

Mechanical transmission 

The host plant Cowpea (C 152) was raised at glass house 

condition in plastic pots. The seeds were obtained from IIHR 

Bangalore. The 7 days old host plants Cowpea (C 152) were 

dusted with celite and mechanicals inoculated with the ice 

cold virus inoculum by applying gently on the plant leaf surface 

using muslin cloth within one min after preparation of 

inoculums and presence of virus was detected by performing 

DAC-ELISA. 

Host range studies 

Chlorotic local lesion that are found on the 13days old 

of post inoculated host plants Cowpea (C 152) sample was 

macerated using 1ml of ice cold 20mM phosphate buffer 

containing 0.025% Sodium thioglycolate. The host plants viz, 

Vigna unguiculata (C 152), Nicotiana benthamiana, 

Nicotiana rustica, Phaseolus vulgaris and Petunia hybrida 

were dusted with celite in order to carry out the mechanical 

inoculation using macerated sap, which was applied gently 

on the plant leaf surface by muslin cloth. The plants were 

maintained at 24 o C in the photo period of 16 hours light and 8 

hours dark period. The symptom appearance was observed 

daily. After two-three passages, the leaves of C 152 were used 

to detect virus by ELISA. 


Screening of PBNV from field collected tomato sample by 

DAC-ELISA 

Primarily DAC-ELISA was performed for the tomato 

samples using different PBNV and TSV antisera. The samples 

which are reacting positively were identified by visual scoring 

and respective ELISA plate was shown with positive result 

with PBNV, TSV and Cucumber mosaic virus (CMV) antisera 

(Figure 3). Among the field samples, 11 samples showed 

positive by visual observation (scoring over buffer controls). 

Optimization of inoculation buffer and Mechanical 

transmission 

The 20 mM phosphate buffer containing in the range 

0.02%-0.09% of sodium thioglycolate are showed efficient than 

the buffer containing -mercaptoethanol infection with 

characteristic symptoms of the PBNV in to the susceptible 

host plant(C 152). Specifically 0.03% of sodium thioglycolate 

are showed more efficient infection than other percentage of 

sodium thioglycolate in phosphate buffer (Table 1). But the 

control (only phosphate buffer) does not showed any 

characteristic symptoms on the host plant (C 152) even after 3 

weeks of post inoculation. The cow pea C 152 genotype 

exhibited characteristic chlorotic and necrotic local lesion on 

the 6 th day after inoculation and positive result for PBNV and 

TSV from 4 th day of post inoculation. 

Host range studies of the PBNV 

The PBNV positive sample subjected to mechanical 

inoculation on host plants Vigna ungiculata, Nicotiana 

benthamiana, Nicotiana rustica, Petunia hybrida and 

Phaseolus vulgaris. was succesfully transmitted to Vigna 

unguiculata, Nicotiana rustica and Nicotiana benthamiana, 

Inoculated leaves of Nicotiana benthamiana plants initially 

showed chlorotic local lesions spreading and resulting in 

systemic infection and apparent death of the whole plant 

(Figure 2a-c) in a span of two weeks. Cowpea C 152 genotype 

exhibited characteristic chlorotic local lesion on the 6 th day of 

after mechanical inoculation. These lesions are expanding 

throughout the leaf as concentric rings. No senescence was 

observed even after 4-5 week of the inoculation. 

The Nicotiana rustica plants also showed initially 

chlorotic local lesions spreading after 20 days of post 

inoculated period, showed stunted growth. The virus was 

not transmitted to Phaseolus vulgaris and Petunia hybrida 

indicating that these are not hosts for PBNV. The virus was 

maintained on cowpea (C 152) and Nicotiana rustica plants 

with periodical re-inoculation in the green house with 15 days 

interval. The host range studies of PBNV are in conformity 

with studies conducted by Reddy, et al., 1991 who studied 

Fig. 1. 

Fig. 2. 

PBNV infected tomato fruit sample showing chlorotic 

concentric rings 

(a-c) Host range study of peanut bud necrosis virus 

(PBNV) 

Table 1. 

Standardization of inoculation buffer 

Control 1µl 2µl 

3µl 

4µl 5µl 6µl 

7µl 

8µl 9µl 

(0.01%) (0.02%) (0.03%) (0.04%) (0.05%) (0.06%) (0.07%) (0.08%) (0.09%) 

- - + +++ ++++ +++ + + + + + + 

a) ++++ = Excellent b) +++ = Very Good c) ++ = Good d) + = Fair e) — = Not Good

MALLU et al., Serological detection and host range studies of Peanut bud necrosis virus on tomato 75 

was detected through ELISA. ELISA has been used to detect 

many plant viruses using either polyclonal or monoclonal 

antibodies (Banttari and Hibi, and Saito, 1985). 

Fig. 2b) 1. Healthy N.rustica p lant 2. 15days o f po st 

inoculation 3. 3 weeks of post inoculation 


The authors acknowledge Dr. Varsha Wesley, ICRISAT, 

Hyderabad, AP, India and Prof. H. Savithri, IISC, Bangalore, 

Karnataka, India, for providing PBNV and TSV antisera. 


Fig. 2c) 1. Healthy N.ben thmian a 2. 3 weeks o f po st 

inoculation 3. 4 weeks of post inoculation 

A1, A2, E1&E2- Blank 

A3, A4, E3&E4- positive control (SeMV) 

A5to C12- TSV positives 

E5 to G2- PBNV positives 

SeMV- Sesbania Mosaic Virus 

Antiserum-PBNV-NP (1:5000) 

Each samples are in duplicates 

Fig. 3. Serological detection of PBNV by DAC-ELISA 

the diagnosis of viral infections based on symptoms through 

sap inoculation to different hosts, Vigna unguiculata, 

Nicotiana benthamiana and Nicotiana glutinosa. The virus 

Banttari, E.E. and Goodwin, P.H. 1985. Detection of potato viruses S, 

X and Y by enzyme-linked immunosorbent assay on nitrocellulose 

membranes (dot-ELISA). Plant Disease, 69: 202-205. 

Hibi and Saito. 1985. A Dot Immunobinding Assay for the detection of 

Tobacco Mosaic Virus in infected tissue. Journal of General Virology, 

66:1191-1194. 

Hobbs, H.A., Reddy, D.V.R., Rajeshwari, R. and Reddy, A.S. 1987. Use 

of direct antigen coating and protein A coating ELISA procedures 

for detection of three peanut viruses. Plant Disease, 71:747-749. 

Mowat, W.P. and Dawson, S. 1987. Detection and identification of 

plant viruses by ELISA using crude sap extracts and unfractionated 

antisera. Journal of Virological Methods, 15(3):233-247. 

Reddy, D.V.R., Wightman, J.A., Beshear, R.J., Highland, B., Black, M., 

Sreenivasulu, P., Dwivedi, S.L., Demski, J.W., McDonald, D., Smith, 

J.W. and Smith, D.H. 1991. Bud necrosis: a disease of groundnut 

caused by tomato spotted wilt virus. Patancheru 502324, Andhra 

Pradesh, India: International Crops Research Institute for the Semi- 

Arid Tropics. Information Bulletin no. 31. 

Reddy, M., Reddy, D.V.R. and Appa Rao, A. 1968. A new record of virus 

disease on peanut. Plant Disease Reporter, 52:494-495. 



Influence of Genotypes on Consumability of the Leaves, Growth and Development 

of Spodoptera litura (Fab.) 

PRADYUMN SINGH, N.S. BHADAURIA AND POOJA CHAUHAN 

Rajmata Vijayaraje Scindia Krishi Vishwa Vidyalaya, Gwalior 474 002 M.P., India 

ABSTRACT 

Experiments on influence of genotypes on consumability of 

the leaves, growth and development of Spodoptera litura (Fab.) 

were conducted during Kharif, 2008-09 and 2009-10 at College 

of Agriculture, Gwalior. On the basis of weight and size of 

larvae, larval and pupal period and growth rate genotype TJM 

12 showed antibiosis mechanism of resistant against pest. On 

the basis of consumption of leaves and consumption index 

genotypes TJM 15 and TJM 12 showed antifeedant affect 

against tobacco caterpillar. Whereas genotypes MI 24-91 and 

MI 181-1 showed highly susceptibility on the basis of 

consumability of leaves. 

Key words 

Tobacco caterpillar, Spodoptera litura, Mungbean 

genotypes 

The tobacco caterpillar, Spodoptera litura (Fab.) is a 

polyphagus insect and in recent years it is causing serious 

losses to pulses and oilseed crops in Madhya Pradesh during 

Kharif season. 

Mungbean is a low economic return crop and grown by 

poor farmers and use of recommended insecticides for the 

control of insect pest, so for are uneconomical. The 

management of pest, with insecticides is not also very effective 

due to difficulty in their application during rainy season. 

Insecticides also cause environmental pollutions, hazard to 

non target species, pest resurgence, secondary pest out break 

residue problem etc. Hence the use of insect resistant varieties 

is important remedies for management of tobacco caterpillar. 

They are stable and economically sound approaches to 

minimize the damage caused by Spodoptera litura (Fab). 

Therefore the present investigations were conducted to find 

out the influence of genotypes on consumption and growth 

of the pest. 


The present studies attempt to know the influence of 

mungbean genotypes and effect on consumability of leaves 

by the larvae and effect on growth and development of tobacco 

caterpillar, Spodoptera litura (Fab.) were carried out during 

Kharif season of 2008-09 and 09-10 in the Department of 

Entomology, College of Agriculture, Gwalior. 

Fifteen genotypes of mungbean viz, TJM 12, TJM 15, 

TJM 47, TJM 59, TJM 65, TJM 234, TJM 268, TM 98-50, TM 

99-58, TM 99-37, MI 24-91, MI 181-1, TARM 18, HUM 1 and K 

851 were grown in the plot size of 15.0 m 2 each with normal 

agronomical practices. The experiment was replicated three 

times. The leaves of each genotypes were picked up from the 

field and kept in petridishes after recording their weight with 

the help of electronic balance. The two first instar larvae were 

released in each petridish. The observation on weight of 

consumed leaves in each genotype was recorded at the time 

of changing the leaves. Fresh leaves were provided in two 

days interval starting from their release. Larval weight was 

recorded at 4, 8, 12 and 16 days after their released. The data 

on larval period, length of full grown larvae, pupal period and 

weight of pupae and adults were also recorded. Total leaves 

weight consumed by the larva, consumption index (C.I.), 

growth rate (G.R.), efficiency of conversion of ingested food 

(ECI) and digestibility were also calculated as per method 

suggested by Waldbauer, 1968. 


Observation recorded on weight of leaves consumed 

by larvae showed that the tested genotypes influence the 

consumption of leaves significantly (Table 1). Minimum 

consumption of leaves (7.680 g) was recorded in genotype 

TJM 15 which found significantly less than rest of the 

genotype, except TJM 12, TM 99-58, TJM 65, TM 98-50, TJM 

47 and K 851. On the other hand significantly higher 

consumption of leaves (10.180 g) was recorded in genotypes 

MI 24-91 which found significantly higher than rest of the 

genotypes except MI 181-1. Bhadauria, et al., 1998 and Saha, 

2005 also reported non-preference mechanism of resistance 

in soybean varieties against tobacco caterpillar. 

The weight gained by larvae reared on different 

genotypes was also significantly influenced by different 

genotypes. It ranged from 0.358 g in genotypes TJM 12 to 

0.815 g in genotypes TM 99-37. The minimum weight gained 

by larvae was recorded in genotypes TJM 12, which indicates 

the presence of antibiosis mechanism of resistance in the 

genotypes, TJM 12 which influence the growth of the larvae. 

The larval period of larvae fed on different genotypes of 

mungbean was differed significantly. The maximum larval 

period (23.0 days) was recorded in genotypes TJM 12 which 

found significantly higher than rest of the genotypes, expect 

TJM 99-58, TJM 65, TJM 59, TJM 15 and TM 98-50. Whereas 

the minimum larval period (16.0 days) was recorded in 

genotypes MI 181-1, which found significantly less than rest 

of genotypes expect TJM 234, MI 24-91 and TM 99-37. 

The larval length was also significantly influenced by 

different genotypes. It ranged from 2.70 cm in genotype TJM 

12 to 3.70 cm in genotype TARM 18. The minimum length of 

larvae was recorded on genotype TJM 12 followed by 

genotype TJM 15, which also indicates the influence of these

Table 1. 

S.N. 

SINGH et al., Influence of Varieties on Consumability of the Leaves, Growth and Development of Spodoptera litura (Fab.) 77 

Effect of the mungbean genotypes on the growth and development of S. litura (Fab.) 

Genotypes 

Weight of leaves consumed (g) Larval weight 

by larvae at 

(g) 

16 Days after release 

ECI-Efficiency of conversion of ingested food *Angular transformed values. 

Table 2. Effect of the mungbean genotypes on consumption, growth rate, ECI and approximate digestibility of S. litura (Fab.) 

genotypes on growth of larvae significantly. Whereas 

maximum length of larvae was recorded on genotypes TARM 

18. 

The pupal period were also influenced significantly by 

different genotypes of mungbean. It ranged from 7.0 to 10.0 

days, respectively. Maximum pupal period recorded in 

genotype TJM 12 showed their antibiosis effect against the 

pest. 

The data computed on consumption index (C.I.), growth 

rate (G.R.) efficiency of conversion of ingested food (ECI) and 

approximate digestibility (A.D.) also showed significant 

influence of different genotypes on consumability, digestibility 

and growth rate of pest (Table 2). 

The minimum consumption index (C.I.) of larvae 

recorded in TJM 15 followed by genotype TJM 12 indicated 

that this genotype influenced the consumability of leaves 

due to their anti-feedancy to the pest. The minimum value of 

Length of larvae 

(cm) 

growth rate efficiency of conversion of ingested food (ECI) 

and approximate digestibility (A.D.) in genotypes TJM 12 

showed their significant influence on digestibility of food and 

growth rate may be due to presence of antibiosis mechanism 

of resistance. The G.R., ECI and A.D. in different genotypes 

were ranged from 0.032 (TJM 12) to 0.075 (TM 99-37), 0.046 

(TJM 12) to 0.094 (TM 99-58), 40.04 (TJM 12) to 61.71 (TM 99- 

37). 


Larval period in 

days 

Pupal Period 

in days 

1. TJM 12 7.710 0.358 2.66 23.0 10.0 

2. TJM 15 7.680 0.649 2.90 21.6 7.3 

3. TJM 47 8.117 0.698 3.38 20.3 8.0 

4. TJM 59 8.377 0.623 3.30 21.6 7.6 

5. TJM 65 7.787 0.603 3.03 21.6 8.6 

6. TJM 234 9.370 0.745 3.36 16.3 7.6 

7. TJM 268 8.893 0.661 3.23 19.0 7.3 

8. TJM 98-50 7.950 0.605 3.16 20.6 8.3 

9. TJM 99-58 7.730 0.732 3.25 22.0 7.6 

10. TJM 99-37 8.757 0.815 3.43 17.6 7.0 

11. MI 24-91 10.180 0.721 3.40 17.3 7.3 

12. MI 181-1 10.127 0.649 3.50 16.0 8.3 

13. TARM 18 9.187 0.810 3.70 19.6 9.0 

14. HUM 1 8.693 0.767 3.40 19.6 8.0 

15. K 851 8.143 0.706 3.60 19.0 8.0 

S.Em(±) 0.193 0.057 0.11 0.8 0.4 

CD at 5% 0.558 0.165 0.32 2.4 1.2 

S.N. Genotypes Consumption index 

(C.I.) 

Growth rate of larvae 

(G.R.) 

Efficiency of conversion of 

ingested food (E.C.I.) 

Approximate 

Digestibility (%) 

1. TJM 12 0.712 0.032 0.046 41.3 (40.04)* 

2. TJM 15 0.710 0.059 0.084 44.5 (41.85) 

3. TJM 47 0.750 0.064 0.085 58.9 (50.16) 

4. TJM 59 0.774 0.057 0.074 55.2 (48.03) 

5. TJM 65 0.719 0.055 0.077 47.4 (43.54) 

6. TJM 234 0.866 0.068 0.079 57.5 (49.33) 

7. TJM 268 0.821 0.061 0.074 56.5 (48.73) 

8. TJM 98-50 0.734 0.056 0.076 50.1 (45.05) 

9. TJM 99-58 0.714 0.067 0.094 75.9 (60.66) 

10. TJM 99-37 0.809 0.075 0.092 77.5 (61.71) 

11. MI 24-91 0.941 0.066 0.070 80.3 (63.69) 

12. MI 181-1 0.936 0.060 0.064 64.2 (53.28) 

13. TARM 18 0.849 0.074 0.088 71.4 (57.72) 

14. HUM 1 0.803 0.070 0.087 66.9 (54.90) 

15. K 851 0.752 0.065 0.086 61.2 (51.48) 

S.Em(±) 0.018 0.006 0.006 0.72 

CD at 5% 0.052 0.016 0.017 2.10 

ECI-Efficiency of conversion of ingested food 

*Angular transformed values. 

Bhadauria, N.K.S., Bhadauria, N.S. and Jakhmola, S.S. 1998. Reaction 

of soybean varieties of bihar hairy cater pillar, Spilosoma obliqua. 

Indian J. Ento., 60(4):419-421. 

Saha, Lopamudra 2005. Reaction of soybean varieties against tobacco 

caterpillar, Spodoptera litura (Fabricious). M.Sc. (Ag.) Thesis 

J.N.K.V.V., Jabalpur (M.P.) 

Waldbauer, G.P. 1968. The consumption and utilization of food by 

insects. Advances in Insect Physiology, 5:229-283. 



Diversity Pattern of Beetles (Insecta: Coleoptera) In and Around the Historic 

Joysagar Tank of Assam, India 

I. RAHMAN 1 , C. SONOWAL AND M. NATH 

Department of Zoology, Sibsagar College, Joysagar 786 665, Assam, India 

email: imdadur123@rediffmail.com 1 

ABSTRACT 

In the present study 10 species of beetles belonging to 8 different 

families viz., Dytiscidae, Gyrinidae, Carabidae, Hydrophilidae, 

Chrysomelidae, Cocc inellida e, Cerambycidae and 

Tenebrionidae were collected and identified in and around the 

Joysagar Tank of the Sivasagar District, Assam. Dytiscidae 

was the predominant family with respect to number and 

abundance. 

Key words 

Coleoptera, Beetles, Insecta, Joysagar Tank 

The order Coleoptera or beetles forms the largest group 

of insects worldwide with about 3,70,000 described species. 

India being situated in tropics is well known for richness of 

coleopteran fauna. Beetles are found in almost every habitat 

and the size varies considerably. The smallest beetle Nanosella 

fungi from South and Central America, hardly measures 0.25 

mm long, while some of the large species often exceed 150 mm 

(Mani, 1994). About 15,088 species of coleopteran insects are 

known from Indian region (Kazmi and Ramamurty, 2004). The 

most important factor in the success of coleopterans is the 

development of forewings into hardened protective covering 

(elytra). Beetles are of variable shape and colour having 

chewing mouthparts belonging to different families. Some 

species have reduced wings (Arnett, 1973). Beetles are 

exceedingly variable both ecologically and biologically. Some 

Beetle species are extremely harmful while others have proven 

beneficial. If the beetle population is healthy in certain complex 

biological environments, we may gauge that the environment 

is also healthy. 

The study of coleopteran diversity in and around the 

Joysagar tank of the Sivasagar District, Assam is lacking 

completely. Therefore, in the present study an effort was made 

to study the diversity pattern of beetles in this area. 


Study area: The study was conducted at monthly interval 

from May, 2011 to April, 2012 at different zones of Joysagar 

tank. The tank is situated at 5 km distance from Sivasagar 

town. It lies between 26° 56' 97'’ and 26° 57' 32'’ N; 94° 37' 38'’ 

and 94° 37' 54'’ E and adjacent to Joy Dole. Joysagar tank is 

the largest man made tank in Asia, built by Ahom King Rudra 

Singha during 1696-1714 AD. It comprises an area of 318 acres 

including four banks. Its water level stays at 14 feet higher 

than ground level. This tank supports diverse flora and fauna. 

The study area was divided into four different habitat zones. 

Zone I - Area containing the men made building. 

Zone II - Dry with shrubs. 

Zone III - Area with man made garden and poultry farm. 

Zone IV - The area containing water bodies. 

Collection and identification of beetles: Beetles were collected 

from different habitats by using different collecting and 

trapping methods. Most beetles were collected by hand and 

with the help of forceps. Insect nets were employed for 

catching flying beetles. Water beetles were collected from 

four sites at four corners of the tank with the help of aquatic 

insect nets. Some beetles were collected during night with the 

help of light traps using light. The collected specimens were 

brought to the laboratory, pinned and identified with the help 

of standard identification manuals and published literatures. 

The collected water beetles were preserved in 4% 

formaldehyde. The specimens were identified with the help of 

available literature (Forey and Fitzsimons, 2007; Castner, 2008). 

Relative abundance (%) and dominance status were also 

determined. 


A total ten species of beetles were recorded during the 

present study. The checklist of beetles, their habitat is given 

in Table 1. These belong to 8 different families viz., Dytiscidae 

(Predaceous Diving Beetles), Gyrinidae (Whirligig Beetles), 

Carabidae (Ground Beetles), Hydrophilidae (Water Scavenger 

Beetles), Chrysomelidae (Leaf Beetles), Coccinellidae 

(Ladybird Beetles), Cerambycidae (Long-Horned Beetles) and 

Tenebrionidae (Darkling Beetles). Dytiscidae was the most 

common family quantitatively representing 43.29% of the total 

beetles in and around the Joysagar Tank. In the present study, 

three aquatic beetle species belonging to three different 

families viz., Dytiscidae, Gyrinidae and Hydrophilidae were 

recorded. In a man made pond Jana, et al., 2009 reported that 

order Odonata was the most common aquatic insect than order 

Coleoptera and order Hemiptera. Khan and Ghosh, 2001 in 

West Bengal and Sharma and Rai, 1991 in Bhagalpur, Bihar 

found Coleoptera to be the most common order quantitatively. 

Species diversity was found maximum in Zone III and 

minimum in Zone I. Zone II and Zone III were similar and other 

Zones were dissimilar in faunal composition which may due 

to presence of similar and dissimilar vegetation in that 

particular zone. 

The relative abundance of different beetle families

Table 1. 

RAHMAN, et al., Diversity pattern of beetles (Insecta: Coleoptera) in and around the historic Joysagar Tank of Assam, India 79 

Checklist of beetles and their habitat in and around the Joysagar Tank of the Sivasagar District, Assam. 

Suborder/Family/ Genus Occurrence Number Relative Abundance (RA) % *Dominance Status 

Suborder: Adephaga 

Family: Dytiscidae 

Eretes sticticus Zone IV 210 43.29 Eudominant 

Family: Gyrinidae 

Dineutus indicus Zone IV 143 29.48 Dominant 

Family: Carabidae 

Anthia sexguttata Zone II, III 11 2.26 Recedent 

Suborder: Polyphaga 

Family: Hydrophilidae 

Hydrophilus olivaceous Zone IV 56 11.54 Dominant 

Family: Chrysomelidae 

Dicladispa armigera 

Leptispa pygmaea Baly. 

Family: Coccinellidae 

Menochilus sexaculatus 

Epilachna dodecastigma 

Zone III 

Zone II, III 

Zone II, III 

Zone II, III 

*RA 31.7% = Eudominant 

8 

12 

11 

14 

1.65 

2.47 

2.26 

2.88 

Recedent 

Recedent 

Recedent 

Recedent 

Family: Cerambycidae 

Batocera rufomaculata Zone I 16 3.29 Subdominant 

Family: Tenebrionidae 

Tribolium castaneum (Herbst) Zone I, III 4 0.82 Subrecedent 

recorded in the study area is given in Fig. 1. Dytiscidae was 

numerically the most common family of beetles. Family 

Tenebrionidae was represented by only 1%. Out of the 18 

families of aquatic Coleoptera known from the world 

representative of 5 families namely Dytiscidae, Gyrinidae and 

Hydrophilidae, Haliplidae, and Elmidae are chiefly represented 

in the India (Deepa and Rao, 2010). Coleoptera belonging to 

40 taxa in 7 families of streams in the Nizke Beskydy Region 

(Slovakia) were studied (Zatovicova, et al., 2004). A total of 12 

species belonging to 5 different families of beetles viz., 

Gyrinidae, Tenebrionidae, Carabidae, Scarabaeidae and 

Meloidae were recorded from the vicinity of Semadoh-Makhala 

Road, Sipna Range, and Melghat Tiger Reserve (Thakare and 

Zade, 2012). 

Fig. 1. 

Relative abundance (%) of the families of beetles 

in and around the Joysagar Tank of the families of 

beetles in and around the Joysagar Tank of the 

Sivasagar District, Assam. 


The authors are grateful to the Head, Department of 

Zoology, Sibsagar College for providing laboratory facilities. 


Arnett, R.H. 1973. Beetles of the U.S.A. Manual for identification, 

American Entomological Institute, Ann Arbar MI. 

Castner, J.L. 2008. Photographic Atlas of Entomology and Guide to 

Insect Identification Published by Feline Press, USA. 

Deepa J. and Rao, C.A.N. 2010. Aquatic Entomofauna of Pocharam 

lake, Andhra Pradesh (Hemiptera & Coleoptera), Wetland ecosystem 

series, Rec. Zool. Surv. India, Kolkata, 13: 37-49. 

Forey, P. and Fitzsimons, C. 2007. Identification guides-European 

Insects, Flame Tree Publishing, London, pp. 194-283. 

Jana, S., Pahari, P., Dutta, T. and Bhattacharya, T. 2009. Diversity and 

community structure of aquatic insects in a pond in Midnapore 

town, West Bengal, India. J. Environ. Biol., 30(2): 283-287. 

Kazmi, S.I. and Ramamurthy, V.V. 2004. Coleoptera (Insecta) fauna 

from the Indian Thar Desert. Rajasthan. Zoos’ Print Journal, 19(4): 

1447-1448. 

Khan, R.A. and Ghosh, L.K. 2001. Faunal diversity of aquatic insects in 

freshwater wetlands of South Eastern West Bengal. Z.S.I. Kolkata. 

Mani, M.S. 1994. General Entomology. 3 rd edition, Published by Mohan 

Primlani for Oxford & IBH Publishing Co. Pvt. Ltd., New Delhi- 

01. 

Sharma, U.P. and Rai, D.N. 1991. Seasonal Variations and species 

diversity of coleopteran insects in a fish pond of Bhagalpur. Journal 

of Freshwater Biology, 3: 241-246. 

Thakare, V.G. and Zade, V.S. 2012. Diversity of Beetles (Insecta: 

Coleoptera) from the vicinity of Semadoh-Makhala Road, Sipna 

Range, and Melghat Tiger Reserve, (M.S.) India. Bioscience 

Discovery, 3(1):112-115. 

Zatovicova, Z., Ciampor, F. and Kodada, J. 2004. Aquatic Coleoptera 

(Insecta) of streams in the Nizke Beskydy Region (Slovakia): 

faunistics, ecology and comparison of sampling methods. Biologia, 

15: 181-189. 



Effect of Ginger and Mint Extract on Microflora of Yoghurt Culture 

DINKER SINGH 1 AND SWASHANKH KUMAR 

Department of Animal Husbandry and Dairying, Institute of Agricultural Sciences, Banaras Hindu University, 

Varanasi 221 005, U.P., India 

e-mail: dinker.bhu@gmail.com 1 

ABSTRACT 

The present research was conducted to study the effect of Ginger 

and Mint extract on the viability of yoghurt culture. The product 

was fermented with traditional yoghurt culture (Streptococcus 

salivarius Thermophilus sp., Lactobacillus delbrueckii, 

Bulgaricus,) and formulated with different levels of Ginger 

and Mint extract at 0 .5%, 1.0 %, 1.5% and 2.0% for 

experimental treatment T 1 

, T 2 

, T 3 

and T 4 

respectively with 10 0 

Brix. On the basis of selective enumeration, Ginger and Mint 

extract with the level of 1.5% in yoghurt has maximum viable 

count (cfu/g) followed by 1.0%, 2.0% and 0.5%. 

Key words 

Yoghurt, Herbal Extract, Yoghurt Cultures and Viable 

Count (cfu/g) 

Globally, yoghurt is one of the most popular fermented 

milk products and has gained widespread consumer 

acceptance as a healthy food due to its therapeutic properties 

beside its high nutritive value (Singh, 2013). Yoghurt contains 

substantial amounts of live lactic acid bacteria (Ghosh and 

Raymond, 2003). Gilliland, 1979, reported that yoghurt can 

inhibit the growth of certain types of tumor. Yoghurt contains 

substantial amounts of live lactic acid bacteria (Ghosh and 

Raymond, 2003) and showed the health benefits for certain 

gastrointestinal conditions, including lactose intolerance, 

constipation, diarrheal diseases, colon cancer, inflammatory 

bowel disease, Helicobacter pylori infection, and allergies 

(Adolfsson, et al., 2004). 

In the present scenario, the herbal products are gaining 

more popularity over synthetic products in the world market. 

This is occurring due to some side effect of synthetic products 

on the body (Amirdivani, 2008). In spite of well-practiced 

knowledge of herbal medicines and occurrence of a large 

number of medicinal plants, the share of India in the global 

market is not up to the mark (Rajkumar, 2009). 

Ginger (Zingiber officinale) has beneficial effects on 

the digestive system, enhancing gastrointestinal motility, and 

is used traditionally for the treatment of stomach ache, 

vomiting and indigestion (Yamahara, et al., 1990). 

Mint (Mentha arvensis) isused to increase the flow of 

digestive juices and bile while relaxing the muscles of the 

digestive tract. Mint is also used to tone the stomach, stimulate 

the mind and body, rid the intestines of gas, and relieve muscle 

spasms (Ingham, et al., 1995). 

The objective of the present study was to assess the 

viability of yoghurt culture with best level of above herbal 

extract for the preparation of improved herbal yoghurt. 


Yoghurt Culture (Streptococcus salivarius Thermophilus 

NCDC 074 and Lactobacillus delbrueckii Bulgaricus NCDC 

009) were obtained from National Collection of Dairy Culture 

(NCDC), Dairy Microbiology Divisionat NDRI Karnal, Haryna, 

India. 

Herbs (Ginger and Mint) were obtained from the local market 

in Allahabad, U.P., India. 

Selection of Herbal Extract 

Herbs viz., Ginger and Mint extract of 10 0 brix were 

selected to enhanced the viability of yoghurt culture by the 

application of Disc Diffusion method (Baur, et al., 1966). 

Procedure Adopted for Manufacturing Frozen Yoghurt 

Both control (T 0 

) and experimental (T 1 

,T 2 

,T 3 

and T 4 

) 

frozen yoghurt mix were standardized to 5.0 % fat, 8.7% serum 

solids, 12% sugar, stabilizer and emulsifier 0.5% and total solids 

adjusted to 26%. Herbal extract (10 0 brix) was added at 0.0%, 

0.5%, 1.0%, 1.5% and 2.0% for treatments T 0 

, T 1 

, T 2 

, T 3 

and T 4 

respectively. 

The mix was homogenized and then pasteurized and 

cooled to 42 o C and yoghurt starter culture was added at 2.5% 

(1:1 ratio). The mix was incubated at 42 o C till we achieve an 

acidity of 0.45%. Before the mix is aged at 5 o C and frozen in a 

batch freezer to an overrun of 70%. The yoghurt was filled in 

cups and stored at refrigerated temperature. 

Analysis of Yoghurt Culture 

Selective enumerations of yoghurt culture 

(Streptococcus salivarius Thermophilus NCDC 074, 

Lactobacillus delbrueckii Bulgaricus NCDC 009) were done. 


The highest mean selective enumeration (10 7 cfu/g) of 

Streptococcus Thermophilus scorewas 60 in group T 3 

followed 

by 35, 22, 6.6 and 2.1 in groups T 2 

, T 4 

, T 1 

and T 0 

respectively 

(Fig. 1). The differences in the values were significant (P< 

0.05), except groups T 0 

-T 1 

. 

The highest mean selective enumeration (10 7 cfu/g) of 

Lactobacillus, Bulgaricusscore was 26.9in group T 3 

followed 

by 18.3, 14.7, 13.8 and 2.5in groups T 2 

, T 4 

, T 1 

and T 0 

respectively (Fig.1). The differences in the values were

SINGH AND KUMAR, Effect of Ginger and Mint extract on Microflora of yoghurt culture 81 

significant (P< 0.05), except groups T 1 

-T 4 

, and T 2 

-T 4 

. 

The highest mean selective enumeration(10 7 cfu/g) of 

sum of yoghurt culture (Lactobacillus, Bulgaricus+ 

Streptococcus, Thermophilus) count was 89.8 in group T 3 

followed by 53.3, 36.7, 20.4 and 4.6 in groups T 2 

, T 4 

, T 1 

and T 0 

respectively (Fig. 2). The differences in the values were 

significant (P< 0.05). 

Rajkumar, 2009, reported that ginger extract and honey 

with cryoprotective agent (Glycerol) enhanced the viable cell 

count of frozen yoghurt. There is an adverse effect on total 

viable count of frozen yoghurt culture during storage due to 

death of cells (Donkor, et al., 2006; Singh, 2009). By analysing 

different levels of herbal extract we have found that viable 

count of frozen yoghurt was enhanced by the use of herbal 

extract. 

Singh, 2009, suggested that herbal extract (Ajowan, 

Cardamom, Garlic and Mint) enhances the growth of yoghurt 

culture. During storage the death of yoghurt culture was found 

non-significant and herbal extract can be recommended as 

growth promoter (Heenan, et al., 2004; Rajkumar, 2009; Radhika, 

2010) for the yoghurt culture. 

Chowdhury, et al., 2008, prepared yoghurt with tulsi 

leaf (Ocimum sanctum), pudina leaf (Mint, Mentha arvensis) 

and coriander leaf (Coriandrum sativum) based. Tulsi yogurt 

had the maximum b-galactosidase activity than other herbal 

yoghurt. 

By analysing different levels of Ginger and Mint extract 

we found that viable count of yoghurt cultures i.e. 

Streptococcus salivarius Thermophilus NCDC 074 and 

Lactobacillus delbrueckii Bulgaricus NCDC 009 was highest 

for treatment containing 1.5% (T 3 

) herbal extract followed by 

Fig. 1. Selective Enumeration of Yoghurt Culture (10 7 cfu/g) 

Fig. 2. Sum of viable count of yoghurt culture. 

1.0% (T 2 

), 2.0% (T 4 

) and 0.5% (T 1 

) in yoghurt. The Ginger and 

Mint extract increases the growth of yoghurt culture and we 

recommend that herbal extract can be used as growth promoter 

for yoghurt culture. 


Adolfsson, O., Meydani, S.N. and Russell, R.M. 2004. Yogurt and gut 

function, American J. Clinical Nutrition, 80:245-56. 

Amirdivani, S. 2008. Inclusion of MenthapiperitaAnethumgraveolence 

and Ocimumbasilicumin Yogurt and their effect on the Inhibition 

of Enzyme Relevant to Hypertension and type-2 Diabetes.M. Sc. 

Thesis, Department of Biochemistry, Institute of Biological Science, 

University of Malaya 50603. 

Bauer, A.W., Kirby, W.M., Sherris, J.C. and Turck, M. 1966. Antibiotic 

susceptibility testing by a standardized single disc method. American 

J. Clinical Pathology. 45:493-496. 

Chowdhury, B.R., Chakraborty, R. and Raychaudhuri, U. 2008. Study 

on b-galactosidase enzymatic activity of herbal yogurt. Int. J. Food 

Sci Nutr., 59(2): 116-22. 

Donkor, O.N., Henriksson, A., Vasiljevic, T. and Shah, N.P. 2006. 

Effect of acidification on the activity of probiotics in yoghurt 

during cold storage. Int. Dairy J., 16(10): 1181-1189. 

Ghosh, S. and Raymond, J.P. 2003. Bioactive natural compounds for 

the treatment of gastrointestinal disorders. Clinical Science, 10(4): 

547-556. 

Gilliland, S.E. 1979. Beneficial interrelationships between certain 

microorganisms and humans: candidate microorganisms for use as 

dietary adjuncts. J. Food Protection, 42:164. 

Heenan, C.N., Adams, M.C., Hosken, R.W. and Fleet, G.H. 2004. Survival 

and sensory acceptability of probiotic microorganisms in a 

nonfermented frozen vegetarian dessert. Food Sci. and Tech., 37(4): 

461-466. 

Ingham, K.E., Linforth, R.S.T. and Andrew, J.T. 1995. The effect of 

eating on the rate of aroma release from mint-flavoured sweets. 

Food Sci. Tech., 28(1): 105-110. 

Radhika. 2010. Study of herbal based probiotic yoghurt. M.Sc. Thesis. 

Department of Animal Husbandry and Dairying, Raja Balwant Singh 

College, Agra, Uttar Pradesh, India. 

Rajkumar, B. 2009. Efficacy of growth factor (Ginger extract & honey) 

and cryoprotective agent (Glycerol) on viable cell count of frozen 

yoghurt. M. Tech Thesis. College of Food and Dairy Technology, 

Allahabad Agricultural Institute-Deemed University, Allahabad, 

Uttar Pradesh, India. 

Singh, J. 2013. Technological and nutritional view of yoghurt.IBDC 

Publishers, Lucknow. 

Singh, R. 2009. Study of herbal extract on viability of yoghurt culture 

during storage. Int. J. Food Sci. Nutr., 61(2): 112-26. 

Yamahara, J., Huang, Q., Li, Y., Xu, L. and Fujimura, H. 1990. 

Gastrointestinal Motility Enhancing Effect of Ginger and its Active 

Constituents, Chem. Pharm. Bull., 38(2):430-1. 

Yamahara, J., Mochizwki, M., Huang, Q.R., Matsuda, H. and Fujimura, 

H. 1988. The anti-ulcer effect in rats of ginger constituents. J. 

Ethnopharmacology, 23(2-3): 299-304. 



Anti-Fungal Evaluation of Calotropis Leaf Extract against Some Plant Pathogenic 

Fungi 

RAJNI SINGH SASODE 1 AND PRADYUMN SINGH 

Department of Plant Pathology, College of Agriculture, Gwalior, M.P. 

Rajmata Vijayaraje Scindia Krishi Vishwa Vidyalaya, Gwalior, M.P. 

e-mail: rnikum_ujjain@yahoo.co.in 1 

ABSTRACT 

Four forms of Calotropis leaf extracts viz.,crude (10%), powder 

(10%), boil (10%) and ethanol (1%) were screened for antifungal 

activity against ten test fungal pathogens by poisoned food 

technique. All the form significantly inhibited the growth of 

the test fungus. The antifungal activity of boil extract @ 10% 

was found more effective than powder, crude and ethanol 

extracts against seven fungal pathogens viz. Fusarium 

oxysporum f.sp. ciceri, Rhizoctonia solani, R. bataticola, 

Colletotrichum gleosporioides, Tolyposporium penicillariae, 

Sclerotinia sclerotiorum ad Phoma sorghina. For the control of 

Fusarium oxysporium f.sp. pallidoroseum, and Sclerotium rolfsii, 

the powdered form of Calotropis leaf extract was found more 

effective. It was observed that the effectivity of boiled, powdered 

and ethanol extract against the respective pathogenic fungi 

was gradually increased with the increase in the concentration 

from 5 to 20%. They are further compared with chemicals viz., 

carbendazim (0.1%) and mancozeb (0.2%), it was observed that 

both the chemicals were found more effective than the Calotropis 

leaf extract (boiled/crude/powder/ethanol). Therefore higher 

concentration of different forms of Calotropis leaf extract should 

be tested under in vitro condition and the effective one may 

also be tested in the field as an alternative to the chemical for 

the eco-friendly management of the disease. 

Key words 

Leaf, boiled, powdered and ethanol extracts, test 

pathogen, chemicals, antifungal activity. 

Calotropis procera (Ait.) R.Br. commonly known, as 

‘Arka’ is a popular medicinal plant found throughout the 

tropics of Asia and Africa covering the arid to semiarid regions 

of the Caribbean, Central America, South America, Africa, India 

and Israel. It is a woody, broadly evergreen shrub (2 m or less, 

sometimes 5 m tall), belongs to family Asclepiadaceae. It 

contains active ingredients such as alkaloids, enzymes and 

other inorganic elements. Botanicals are gaining importance 

in crop protection in view of their selective properties, low 

cost and safety to ecosystem. Many botanicals have been 

identified to be effective in the control of plant diseases. 

Among the 5280 species tested, 1134, 346 and 92 plant species 

possessed insecticidal, fungicidal and bactericidal properties 

respectively (Ahmed and Grainge, 1982). Plant extracts and 

essential oils are known to prossess antifungal activity against 

a wide range of fungi (Grane and Ahmad, 1988, Wilson, et al., 

1997 and Abd-Alla, et al., 2001). Plant metabolites and plant 

based pesticides appear to be one of the better alternatives as 

they are known to have minimal environmental impact and 

danger to consumer in contrast to synthetic pesticides (Verma 

and Dubey, 1999). Therefore, the development of bio-pesticides 

has been focused as a viable disease control strategy. In view 

of these the present investigation was undertaken to screen 

for the efficacy of antifungal potency of Calotropis leaf extract 

against important phytopathogenic fungi. 


The selected ten fungal pathogens viz., Rhizoctonia 

solani, Phoma sorghina, Colletotrichum gloeosporioid, 

Fusarium oxysporum f.sp. pallidorosem, Fusarium 

oxysporum f.sp. ciceri, Rhizoctonia bataticola, Sclerotium 

rolfsii, Sclerotinia sclerotiorum, Alternaria cyamopsidis and 

Tolyposporium penicillariae were isolated from infected host 

and the selection of media is based on the standard 

recommendation for culturing these fungi. 

Preparation of extracts 

(i) 

(ii) 

(iii) 

Powdered extract: Thoroughly washed fresh plant 

leaves are oven dried at 60 0 C for two consecutive days. 

After drying the leaves were easily crushed by mixer. 

The powder of crushed leaves was stored in the airtight 

plastic bottles. The powder was used at the 

concentration of 10% for this 10 gm powder was 

incorporated into 50 ml of distilled water then it was 

kept for 24 hours. Thereafter, it was filtered into a 

measuring cylinder and 500 ml volume of the extract was 

maintained by adding the water and finally it was 

incorporated into the PDA flask containing concentrated 

potato dextrose agar media. Thus, after incorporation of 

500 ml extract (20%) in 500 ml PDA the final concentration 

of the powdered extract into the PDA was 10%. 

Fresh extracts (Crude): Fresh plant leaves (50g) 

thoroughly washed with running tap water was macerated 

with 50 ml sterile water in a warning blender for 10 

minutes. The macerate was first filtered through double 

layered muslin cloth and centrifuged at 4000g for 30 

minutes the supernatant was filtered through Whatman 

No.1 filter paper and sterilized at 120°C for 30 minute. 

The extract was preserved aseptically in a brown bottle 

at 5°C until further use (Satish, et al., 1999). The obtained 

extract served as the crude extract (100% concentration) 

Boiled extracts: The fresh leaves of Calotropis were 

dried in shadow, washed, weighted and boiled for two

SASODE AND SINGH, Anti-Fungal Evaluation of Calotropis Leaf Plant Extracts Against Some Plant Pathogenic Fungi 83 

(iv) 

hours, thereafter it was filtered and water were 

incorporated into it to maintained 1:1 weight/volume 

basis. The extract was stored and used for bioassay of 

test fungus at the concentration 10%. 

Ethanol extracts: 100 gm powder of Calotropis leaves 

was incorporated into flask containing 500 ml of ethanol 

then it was kept open on hot plate at the temperature of 

42 0 C±2 for the evaporation of ethanol after then the crust 

of the extract remained in the flask was scraped, weighted 

and used for bioassay @ 1% concentration. 

For standardization of the concentration of the effective 

form the crude, boil and powdered extracts were used @ 5, 10, 

15 and 20% concentrations while the ethanol extract was used 

@ 1, 2, 3 and 4% concentrations, respectively. 

The plant extract were amended aseptically to melted 

potato dextrose agar medium in appropriate proportions and 

sterilized at 121°C, 15Ib/inch 2 pressure for 15 minutes and 

allowed to cool. Twenty ml of the medium is poured in each 10 

cm diameter Petridish and solidified. One disc (7 mm) of the 

medium containing fungal culture of the pathogen was cut 

from the 7 days old culture and was transferred in the centre 

of the Petridish under aseptic condition. The plates were then 

incubated at temperature 28±1 o C. Average growth of the 

pathogen colony was measured 5 days after incubation. 

Petridishes containing media devoid of the plant extract but 

with the same amount of distilled water served as control. 

Four replicates were maintained for each treatment. 

All the leaf extract forms were subjected to antifungal 

activity assay with crude (10%), powdered (10%), boil (10%) 

and ethanol (1%) extracts and was further subjected to 

poisoned food technique. Among all the tested extracts the 

effective extract against the respective pathogens were 

selected and again subjected to different four concentration 

of boil, powder (5, 10, 15 and 20%) and ethanol at (1, 2, 3 and 

4%) to find out its appropriate form respectively. 

The effective forms under different concentrations (boil 

and powder at 15 and 20% and ethanol at 3and 4%) were 

further evaluated against test pathogens. On the basis of 

significance, the concentrations of respective forms were 

standardized against the pathogens and were also compared 

with the recommended chemicals viz., carbendazim (0.1%) and 

mancozeb (0.2%), under in-vitro condition. 


Result presented in the Table 1 shows that all the four 

forms of Calotropis leaf extracts significantly inhibited the 

growth of fungal mycelium but none of them could absolutely 

inhibited the growth. However, the boil form was found 

effective against seven phytopathogenic fungi viz., Phoma 

sorghina(21.00), Fusarium oxysporium f.sp. ciceri (23.24), 

Tolyposporium penicillariae (25.00), Colletotrichum 

gleosporioides (27.50), Sclerotinia sclerotiorum (30.00), 

Rhizoctonia solani (32.50), and Rhizoctonia bataticola 

(34.50). The powder form also effectively inhibited the growth 

of Fusarium oxysporum f.sp. pallidorosem (45.50) and 

Sclerotium rolfsii (35.50). The ethanol form of Calotropis leaf 

extract was found more effective than crude, boil, powder 

extract form for the growth inhibition of Alternaria 

cyamopsides (26.50). In respect of growth inhibition the boil 

extract was significantly superior over the other forms. 

Powdered extract was significantly superior over crude and 

ethanol extract. Ethanol extract was found least effective but 

it was statistically at par with crude extract. 

Results presented in the Table 2 reveal that all the four 

concentrations significantly inhibited the growth of all the 

fungal pathogens but none of the concentrations absolutely 

inhibited the growth, however the effectivity against all the 

above tested fungal pathogens increases with the increase in 

the concentration from 5 to 20%. The growth of Fusarium 

oxysporum f.sp. ciceri and Rhizoctonia solani under 20% 

concentration of boiled leaf extract was 8 mm and 7 mm, 

Table 1. Efficacy of different forms of Calotropis leaf extracts against test pathogens 

S.No. Treatments 

Radial growth of fungal mycelium (mm) 

Fusarium oxysporium 

f.sp. ciceri 

F.o. f.sp. 

pallidoroseum 

Rhizoctonia solani 

1 Powder @ 10% 46.50 45.50 50.50 44.50 43.50 60.50 33.25 33.50 40.00 37.00 

2 Crude @ 10% 54.50 61.50 41.00 56.00 65.50 53.00 64.00 62.50 61.50 30.00 

3 Boil @10% 23.24 52.00 32.50 34.50 27.50 33.50 25.00 54.00 30.00 21.00 

4 Ethanol @ 1% 60.00 51.65 55.00 63.50 69.00 26.50 60.00 56.00 70.00 50.00 

5 Control 74.00 76.50 82.00 77.00 83.00 71.50 73.00 80.00 80.50 60.00 

SE(m)± 

CD at 5% 

1.553 

4.839 

1.438 

4.481 

1.171 

3.648 

R. bataticola 

1.271 

3.959 

Colletorichum 

gleosprioides 

1.084 

3.378 

Alternaria 

cyamopsides 

1.309 

4.072 

Tolyposporium 

penicillariae 

0.536 

1.670 

Sclerotium rolfsii 

0.841 

2.620 

Sclerotinia 

sclerotiorum 

1.094 

3.407 

Phoma sorghina 

1.676 

5.221


Table 2. 

Comparative efficacy of powdered extracts of 

Calotropis leaf under four concentrations against 

test pathogens 

Treatments 


Fusarium oxysporum f. sp. 

pallidoroseum 

Sclerotium 

rolfsii 

Powder leaf extract @ 5% 47.00 64.00 




Control 72.00 79.00 

SE(m)± 

CD at 5% 

1.463 

4.559 

1.571 

4.893 

and Singh, 2004. Calotropis in form of the ethanol extract 

(Table 4) was found more effective than the other forms for 

the inhibition of Alternaria cyamopsides under in vitro 

condition. 

On the basis of significance, the effective concentrations 

of respective forms were standardized against the pathogens 

and were also compared with the recommended chemicals 

under in-vitro condition. The results (Table 5) reveal that the 

growth of Fusarium oxysporum f.sp. ciceri (19.50), 

Rhizoctonia solani (22.00), Rhizoctonia bataticola (9.00), 

Colletotrichum gleosporioides (13.00), Tolyposporium 

penicillariae (0.00) Sclerotinia sclerotiorum (19.50) and 

Table 3. Comparative efficacy of boil extracts of Calotropis leaf under four concentrations against test pathogens 

Treatments 


Fusarium oxysporum 

f.sp. ciceri 

Rhizoctonia solani 

R. bataticola 

Colletorichum 

gleosprioides 

Tolyposporium 

penicillariae 

Sclerotinia 

sclerotiorum 


Boil leaf extract @ 5% 48.00 58.00 41.50 60.00 43.00 54.00 55.00 




Control 81.00 72.00 75.00 70.00 70.00 75.00 72.00 

SE(m)± 

CD at 5% 

1.408 

4.387 

1.326 

4.131 

1.271 

3.961 

1.678 

5.229 

1.461 

4.550 

1.618 

5.040 

1.483 

4.621 

respectively as compared to 81 and 72 mm of control, while 16 

and 25 mm fungal growth of the above two pathogens was 

recorded under its 15% concentration. 20% showed significant 

inhibition over 15%, this indicate that there is a need to test 

the higher concentrations (>20%) of Calotropis leaf extract 

under boil form to obtain complete inhibition of fungal growth 

against Fusarium oxysporum f.sp. ciceri and Rhizoctonia 

solani, similarly 20% concentration of Calotropis leaf extract 

under boil form showed significant inhibition of Rhizoctonia 

bataticola, Colletotrichum gleosporioides and 

Tolyposporium penicillariae over its 15% concentration. 

These also reflect to go for its higher concentration to obtain 

the complete/better inhibition. Though the minimum growth 

of Sclerotinia sclerotiorum and Phoma sorghina was recorded 

under 20% concentration, but it was statistically at par with 

its 15% concentration. This supports the use of 15% 

concentration of boiled extract instead of 20% for the control 

of Sclerotinia sclerotiorum and Phoma sorghina. 

The results presented in Table 3 reveal that the 

fungitoxicity of powder form of Calotropis leaf @ 20% was 

significantly higher over its 15% concentration for the 

inhibition of Fusarium oxysporum f.sp. pallidoroseum and 

Sclerotium rolfsii. This indicate that higher concentration 

(>20%) may show complete inhibition of the growth. The above 

finding is supported by the work of Singh and Chand, 2004 

Phoma sorghina (9.50) effectively inhibited by the boiled 

extract at 20% of Calotropis leaf but the boiled extract even at 

20% concentration is significantly less effective than 

carbendazim (0.1%) and mancozeb (0.2%). It means that the 

boil extract @ 20% concentration may not act as an alternative 

to the carbendazim (0.1%) and mancozeb (0.2%), however as 

per the effectively trend it seems that they may act as an 

alternative source to the chemical under much higher 

concentration. 

Like boil extract the powder extract also effectively 

inhibited the growth of Fusarium oxysporum f.sp. 

pallidoroseum (25.00) and Sclerotium rolfsii, (12.50) at 20% 

concentration but these extracts could not show absolute 

Table 4. 

Comparative efficacy of ethanol extracts of 

Calotropis leaf under four concentrations against 

Alternaria cyamopsides 

Treatments 


Ethanol leaf extract @ 1% 53.00 




Control 75.00 

SE(m)± 

CD at 5% 

1.012 

3.154

SASODE AND SINGH, Anti-Fungal Evaluation of Calotropis Leaf Plant Extracts Against Some Plant Pathogenic Fungi 85 

Table 5. 

Comparison of in vitro efficacy of different forms of Calotropis leaf extracts with chemicals against test pathogens 

Fusarium 

oxysporium f.sp. 

ciceri 

F.o. f.sp. 

pallidoroseum 

Rhizoctonia 

solani 

Powder leaf extract @ 15% N.E 32.00 N.E N.E N.E N.E N.E 24.00 N.E N.E 

Powder leaf extract @ 20% N.E 25.00 N.E N.E N.E N.E N.E 12.50 N.E N.E 

Boil extract @ 15% 29.00 N.E 28.00 24.00 22.00 N.E 12.75 N.E 30.50 20.50 

Boil extract @ 20% 19.50 N.E 22.00 13.00 13.00 N.E 0.00 N.E 19.50 9.50 

Ethanol leaf extract @3% N.E N.E N.E N.E N.E 31.00 N.E N.E N.E N.E 

Ethanol leaf extract @ 4% N.E N.E N.E N.E N.E 14.00 N.E N.E N.E N.E 

Mancozeb @ 0.2% 00.00 0.00 0.00 9.00 4.50 8.00 0.00 0.00 0.00 0.00 

Carbendazim @ 0.1% 00.00 0.00 7.00 00.00 2.50 0.00 8.00 2.50 0.00 8.50 

Control 80.00 87.00 74.00 80.00 78.00 80.00 80.00 64.00 74.00 78.00 

SE(m)± 

CD at 5% 

2.167 

6.751 

1.413 

4.403 

1.661 

5.174 

R. bataticola 

1.620 

5.048 

Colletorichum 

gleosprioides 

0.980 

3.052 

Alternaria 

cyamopsides 

1.095 

3.413 

Tolyposporium 

penicillariae 

1.220 

3.802 

Sclerotium 

rolfsii 

0.801 

2.495 

Sclerotinia 

sclerotiorum 

1.777 

5.537 


1.114 

3.472 

inhibition and were significantly inferior to carbendazim (0.1%) 

and mancozeb (0.2%), though the effectiveness of powder 

extract gradually increased in the concentration from 5 to 20%. 

It indicates that the absolute inhibition in the growth of above 

test fungi may be achieved under > 20% concentration of 

powder form of Calotropis leaf extract. 

The ethanol form of Calotropis leaf extract was found 

more effective than the crude, boil and powder extract form 

for the growth inhibition of Alternaria cyamopsides (14.00) at 

4% concentration, but like boil, powder extract it could not 

show the absolute inhibition while the carbendazim (0.1%) 

completely inhibited the growth. 

Therefore in the direction of eco-friendly management 

there is a need to test the extracts against the fungal pathogens 

under much higher concentration so that possibilities may be 

created for the use of the above forms of Calotropis extracts 

as an alternative to the chemical for the management of above 

fungal pathogens. 


Abd-Alla, M.S., Atalla, K.M. and Elp-Sawi, M.A.M. 2001. Effect of 

some plant waste extracts on growth and aflatoxin production by 

Aspergillus flavus. Annals Agric. Sci., Ain Shams Univ., Cairo., 46: 

579-592. 

Ahmed, N. and Grainge, M. 1982. Some promising plants for pest 

control under small scale forming operation in developing countries 

working paper resource system institute, East-West Centre, 

Honolulu, Hawai, USA. pp.55. 

Grane, M. and Ahmad, S. 1988. Handbook of Plants with Pest Control 

Properties. John Wiley and Sons, New York. 

Satish, S., Raveesha, K.A. and Janardhana, G. R. 1999. Antibacterial 

activity of plant extracts on phytopathogenic Xanthomonas 

campestris pathovars. Letters in Applied Microbiology, 28: 145- 

147. 

Singh, P.P. 2004. Management of Alternaria blight of mustard by use of 

botanicals. M.Sc. (Ag) Thesis, JNKVV, Jabalpur. 

Singh, Surendra and Chand, Hari 2004. Effect of extracts of some 

medicinal plants on spore germination of chickpea wilt pathogen 

(Fusarium oxysporum f.sp. (Pad.) Snyd. and Hans.). Indian J. Pl. 

Prot., 32(1): 162-163. 

Verma, J. and Dubey, N.K. 1999. Prospecdtives of botanical and 

microbial products aspesticides of tomorrow. Current Science, 76: 

172-179. 

Wilson, C.L., Solar, J.M., Ghaouth, A.El. and Wisniewski, M.E. 1997. 

Rapid evaluation of plant extracts and essential oils for antifungal 

activity against Botrytis cinerea. Pl. Dis., 81: 201-210. 



Studies on Status of Leaf Spot of Groundnut in Chunar and Nearby Areas of Mirzapur 

District of Uttar Pradesh 

BHARAT CHANDRA NATH, R.B. SINGH AND LAXMAN PRASAD BALAI 

Department of Mycology and Plant Pathology, Institute of Agricultural Sciences, Banaras Hindu University, 

Varanasi 221 005, India 

e-mail: bharatpal05@gmail.com 

ABSTRACT 

Roving surveys conducted from 2009-10 and 2010-11 kharif 

season (August to October) indicated that groundnut plants in 

Chunar and its neighbouring areas of Mirzapur district of Uttar 

Pradesh averaged 40 and 39% in two years, respectively and 

groundnut crops averaged 13% loss due to late leaf spot (LLS) 

fungus, Phaeoisariopsis personata. The late leaf spot disease 

was more prevalent than early leaf spot (ELS) caused by 

Cercospora arachidicola in these areas. Intensity of the disease 

in different areas surveyed, ranged from 13 to 22% in the month 

of September and 35 to 45% in October during 2009. In the 

year 2010 disease intensity ranged from 13 to 24% in September 

and 33 to 42% in October respectively. 

Key words 

Leaf spat, survey, cercospora, disease intensity 

Groundnut (Arachis hypogea L.) also known as peanut 

and native to South America, it is grown over an area of 8 

million ha with an annual production of 7.5 million tonnes. A 

production of 83.32 lakh tonnes has been recorded during 

2003-04 (Hegde, 2005). The major states that grow this crop 

are Gujarat, Andhra Pradesh, Karnataka, Tamil Nadu, 

Maharashtra, Rajasthan, Madhya Pradesh, Uttar Pradesh and 

Orissa. Diseases, pests and unpredictable rainfall patterns 

are one of the major constraints in achieving high production 

of groundnut. A large number of diseases attack groundnut in 

India (Mayee and Dater, 1988). The majority is caused by 

fungi and several of them are yield reducer in certain regions 

and seasons. There are a few economically important foliar 

fungal diseases, such as early (Cercospora arachidicola) 

and late leaf spots (Phaeoisariopsis personata (Berk and Curt 

v. Arx,) commonly called as tikka diseases and groundnut 

rust. As the area under groundnut is predominant in kharif 

(rainy) season (81%), the foliar diseases like late leaf spot and 

rust may cause yield losses up to 50% in the semi - arid tropics. 

In India, late leaf spot is more severe than early leaf spot 

(Ghewande, 1990; Anon., 1993). It causes severe defoliation 

and reduces pod yields by more than 50% if the crop is not 

protected with chemicals (Shew, et al., 1988). The symptoms 

are characterised by circular necrotic lesions of 1-6mm diameter 

on both surfaces of leaf, later become dark brown to black 

without any yellow halo (Rangaswami and Mahadevan, 2006). 

Yield losses of about 10 per cent kernels have been estimated 

from the southern USA where fungicide application is normally 

practiced. Over much of the semi-arid tropics where chemical 

control of leaf spots is rarely practiced, losses in excess of 

50% are common (Jackson and Bell, 1969; Garren and Jackson, 

1973). In India, losses in yield due to the leaf spots have been 

estimated to be in the range of 15 to 59% (Sulaiman, 1966; 

Ramakrishnan and Rao, 1968 and Chohan, 1974). Sulaiman, 

1966 also recorded a reduction in groundnut yield of 40% due 

to leaf spot in Maharashtra and Siddaramaiah, et al., 1977 

found a loss of more than 50% in Karnataka. Sundaram, 1965 

observed a loss up to 22% in yield compared plots receiving 

fungicide sprays. Hegde, et al., 1995 observed that the late 

leaf spot disease of groundnut appeared at 55 to 60 days after 

sowing and would cause more than 50% loss in pod and haulm 

yield in groundnut producing areas of Karnataka. Keeping in 

view the above facts, a survey was conducted to elucidate 

the status of leaf spot of groundnut disease. 


Groundnut fields were randomly roving survey of Chunar 

and nearby areas of Mirzapur District of Uttar Pradesh were 

conducted for the occurrence and severity of of Cercospora 

leaf spot. Survey was conducted during kharif season 

(August-October) at fourteen locations of the area in the year 

2009-10 and 2010-11. For assessing disease intensity five fields 

from each location, situated 500 m apart from each other, were 

selected. From each field ten plants were selected to calculate 

disease intensity. Disease scoring was done based on the 1-9 

point scale was adopted for recording the severity similar to 

field screening of groundnut genotypes for resistance to late 

leaf spot (Subrahmanyam, et al., 1995). Thirty leaves were 

collected from 10 randomly selected plants from top, middle 

and bottom of the crop canopy in each field and scored 

individually taking into account the leaf area damaged by the 

disease and the extent of defoliation in the plants where 1 = 

0%, 2 = 1-5%, 3 = 6 -10%, 4 = 11-20%, 5 = 21-30%, 6 = 31-40%, 

7 = 41-60%, 8 = 61-80%, 9 = 81- 100% disease severity. The per 

cent disease severity index (PDI) was then calculated. Disease 

rating was done on each trifoliate leaves on selected plants. 

Later these ratings were converted into percent disease index 

(PDI) by using the following formula. 

Sum of individual rating scale 

Per cent disease index (PDI) = —————————— x 100 

No. of disease plant observation × maxi. dis. rating 

Leaves showing typical Cercospora leaf spot symptom 

were collected and isolation were made under laboratory 

conditions.

NATH, et al., Studies on status of leaf spot of groundnut in Chunar and nearby areas of Mirzapur 87 


Groundnut crop grown in Chunar and nearby areas was 

infected by Phaeoisariopsis personata and produced typical 

Cercospora leaf spot that was noticed in the 4 th week of August 

in both the year 2009 and 2010. The severe incidence of the 

disease was seen in last week of September to October. The 

symptom of the disease appeared mainly on the leaf, but on 

later stage disease spread to stem portion also. The disease 

first appeared on upper surface of the lower leaf and then 

moved towards up in favorable conditions. In severe 

conditions defoliation of leaves occurs. Late leaf spot caused 

by P. personata was more prevalent in these areas than early 

leaf spot caused by Cercospora arachidicola. 

The intensity of the disease in different surveyed 

villages of Chunar and nearby area ranged from 12.57 to 22.12% 

in the month of September and 34.67 to 44.95% in the month 

of October in 2009. In the year 2010 disease intensity ranged 

from 12.63 to 23.88% in the month of September and 33.22 to 

41.84% in the month of October. Average intensity was found 

as 17.84, 40.14, 18.96 and 38.61% in the month of September 

and October and in the year 2009 and 2010, respectively (Table 

1). These results confirm the findings (Ghewande, 1990; 

Anon., 1993). In India, late leaf spot is more severe than early 

leaf spot. It causes severe defoliation and reduces pod yields 

by more than 50% if the crop is not protected with chemicals 

(Shew, et al., 1988). Hegde, et al., 1995 observed that the late 

leaf spot disease of groundnut appeared at 55 to 60 days after 

sowing and would cause more than 50% loss in pod and haulm 

yield in groundnut producing areas of Karnataka. Current 

studies on occurrence of Cercospora leaf spot (CLS) disease 

was recorded throughout kharif season starting from 4 th week 

of August on most of the cultivars grown in the Chunar and 

Table 1. 

Disease index of Cercospora leaf spot of groundnut 

in Chunar and nearby areas of Mirzapur district 

of Uttar Pradesh 

Village 

Per cent disease index (PDI) 

2009 2010 

September October September October 

Nakahara 18.26 39.49 20.36 33.22 

Jalalpur 16.57 41.32 22.10 41.84 

Keshavpur 15.39 43.79 17.39 36.87 

Pachevara 13.20 41.27 12.63 39.40 

Naugaraha 22.12 44.95 19.84 40.50 

Bishnupur 20.03 42.73 14.43 38.39 

Chandapur 19.70 37.35 21.20 38.70 

Shivpur 15.89 36.26 13.93 39.29 

Gangpur 19.32 38.37 23.88 35.25 

Bhavani 20.33 39.71 21.59 33.60 

Lohra 12.57 34.67 19.63 40.32 

Kailahat 21.38 40.54 17.88 38.82 

Sikandarpur 16.76 38.98 20.30 41.51 

Samauli 18.34 42.64 20.34 42.94 

Average 17.84 40.14 18.96 38.61 

nearby areas of Mirzapur district, Uttar Pradesh. The severe 

incidence of the disease was seen in last week of September 

to October. The intensity of the disease reached up to 44.95% 

on most of the local cultivars of groundnut grown. In severe 

condition defoliation of leaves takes place and disease spread 

to stem portion also. So it can be concluded that Cercospora 

leaf spot (CLS) disease is a very important disease of study 

and subsequently management practices to be adopted for 

betterment of the farming community. 


Anonymous 1993. Annual progress report on kharif Groundnut. All 

India co-ordinated research project on Oilseeds, Directorate of 

Oilseeds Research, Rajendra Nagar, Hyderabad, pp.491. 

Chohan, J. 1974. Recent advances in diseases of groundnut in India. In: 

Current trends in Plant Pathology, Lucknow University press, 

Lucknow, Uttar Pradesh, pp.171-184. 

Garren, K.H. and Jackson, C.R. 1973. Peanut diseases. La: Peanutsculture 

and uses. American Peanut Research and Education 

Association, stone printing Co, Roanoke, Viginia, USA. pp. 429- 

491. 

Ghewande, M.P. 1990. Diseases of groundnut and their management. 

Journal of Oilseeds Research., 7: 78-97. 

Hegde, D.M. 2005. Striving for self-sufficiency. The Hindu Survey of 

Indian Agriculture, pp.55-63. 

Hegde, V.M., Subrahmanyam, K., Gowda, M.V.C. and Prabhu, T.G. 1995. 

Estimation of yield loss due to late leaf spot disease in Spanish 

groundnut in Karnataka. Karolinska Journal of Agricultural Science, 

8: 355-359. 

Jackson, C.R. and Bell, D.K. 1969. Diseases of peanut (groundnut) 

caused by fungi. University of Georgia, College of Agriculture 

Experiment Station, Research Bulletin No. 56, pp.137. 

Mayee, C.D. and Dater, V.V. 1988. Diseases of groundnut in the tropics. 

Review of Tropical Plant Pathology, 5: 169-198. 

Ramkrishnan, V. and Appa Rao, A. 1968. Studies on tikka disease of 

groundnut. Indian Phytopathology, 21: 31-36. 

Rangaswami, G. and Mahadevan, A. 2006. Diseases of crop plants in 

India. 4 th Ed. Prentice Hall of India Pvt. Ltd. New Delhi. pp.335- 

336. 

Shew, B.B., Beute, M.K. and Wynne, J.C. 1988. Effect of temperature 

and relative humidity on Expansion of resistance to Cercosporidium 

personatum in Peanut. Phytopathology, 78: 493-498. 

Siddaramaiah, A.L., Hiramath, R.V. and Krishan, K.S. 1977. Control of 

tikka disease of groundnut with Detur. Journal of Plant Protection, 

5: 193-194. 

Subrahmanyam, P., Mc Donald, D., Waliyar, F., Reddy, L.J., Nigam, S. 

N., Gibbons, R.W., Rao, V.R., Singh, A.K., Pande, S., Reddy, P.M. 

and Subba Rao, P.V. 1995. Screening methods and sources of 

resistance to rust and late leaf spot of groundnut. Information 

Bullatin No. 47, ICRISAT, Patancheru, Andhra Pradesh, India, pp.19. 

Sulaiman, M. 1966. Investigations on the control of tikka disease of 

groundnut in the Maharashtra State. Abstract. Review of Applied 

Mycology, 45: 305. 

Sundaram, N.V. 1965. Note on creation of tikka leaf spot of groundnut, 

Indian Oilseeds Journal, 9: 98. 



Studies on the Effect of Scent Components on the Somatic Cells of Allium sativum L 

CH. SRINIVASULU 

Department of Zoology, Govt Degree College, Peddapalli, Karimnagar Distt. A.P. India 

e-mail: chsri39@gmail.com 

ABSTRACT 

The mutagenic efficiency of certain secretions (1-Butanol and 

1-Hexanol) of heteropteran, bugs are assayed on the root tips 

cells of Allium sativum L 1-Butanol and 1-hexanol (alcohols) 

when applied on the root tips cells of A.satium,caused 

considerable disturbance during the course of mitotic division. 

The dec line in mito tic inde x wa s greate r at hig her 

concentrations. The spectrum of mitotic abnormalities included 

disturbed chromosomes at prophase, un-orientation of 

chromosomes at metaphase,due to non-formation of spindles. 

Unequal separation of chromosomes, formation of anaphase 

bridges, tripolar grouping and unoriented chromosomes, 

fragmented chromosomes at prophase, disturbed chromosomes 

at metaphase, chromosomal stickiness, and disturbed anaphase 

due to un-orientation spindle.The chromosomal aberrations 

induced by these test compounds were gaps, chromatids breaks, 

pulverizations,clumping and cells with sticky chromosomes 

dice ntric chromo somes and ce ntrome ric breaks,more 

frequently and regularly appeared in all the time intervals. 

Key words 

Heteropteran bugs,scent components,1-Butanol,1- 

Hexanol, Allium sativum 

Most of the heteropteran bugs secrete a pungent volatile 

compounds from the abdominal and metathoracic scent 

glands. In order to test the mutagenicity of these scent 

compounds from insects. Allium is being used as an 

experimental materials. Certain aldehydes like octenal,2- 

octenal, and 4-hydroxy-2-octenal acted as antirespiratories, 

cytological aberrations they induce are very few (Canuto, et 

al., 1985). n-butyl butyrate, n-dodecane and n-pentadecane 

are the scent components of C.purpureus causing cytological 

aberrations in the root tips cells of Allium sativum (Surender, 

et al., 1987). They showed inhibition effect may be due to the 

blockage of DNA synthesis (Digamber Rao, et al., 1987). 1- 

Butanol, 1-Hexanol were the scent components reported from 

the scent glands of Libyaspis angoleunsis (Cmelic, 1969) and 

Gelastocoris oculatus (Staddon and Thorne, 1973). 

Most of the chemicals were previously treated for 

inducing chromosomal aberrations (Sagoo, et al., 1993). 

Sadasivaiah, et al., 1973, studied the effect of lysergic acid, 

diethyl amide on onion, rye and barley. Action of 

antihistamine, dimecras and leaf extracts of Azadirachta indica 

were observed as somatic cells of Allium sativum L 

Chandrakala and Vidyavathi, 1992). In tumor cells, Aliphatic 

aldehydes react with cellular thiol compounds (Sessa, et al., 

1977) and cause ultra structural alterations of plasma 

membranes of Golgi vesicles, mitochondria and nuclei (Bassi, 

1967). 

Effect of mitomycino-c was studied on root tips cells of 

Cuminium cyminum (Mogili, et al.,1984) and BIT-4 on root 

tips of A.sativum. (Sadanandam, et al., 1984). Teratogenicity 

and abnormal chromosomal recombination by selenium 

compounds have been reported (Ray, et al., 1978; Morimoto, 

et al., 1982). Effect of polluted water, heavy metals, fertilizer, 

pesticides as chromosomal of Allium sps. Roots were also 

studied. 


Actively growing root tips of A.sativum L were 

harvested and treated with various concentrations (0.01, 0.5, 

1.0 and 2.0%) of scent components from heteropteran bugs 

for a duration of 2,4 and 6 hours. The required concentrations 

were prepared in acetone as these components do not dissolve 

in distilled water. 

After each duration of treatment of the root tips from all 

the samples were fixed in carnoys fluid (1 part of acetic acid 

and 3 parts of absolute alcohol) and kept in the refrigerator. 

The root tips were allowed to remain in the fixative for 24 

hours and then transferred into 70% alcohol. About 2000 

randomly selected dividing cells from ten different root tips 

were washed thoroughly with distilled water and then 

temporary squash preparations were prepared for cytological 

observations employing the acetocarmine as the nuclear 

stain.Various mitotic aberrations of metaphase, anaphase and 

telophase stages have been observed under meopta 

compound microscope (Czechoslovakia) with the aid of X10 

as eye piece and X 60 as objective. 

Number of cells in division 

Mitotic index (MI)= ——————————————— x100 

Total number of cells scored 

Estimation of mitotic index is an important protocol as it 

reflects the action of a given compound on cell division and 

DNA synthesis. 1500 to 2000 cells were scored from each time 

interval i.e., 24,48 and 72 hours. 


1-Butanol, the alcoholic component from the scent 

secretions of abdominal scent glands of Libiaspis angolensis 

(Cmelik, 1969) and 1-Hexanol, the alcoholic component in the 

scent secretions of metathoracic scent glands of Gelastocoris 

oculatus (Staddon, 1973) were used. These two alcoholic 

compounds were employed for mutagenic efficiency on the 

chromosomes of Allium sativum. 1-Butanol and 1-hexanol

SRINIVASULU, Studies on the effect of scent components on the somatic cells of Allium sativum L 89 

alcoholic compounds were proved to be irritants. 

Various concentrations (0.01,0.5, 1.0 and 2.0) of these 

scent components were prepared in acetone and treated with 

the Allium sativum root tips.It has been observed that these 

components caused considerable disturbance during the 

course of mitotic divisions. The data of chromosomal 

aberrations and mitotic index, presented in Table 1, 2, showed 

clearly the lower values compared to control in all the 

concentrations and durations (2, 4 and 6 hours) for both 1- 

butanol, 1-hexanol compounds and further decrease in values 

were found in higher concentrations.The maximum decline in 

mitotic index was recorded in 2.0% concentration of 1-butanol 

treated for 6hours. The fall in mitotic induces soon after 1- 

butanol and 1-hexanol treatments indicated that many cells of 

proceeding G2 phase entering in the mitotic cycle were effected 

and may be due to the chronic effect on all or on some of the 

proceeding stages of mitotic divisions. 

The most frequent abnormalities uncounted were 

stickiness of chromosome at higher concentrations (1.0%, 

2.0%) due to the effect of alcoholic compound, 1-butanol the 

spectrum of mitotic abnormalities included disturbed 

chromosomes at prophase (Fig.1), unorientation of 

chromosomes at metaphase due to non formation of spindle 

(Fig. 2) and unequal separation of chromosomes (Fig. 3). 

The root tips of A.sativum treated with 1-butanol with 

different concentration showed chromosomal stickiness, 

fragments and bridges at metaphase, anaphase and telophase. 

Moreover, the chromosomal abnormalities observed were 

found to be increased with increased concentrations of 1- 

butanol. The compound had more inhibitory effect on the 

percentage of mitosis in the root tips. Simultaneously, delayed 

mitosis was observed in the root tips treated with higher 

concentrations and the mitotic index gradually decreased with 

increased concentrations. 

The formation of anaphase bridge may be due to the 

development of dicentric chromosomes with two centromeres 

of each dicentric chromosomes being oriented towards 

opposite poles (Aly Mohammed, et al., 1966). The dicentric 

chromosomes, presumably, arose by the reunion of two sister 

chromatids of the broken chromosomes. (Koller, 1952, Grant 

and Harnery, 1960). Fragmentation of Chromosomes in A. 

sativum has earlier been reported (Digamber Rao, et al ., 1987). 

Mitodepressive and Clastogenic activity of crude drug 

Combination on the somatic cells of Farniculum vulgare is 

reported (Krishna Reddy, et al., 1984), and also earlier reported 

the Influence of Floral pigments of Periwinkle plant on Cell 

division (Mogili, et al., 1985). Diagonal spindles were 

frequently observed in all concentrations for both these 

chemicals under study. Antimutagenic potential of curcumin 

on chromosomal aberrations in Allium cepa (Ragunathan and 

Pammeer, 2007), cytotoxic and genotoxic impact of silver 

nanoparticle using root tips cells of Allium cepa as an indicator 

(Kumari, et al., 2009). 

Similar results were identified for treatment of 1-hexanol 

with the root tips of A. sativum in different concentrations. 

The chromosomal abnormalities showing stickiness, 

fragments and bridges were observed in higher concentrations 

(0.5%, 1.0% and 2.0%). The spectrum of mitotic abnormalities 

included, formation of anaphase bridges (Fig.4), tripolar 

groupings and un-oriented chromosomes (Fig.5). Fragmented 

chromosomes and bridges (Fig.6). These chromosomal 

aberrations were found to be increased with higher 

concentration of 1-hexanol (1.0+2.0 %). These results show 

that among these two alcoholic compounds the 1-butanol 

was more effective than that of 1-hexanol for the root tips of 

A.sativum. 

If certain heteropteran bugs on being disturbed they 

discharge a pungent, volatile liquid from both,abdominal and 

Table 1. 

Concentration 

Percentage 

Mitotic Index (MI) and percentage aberrations in Allium sativum L. with 1-Butanol a scent component of Heteropteran 

bugs 

Duration of 

treatment 

(Hr) 

Types of Abnormalities 

Mitotic 

Metaphase Anaphase Telophase 

index 

(MI) Stickiness Fragments Stickiness Bridges Stickiness Bridges 

Control 12.4 0.05 -- -- -- -- -- 0.04 0.05 

2 12.0 -- 0.2 -- -- -- -- 0.2 0.4 

0.01 4 12.1 0.3 0.3 -- -- -- -- 0.2 0.8 

6 12.3 0.4 0.5 0.3 0.2 -- 0.1 0.4 1.9 

2 12.1 0.3 0.3 0.1 0.1 0.2 0.1 0.3 1.4 

0.5 4 12 0.4 0.5 0.4 0.1 0.2 0.2 0.3 2.1 

6 12.2 0.5 0.7 0.6 0.2 0.3 0.4 0.6 3.2 

2 12.2 0.4 0.5 0.3 0.3 0.3 0.6 0.2 2.6 

1.0 4 11 0.6 0.7 0.5 0.6 0.5 0.7 0.4 4.0 

6 11.4 0.8 1.0 0.5 0.8 0.7 0.9 0.8 5.5 

2 11.0 0.8 0.6 0.7 0.8 0.7 0.6 0.8 5.0 

2.0 4 10.2 1.2 1.4 1.2 1.4 0.9 0.7 1.0 7.8 

6 9.6 1.8 2.1 1.8 1.8 1.3 1.0 1.2 11.0 

Diagonal 

spindles 

Total 

aberrations


metathoracic scent glands.Some hydrocarbons and 

unsaturated aliphatic aldehydes have been reported from the 

scent secretion of dorsal abdominal scent glands of certain 

heteropteran bugs (Baggini, et al., 1966) Watanuki and 

Sakaguchi, 1981 demonstrated the antitumor activity of 

benzaldehyde. Canuto, et al., 1985 reported some of the 

aldehydes like octanal, 2-octenal, 4-hydroxy-2-octenal (HOE), 

nonal, 2-nonenal, 4-hydroxy-2-nonenal (HNE) acted as 

antirespiratory. Some of the scent components are 

carcinostatic (Schaurenstein, et al., 1987), anti fungal 

(Surender, et al., 1987) and antibacterial; (Surender, et al., 1988) 

sex-attractant (Park and Sutherland, 1962) and neurotoxins 

(Escousbas and Nakajime, 1994). Surender, et al., 1987 reported 

the mitotic effect due to n-dodecane and n-pentadecane, the 

hydrocarbons which act as contact poision reported from the 

abdominal scent glands of larvae Chrysocoris purpureus 

(Janaiah, et al., 1988). Watanuki and Sakaguchi, 1981 

demonstrated the antitumor activity of benzaldehyde. Canuto, 

et al., 1985 reported some of the aldehydes like octanal, 2- 

octenal, 4-hydroxy-2-octenal (HOE), 2-nonenal acted as 

antirespiratory. In recent years some of the antibiotic have 

proved to be effective, cytotoxic and mutagenic in both animals 

and plants many antibiotics. Which interfer with DNA 

synthesis, also effect cell division process and induce 

chromosomal aberrations (Rendi and Ochae, 1962). Digambar 

Rao, et al., 1987 studies the mutagenic efficiency of n-butylbutyrate. 

The ester component,found in the scent secretion 

of Amorbus rhombifer on the root tips of Allium sativum with 

different concentrations.it was found that higher 

concentration of n-butyl-butyrate showed greater effect on 

the mitosis of A.sativum, inhibition of cell division, spindle 

apparatus and cell wall development. The inhibitory effect 

may be due to the blockage of DNA synthesis. (Heiner, 1971). 

Action of 1-butanol, 1-hexanol on mitotic cells (Table 

1,2) division of A.sativum caused considerable mitodepression. 

The spectrum of mitotic abnormalities includes 

disturbed chromosomes at prophase, un-orientation of 

chromosomes at metaphase due to non-formation of spindle, 

unequal separation of chromosomes, formation of anaphase 

bridges, tripolar grouping and un oriented chromosomes, 

fragmented chromosomes and bridges. The formation of 

anaphase bridges may be due to the development of dicentric 

chromosomes with two centromeres of each dicentric 

chromosomes being oriented towards opposite poles (Aly 

Mohammed, et al., 1966). Formation of chromosomes in Allium 

cepa has been earlier reported by Sagoo, et al., 1993. The 

dicentric chromosomes presumably, arose by the re union of 

two sister chromatids of the broken chromosomes (Koller, 

1952, Grant and Harney, 1960, Unoriented chromosomes were 

frequently observed in all concentrations for both alcoholic 

compounds. These treated root tips cells showed considerable 

decline in mitotic indices at all concentrations compared to 

the control the fall in the mitotic indices soon after the 

treatments may be due to affect of proceeding G2 phase 

entering in mitosis. 

Thus it may be observed that the scent components 

induced cytological aberrations and possibly acted as mitotic 

inhibitor; The inhibitory effect may be due to the blockage of 

DNA synthesis in general. The spindle inhibiting action may 

mostly be caused by other C-mitotic agents (Deysson Gay, 

1975). 


I thanks, Head, Department of Zoology, Kakatiya 

University Warangal, A.P. for providing his laboratory facilities 

to perform some experiments during this work. I express my 

sincere gratitude to my beloved research guide Dr. C. Janaiah, 

Retired Professor, Department of Zoology, KU. for his 

invaluable support and encouragement in doing this article. I 

Table 2. 

Concentration 

percentage 

Mitotic Index (MI) and percentage aberrations in Allium sativum L. with 1-Hexanol a scent component of Heteropteran 

bugs 

Duration of 

treatment (hr) 

Mitotic index 

(MI) 

Types of abnormalities 

Metaphase Anaphase Telophase 

Stickiness Fragments Stickiness Bridges Stickiness Bridges 

Control 12.4 0.05 -- -- -- -- -- 0.04 0.05 

2 12.3 -- 0.3 -- -- -- -- -- 0.3 

0.01 

4 12.1 0.1 0.2 -- -- -- -- 0.2 0.5 

6 12.2 0.2 0.5 -- -- -- -- 0.4 1.1 

2 12.0 0.2 0.4 -- 0.2 0.4 -- 0.3 1.5 

0.5 

4 11.8 0.4 0.6 0.2 0.2 0.2 0.3 0.3 2.2 

6 11.6 0.6 0.2 0.3 0.2 0.6 0.2 0.3 2.4 

2 11.2 0.5 0.6 0.4 0.4 0.3 0.3 0.2 2.7 

1.0 

4 11.0 0.3 0.4 0.4 0.5 0.6 0.5 0.2 2.9 

6 11.0 0.4 1.0 0.5 0.8 0.5 0.4 0.6 4.2 

2 11.0 0.2 0.8 0.5 0.8 0.5 0.8 0.7 4.3 

2.0 

4 10.0 1.2 1.2 1.5 1.2 0.6 0.7 1.2 7.6 

6 9.8 1.0 2.0 1.3 1.6 1.2 1.2 1.4 9.7 

Diagonal 

spindles 

Total 

aberrations

SRINIVASULU, Studies on the effect of scent components on the somatic cells of Allium sativum L 91 

thank Prof. Vidyavathi, Department of Botany, Kakatiya 

University Warangal for her critical and invaluable 

suggestions and allowing us to use laboratory space to perform 

photography. 


Fig. 1-6: 1-Butanol and 1-Hexano l induc ed c ytologic al 

aberrations in the root tips of Allium sativum (x2000) 

Fig. 1: Disturbed chromosomes at prophase 2% 1-Butanol for 

4 hrs. 

Fig. 2: Un orientation of chromosomes at metaphase due to 

Non-formation of spindle at 1% 1-Butanol treated for 6 

hours 

Fig. 3: Unequal separation of Chromosomes at 1% 1-Butanol 

treated for 6 hours 

Fig. 4: Formation of anaphase bridges at 2% 1-Hexanol treated 

for 6 hours 

Fig. 5: Tripolar groupings and Un oriented Chromosomes at 

2% 1-Hexanol treated for 4 hours. 

Fig. 6: Fragmented Chromosomes and bridges 1% 1-Hexanol 

treated for 6 hours. 

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A Study on the Quality of Milk in Gwalior 

CHANDANA JAIN AND RITU MEDIRATTA 

Department of Life Science, Boston College for Professional Studies, Gwalior, M.P. 

ABSTRACT 

The study was carried out to know the quality of milk distributed 

and consumed in Gwalior. Area of study was Gwalior city and 

adjoining rural area. A total number of 80 samples were 

collected of which 32 were from Gwalior city and 48 were from 

adjoining rural area which feeds milk to Gwalior city. All the 

samples were analyzed for different adulterants. 40% samples 

in urban area and 20.83% samples in rural area were found to 

be adulterated with water. The milk consumed in urban area 

was highly adulterated as compared to rural area milk. In 

Gwalior urban area 6.25% samples for Ammonium Sulphate, 

15.62% samples for added urea, 18.75% samples for 

Formaldehyde, and 15.62% samples for detergent were found 

to be positive. 25% samples in Gwalior city found to be 

adulterated with skimmed milk as compared to 2.08% in rural 

area. None of the sample was found to be adulterated with cane 

sugar, starch and added glucose. 

Key words 

Quality, milk, Gwalior, adulteration 

Milk is almost a complete food. It is an excellent source 

of protein, fat, carbohydrates, minerals and vitamins. 

Adulterated milk is harmful for human consumption. In most 

developing countries, food safety systems are dysfunctional 

and despite increasing concern most of rural populations are 

unaware of adulteration. India is no exception. Despite the 

laws governing the control of quality and sale of milk existing 

in India for decades, the adulteration of milk has not been 

checked completely. Adulterant like cane sugar, starch, 

glucose, ammonium sulphate, urea, NaCl, skimmed milk powder, 

pulverized soap, detergents and chemical like formalin, 

hydrogen peroxide are added to the milk. These adulterants 

are poisonous and cause health hazards. Milk is mixed with 

toxic additives like Urea to increase the consistency and it is 

also said that such adulterated milk remains intact even for 2 

or more days while ordinary milk goes sour in a day. Urea is a 

natural constituent of milk is present extent of 700 ppm so it’s 

detection as a adulterant is not easy. Above 700 ppm of urea 

indicate the urea is added in milk as adulterant. Pal, 1963 

conducted a survey in Ludhiana showed the adulteration 

figure as 15.9% out of 4597 samples of milk. Other surveys 

conducted by Zariwala, et al., in Bombay, Karpude, et. al., 

1987 in Parbhani town also showed the extensive problem of 

adulteration in milk. Hence this study has been planned to 

know the extent of adulteration of milk received from various 

milk outlets of the city. These adulterant are not allowed 

according of P.F.A. (Prevention of Food Adulterants) 


80 milk samples were collected from different outlets of 

milk in the city from 5 location, including co-operative dairy 

outlets and private organization from of the Gwalior city i.e. 

urban areas and adjoin rural area of Gwalior city. This study 

was carried out in the summer of year 2010 and 2011 (40 

samples each year). Sample study had been done on the same 

day. Samples were collected randomly in the morning and 

transported to the lab. After the preparation of sample organo 

leptic test, correct lactometer reading (C.L.R), clot on boiling 

test and alcohol test for stability of milk were performed 

immediately in the lab. All the tests of detection of adulterates 

i.e., cane sugar, starch, added glucose, ammonium sulphate, 

urea, sodium chloride, skimmed milk powder, pulverized soap, 

detergents, preservatives like hydrogen peroxide, formalin 

were carried out qualitatively. 


Presence of adulterants in milk samples collected from 

different sources of each location of Gwalior presented in 

Table 1. It is evident from the Table 1 that a many of the 

samples were found to be adulterated. It has been observed 

that the maximum milk samples collected from both urban areas 

and nearby rural areas were found to be adulterated with water. 

None of the sample was detected positive for cane sugar, 

glucose and starch (Table 1). These observations are in 

agreement with those by Mishra, et al., 1997, Patel, 1997 and 

Wadekar, et.al., 2011 observed that none of the milk simples 

was found to be adulterated with starch and glucose. 

Urea and detergent were detected as an adulterant and 

it was found that urea and detergent were present in 15.62% 

samples in urban area and 10.41% samples in rural area were 

found to be adulterated with urea. It showed that more number 

of sample from urban area were adulterated with urea and 

detergent than rural of nearby city. The higher number of urea 

and detergent was detected in Morar area of the city. 

Use of preservatives like hydrogen peroxide and 

formaldehyde to reduce bacteria count was also observed in 

samples both from urban as well rural areas to increase keeping 

quality of milk. Formaldehyde was detected in 18.75% samples 

both in urban and rural area. Whereas hydrogen peroxide was 

detected in 3.10% samples in urban area only. No sample tested 

positive in rural area. Arora, et al., 2004 observed that the 

0.4% milk samples collected form organized and unorganized 

sector were positive for formalin test. Bansal and Singhal, 

1991 reported that the addition of formalin even at low levels 

completely inhibited the proliferation of any bacteria in the 

milk. In a study Wadekar, et al., 2011 also observed that 12% 

samples in summer and 10% samples in rainy season in Nanded 

were adulterated with formaldehyde. Maximum milk samples


Table 1. 

Tested for 

Presence of adulterants in milk (No. of samples) 

Lashkar(U) Morar(U) Sirol(R) Banmor(R) Alapore(R) 

No. of No. of No. of No. of No. of No. of No. of No. of 

samples samples samples samples samples samples samples samples 

found found found found found found found found 

negative positive negative positive negative positive negative positive 

No. of 

samples 

found 

positive 

No. of 

samples 

found 

Total No. of 

samples found 

positive 

negative Urban Rural 

COB 0 16 0 16 0 16 0 16 0 16 0 0 

Stability test 16 0 16 0 16 0 16 0 16 0 32 48 

Water test 7 9 6 10 4 12 2 14 4 12 13 10 

(CLR) 

Cane Sugar 0 16 0 16 0 16 0 16 0 16 0 0 

Starch 0 16 0 16 0 16 0 16 0 16 0 0 

Glucose 0 16 0 16 0 16 0 16 0 16 0 0 

Ammonium 1 15 1 15 0 16 0 16 1 15 2 1 

Sulphate 

Added Urea 3 13 2 14 1 15 3 13 1 15 5 5 

Sodium 

1 15 0 16 1 15 3 13 1 15 1 5 

Chloride 

Hydrogen 0 16 1 15 0 16 0 16 0 16 1 0 

peroxide 

Formalin 4 12 2 14 2 14 5 11 2 14 6 9 

Skimmed milk 5 11 3 13 0 16 1 15 0 16 8 1 

powder 

Pulverized soap 2 14 0 16 0 16 1 15 0 16 2 1 

Detergent 2 14 3 13 1 15 2 14 1 15 5 4 

detected positive for formalin were from Banmore rural area 

adjacent to Gwalior. From Banmore 5 out of 16 samples were 

positive. 

Results indicate that skimmed milk powder is more 

common adulterant in urban area as compared to rural area. 

Presence of skimmed milk powder was detected in 25% milk 

samples collected from Gwalior urban area i.e. Lashkar and 

Morar. Whereas only 2.08% i.e. only one milk sample found 

to be adulterated with skimmed milk powder in rural area. It 

showed that milk brought from rural area was reprocessed to 

increase SNF value of milk and then sold in city. Arora, et al., 

2004 also agreed with the results that milk is collected from 

vendors and then reprocessed to increase SNF value. 

Table 2. 

Incidence of adulterants (% positive samples) in 

milk obtained from various sources. 

Tested for 

Samples found positive Over all 

Urban (U) 

% 

Rural (R) 

% 

percentage 

of positive samples 

COB 0.00 0.00 0.00 

Stability test 100.00 100.00 100.00 

Water test (CLR) 40.00 20.83 31.41 

Cane Sugar 0.00 0.00 0.00 

Starch 0.00 0.00 0.00 

Glucose 0.00 0.00 0.00 

Ammonium Sulphate 6.25 2.08 4.16 

Added Urea 15.62 10.41 13.01 

Sodium Chloride 3.12 10.41 6.76 

Hydrogen peroxide 3.12 0.00 1.56 

Formalin 18.75 18.75 18.75 

Skimmed milk powder 25.00 2.08 13.54 

Pulverized soap 6.25 2.08 4.16 

Detergent 15.62 8.33 11.97 

Samples were also tested for adulteration with pulverized 

soap. For pulverized soap only one sample in Alapore rural 

area of Gwalior was found to be positive out of total 80 samples. 

Results indicate that ammonium sulphate and sodium chloride 

are less frequently used as an adulterant in milk as three 

samples tested positive for Ammonium sulphate where 6.75% 

samples found adulterated with sodium chloride. 


Arora, S., Sharma, V. Des Raj, Motiram and Kishore, K. 2004. Status of 

milk adulteration in some states of North India. Indian J.Dairy 

Sci., 57(1): 65. 

Bansal, A. and Singhal, O.P. 1991. Preservation of milk samples with 

formalin-Effect on Acidity. Indian J. Dairy Sci., 44(9): 573. 

Karpude, A.A., Rathi, S.D., Joglekar, N.V. and Ingle, U.M. 1987. 

Adulterated Milk sold in Parbhani town. Asian J. Dairy Res. 6(2): 

83-86. 

Mishra, M., Dehury, M., and Nayak, J.B. 1977. Adulteration of market 

milk at Bhubaneswar. Indian J.Vet. Sci. pp. 378-380. 

Pal, R.N. 1963. Milk adulteration in Ludhiana city. Indian J. Dairy 

Sci., 16:92-97. 

Patel, R.K. 1979. A study on the quality of milk collected at different 

collection centers. Dairy guide, pp.27. 

Wadekar, S.B., Chavan, B.R. and Menkunale, G.V. 2011. Survey on 

adulteration of the milk received from Government milk scheme 

in Nanded town. Interlink Research Analysis, 1(4): 32-35. 

Zariwala, T.T., Sharma, U.P. and Gayakwad, K.S. 1976. Survey of quality 

of Milk in Bombay. Indian J. Dariy Sci. pp. 105-118. 



Prediction of Lifetime Milk Production from Early Lactation Traits in Crossbred 

Cattle 

HEMANT KUMAR 1 AND B.K. HOODA 

CCS Haryana Agricultural University, Hisar 125 001 

e-mail: rushtohemant@rediffmail.com 1 

ABSTRACT 

The first and second lactation traits of 158 crossbred cattle has 

been analyzed for prediction of life time milk production (LTP3) 

of crossbred cattle. On the basis of individually first and second 

lactation traits the lactation milk yield (LMY) and lactation 

length (LL) are important traits for prediction of lifetime milk 

production. When we consider all the first and second lactation 

traits including age at first calving the R 2 and R A 

2 

were found 

0.872 and 0.862 respectively, and finally traits LMY 1, LMY 2, 

LL 1, LL 2, SP 1, SP 2, DP 1 and DP 2 were selected in the model 

by using backward elimination method of regression analysis. 

Key words 

Crossbred cattle, backward elimination, regression, 

coefficient of determination (R 2 ), Correlation. 

The overall productivity of dairy animals depends upon 

their lifetime performance rather than on a single lactation 

performance. Ultimate aim of the dairy producers is to maximize 

milk production and profitability, therefore lifetime milk 

production is an important economic trait. Improvement of 

lifetime milk production through early selection of females 

would be desirable because waiting for an actual estimate of 

this trait not only delays selection decision but also reduces 

rate of genetic progress, in addition to high cost of maintaining 

poor producers. Generation interval and expenses involved 

in maintaining less productive animals could be reduced if the 

animals are selected for lifetime productivity on the basis of 

traits expressed in their early lifetime. Many workers have 

predicted the lifetime production on the basis of early 

performance traits in different breed of cattle and buffalo e.g. 

Pathodia, et al., 2003, Dalal, et al., 204, Gandhi, et al., 2009, 

and Shinde, et al., 2010. Hence this study was planned to 

project the lifetime milk production from first and second 

lactation traits in crossbred cattle. 


Data pertaining to 158 crossbred cattle from the history cum 

pedigree sheet maintained in the department of Animal 

Genetics and Breeding, Lala Lajpat Rai University of veterinary 

and Animal Sciences, Hisar, Haryana for a period of 24 years 

(1985 to 2009) were used for this study. 

Lactation traits were taken this study were age at first calving 

(AFC), lactation milk yield (LMY1, LMY2), lactation length 

(LL1 and LL2), service period (SP1 and SP2), dry period (DP1 

and DP2), calving interval (CI1 and CI2). Only those animals 

were included in this study which has completed three 

lactations milk yield. The lifetime milk production (LTP3) was 

defined as total amount of milk produced by cattle from the 

initiation of first lactation till the completion of third lactation. 

The pearsonian correlation of first and second lactation traits 

with LTP3 was calculated and it was predicted using backward 

elimination method of multiple regression analysis (Draper 

and Smith, 1987) by SPSS software. To predict LTP3, we 

consider 3 set a explanatory variables in the models and 

analyzed using backward elimination procedure of multiple 

regression analysis. In first set only first lactation traits and 

AFC are included in the prediction equation, in the second 

set only second lactation traits and AFC were included and 

finally in third set all the first and second lactation traits 

including AFC were considered. 


The Correlation coefficients (Table 1) of LTP 3 with 

LMY1, LMY2, LL1, LL2, and SP2 were positive and significant, 

and with DP1 and DP2 were negative significant. The 

correlation of LTP3 with AFC is negative but not significant 

and with other variables i.e. CI1, CI2 and SP1 were positive 

and non-significant. 

To predict LTP3, when all the traits of first set were 

incorporated in the model, the accuracy of prediction (R 2 ) and 

adjusted R-square (R A2 

) were 0.645 and 0.631 respectively and 

only LMY1 was signifianct. Finally LMY1 and LL1 were 

selected in the model by backward eliminatio method in the 

first set and R 2 and R A 

2 

was estimated 0.633 and 0.628 

respectively. Similar result was found for the second set 

Table 1. 

Phenotypic correlation of early lactation traits with 

LTP3 

Early lactation traits 

Correlation with LTP3 

LMY1 0.791** 

LMY2 0.855** 

AFC -0.099 

LL1 0.589** 

LL2 0.440** 

CI1 0.144 

CI2 0.120 

SP1 0.145 

SP2 0.178* 

DP1 -0.361** 

DP2 -0.170 

*Significant at 5%, **Significant at 1%


Table 2. 

Regression equation estimated by backward elimination method 

Set Step Equation R 2 2 

R A 

One First LTP3 = 5934.280 + 2.145LMY1** - 0.065AFC - 4.321LL1 - 6.554CI1 + 7.998SP1 - 2.768DP1 0.645 0.631 

Final LTP3 = 3359 + 2.208LMY1** - 3.178LL1 0.633 0.628 

Two First LTP3 = 2222.046 + 2.740LMY2** + 0.139AFC - 6.557**LL2 – 0.229CI2 + 2.394SP2 - 1.584DP2 0.754 0.744 

Final LTP3 = 1934.768 + 2.754LMY2** - 4.983LL2 0.748 0.745 

Three First LTP3 = 3103.855 + 1.263LMY1** + 1.731LMY2** + 0.187AFC - 4.4514LL1* - 4.081Ll2* - 822CI1 + 0.14CI2 + 3.823SP1 + 0.872 0.862 

2.9SP2* - 2.827DP1* - 3.551DP2* 

Final LTP3 = 2899.237 + 1.249LMY1** + 1.725LMY2** - 4.630LL1** - 3.981LL2** + 2.194SP1 + 2.984SP2** - 3.044DP1* - 0.871 0.864 

3.517DP2** 

*Significant at 5%, **Significant at 1% 

regarding significance of traits when all the traits included in 

the model i.e. only LMY2 was signifianct and R 2 and R A 

2 

was 

estimated 0.754 and 0.744. Finally LMY2 and LL2 were selected 

in the model for the second set and R 2 and R A 

2 

were 0.748 and 

0.745. So we can say that on the basis of individually first and 

second lactation traits the lactation milk yield and lactation 

length are important traits for prediction of lifetime milk 

production. When we consider third set in which all the first 

and second lactation trait were involved including AFC the R 2 

and R A 

2 

were 0.872 and 0.862 respectively, and finally LMY1, 

LMY2, LL1, LL2, SP1, SP2, DP1 and DP2 were selected in the 

model and the R 2 and R A 

2 

were 0.871 and 0.864 respectively. 

From Table 2 it reveals that there is numerous improvement in 

R 2 and R A 

2 

in third set as compared to first and second set, so 

we recommend final model of set 3 for prediction of lifetime 

milk production. 


Dalal, D.S., Malik, Z.S., Chhikara, B.S. and Ramesh Chander. 2004. 

Prediction of lifetime milk production from early lactation trait in 

Hariana cattle. Indian Journal of Animal Sciences, 74(11): 1145- 

1149. 

Darper, N.R. and Smith, H. 1987. Applied Regression Analysis. John 

Wiley and Sons Inc., New Yark. 

Gandhi, R.S., Raja, T.V., Ruhil, A.P. and Kumar, A. 2009. Prediction of 

lifetime milk production using artificial neural network in sahiwal 

cattle. Indian Journal of Animal Sciences, 79(10): 1038-1040. 

Pathodia, O.P., Tailor, S.P. and Jain, L.S. 2003. Projection of lifetime 

prediction from early available traits in Surti buffalo for Food Security 

and Rural Employment held at Delhi, 2:107. 

Pundir, R.K. and Raheja K.L. 1995. Prediction of lifetime milk yield by 

early traits in sahiwal and Hariana cows. Indian Journal of Dairy 

Science, 48: 146-49. 

Shinde, N.V., Mote, M.G., Khutal, B.B. and Jagtap, D.Z. 2010. Prediction 

of lifetime milk production on the basis of lactation traits in Phle 

Triveni crossbred cattle. Indian Journal of Animal Sciences, 80(10): 

986-988. 



Fungal Growth on C1 Compounds: A Study of the Amino Acid Composition of a 

Methanol-utilizing Strain of Trichoderma viride 01 PP 

MOHD. SHAHID, MUKESH SRIVASTVA, ANURADHA SINGH AND 1 NEELAM PATHAK 

Department of Plant Pathology, C.S. Azad University of Agriculture and Technology, Kanpur 208 002 

1 

Department of Biosciences, Integral University, Kursi Road, Lucknow 

email: shahid.biotech@rediffmail.com 

ABSTRACT 

The amino acid composition of the C,-utilizing fungus 

Trichoderma viride 01 PP, growing at the expense of methanol 

as the sole source of carbon, was determined. With the exception 

of an insufficient content of methionine, the essential amino 

acid composition of this novel protein source appears adequate 

in terms of the Food and Agricultural Organization Reference 

Protein for both direct and indirect use in the human diet as a 

food or animal feed, respectively. 

Key word 

Trichoderma, amino acid, protein, FAO 

The exploitation of Cl-utilizing bacteria as a potential 

source of dietary protein for animals is under active 

development (Champagnat, 1974, Cooneyad and Levine, 1972). 

As a consequence of this interest, the amino acid composition 

of the protein from several methanol-utilizing and methaneutilizing 

bacteria (D’Mello, 1972) has been determined: almost 

all of these prokaryotes contained an essential amino acid 

complement compatible with their successful adaption as a 

nutritionally beneficial dietary protein for animals. Particularly 

significant in this respect was the fact that several such 

prokaryotic protein sources contained a higher percentage of 

methionine than conventional dietary plant proteins, such as 

soyabean meal, or novel proteins, such as British Petroleum 

(BP) yeast concentrate. The recent isolation of a methanolutilizing 

strain of the fungus Trichoderma viride 01 PP (10; R. 

J. Tye and A. J. Willetts, J. Appl. Bacteriol.) presents the 

possibility of exploiting mycelial protein produced by this 

eukaryotic microorganism as a source of dietary protein for 

animals. Fungi offer several advantages over bacteria in this 

respect, including a significantly lower nucleic acid-protein 

ratio and comparatively lower harvesting costs from spent 

media. The present investigation, conducted to examine the 

protein and amino acid composition of the methanol-utilizing 

fungus T. viride 01 PP, was a pertinent contribution towards a 

rational assessment of this possibility. 


The isolation, maintenance, and growth of the fungus, 

identified as a strain of T. viride 01 PP, were as previously 

described (10; Tye and Willetts, in press). Analytical methods. 

Amino acid analyses were made using a modification of the 

method described by D’Mello, 1972: 10 mg of methanol-grown 

fungal mycelium was hydrolyzed by autoclaving for 15 h at 

115 C in 5 ml of 6 N HC1 in a sealed glass ampoule under 

nitrogen. The hydrolysate was made up to 100 ml in a 

volumetric flask with deionized water and bsequently filtered 

twice through a sintered glass funnel. A 10-ml aliquot was 

evaporated to dryness in a rotary evaporator. The residue 

was taken up in 2 ml of 10% sucrose in 0.1 N HCl and analyzed 

for amino acids using a autoanalyzer. 2,4,6-Trinitrobenzene 

sulphonic acid was used as the developing reagent. Cystine 

was determined by the method of Moore, 1963 and tryptophan 

was analyzed in alkaline hydrolysates (Inglis and Leaver, 1964). 

The nitrogen content of the fungal mycelium was established 

by the classical Kjeldahl method (Kilbrg, 1972). Chitin, 

Bluementhal and Raseman, 1957 and nucleic acid Ohta, et al., 

1971 levels were assayed as previously described. 

The nitrogen content of the mycelium of methanol-grown 

T. viride 01 PP was 9.82% of N, which was equivalent to a 

protein content of 61.4% (N x 6.25). Growth conditions which 

promote a high mycelial protein content are currently under 

investigation. The content of all of the individual amino acids 

in the protein of the fungal mycelium after growth on methanol 

was remarkably consistent (Table 1). The essential amino acid 

content compared favorably with the 1957 Food and 

Agricultural Organisation (FAO) standard reference protein 

in most aspects; the one notable exception was a deficiency 

with respect to methionine (Table 2). The mycelial protein of 

methanol-grown T. viride 01 PP also compared favorably with 

whole wheat protein, domestic fowl egg protein, and the 

proteins from several other microbial sources (Table 2). Over 

80% of the total nitrogen compounds present in the mycelium 

of methanol-grown T. viride 01 PP was recovered as amino 

acids during a quantitative analysis after acid hydrolysis. The 

chitin and nucleic acid contents of the mycelium of methanolgrown 

T. viride 01 PP were 3.8 and 5.1%, respectively. 

RESLULTS AND DISCUSSION 

The results indicate that the fungus T. viride 01 PP 

growing at the expense of methanol as the sole source of 

utilizable carbon produces mycelia protein with a spectrum of 

essential amino acids superior in all but one respect to the 

FAO Reference Protein: the one major deficit ofmethanolgrown 

fungal protein is a subminimal methionine content, 

which shows a 15% shortfall with respect to the FAO standard. 

However, as indicated in Table 2, the methionine content of 

the mycelial protein of methanol-grown T. viride 01 PP (1.85


g/16 g of N), although below the level in FAO Reference 

Protein, is nevertheless in excess of the content of many other 

sources of dietary protein including conventional plant 

proteins such as whole wheat protein (1.5 g/16 g of N) and 

soyabean meal (1.2 to 1.4 g/16 g of N), as well as other novel 

proteins such as BP yeast concentrate (1.4 to 1.6 g/16 g of N) 

and gas oilgrown Candida lipolyticum (1.6 g/16 g of N). Any 

dietary deficiency resulting from this subminimal content of 

methionine could be avoided by using mycelial protein from 

methanol-grown T. viride 01 PP as the basis for a balanced 

food or feed incorporating compensatory amounts of either 

pure methionine or methionine-rich proteins such as domestic 

chicken egg protein (Table 2). 

The mycelial protein of methanol-grown T. viride 01 PP 

has an acceptable complement of all other essential amino 

acids, including lysine,as assessed against the FAO standard, 

as well as a full complement of non-essential amino acids 

(Table 1). The complement of arginine and histidine, two amino 

acids which are not catagorized as essential human dietary 

components, but supplementary sources of which are widely 

considered necessary for the normal growth of children, is 

similar to that of many conventional dietary proteins. The 

amino acid composition of the mycelia protein of methanolgrown 

T. viride 01 PP is similar to that of several methanegrown 

bacteria; the comparatively high content of tryptophan 

is particularly significant in this respect. 

Table 1. Amino acid content of methanol-grown T. viride 

01 PP 

Mean amino acid con- 

Amino acid tent (g/16 g of N) ± 

standard deviation 

Alanine ...................... 6.80 ± 0.20 

Arginine ..................... 4.92 ± 0.16 

Aspartic acid ................. 8.63 ± 0.19 

Cystine ...................... 0.90 ± 0.08 

Glutamic acid ................ 10.03 ± 0.22 

Glycine ...................... 4.87 ± 0.09 

Histidine ..................... 2.03 ± 0.12 

Isoleucine .................... 4.48 ± 0.09 

Leucine ...................... 7.88 ± 0.11 

Lysine ....................... 5.13 + 0.09 

Methionine ................... 1.85 ± 0.08 

Phenylalanine ................ 4.38 ± 0.08 

Serine ....................... 3.48 ± 0.09 

Threonine .................... 4.62 ± 0.11 

Tryptophan .................. 1.82 ± 0.08 

Tyrosine ..................... 3.65 ± 0.08 

Valine ....................... 5.72 ± 0.10 

a Average of six experiments. 

Table 2. Essential amino acid content ofmethanol-grown 

T. viride 01 PP, FAO Reference Protein, wheat, egg,and some 

other established microbial protein sources (expressed as g116 

g of N)Scene- Saccha- C. lipo- Psu FAO Amino acid lignTo.rum 

deosbm-us mrausxliinmaa roccmeeyreec-es ly (gtiscgaas 

(dmoemtohnana- s Wwhhoelaet Wheoglge Reenfecre- 

(methanol) Iiquus (6) visiae oil) (6) (6) (6) Protein 

(6) (6) (6) (6) 

Isoleucine 4.48, 3.8, 6.0, 4.6, 5.3, 3.9, 3.3 ,6.7 ,4.2 

Leucine 7.88, 8.4, 8.0, 7.0, 7.8, 7.0, 6.7, 8.9, 4.8 

Lysine 5.13, 5.7, 4.6, 7.7, 7.8, 5.3, 2.8, 6.5, 4.2 

Methionine 1.85 ,1.7, 1.4, 1.7, 1.6, 1.8, 1.5, 3.2, 2.2 

Phenylalanine 4.38, 5.1, 5.0, 4.1, 4.8, 4.2, 4.5, 5.8, 2.8 

Threonine 4.62, 5.1, 4.6, 4.8, 5.4, 4.5, 2.9, 5.1, 2.8 

Tryptophan 1.82 ,1.5 ,1.4, 1.0, 1.3 ,1.1, 1.6, 1.4 

Valine 5.72, 5.7, 6.5, 5.3, 5.8, 5.9, 4.4, 7.3, 4.2 

mycelial protein from methanol-grown T. viride 01 PP as 

a food or feed remain to be assessed directly. 


The paper forms a part of the Ph.D work of the first 

author and the facilities provided by Department of Plant 

Pathology, CS Azad University of Technology & Agriculture, 

Kanpur for the conduct of this study is gratefully 

acknowledged. 


Blumenthal, H.J., and Roseman, S. 1957. Quantitative estimation of 

chitin in fungi. J. Bacteriol., 74:222-229. 

Champagnat, A. 1974. Proteins from hydrocarbons, In: Industrial 

aspects of biochemistry. (ed. B. Spencer), Federation of European 

Biochemical Societies. pp.342-353. 

Cooney, C.L. and Levine, D.W. 1972. Microbial utilization of methanol. 

Adv. Appl. Microbiol., 15:337-365. 

D’Mello, J.P.F. 1972. A study of the amino acid composition of methane 

utilising bacteria. J. Appl. Bacteriol., 35:145-148. 

Inglis, A.S. and Leaver, I.H. 1964. Studies in the determination of 

tryptophan. Modified Fischl procedure. Anal. Biochem., 7:10-14. 

Kilberg, R. 1972. The microbe as a source of food. Annu. Rev. Microbiol., 

26:427-466. 

Miller, L. and Houghton, J.A. 1945. The micro-Kjeldahl determination 

of the nitrogen content of amino acids and proteins. J. Biol. Chem., 

159:373-383. 

Moore, S. 1963. On the determination of cystine as cysteic acid. J. 

Biol. Chem. 238:235-237. 

Ohta, S., Maul, S., Sinskey, A.J. and Tannenbaum, S.R. 1971. 

Characterization of a heat-shock process for reduction of the nucleic 

acid content of Candida utilis. Appl. Microbiol., 22:415-421. 



Varietal Evaluation of Cauliflower (Brassica oleracea var. botrytis L.) In Allahabad 

Agro – climatic Condition 

MUKESH YADAV, V.M. PRASAD, AND CHANDAN SINGH AHIRWAR 

Department of Horticulture Allahabad School of Agriculture 

Sam Higginbottom Institute of Agriculture, Technology & Sciences, Allahabad 211 007 

ABSTRACT 

The investigation consisted 15 cauliflower hybrid variety and 

laid out in randomized block design with three replication. 

Observation recorded on plant height, leaves, spread curd, 

size and yield attributes during the investigation. Amongst all 

the cauliflower hybrid variety evaluated for growth, yield and 

quality trails is present investigation wich revealed that the 

variety poorninma had highest plant height (24.64cm), Number 

of leaves per plant (20.83) plant spread (65.79cm), diameter of 

curd (18.92cm), weight of untrimmed curd (2.450 kg), weight of 

trimmed curd (1001.67g), curd yield (345.083 ha -1 ) vitamin – C 

(53.93mg/100g each) and moisture – dry matter ratio (7.85). 

For growth yield and quality character hybrid variety poornima 

is recommended for commercial cultivation in winter season 

of Allahabad agro – climatic condition. 

Key words 

Brassica oleracea. var. botrytis, vitamin – C, moisture 

dry matter ratio 

Cauliflower (Brassica oleracea, var botrytis L.) is a very 

popular vegetable belonging to cole group of vegetable. It is 

a member of crucifarae family and is characterized by petals, 

standing opposite to each other in a square cross. 6 stamens 

of which 4 are long and 2 are short. It has basic chromosome 

number (n=9). Cauliflower has both annual and biennial types, 

but in India only annuals are cultivated. 

Cauliflower (Brassica oleracea var. botrytis L.) is another 

popular crucifer grown mainly in cooler areas. It is steamed, 

stir fried, or pickled. Cauliflower is grown in 1,017 ha, mainly in 

llocos Sur (450 ha) and Benguet (340 ha) (Bureau of 

Agricultural Statistics 2005). Nutritional Value Per 100 g fresh 

edible portion, Cauliflower curd contains: Water (g) – 88.0, 

Protein (g) – 4.0, Fat(g) – 0.30, Carbohydrates (g) – 6.0, Fiber 

(g) – 1.5, Calcium (mg) – 1 50, Potassium (mg) – 325, Carotene 

(mg) – 800, Vitamin C (mg) – 100, Energy Value (kj) – 245. 

Values are similar for cauliflower except for its lower calcium 

(25 mg), carotene (200 mg), and vitamin C (40 mg) contents. 

MATERIALS OF AND METHODS 

The experiment was carried out in Vegetable Research 

Farm, Department of Horticulture, Allahabad School of 

Agriculture, SHIATS, Allahabad. The 15 cauliflower hybrids 

viz., Aghani, Kinaya, Snow crown, Shalakha, Cashmere, 

Easley show, Kavita, Madhvi, NHB-Saritha, Sharad Safedhi 

70, Shigra, NHB 1011, Poornima, GS 75 and Girija were laid 

out in Randomized complete Block design with three 

replication. The spacing which was used in (50×45) all the 

agronomic packages and practices were adopted to grow good 

and healthy crop to get maximum yield. The observation 

recorded on Plant height (cm), Number of leaves (15, 30 and 

45DAT) , Plant spread (cm) , Diameter of curd (cm), Weight of 

untrimmed curd (kg), Weight of trimmed curd (g), Curd yield 

per plot (kg), Curd yield (q/ha.), Vitamin C (mg/100g of edible 

portion), Moisture – dry matter ratio and Economics of 

cultivation. 


Among all the hybrid of cauliflower, Poornima was 

found best for all the growth parameters viz. maximum plant 

height (26.64 cm), max. no. of leaves (20.83), max. plant spread 

(65.79 cm). followed by the Kimaya had plant height (24.56 

cm), max. no. of leaves (20.75) and the max. plant spread (64.21 

cm). The above findings are supported by the findings of 

Ahmad, et al., 2003 for plant height and number of leaves and 

Tripathi and Sharma, 1991 for plant spread. 

Poornima also found superior over other hybrids in 

respect to yield parameter. The maximum curd diameter (18.92 

cm), max wt. of untrimmed and (2.450 kg), max at of trimmed 

used (1001.67 gm), and yield per plant (5.176 kg), total and 

yield per hac (345.083 q), were recorded for the Poornima. 

Followed by the Kimaya found good for the Kimaya had The 

maximum curd diameter (18.00 cm), max wt. of untrimmed (2.302 

kg) and max at of trimmed used (864.00 gm), and yield per 

plant (4.914 kg), total and yield per hac (327.00 q).The above 

findings are similar to the findings El-Rehim, et al., 2003 for 

diameter of curd (cm), weight of untrimmed and weight of 

trimmed curd (g) and Daljeet and Singh, 1990 for curd yield 

per plot (kg) and Vanparys, 1998 and Thamburaj, et al., 1980 

for curd yield (q ha -1 ). 

In respect to quality parameter i.e. vitamin – c and 

moisture-dry matter ratio the hybrid poornima showed 

superiority over all the hybrid i.e. 53.93 mg/100g edible portion 

and 7.85 respectively. Followed by the hybrid Kimaya had 

max. vit-c contain (53.57 mg/100g edibleportion) and moisturedry 

matte7.80. The above findings are supported by the finding 

of Thamburaj, et al., 1980 ascorbic acid (mg/100g of edible 

portion) and Wurr, et al., 1981 for moisture dry matter ration. 

Poornima recorded maximum gross return (172,542 Rs.


Table 1. 

Varieties 

Varietal evaluation of cauliflower (Brassica oleraceA var. botrytis L.) in Allahabad Agro-Climatic Condition 

Plant 

height 

(cm) 45 

DAS 

Number of 

leaves per 

plant 45 

DAS 

Plant 

spread 

(cm) 45 

DAS 

VI-VIS : Varites name given in Materials and Methods 

Diameter of 

curd (cm) 

Weight of 

untrimmed 

curd (kg) 

Weight of 

trimmed 

curd (g) 

Curd 

yield per 

plot (kg) 

Curd 

yield 

(q ha -1 ) 

Vitamin C 

(mg / 100 g 

edible 

portion) 

Moisture - 

dry matter 

ratio 

V 1 21.23 18.50 57.41 16.08 2.124 793.33 4.538 302.507 50.97 7.35 

V 2 24.56 20.75 64.21 18.00 2.302 864.00 4.914 327.597 53.57 7.80 

V 3 17.06 15.50 49.34 13.88 0.963 461.67 3.093 206.197 48.77 6.98 

V 4 21.75 19.17 58.66 16.25 2.125 795.33 4.586 305.730 51.80 7.40 

V 5 19.16 16.50 55.01 15.86 1.892 730.00 4.369 291.283 50.43 7.26 

V 6 18.16 15.50 49.92 15.15 1.152 464.00 3.230 215.353 49.77 7.02 

V 7 22.45 19.58 62.30 17.45 2.225 855.67 4.808 320.527 52.93 7.78 

V 8 21.96 19.17 59.18 16.40 2.143 810.67 4.658 310.533 52.20 7.72 

V 9 19.86 16.67 55.08 15.88 2.030 733.00 4.399 293.240 50.53 7.29 

V 10 18.72 15.83 54.44 15.78 1.870 707.67 4.188 279.220 50.27 7.18 

V 11 21.99 19.50 59.35 17.29 2.196 823.67 4.698 313.217 52.43 7.73 

V 12 18.35 15.58 52.43 15.56 1.785 700.67 4.182 278.773 49.80 7.18 

V 13 24.64 20.83 65.79 18.92 2.450 1001.67 5.176 345.083 53.93 7.85 

V 14 20.19 16.83 56.59 15.96 2.052 740.00 4.459 297.263 50.60 7.32 

V 15 20.85 17.17 56.75 15.97 2.102 751.33 4.537 302.487 50.70 7.34 

ha -1 ), net return (125,215 Rs. ha -1 ) and Cost: Benefit ratio (1: 

3.65). Treatment (T 3 

) Snow Crown recorded the minimum 

gross return (103,098 Rs ha -1 ) and minimum net return ( 53,609 

Rs. ha -1 ) and Cost: Benefit ratio1: 2.08). 

Among all the treatments, (T 13 

) Poornima emerged as 

superior over all the other treatments, in relation to plant 

growth, yield attributing characters, quality parameters and 

economics of couliflower. The above findings are supported 

by the findings of Batra and Singh, 2002 and Sharma and 

Chandra, 2002 for Economics of different varieties. 


Ahmad, S., Saha, S.R., Uddin, M.N., Choudhury, S.S., Awal, M.A. and 

Salam, M.A. 2003. Performance evaluation of some cauliflower 

genotypes in the eastern region of Bangladesh. Pakistan J. Bio. 

Sci., 6(21):1792-1794. 

Batra, V.K. and Singh, Jitendra. 2000. Evaluation of some cauliflower 

varieties at Hisar. Haryana J. Hort. Sci., 29(½):125-126. 

Singh, Daljeet, Thakur, J.C. and Singh, D. 1990. Performance of 

Cauliflower varieties. Haryana J. Hort. Sci., 19(3-4): 337-341. 

El-Rehim, G.H.A. 2003. Evaluation of yield and quality of newly 

produced cauliflower (Brassica oleracea var. botrytis L.) genotype 

under Assiut conditions. Assit J. Agri. Sci., 34(5): 225-239. 

Sharma, A. and Chandra, A. 2002. Economic evaluation of different 

treatment combinations of plant spacing and nitrogen in cabbage 

and cauliflower. Current Agri., 26(½): 103-105. 

Thamburaj, S., Pillai, O.A.A., Anbu, S. and Shanmugavelu, K.G. 1980. 

Preliminary studies on the performance of certain varieties of 

Cauliflower (Ioleracea Var. Botrytis L.) at Coimbatore, South India 

Hort., 28(3): 82-84. 

Tripathi, P.K. and Sharma, R.K. 1991. Studies on the effect of Age of 

Seedling and Spacing of the growth and yield of cauliflower (Brassica 

oleraea Var. Botrytis. L.). New Agri., 1(2): 177-182. 

Vanparys, L. 1998. Cultivars studies for winter Cauliflower. Mededeling 

Provincial Onderzocken Voorlichtings Centrum Voor Land en 

Tuinbouw Beitem Roeselare No. 400, pp.4. 

Wurr, D.C.E., Kay, R.H., Allen, E.J. and Patel, J.C. 1981. Studies of the 

growth o and Development of Winter heading Cauliflower J. Agri 

Sci, U.K., 97(2): 409-419. 



Screening of Chickpea Genotypes for Resistant against Pod Borer, Helicoverpa 

armigera Hubn. 

JEEWESH KUMAR, D.C. SINGH, A.P. SINGH AND S.K. VERMA 

Department of Entomology, Narendra Deva University of Agriculture Technology, Kumarganj, Faizabad, 

U.P. 224 229 

email: singhjeewesh@gmail.com 

ABSTRACT 

Fifty genotypes of chickpea were screened against gram pod 

borer under field condition. Each genotype was sown in two 

rows of 2 m length in Augmented Design, Susceptible (H82-2) 

and resistant (C 235) genotypes as checks were sown after every 

10 genotypes. The populations of larvae of gram pod borer were 

recorded on each test genotype at 50% pod filling stage. On the 

basis of pod damage throughpod borer were grouped in three 

categories (Tolerant, Moderate resistant, Susceptible). Nine 

genotypes registered as rating scale 1 were graded as tolerant 

because their pod damage (5.50-8.50 per cent) were moderate 

between resistant and susceptibleand these were DGP 15, GIG 

0312, ICCL 87315, ICCV 7, RIL 115, ICC 29, ICC 12470, ICCV 

10 and PG 23. Seven genotypes (DGP 15, GIG 0312, ICCL 87315, 

ICCV 7, RIL 115, ICC 29, ICC 12470, ICCV 10 and PG 23) had 

showed pod damage varies from 20.00-23.00 per cent and rating 

scale 3 were placed under susceptible grade because they were 

at par with check susceptible (H82-2), remaining 34 genotypes 

had showed pod damage varies from 10.50-1900 per cent rating 

stage scored 2, were placed under moderately resistant grade. 

Key words 

Chickpea, H. armigera, Screening 

India is the largest producer of chickpea with 67 % of 

the global production It occupies nearly 31 % of total pulse 

area in the country and contributes over 37 % to the national 

pulse production. One of the most practical means of 

increasing chickpea production is to minimize losses caused 

by insect-pests. Gram pod borer, H. Armigera is known to be 

the key pest due to high reproduction rates, a fast generation 

on turn over, wide genetic diversity occurs location and an 

ability to withstand, metabolize and avoid toxic chemicals. 

The young larvae feed on the tender portion of the leaves on 

shoots by making scratches. Second and subsequent grown 

up larvae consume whole leaf, leaf buds, flower buds and 

flower. Under severe pest infestation, whole crop may get 

defoliated. With the availability of pods, the 3 rd instar larvae 

make a hole in the pods and move inside to feed on the grains, 

where as fully grown larvae feed after making a hole in the 

pods and thrusting its head, there in while keeping hind parts 

of the body outside. The yield loss in chickpea due to H. 

armigera was reported as 10-60% in the normal weather 

condition (Vaishmpayan and Veda, 1980). The single larva of 

H. armigera may destroy 30-40 pods in its lifetime (Atwal and 

Dhaliwal, 2005). Keeping the aforesaid fact in view, the present 

study was undertaken to develop effective, economic and 

feasible management of this pest. 


The experimental site lies between 26.47 0 N latitude, 

82.12 0 E longitude and 113 m above from the sea level in the 

sub-tropical belt of the country and soil is alkaline to normal. 

The present investigation was carried out at Students 

Instructional Farm of Narendra Deva University of Agriculture 

Technology, Kumarganj, Faizabad during Rabi, 2007-08. Fifty 

genotypes of chickpea were screened against gram pod borer 

under field condition. Each genotype was sown in two rows 

of 2 m length in Augmented Design, Susceptible (H82-2) and 

resistant (C 235) varieties as checks were sown after every 10 

genotypes. The populations of larvae of gram pod borer were 

recorded on each test genotype at 50 per cent pod filling 

stage. The pod damage was recorded at pod maturity stage 

using 1 to 9 damage rating scale. (1 = > 10 % pod damage, 2 = 

11-20 % pod damage, 3 = 21-30% pod damage, 4 = 31-40% pod 

damage, 5 = 41-50% pod damage, 6 = 51-60% pod damage, 7 = 

61-70% pod damage, 8 = 71-80% pod damage, 9 = > 80% pod 

damage). 


The population of H. armigera and pod were damage 

observed on fifty genotypes sown in 4 th week of October, 

2007 in 2 rows of 2 m length in augmented design. Susceptible 

(H82-2) and resistant (C 235) genotypes as check were sown 

after every 10 genotypes. The larval population could be not 

recorded at 50% flowering stage of crop in any genotypes 

due to non presence of larvae then larval population of pod 

borer was recorded at 50% at flowering stage. 

The maximum number of larvae 3.0/ 10 plants was at 

50% pod filling stage and highest per cent pod damage 23% 

was recorded in susceptible genotype H82-2 at pod maturity 

stage. The lowest larval population 0.5/ 10 plants was noted 

at 50% pod filling stage and lowest pod damage 5.5% was 

recorded at pod maturity stage in resistance genotype C 235. 

The seven varieties NDG 5-32, ICCC 37, RIL 27, DCP 8, BDNG 

9-3, Udai and ICC 12479 were at par with check H 82-2 on the 

basis of larval population varies from 2-3 per 10 plants and per 

cent pod damage varies from 20-23%. The nine genotypes 

DGP 15, GIG 0312, ICCL 87315, ICCV 7, RIL 115, ICC 29, ICC


12470, ICCV 10 and PG 23 were at par with check C 232 based 

on larval population (0-1 larva/10 plants) and 5.50-8.50% pod 

damage. Rest 34 genotypes were found moderate level of larval 

population (1-2 larvae/10plants) and pod damage (10.50- 

19.00%). Gumber, et al., 2000 recorded one number of pod per 

plant, number of pods damaged by pod borer, pod length, 

pod width and seed yield per plant showed that accessions 

ICC 93512, ICC 93515 and ICC 93212 were the most promising 

with higher seed yield and low pod borer damage. Similarly 

Afzal, et al., 2001 screened eight test lines of chickpea against 

H. armigera and observed that two genotypes (Pb 2000 and 

96051) were highly resistant, three advanced lines (9075, 96052 

and BC 6-5) were resistant, two advanced lines (90395-K and 

97047) were moderately resistant and only advanced line 

(88194) was susceptible. Mandal, 2003 evaluated 18 chickpea 

genotypes against H. armigera in field condition. Pod damage 

varied from 9.43 to 24.80%. Two genotypes (PS 83149 and 

Bhawanipatna) were resistant, 6 were moderately resistant, 7 

were moderately susceptible and 3 were insusceptible to the 

pest. Gowda, et al., 2005 recorded that genotype BGD 237 had 

significantly lowest per cent pod damage of 11.86 and 10.84 

during 2000-01 and 2001-02, respectively. BGD 237 recorded a 

pest susceptibility rating of 5, indicating resistant in both 

years. 

On the basis of pod damage through pod borer were 

grouped in three categories (Tolerant, Moderate resistant, 

Susceptible). Nine genotypes registered as rating scale 1 were 

graded as tolerant because their pod damage (5.50-8.50 per 

cent) were moderate between resistant and susceptibleand 

these were had showed pod damage varies from 20.00-23.00% 

andrating scale 3 were placed under susceptible grade because 

they were at par with check susceptible (H82-2) (Table 2), 

remaining 34 genotypes had showed pod damage varies from 

10.50-1900 per cent rating stage scored 2, were placed under 

moderately resistant grade and Singh and Yadav, 1999 also 

found the genotype ICC 29 as tolerant to pod borer. Rai and 

Ramujagir, 2005 found four genotypes (ICCV 93929, ICCV 

96029, ICCV 96030 and ICCV 2) as resistant to pod borer with 

a pest damage of 3 scale on 1-9. On the other hand, ICCV 10, 

ICCV 97115, ICCV 97119 and ICCV 16381 proved tolerant. 

Bhagwat and Sharma, 2000 tested twenty one chickpea 

genotypes for resistance to H. armigera in glasshouse in 

field condition. Genotypes ICC506, ICCV, ICCL86102 and 

ICCV95992 had a low pod damage rating of 3 (1 to 9 scale). 

ICC86102, a short duration genotype and ICCL86106, a medium 

Table 1. 

Screening of chickpea genotypes for resistance to gram pod borer, H. armigera 

S. No. Genotypes 

larvae/ 10 plants at Pod damage 

larvae/ 10 plants at pod damage 

S. No. Genotypes 

filling stage 

(%) 

filling stage 

(%) 

# H82-2 2.0 19.0 26 RIL 85 1.5 14.0 

* C 235 0.5 6.0 27 ICC 4934 1.0 16.0 

1 DGP 15 1.0 8.5 28 ICC 1433 1.5 12.0 

2 ICCV 96752 1.5 13.5 29 BG 2044 2.0 16.0 

3 PG 700 1.5 15.5 30 BDNG 9-3 2.5 20.0 

4 NDGS 32 2.0 20.5 # H 82-2 0.5 23.0 

5 GL 22067 1.5 14.5 * C 235 1.0 7.5 

6 PG 114 1.5 14.0 31 RIL 115 1.0 8.0 

7 Annigeri 1.0 10.5 32 GL 1362 1.0 12.5 

8 ICC 15894 1.5 11.5 33 Vijay 1.5 15.5 

9 BGM 555 2.0 15.5 34 ICC 79037 1.5 14.5 

10 ICCC 37 2.5 21.5. 35 ICC 506 1.5 15.0 

# H82-2 3.0 21.5 36 ICC 29 1.0 7.5 

* C 235 1.0 6.5 37 ICC 3935 1.0 12.0 

11 GIG 0312 1.0 8.0 38 ICCV 95992 1.5 12.5 

12 ICCL 86111 1.5 12.0 39 BG 256 1.0 11.0 

13 ICC 3137 1.5 14.0 40 BGP 1033 1.5 11.5 

14 Vishwash-12 1.5 13.0 # H 82-2 2.0 19.0 

15 RIL 27 2.0 21.5 * C 235 1.0 5.5 

16 ICL 2000-6 1.5 14.5 41 H 86-18 1.5 12.5 

17 ICCL 87315 1.0 8.0 42 IC 14872 1.5 10.5 

18 ICC 8923 1.5 12.5 43 ICCL 87316 1.5 14.0 

19 ICC 5780 2.0 14.5 44 ICC 12470 1.0 8.0 

20 ICCV 10 1.02.5 7.5 45 RSG 888 1.0 13.0 

# H82-2 1.0 21.0 46 DCP 8 2.0 20.5 

* C 235 1.0 6.0 47 PG 32 1.5 12.0 

21 ICCV 7 1.5 6.5 48 ICCV 4959 1.0 8.0 

22 RIL 81 1.0 12.5 49 Udai 2.0 20.5 

23 ICC 15869 1.5 10.5 50 ICC 12479 2.0 20.0 

24 ICCL 87314 1.5 15.5 # H 82-2 2.0 21.0 

25 Radhe 1.5 13.5 * C 235 1.0 7.0 

#Check susceptible genotype 

*Check resistant genotype

Table 2. 

KUMAR et al., Screening of chickpea genotypes for resistant against pod borer, Helicoverpa armigera Hubn. 103 

Identification of promising chickpea genotypes against H. armigera 

Grade Damage rating stage Number of genotypes Genotypes identified 

Tolerant 1 9 DGP 15, GIG 0312, ICCL 87315, ICCV 7, RIL 115, ICC 29, ICC 12470, 

ICCV 10 and PG 23 

Moderate resistant 2 34 ICCV 96752, PG 700, GL 22067, PG 114, Annigeri, ICC 15894, BGM 555, 

ICCL 86111, ICC 3137, Vishwash 12, ICL 2000 6, ICC 8923, ICC 5780, 

RIL 81, ICC 15869, ICCL 87314, Radhe, RIL 85, ICC 4934, ICC 1433, BG 

2044, GL 1362, Vijay, ICC 79037, ICC 506, ICC 3935, ICCV 95992, BG 

256, BGP 1033, H 86-18, IC 14872, ICCL 87316, RSG 888, PG 32 

Susceptible 3 7 NDGS 32, ICCC 37, RIL 27, DCP 8, BDNG9-3, Udai and ICC12479 

duration genotype were identified as a promising source of 

resistance to H. armigera. The lowest yield (772 kg/ ha) was 

produced by the genotype Chaffa due to its smaller grain size 

and lowest yield potential. Sanap and Jamadagni, 2005 

screened 25 genotypes and observed that resistant genotypes 

ICC506EB, ICC13, ICCX730014, ICC9854, ICC5800 and 

ICC12493, with a rating of 7 and ICC926, ICC19492 and 

ICCL87220 with a rating 8, were susceptible to H. armigera. 

Rai and Ramujagir, 2005 screened chickpea genotypes and 

reported that two genotype Avrodh and L550 as susceptible, 

four genotypes (ICCB93929, ICCB96029, ICCE96030 and ICCB2 

as resistant to gram pod borer with a pest damage score of 3 

on a scale of 1-9. On the other hand, ICCV10, ICCV97115, 

ICCV97119 and ICC16381 proved tolerant against H. armigera. 


Afzal, M., Ghafoor, H.A., Javed, A. and Sabri, M.A. 2001. Role of 

physico-morphic characters in different varieties and advanced 

lines of chickpea towards resistance against Helicoverpa armigera 

(Hub.). Pakistan Entomologist, 23(1/2): 71-74. 

Atwal, A.S. and Dhaliwal, G.S. 2005. Agricultural Pests of South Asia 

and their Management, Kalyani Publisher, New Delhi. 

Bhagwat, V.R. and Sharma, S.B. 2000. Evaluation of chickpea genotypes 

for resistance to legumes pod borer, Helicoverpa armigera and 

root-knot nematode, Meloidogynejavanica. Indian J. Plant Prot., 

28(1): 69-73. 

Gowda, D.K.S., Sharanabasappa and Halle, D. 2005. Screening of 

resistant chickpea genotypes for Helicoverpa armigera (Hub.) 

Karnataka J. Agric. Sci., 18(4): 1107-1108. 

Gumber, R.K., Singh, Sarvjeet, Kular, J.S., Singh, Kuldip, Singh, S. and 

Singh, K. 2000. Screening of chickpea genotypes for resistance to 

Helicoverpa armigera. Intl. Chickpea, Pigeonpea, Newsl., 7: 20- 

21. 

Mandal, S.M.A. 2003. Screening of some chickpea genotypes against 

Helicoverpa armigera. Environment and Ecology, 21(1): 240-241. 

Rai, D. and Ramujagir 2005. Screening of chickpea genotypes for 

resistance to pod borer Helicoverpa armigera. Indian J. Agric. 

Sci., 7(2): 120-122. 

Sanap, M.M. and Jamadagni, B.M. 2005. Screening of chickpea for 

resistance to pod borer, Helicoverpa armigera (Hub.) at Rahuri, 

Maharashtra, India. Intl. Chickpea and Pigeonpea Newsl., (12): 

37-39. 

Singh, B. and Yadav, R.P. 1999. Field screening of chickpea (Cicer 

arietinum L.) genotypes against gram pod borer (Helicoverpa 

armigera Hub.) under late sown conditions. J. Ent. Res., 23(2): 

133-140. 

Vaishampayan, S.M. and Veda, O.P. 1980. Population dynamics of 

gram pod borer (Helicoverpa armigera) in chickpea at Pantnagar 

(U.P.), Indian J. Pl. Prot., 15: 39-41. 



Knowledge Level of Farmers About Recommended Cultivation Practices of 

Groundnut in Panchayat Samiti Govindgarh of District Jaipur (Rajasthan) 

H.N. VERMA AND J.P. YADAV 

Department of Extension Education, S.K.N. College of Agriculture, Jobner, Jaipur 303 329, 

S.K. Rajasthan Agricultural University, Bikaner 334 006 

email: kataria.exted@gmail.com 

ABSTRACT 

Despite of the facts, the groundnut contributes about two-fifth 

of the total area and production of oilseeds in country. India is 

facing acute shortage of vegetable oils. This is mainly due to 

the low productivity of these crops. This may be attributed to a 

number of factors and constraints, which hinders the adoption 

of the innovations. These factors and constraints should be 

identified and remove by developing an effective strategy for 

farmers. 

Key word 

Constraints, Adoption, Groundnut Cultivation, 

innovation 

Indian Economy depends upon Indian agriculture. 

About 72.20% population lives in rural areas. The main 

occupation of rural people is agriculture. About 30% of the 

national income originates from the agriculture sector. About 

75% of its population and 66.67% of labour force directly or 

indirectly is dependent on agriculture for livelihood. Large 

number of important industries like jute, textiles, edible oils, 

tobacco, sugar etc. receive the raw materials produced by 

agriculture sector. All type of crops representing cereals, 

pulses, oilseeds, fibers, species and condiments and many 

others are gown in our country. 

Groundnut oil is used in manufacturing of soap, hair oil, 

vanaspathi, lubricants, textile, auxiliaries etc. The oil find 

extensive use as a cooking medium both in its refined form 

and as vanaspati ghee. The oil cakes obtained after the 

extraction of oil from the groundnut are valuable organic 

manure and used as animal feed. 

Groundnut is used for edible and non-edible purposes. 

About 81.6% of the total production is crushed and used for 

edible purpose. The remaining production goes for seed 

(12%), feed (5.3%) and exports (1.1%). (Anonymous, 2006-07) 


The study was conducted in Govindgarh Panchayat 

Samiti of Jaipur district. Eight villages namely Ghinoi, Bailabas, 

Jheera, Samod, Nindola, Khejroli, Hathnoda and Nangal 

Bharda from four selected Gram Panchayat were selected 

with the help of simple random sampling technique. From each 

selected village, a list of all groundnut growers was prepared 

and 20% groundnut growers were selected from each selected 

village by using proportionate random sampling technique. 

In total, a sample of 124 farmers were drawn for the study 

purpose. The adoption level of farmers of recommended 

cultivation practices of groundnut is directly or indirectly 

related to knowledge level of farmers about different 

recommended cultivation practices of groundnut. Hence, it 

was considered necessary to assess the knowledge level of 

farmers about groundnut cultivation. The knowledge about 

the technology had influence on the decision making about 

its adoption keeping this view in mind the study “Knowledge 

level of farmers about recommended cultivation practices of 

groundnut in Panchayat Samiti Govindgarh of district Jaipur 

(Rajasthan)” was undertaken with the specific objectives : To 

measure the knowledge level of farmers about recommended 

cultivation practices of groundnut. 

In all, questions were included in the schedule to test 

the knowledge of the respondents. Equal weightage was given 

to each item, assuming that all the items included were equal 

in difficulty to understand, apply and recall. One mark was 

given to every right answer and zero for wrong answer. The 

following formula was used to work out the knowledge index. 

Knowledge index = X 1 

+ X 2 

+ X 3 

+ ——————— X n 

Where X 1 

, X 2 

, X 3 

, …………….X n 

are correct answer for 

first, second, third ………….. X n 

questions and n is the 

maximum score possible to secure or the number of question 

i.e. 12 and the minimum was zero. 

The mean and standard deviation of scores secured by 

all the respondents were computed for classifying the 

knowledge level in different categories. Based on the 

knowledge scores, three levels of knowledge of farmers were 

categorized as under 

(a) Low knowledge level : obtained score below (Mean – 

Standard deviation) 

(b) Medium knowledge level : obtained score from (Mean – 

Standard deviation) to (Mean + Standard deviation) 

(c) High knowledge level: Obtained score above (Mean + 

Standard deviation). 


Interview schedule consisting of face data of the 

respondents measuring device of knowledge level was 

prepared for the investigation in light of the suggestions of 

the experts. The researcher himself personally interviewed 

the respondent and the statistical data regarding the 

knowledge of respondents about recommended cultivation 

practices of groundnut have been given in Table 1.

VERMA AND YADAV, Knowledge Level of Farmers about Recommended Cultivation Practices of Groundnut 105 

Table 1. 

Knowledge level of farmers about recommended 

cultivation practices of groundnut N = 124 

S.No. Knowledge categories Frequency Per cent of 

farmers 

1 Low knowledge (score below 27.49) 22 17.74 

2 Medium knowledge (score from 74 59.68 

27.49 to 35.55) 

3 High knowledge (score above 33.55) 28 22.58 

X = 30.52, = 3.02, 2 = 39.16, Expected frequency = 41.33 

The data in Table 1 state that on the whole, 59.68% (74) 

groundnut growers were having medium knowledge level 

about groundnut cultivation practices and 22.58% (28) 

respondents were having high knowledge level, whereas 

17.74% (22) farmers were having low knowledge level about 

recommended cultivation practices of groundnut. Hence the 

null hypothesis (H 01.1 

) ‘There is no difference in the level of 

knowledge of the farmers about recommended cultivation 

practices of groundnut’ was rejected. It means there was a 

difference in the knowledge level of the farmers growing 

groundnut. 

Further, more the percentage of farmers having 

knowledge about different aspects of recommended 

cultivation practices of groundnut were analysed separately. 

The relative importance of the 10 aspects of recommended 

cultivation practices of groundnut was highlighted by ranking 

them on the basis of the mean per cent scores of farmers 

having knowledge about these recommended cultivation 

practices of groundnut (Table 2). 

The data in Table 2 indicate that 80.50% farmers had 

highest knowledge about ‘Sowing time’ and hence, it was 

ranked first. The second highest per cent of farmers (75.50) 

had knowledge about ‘Recommended seed rate’ followed by 

75.00% farmers who had knowledge about ‘Irrigation 

management’ were ranked third whereas, 70% farmers had 

knowledge about ‘Spacing’ were ranked fourth, while 68.01% 

farmers were having knowledge about ‘Seed treatment’ were 

ranked fifth. Furthermore, sixth, seventh, eight and ninth ranks 

were awarded to weed management (65.10 MPS), plant 

protection measures (50.22 MPS), depth of sowing (50.03 MPS) 

and use of high yielding variety (50.02 MPS), respectively. 

The lowest knowledge of farmers was found about fertilizer 

application with 40.21 MPS, hence it was awarded lowest rank 

i.e. tenth. 

It leads to the conclusion that farmers in general had 

medium knowledge about the recommended cultivation 

practices of groundnut. This low knowledge may be attributed 

due to low literacy, low exposure to mass media, unavailability 

of extension literature and less contact with extension 

personnels. 

From the findings, it is also evident that majority of the 

farmers were having higher knowledge about the ‘Sowing 

time’, ‘Seed rate’, ‘Irrigation management’, and ‘Spacing’. This 

might be due to the reason that majority of the farmers were 

regularly growing groundnut for market purpose and these 

Table 2. 

Practicewise knowledge level of farmers about 

recommended cultivation practices of groundnut 

N = 124 

S.No. Recommended practices Knowledge in mean per cent Rank 

1. Use of high yielding variety 50.02 IX 

2. Seed rate 75.50 II 

3. Seed treatment 68.01 V 

4. Spacing 70.04 IV 

5. Sowing time 80.50 I 

6. Depth of sowing 50.03 VIII 

7. Fertilizer application 40.21 X 

8. Irrigation management 75.00 III 

9. Weed management 65.10 VI 

10. Plant protection measures 50.22 VII 

Overall 62.46 

practices are most critical from the point of view of the 

groundnut production. A slight carelessness in these practices 

may reduce the production of groundnut drastically, so the 

farmers remain most careful about these practices. Also for 

producing good quality groundnut, they mostly remain in 

contact with the extension agencies, sale agents etc. resulting 

gain in knowledge about these recommended cultivation 

practices. Most of the farmers under study were literate hence 

they might have knowledge about these practices by reading 

the related literature. The farmers had medium knowledge 

about ‘Seed treatment’, ‘Weed management’, ‘Plant protection 

measures’, ‘Depth of sowing’, ‘Use of high yielding varieties’ 

and ‘Fertilizer application’. This might be due to the reason 

that the farmers might not understand the intructions written 

on the pack of chemical because of its complex language, as 

the instructions are mostly written in typical Hindi or English 

language or in the language of the state, where the insecticides, 

fungicides, weedicides etc. are manufactured. 

Majority of the groundnut growers had medium 

knowledge level about the recommended cultivation practices 

of groundnut. Among the various aspects of different 

cultivation practices, majority of the farmers had knowledge 

about ‘Sowing time’, ‘Seed rate’ ‘Irrigation management’, 

‘Spacing’, ‘Seed treatment’, ‘Weed management’ and ‘Plant 

protection measures’. 


Agarwal, J.K. 2000. Knowledge and adoption of improved pea 

cultivation practices in Jaipur district of Rajasthan. M.Sc. (Ag.) 

Thesis, R.A.U., Campus-Jobner. 

Anonymous. 2006-07. Economic Survey. Ministry of Agriculture, Govt. 

of India. 

Kumar, M. 2004. Adoption of improved cultivation practices of cabbage 

in Jaipur district of Rajasthan. M.Sc. (Ag.) Thesis, R.A.U, Campus- 

Jobner. 

Subhash, C. 2001. Factors affecting adoption of recommended production 

technology of groundnut in panchayat samiti Sambhar lake of district 

Jaipur (Rajasthan). M.Sc. (Ag.) Thesis, R.A.U., Campus-Jobner. 

Yadav, B.C. 2006. A study on knowledge and adoption of improved 

production technology of mandarin by the farmers in Jharapatan 

panchayat samiti of Jhalawar district of Rajasthan. M.Sc. (Ag.) 

Thesis, R.A.U., Campus Jobner. 



Stability Studies on Sewage Irrigation Effect on Grain Yield and Protein-content in 

Mungbean [Vigna radiata (L.) Wilczek 

DYUTI PANDEY, P.S. SHUKLA AND D.N. SHUKLA 

Department of Botany, University of Allahabad, Allahabad 211 002 

email: dyutipandey@gmail.com 

ABSTTRACT 

Ten genotypes of mung bean viz., Samrat (PDM 139), Kopergaon, 

MUM 1, RMG 62, Phule M-2, Pusa 9531, Vamban 1, PDM 11, 

PDM 96-262, PDM 99-284, were evaluated in four environments 

i.e. plain water(E 1 

) , 

sewage water(E 2 

), plain water+15 kg N/ha(E 3 

) 

and sewage water +15 kg N/ha(E 4 

) . The stability parameters, 

mean value (x), regression coefficient (b i 

) and deviation from 

regression (S 2 ) were studied for yield per plant and protein 

di 

content in various environments. In the study significant 

difference s we re o bserved for geno type s in differe nt 

environments. Treatment E 1 

gave the highest yield and E 3 

gave 

the highest protein- content. Variety PDM 11 showed high mean 

values for yield per plant in treatment E1, followed by PDM 96- 

262 and for protein- content. PDM 11 and Phule M-2 both were 

found highly significant, they were considered to be stable 

genotypes in different environment. 

Key words 

Genotypes, environment, mung bean, sewage water, 

protein content 

Mungbean (green gram), which is grown throughout 

the year in one or the other part of the country, has array of 

variability for maturity duration and plant type. In North India, 

it is grown in rainy as well as in summer season. Sewage water 

irrigation provides organic matter with macro- and micronutrients 

which are essential for the growth and development 

of mung bean. Commercial agricultural procedures, poor 

genetic variability and lack of authentic genetic information 

are the main constraints in the way to select suitable varieties 

which can be grown successfully if only sewage water 

irrigation facility is available their. Under these circumstances, 

it becomes imperative to find out genotypes with their stability 

possessing wider adaptability when irrigated with sewage 

water. Genetic, Environmental and Genetic x Environmental (G 

x E) interactions are of considerable importance in the selection 

of desirable genotypes. In the present study stability 

parameters for yield per plant and protein content were studied 

for ten varieties of mung bean in four environments (irrigations) 

to find out the effect for the same. 


The experimental material consisted of 10 promising 

genotypes of mungbean–Samrat (PDM 139), Kopergaon, 

MUM 1, RMG 62, Phule M 2, Pusa 9531, Vamban 1, PDM 11, 

PDM 96-262, PDM 99-284, which were grown in four 

environments viz., plain water (E 1 

), sewage water (E 2 

), plain 

water +15 kg N/ha (E 3 

) and sewage water +15 kg N/ha (E 4 

) 

under randomized block design (RBD) with three replications, 

keeping single row of genotypes with inter- and intra-row 

distances of 45 cm and 15 cm, respectively .The entire 

experiment was carried out in two parts. In the first part, twenty 

genotypes were grown in sewage water and plain water, 

simultaneously. Out of twenty varieties, ten varieties, which 

were found significant regarding tolerance to sewage water 

and were comparatively better in yield as well as in other yield 

attributes, were selected for the second experiment consisting 

of stability study. The weight of seeds of five randomly 

selected plants of each genotype was recorded and the final 

grain yield per plant was worked out. The protein content was 

estimated in per cent by chemical analysis of grains. Stability 

analysis was carried out. 


It is evident from the ANOVA (Table 2) that there were 

significant differences among genotypes, environments and 

genotypes x environment (GxE) interaction. The significant G 

x E interaction revealed that genotypes responded differently 

to the changes in the environment. Different environments 

were provided at one location (Luthra, et al., 1974). The 

average performance of genotypes with respect to both the 

traits varied significantly. The environmental interactions, both 

linear and non-linear, were significant for both the characters. 

A genotype may be considered to be stable over different 

environments if it shows unit or less than unit regression 

coefficient (b i 

) with the lowest deviation (non-significant) from 

the linear regression (S 2 d i 

) (Rai, et al., 1989). With this view, 

high and desirable mean performance of a variety over 

environments is also supposed to be a positive consideration 

to rank the variety as a better and stable genotype. 

The genotypes PDM 11 was found having highest mean 

value for grain yield per plant and protein-content. For yield 

per plant, unit or less than unit regression coefficient and 

non-significant deviation from the linear regression (S 2 di) was 

observed for Kopergaon, MUM 1 and PDM 99-284 (Table 2). 

Regarding protein-content, genotypes Samrat, Kopergaon, 

MUM 1, RMG 62 and PDM 11 were found having regression 

coefficient (bi) less than one with non-significant deviation 

from linear regression (S 2 di). Therefore, on the basis of these 

results, genotypes Kopergaon and MUM 1 can be 

recommended as stable for wide range of environments i.e. 

non-sewage and sewage irrigation.

Table 1. 

PANDEY et al., Stability Studies on Sewage Irrigation Effect on Grain Yield and Protein-content in Mungbean 107 

Estimates of Mean, Range, Standard Error and Coefficient of Variation in respect to grain yield per plant and 

proteincontent for 10 genotypes of Mungbean over four environments. 

S. Genotypes 

Grain yield/plant 

Protein content 

No. 

E 1 E 2 E 3 E 4 Pooled E 1 E 2 E 3 E 4 Pooled 

1 Samrat (PDM139) 4.5 4.84 5.24 5.43 5.01 22.89 23.19 23.74 23.73 23.39 

2 Kopergaon 5.58 5.41 5.36 5.59 5.48 23.78 23.86 24.41 24.61 24.16 

3 MUM 1 6.8 6.81 6.84 6.91 6.84 24.81 25.11 25.4 25.16 25.12 

4 RMG 62 4.98 5.38 5.38 5.53 5.32 22.47 23.1 23.43 23.43 23.11 

5 Phule M 2 6.54 5.07 4.98 5.19 5.44 24.77 25.54 25.88 25.86 25.51 

6 Pusa 9531 4.55 5.21 5.68 5.59 5.26 23.12 23.4 24.16 24.23 23.73 

7 Vamban 1 4.51 4.69 4.7 4.78 4.67 25.18 25.97 26.78 26.45 26.09 

8 PDM 11 10.42 8.36 7.49 8.33 8.65 26.08 26.22 26.51 26.5 26.33 

9 PDM 96-262 8.91 8.2 7.53 8.12 8.19 24.69 25.26 26.2 24.82 24.48 

10 PDM 99-284 7.05 7.03 6.88 7.26 7.04 23.83 25.27 24.99 24.82 24.48 

Range among genotypes 4.50-10.42 4.69- 8.36 4.70- 7.53 4.78-8.33 4.67-8.65 22.47-26.08 23.10 -26.22 23.73 - 26.78 23.43-26.50 23.10 -26.33 

G. Mean 6.38 6.09 6.01 6.26 6.18 24.16 24.59 25.14 25.08 24.75 

S.E. Diff. of Mean± 0.66 0.55 0.18 0.2 0.18 0.12 0.17 0.67 0.23 0.79 

Coefficient of Variation 1.48 1.28 4.36 4.33 8.23 0.68 0.97 0.38 1.32 0.9 

Table 2. ANOVA for stability parameters for different 

characters of mungbean genotypes. 

Source of variation d. f. Protein content Grain yield per plant 

Genotypes (G) 9 5.18** 7.82** 

Environments (E) 3 2.13** 0.28* 

G X E 27 0.038** 0.29** 

Environments + ( G X E) 30 0.25** 0.30** 

Environments (Linear) 1 6.4** 0.85** 

G X E (Linear) 9 0.081** 0.58** 

Pooled deviations 20 0.014 0.14** 

Pooled error 108 0.05 0.25 

*Significant at p=0.05; **Significant at p=0.01 


Luthra, O.P., Singh, R.K. and Kakar, S.N. 1974. Comparison of different 

stability models in wheat. Theoretical Applied Genetics, 45: 143- 

49. 

Rai, M., Kerkhi, S.A., Pandey, S., Naqvi, P.A. and Vashistha 1989. 

Stability analysis for some quality components of seed and oil in 

linseed (Linum usitatissimum L.) Indian Journal of Genetics. 49(3): 

291-295. 



In Vitro Screening and Identification of Putative Sunflower (Helianthus annus L.) 

Transformants Expressing ECNAC1 Gene by Salt Stress Method 

K.C. MANJUNATH 1&2 , A. MAHADEVA 2 , ROHINI SREEVATHSA 2 , N. RAMACHANDRA SWAMY 1 AND 

T.G. PRASAD 2 

1 

Department of Biochemistry, Bangalore University, Central College, Bangalore 560 001, Karnataka, India 

2 

Department of Crop Physiology, University of Agricultural Sciences, GKVK, Bangalore 560 065, Karnataka, India 

email: kcmanjunath81@gmail.com 1 

ABSTRACT 

A simple and effective screening methodology to identify 

transformants at plant level under NaCl stress has been 

established. Leaf disc of putative transformants (T 1 

individual, 

6-8 weeks old) and wild type plants were placed on filter paper 

in the presence of NaCl (200 mM) for 48 hr at room temperature. 

The leaf discs of EcNAC1 gene transformed plants showed salt 

tolerance by retaining green colour after 48 hours, while leaf 

discs from non-transformed plants turned dark brown or black 

in colour. Further, molecular analysis of these salt tolerant 

plants showed PCR positive to EcNAC1 gene specific and HPT 

II specific primers, and digestion of amplified EcNAC1 gene 

product with Sac I restriction enzyme showed expected bands 

size on agarose gel. Our data suggest that in vitro screening 

strategy at plant level based on the target gene incorporated 

would result in the initial identification of promis ing 

transformants for further analysis. 

Key words 

Abiotic stress, EcNAC1, Transcription Factor, 

Methodology, Transgenics 

The salt stress and dehydration stress show a high 

degree of similarity with respect to physiological, biochemical, 

molecular and genetical effects (Cushman, et al., 1990). 

Introducing a single regulatory gene improved both drought 

and salt tolerance in plants reported by many researchers 

(Yong Xiang, et al., 2008 and Xingnan Zheng, et al., 2009). 

Transcription factors have proven quite useful in improving 

stress tolerance in transgenic plants, through influencing 

expression of a number of stress-related target genes. Among 

this, the NAC transcription factor proteins constitute one of 

the largest families of plant-specific transcription factors 

involved in response to various environmental stresses. 

Collectively, many reports indicate that overexpression of NAC 

transcription factor conferred abiotic stress tolerance in plants 

(Honghong, et al., 2006 and Venkategowda Ramegowda, et 

al., 2012). 

The GUS assay (beta glucuronidase assay) is an easy 

method to determine the transient expression of the reporter 

gene (Jefferson, et al., 1987). This assay is based on 

fluorescence emitted by the product of GUS substrate X-gluc, 

which makes it very expensive. The selectable marker gene 

activity such as spraying on the whole plants or leaf with 

herbicide (Datta, et al., 1992), germination of seeds on selective 

media (Hiei, et al., 1994) are being used to screen putative 

transformants. Screening the putative transformants based 

on the function of target gene incorporated will give the 

confirmation of the gene incorporated along with the abiotic 

stress tolerance. In this paper, we report a simple and efficient 

in vitro screening strategy for the selection of putative 

transformants based on the performance of target gene by 

salt stress method. 


Plant material 

The sunflower (Helianthus annuus L.) inbreds seeds 

(IB20) were procured from Sunflower Scheme, UAS, GKVK, 

Bangalore, India. The Agrobacterium culture harboring 

EcNAC1 (pG-CaMV 35S-EcNAC1 with hpt II) gene was 

procured from Dept of Crop Physiology, UAS, GKVK, 

Bangalore, India. The Agrobacterium culture was used for 

plant transformation in sunflower inbred line by following in 

planta transformation protocol (Sankara Rao and Rohini, 1999). 

The seedlings with just emerging plumule were infected by 

pricking at the meristems with a sterile needle and immersed in 

a suspension of Agrobacterium for 1 hr. Then, the seedlings 

were washed with sterile water and transferred onto autoclaved 

soilrite (vermiculite equivalent) moistened with water for 

germination under aseptic conditions in glass jars of 200 ml 

capacity with 3 seedlings per jar. After 5 to 6 days, the seedlings 

were transferred to pots and allowed to grow in green house 

condition. The T 1 

seeds of the above transformants were used 

for the development of screening strategies. 

Standardization of stress inducers concentration 

Leaf discs from fully expanded leaves of 6-8 weeks old 

wild type sunflower (IB20) plants were subjected to 50, 100, 

150, 200 and 250 mM NaCl concentration, incubated for 48 hr 

at room temperature. The induction of chlorosis in the leaf 

discs were recorded after 48 hr incubation. The final screening 

concentration was fixed based on the concentration at which 

the green colour leaf discs turns to complete dark brown or 

black. 

Screening putative transformants at plant level by stress 

induced leaf damage 

Leaf discs from fully expanded leaves of 6-8 weeks old 

putative transformants and wild type were placed on filter

MANJUNATH et al., In Vitro Screening and Identification of Putative Sunflower (Helianthus annus L.) 109 

paper dampened with 200 mM NaCl. They were incubated for 

48 hr at room temperature and the symptom of bleaching or 

browning was documented. The salt tolerant plants were 

selected for further molecular analysis. 

Molecular Analysis 

PCR Analysis 

DNA isolated from the leaves of salt tolerant and wild 

type plants used as templet DNA for polymerase chain 

reaction along with the primers EcNAC1 FP 5’- 

CAGGATCCATGACCATGGGAGGAGGG-3’ and RP 5’- 

CCTAGAATTCTGTATTTACAGAGGTCGC-3’, HPT II FP 5’- 

CCTAGAATTCTGTATTTACAGAGGTCGC-3’ and RP 5’- 

ATCGCCTCGCTCCAGTCAATG-3’. Amplification was 

performed in the total volume of 15 ìl containing 1.5 ìl of 10X 

buffer (Fermentas, U.S.A), 1.5 ìl of 2 mM dNTP, 1 ìl of genomic 

DNA (100 ng), 5 pmol of each forward and reverse primer, and 

1ìl (2.5 U/ìl) unit of Taq polymerase. PCR were carried out with 

a Eppendorf thermal cycler by using initial denaturation at 

94 o C for 5 min followed by 30 cycles consisting of 94 o C for 30 

sec, 55 o C for 30 sec and 72 o C for 90 sec, a final extension step 

consisting of 72 o C for 10 min. The resulting amplified product 

resolved on 1% agarose gel. 

Restriction Digestion Analysis 

The Gene specific PCR products were purified by PCR 

purification kit (Qiagen, Germany) according to the 

manufacturer’s instruction and the PCR product was digested 

with Sac I restriction enzyme (Fermentas, USA). The restriction 

digestion were performed in the total volume of 20 ìl containing 

2 ìl of gene specific PCR product, 2 ìl of Sac I buffer B (Blue) 

1x, 1 ìl of Sac I restriction enzyme and 15 ìl of nuclease free 

water. The digestion was carried out for 1 hour and the resulting 

digested products were resolved on 1% agarose gel. 


The development of transgenic plants is a potential 

option to improve abiotic stress tolerance, the screening for 

superior transgenic lines is therefore a crucial step. The use 

of an inplanta transformation protocol results in the 

development of a large number of T 0 

plants and in turn a large 

number of T 1 

generation plants. Screening of a large number 

of plants requires standardization of high throughput 

screening procedures based on either the selectable marker 

or the target gene incorporated. The leaf tip assay was reported 

by Noor, et al., 2000 to screen putative transgenic rice and 

cotton based on selectable marker hpt II or npt II. This leaf tip 

assay doesn’t give information about the performance of the 

target gene incorporated in the plant to combat abiotic stress. 

In this study, screening the putative transformants based on 

target gene incorporated. This strategy utilized leaves of 6-8 

week old plants. The leaf disc bioassay is an example of a 

quick method to measure salt induced damage (Singla-Pareek, 

et al., 2003 and Sanan-Mishra, et al., 2005). This technique 

reveals the ability of the tissue to tolerate Na + accumulation 

and eliminates the consequence of altered ion transport from 

root to shoot. 

Standardization of Salt concentration 

Detection of salt concentration to screen putative 

transformants was carried out by subjecting the leaf discs of 

wild type sunflower plant (IB20) on 50, 100, 150, 200, 250 mM 

Fig. 1. 

Maximum salt tolerance limits of the wild type sunflower (IB20) plants detected in different salt concentration. (a) 0 mM 

(Distil water); (b to f): 50, 100, 150, 200, 250 mM NaCl concentration. 

Fig. 2. 

Screening putative transformants at plant level by 200 mM NaCl induced leaf damage (a-d). Wt: wild type, 1-15: putative 

transformants.


NaCl, incubated for 48 hr at room temperature. No colour 

change was observed in 50 mM NaCl, while gradual colour 

change appeared in 100 and 150 mM NaCl concentration, while 

the leaves turned complete dark brown or black colour in 200 

and 250 mM NaCl concentration (Fig 1). This indicated that 

200 mM NaCl concentration was ideal for screening putative 

sunflower transformants. 

Screening putative transformants at plant level by stress 

induced leaf damage 

A total of 860 T 1 

plants were raised from the 24 T 0 

individual plants and the leaf discs of each individual plant 

were subjected to salt stress. Leaf discs of putative 

transformants along with the wild type plants were placed on 

filter paper dampened with 200 mM NaCl and incubated for 48 

hr at room temperature and scored subsequently. The leaves 

of tolerant plant retained green colour, while susceptible and 

wild type treated plant leaves turned dark brown or black 

colour after 48 hr incubation (Fig. 2). Out of 860 T 1 

plant only 

10 plants showed salt tolerance and selected for further 

molecular analysis. 

PCR Analysis 

The gene integration was confirmed in 10 salt tolerant 

plants. The selected 10 salt tolerant plants showed PCR 

positive for both Gene specific and HPT II primers with the 

product size of 1.1 kb and 0.5 kb respectively (Fig. 3a & 3b). 

The transgenic nature of the tolerant plants was further 

confirmed by the polymerase chain reaction with gene specific 

primer for salt susceptible plants that showed absence of 1.1 

kb band (Fig. 3c). 

Restriction Digestion Analysis 

Restriction digestion analysis were carried out in 5 PCR 

amplified EcNAC1 gene product with Sac I restriction enzyme. 

The digested product showed expected band size of 700 bp 

and 400 bp on 1% agarose gel (Fig. 4). 

In this assay both wild type and putative transformants 

leaf discs placed on single filter paper to reduce the variations 

in the treatment and also easy to compare colour change 

between wild type and transgenics. The salt tolerance 

exhibited by transgenic plants expressing EcNAC1 

transcription factor will be due to up-regulation of downstream 

salt responsive gene. Ramegowda, et al., 2012 reported that 

EcNAC1-confers abiotic stress tolerance in tobacco. The leaf 

discs of wild type plant exhibits tolerance to lower salt 

concentration (50 mM), this may be due to transporting Na + 

ion into the vacuole, but high salt concentration exceed the 

limit of vacuolar storage capacity or transport efficiency, which 

in turn causes oxidative stress. The accumulation of reactive 

oxygen species (ROS) increases within the leaf discs and 

damage the chloroplast, which turns green colour to dark 

brown or black. Therefore, the plants retained green colour 

was considered as putative transformants and selected for 

further molecular analysis. Salt tolerant plants showed 

amplification of EcNAC1 and HPT II genes, which indicates 

stable gene integration. Further confirmation was done by 

digesting the EcNAC1 gene products of 5 plants with Sac I 

restriction enzyme which resulted in the expected band size 

on agarose gel. The leaf disc assay is being used in general to 

assess whether the transgene brings about tolerance to the 

stress and to assess the relative difference. 

Fig. 3. 

PCR analysis for Salt tolerant Plants: (a) PCR amplified 1.1 Kb EcNAC1 gene products (b) PCR amplified 0.5 Kb HPT II 

gene products. M: 1kb DNA ladder (Fermentas U.S.A); P: Plasmid (Positive control); Salt Susceptible plants (c) PCR reaction 

with gene specific primer. [B: blank (PCR cocktail without Taq polymerase); W: Wild type (Negative control); 1-10: Putative 

transformants.

MANJUNATH et al., In Vitro Screening and Identification of Putative Sunflower (Helianthus annus L.) 111 

Fig. 4. 

PCR amplified five EcNAC1 gene products digested 

with Sac I restriction enzyme M: 1kb DNA ladder; 1- 

5: Putative transformants. 

In this study the emphasis was to screen the transformed 

segregants to identify putative transformants with stable 

integration and expression. Understanding the molecular basis 

will be helpful in developing selection strategies for improving 

abiotic stress tolerance. The stringent primary screening 

strategy as a part would help in identification and better 

understanding of the phenotypes associated with abiotic 

stress tolerance. In vitro Screening of putative transformants 

based on target gene incorporated is an effective methodology 

to drastically narrow down the potential tolerant lines. 


Program supported by: ICAR –NAE: 10-(6)/2005 EP & D 

and are gratefully acknowledge DBT-COE: BT/01/COE/05/03 


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during adaptation to salt stress. In: Environmental Injury of Plants 

(ed. Kalterman, F.), Academic Press, San Diego, pp.173-203. 

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after PEG-mediated transformation of protopiasts. Plant. Mol. 

Biol, 20: 619-629. 

Freyssinet, M. and Freyssinet, G. 1998. Fertile plant regeneration from 

sunflower (Helianthus annuus L.) immature embryos. Plant Sci., 

56: 177-181. 

Hiei, Y., Ohta, S., Komari, T. and Kumashiro, T. 1994. Efficient 

transformation of rice (Oryza sativa L.) mediated by Agrobacterium 

and sequence analysis of the boundaries of the T-DNA. Plant J, 6: 

271-282. 

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Qifa Zhang, Lizhong Xiong. 2006. Overexpressing a NAM, ATAF, 

and CUC (NAC) transcription factor enhances drought resistance 

and salt tolerance in rice. Proc. Natl. Acad. Sci., 103(35): 12987- 

12992. 

Jefferson, R.A., Burgess, S.M. and Hirsh, D. 1987. Beta glucuronidase 

from E. coli as a gene fusion marker. Proc. Natl. Acad. Sci. USA, 

88: 8447-8451. 

Rohini, V.K. and Sankara Rao, K. 2000. Transformation of peanut 

(Arachis hypogaea L.): A non-tissue culture based approach for 

generating transgenic plants. Plant Sci., 150(1): 41-49. 

Samina Noor, Tayyab Husnain, Sheikh Riazuddin. 2000. Screening for 

putative transgenic rice and cotton plants: A simple and easy method. 

Pakistan Journal of Pakistan, 3(12): 2226-2228. 

Sankara Rao, K. and Rohini, V.K. 1999. Agrobacterium-mediated 

transformation of sunflower (Helianthus annuus L.): A simple 

protocol. Ann. Bot., 83: 347-354. 

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cultivars of safflower (Carthamus tinctorius L.) using Agrobacterium 

tumefaciens. Plant Biotechnology, 16(3): 201-206. 

Sanan-Mishra, N., Pham, X.H., Sopory, S.K. and Tuteja, N. 2005. Pea 

DNA helicase 45 overexpression in tobacco confers high salinity 

tolerance without affecting yield. Proc. Natl. Acad. Sci. USA, 102: 

509-514. 

Singla-Pareek, S.L., Reddy, M.K. and Sopory, S.K. 2003. Genetic 

engineering of the glyoxalase pathway in tobacco leads to enhanced 

salinity tolerance. Proc. Natl. Acad. Sci. USA, 100: 14672-14677. 

Venkategowda Ramegowda, Muthappa Senthil-Kumar, Karaba, N. 

Nataraja, Malireddy K. Reddy, Kirankumar, S. Mysore, Makarla 

Udayakumar. 2012. Expression of a finger millet transcription 

factor, EcNAC1, in tobacco confers abiotic stress-tolerance. PLoS 

ONE, 7(7): e40397. 

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of a NAC transcription factor enhances reice drought and salt 

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Characterization of OsbZIP23 as a key player of the basic leucine 

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Physiology, 148(4): 1938-1952. 



Efficacy of Different Herbicides for Weed Control in Transplanted Rice (Oryza sativa. 

L) Under Vertisols of Rajasthan 

CHANDRA PRAKASH, R.K. SHIVRAN, N.R. KOLI AND J.C. SHARMA 

Agricultural Research Station, Ummedganj, Kota, Maharana Pratap University of Agricultural &Technology, 

Udaipur 

email: rshivranars2007@gmail.com 

ABSTRACT 

The study was carried out during kharif season of 2007 and 

2008 at the Agricultural Research Station, Ummedganj, Kota 

(Rajasthan). The experiment was laid out in randomized block 

design with tweleve treatments replicated four times. The major 

weeds observed in the experimental plots included grasses 

Echinochloa colonum and Echinochloa crusgalli sedges like 

Cyperus rotundus, Cyperus difformis and Cyperus iria and broad 

leaf weeds Eclipta alba, Euphorbia hirta L, Commelina diffusa 

and Ammania baccifera. Among the herbicides tested, 

Pretilachlor 50 EC @ 0.750 kg a.i.ha -1 at 3-5 days after 

transplanting registered significantly lowest weed density (16 

& 15), weed dry weight (29.34 & 22.10 g m -2 ) at 60 DAT over 

other herbicidal treatments, leading to highest weed control 

efficiency of 79.73 and 90.23% and lowest weed index value of 

(3.95 and 14.52) and highest herbicidal efficiency index (1.82 

and 2.21), higher number of panicles m -2 (381 and 278), panicle 

weight (3.70 and 3.80 g), grain (4.96 and 4.23 t ha -1 ) and straw 

yield (6.61 and 5.81 t ha -1 ) in 2007 and 2008, respectively. The 

per cent increase in grain yield was 34.78 and 42.42 with the 

application of pretilachlor 50 EC @ 0.750 kg a.i. ha -1 at 3-5 days 

after transplanting over non weeded control in 2007 and 2008, 

respectively 

Key words 

Efficacy, Weed flora, Pretilachlor, Transplanted rice. 

Weed control 

Rice (Oryza sativa.L) is one of the most important food 

crops in the world. In India, it is cultivated on an area of 44 

million ha, which accounts for about 45% of food grain 

production and 55% of cereal production in the country. The 

weed flora under transplanted condition is very much diverse 

and consists of grasses, sedges and broad leaved weeds The 

effective control of weeds at initial stages (0-40 DAT) can 

help in improving the productivity of rice. However, the crop 

is subjected to greater weed competition for various growth 

resources, viz., nutrients, light and space because both crop 

and weed seeds emerge at the same time and its yield is reduced 

up to 50–100% (Singh, et al., 2004). Traditionally weed control 

is done by hand weeding, which is drudgery and also during 

the peak periods, weeding become rather difficult due to costly 

and scarcity of labour. In view of the above facts, the present 

study was undertaken to evaluate bio efficacy of different 

herbicides on weed dynamics and yield of transplanted rice in 

south east plain zone of Rajasthan. 


The study was carried out during kharif season of 2007 

and 2008 at the Agricultural Research Station, Ummedganj, 

Kota (Rajasthan).which is situated in between 25 0 11 ‘N latitude 

and 75 0 54’ E longitude with an altitude of 273 m mean sea 

level The soil was clayey in texture, slightly alkaline in reaction 

(pH 7.5), low in organic carbon (0.49 %) and medium in available 

nitrogen (278 kg ha -1 ), medium in available phosphorus (14.3 

kg ha -1 ) and high in available potassium (305 kg ha -1 ). The 

experiment was laid out in randomized block design with 

tweleve treatments comprises of Pretilachlor 50 EC @ 0.500 kg 

a.i.ha -1 at 3-5 DAT, Pretilachlor 50 EC @ 0.750 kg a.i.ha -1 at 3- 

5 DAT, Triasulfuran 20 WSG @ 0.006 kg a.i ha -1 at 5-7 DAT, 

Triasulfuran 20 WSG @ 0.009 kg a.i. ha -1 at 5-10 DAT, 

Triasulfuran 20 WSG @ 0.006 kg a.i. ha -1 at 12-15 DAT, 

Triasulfuran 20 WSG @ 0.009 kg a.i ha -1 at 12-15 DAT, 

Triasulfuran 20 WSG + Pretilachlor 50 EC @ 0.006 + 0.005 kg 

a.i ha -1 at 5-7 DAT, Triasulfuran 20 WSG + Pretilachlor 50 EC 

@ 0.009 + 0.005 at 5-7 kg a.i ha -1 DAT, Bensulfuron-methyl 60 

DF @ 0.05 kg a.i ha -1 at 20-25 DAT, Bensulfuron-methyl 60 DF 

@ 0.05 kg a.i.ha -1 at 20-25 DAT, Two hand weeding (20 and 40 

DAT) and Non weeded control replicated four times. Fertilizers 

were applied to the plots as N-P-K @ 120-60-40 kg ha -1 through 

Urea, SSP, Muriate of potash respectively. The whole amount 

of P and K was applied as basal dose during final land 

preparation. N was top-dressed in three equal splits at 20, 40 

and 55 DAT. The variety ‘Ratna’ was used as the test crop. 

Thirty-day old rice seedlings were transplanted 20 cm x 10 cm 

apart on 23 rd July, 2007 and 25 th July, 2008 at the seed rate of 

25 kg in nursery for one hectare. Two sprays of 

Monocrotophos @ 1 litre ha -1 were applied as prophylactic 

measure against insects-pests. The crops were kept under 

constant observation from transplanting till harvesting. The 

data on weed infestation and weed density were collected 

from each plot at 60 DAT. A quadrate of 0.25 m 2 was placed 

randomly at three different spots outside an area of 12 m 2 in 

the middle of the plot. The average number of three samples 

was then multiplied by 4 to obtain the weed density per m 2 . 

The collected weeds were first dried in the sun and then in an 

electric oven for 72 hours maintaining a constant temperature 

of 80°C. After drying, weight of each species was taken and 

expressed in g m -2 . Weed control efficiency and weed index 

was calculated with the following formula:

CHANDA PAKASH et al., Efficacy of Different Herbicides for Weed Control in Transplanted Rice (Oryza sativa. L) 113 

Weed control efficiency (WCE) = DMC –DMT x 100 

DMC 

Where: DMC = Weed dry matter production in unweeded 

treatment 

DMT = Weed dry matter production in weed control treatment 

Weed index = Yield in weed free plot- Grain yield in treated plot X 100 

Yield in weed free plot 

Herbicide Efficiency Index = YT-YC X 100/YC divided 

by DMT x 100/DMC, 

Where, YT and YC stand for the yields of treated and 

weedy control, respectively, while DMT and DMC refer to 

respective weed dry matter. The Weed Persistency index and 

HEI denote the tolerance of weeds to a particular treatment 

and efficiency of various herbicide doses in eradicating weeds, 

respectively. A lower value of WPI and higher value of HEI 

depict satisfactory levels of weed control. Results of both the 

years were analysed statistically and data which did not show 

the homogeneity hence were given individual year- wise Least 

Significant Differences (LSD), was used for means verification 

and for discussion of the results under probability level of 

0.05. 


Weed flora 

The major weeds observed in the experimental plots 

included grasses Echinochloa colonum and Echinochloa 

crusgalli sedges like Cyperus rotundus, Cyperus difformis 

and Cyperus iria and broad leaf weeds Eclipta alba, 

Euphorbia hirta L, Commelina diffusa and Ammania 

baccifera. Emergence of these weeds was observed during 

20–30 days after transplanting and there after it continuously 

emerged throughout the growth stages. 

Effect on weed density, weed dry weight and weed control 

efficiency 

Experimental results (Table 1) revealed that among the 

herbicides tested, Pretilachlor 50 EC @ 0.750 kg a.i.ha -1 at 3 

days after transplanting registered significantly lowest weed 

density (16 and 15), weed dry weight (29.34 and 22.10 g m -2 ) at 

60 DAT over other herbicidal treatments, leading to highest 

weed control efficiency of 79.73 and 90.23% and lowest weed 

index value of (3.95 and 14.52) and highest herbicidal efficiency 

index (1.82 and 2.21) in 2007 and 2008, respectively. These 

results reflected its selectivity and high bio-efficacy in 

controlling and suppressing weed growth effectively at the 

initial growth stage and no phytotoxicity on rice plant. The 

similar results have also been reported by Ravisankar, et al., 

2008, Bhambri and Kolhe, 2006 and Duary, et al., 2005. However, 

two hand weeding at 20 and 40 DAT registered lowest weed 

density (9 and 8), weed dry weight (5.43 and 3.64 g m -2 ) at 60 

DAT, respectively in 2007 and 2008. 

Effect on Yield attributes 

Results revealed (Table 2) that the highest yield 

attributes viz., number of panicle (398 and 302) and panicle 

weight (3.88 and 3.90 g) was recorded in two hand weeded 

plot respectively in 2007 and 2008. This might be due to 

minimum crop-weed competition for growth factors. Among 

the herbicide tested significantly higher number of panicles 

m -2 (381 and 278) and panicle weight (3.70 and 3.80 g) were 

recorded with pretilachlor 50 EC @ 0.750 kg a.i./ha at 3-5 days 

after transplanting over other treatments viz., Pretilachlor 50 

Table 1. 

Effect of weed control practices on weed density, weed control efficiency, weed index and herbicidal efficiency index in 

rice. 

Treatments 

Weed density Dry weight of weeds 

(g m -2 ) 

Weed control 

efficiency 

Weed 

index 

Herbicide 

efficiency index 

2007 2008 2007 2008 2007 2008 2007 2008 2007 2008 

Pretilachlor @ 0.500 kg ha -1 at 3-5 DAT 32 30 83.21 66.00 61.39 80.13 13.74 27.57 0.31 0.36 

Pretilachlor 0.750 kg ha -1 at 3-5 DAT 16 15 29.34 22.10 79.73 90.23 3.95 14.53 1.82 2.21 

Triasulfuran @ 0.006 kg ha -1 at 5-7 DAT 38 42 96.76 64.24 54.15 72.01 17.83 26.65 0.24 0.40 


Triasulfuran @ 0.006 kg ha -1 at 12-15 55 65 97.21 85.00 33.64 57.13 21.23 29.35 0.15 0.24 

DAT 


Triasulfuran + Pretilachlor @ 0.006 +0.500 42 46 68.46 52.16 48.72 69.37 18.23 22.63 0.33 0.64 

kg ha -1 at 5-7 DAT 

Triasulfuran + Pretilachlor @ 0.009 +0.500 58 68 35.78 30.55 29.78 54.97 24.33 15.46 0.27 1.54 

at 5-7 kg ha -1 DAT 

Bensulfuron-methyl @ 0.05 kg ha -1 at 20-25 26 24 98.23 92.73 68.27 84.00 12.20 30.62 0.32 0.19 

DAT 

Bensulfuron-methyl @ 0.05 kg ha -1 at 20-25 52 61 93.47 78.24 36.53 59.60 19.69 27.88 0.21 0.30 

DAT 

Two hand weeding (20 and 40 DAT) 9 8 5.43 3.64 - - - - - - 

Non weeded control 83 89 115.0 131.2 - - - - 

SEm ± 1.50 1.41 2.42 2.27 

CD (P=0.05) 4.89 4.40 7.23 6.88


Table 2. 

Effect of weed control practices on yield attributing characters and yield of rice 

Treatments 

No. of panicles 

m -2 

Panicle weight 

(g) 

Grain yield 

( t ha -1 ) 

Straw yield 

(t ha -1 ) 

Harvest 

Index 

2007 2008 2007 2008 2007 2008 2007 2008 2007 2008 

Pretilachlor @ 0.500 kg ha -1 at 3-5 DAT 358 252 3.40 3.52 4.46 3.59 6.32 5.08 41.35 41.38 

Pretilachlor 0.750 kg ha -1 at 3-5 DAT 381 278 3.70 3.80 4.96 4.23 6.61 5.81 42.89 42.92 





Triasulfuran + Pretilachlor @ 0.006 +0.500 kg ha -1 at 5-7 DAT 336 266 3.34 3.63 4.22 3.83 5.57 5.05 43.13 43.16 

Triasulfuran + Pretilachlor @ 0.009 +0.500 at 5-7 kg ha -1 DAT 327 275 3.20 3.76 3.91 4.19 5.43 5.63 41.85 41.88 

Bensulfuron-methyl @ 0.05 kg ha -1 at 20-25 DAT 369 242 3.45 3.18 4.54 3.44 6.47 4.90 41.20 41.23 

Bensulfuron-methyl @ 0.05 kg ha -1 at 20-25 DAT 325 248 3.28 3.27 4.15 3.57 5.66 4.87 42.30 42.33 

Two hand weeding (20 and 40 DAT) 398 302 3.88 3.90 5.35 4.72 7.51 6.81 41.60 41.63 

Non weeded control 317 235 3.15 3.12 3.68 2.97 5.01 4.05 42.30 42.33 

SEm ± 7.7 6.0 0.04 6.07 0.10 0.10 0.09 0.12 0.64 0.66 

CD (P=0.05) 22.1 17.8 0.12 0.22 0.29 0.30 0.28 0.35 NS NS 

EC @ 0.500 kg a.i.ha -1 at 3-5 DAT, Triasulfuran 20 WSG @ 

0.006 kg a.i ha -1 at 5-7 DAT, Triasulfuran 20 WSG @ 0.009 kg 

a.i. ha -1 at 5-10 DAT, Triasulfuran 20 WSG @ 0.006 kg a.i. ha - 

1 

at 12-15 DAT, Triasulfuran 20 WSG @ 0.009 kg a.i ha -1 at 12- 

15 DAT, Triasulfuran 20 WSG + Pretilachlor 50 EC @ 0.006 + 

0.005 kg a.i ha -1 at 5-7 DAT, Triasulfuran 20 WSG + Pretilachlor 

50 EC @ 0.009 + 0.005 at 5-7 kg a.i ha -1 DAT, Bensulfuronmethyl 

60 DF @ 0.05 kg a.i ha -1 at 20-25 DAT, Bensulfuronmethyl 

60 DF @ 0.05 kg a.i.ha -1 at 20-25 DAT, 

Effect on Yield 

Results obtained from the experiment revealed that 

among the chemical weed management practices, Application 

of pretilachlor 50 EC @ 0.750 kg a.i. ha -1 at 3-5days after 

transplanting produced significantly higher grain (4.96 and 

4.23 t ha -1 ) and straw yield (6.61 and 5.81 t ha -1 ) respectively, in 

both the years of experimentation over all other herbicidal 

treatments. The per cent increase in grain yield was 34.78 & 

42.42 due to the weed control with pretilachlor 50 EC @ 0.750 

kg a.i. ha -1 at 3-5 days after transplanting over non weeded 

control in 2007 and 2008, respectively. This was owing to the 

fact that pretilachlor 50 EC @ 0.750 kg a.i. ha -1 at 3-5 days after 

transplanting produced maximum crop growth, and thereby 

the increased accumulation of photosynthates in reproductive 

parts, which ultimately brought about marked improvement in 

yield. Gnanasambandan and Murthy, 2000 and Singh and 

Singh, 2010 have also reported that treatments which had 

better growth and yield attributes had resulted higher grains 

yields. However, the maximum grain yield (5.35 & 4.72 t ha -1 ) 

was obtained with two hand weeding at 20 & 40 DAT 

respectively in 2007 & 2008. 

Thus the present findings indicated that pretilachlor 50 

EC @ 0.750 kg a.i ha -1 at 3-5 days after transplanting can be 

recommended for effective control over weeds as well as 

getting higher yield during kharif season in transplanted rice 

ecosystem under south east plain zone of Rajasthan. 


Bhambri, M.C. and Kolhe, S.S. 2006. Possibilities of green manuring in 

direct seeded rice (Oryza sativa) and its impact on weed dynamics. 

In: Extended summaries. National Symposium on Conservation 

Agriculture and Environment, Indian Society of Agronomy, held 

during 26–28 October, Banaras Hindu University, Varanasi, pp. 

314. 

Duary, B., Hossain, A. and Mondal, D.C. 2005. Integrated weed 

management in direct seeded dry sown rice in lateritic belt of West 

Bengal. Indian Journal of Weed Science 37(1/2): 101–2. 

Gnanasambandan, S. and Murthy, P.B. 2000. Effect of tillage practices 

and pre-emergence herbicides application for weed control in wetseeded 

rice. Advances in Agricultural Research in India, 14: 115- 

20. 

Ravisankar, N., Chandrasekaran, B., Raja, R., Din, M. and Chaudhuri, 

S.G. 2008. Influence of integrated weed management practices on 

productivity and profitability of wet seeded rice (Oryza sativa). 

Indian Journal of Agronomy, 53(1): 57-61. 

Singh, M. and Singh, R.P. 2010. Efficacy of herbicides under different 

methods of direct-seeded rice (Oryza sativa) establishments. Indian 

Journal of Agricultural Sciences, 80(9): 815–819. 

Singh, V.P., Singh, Govindra and Singh, Mahendra. 2004. Effect of 

fenoxaprop-P-ethyl on transplanted rice and associated weeds. 

Indian Journal of Weed Sciences, 36: 190-92. 



Bio-efficacy of Herbicide Combination on Weed Control and Yield Performance of 

Transplanted Rice (Oryza sativa. L) 

CHANDRA PRAKASH, R.K. SHIVRAN, N.R. KOLI AND J.C. SHARMA 

Agricultural Research Station, Ummedganj, Kota, Maharana Pratap University of Agricultural and Technology, 

Udaipur 

email: rshivranars2007@gmail.com 

ABSTRACT 

A field experiment was conducted during kharif season of 2008 

and 2009 at Agricultural Research Station, Ummedganj, Kota 

(Rajasthan) to study the efficacy of herbicide combination on 

weed control and yield performance of transplanted rice. 

Application of Glyphosate @ 0.75 kg a.i. ha -1 at 15 days before 

crop establishment + Bensulfuron - methyl + pretilachlor 

(6.6GR) @ 0.06 + 0.60 kg a.i.ha -1 at 8-15 DAT was found most 

effective in lowering the weed density of grassy and non-grassy 

weeds and their dry weight, and thus enhancing yield attributes 

and yield (6.06 and 5.76 t ha -1 ) of rice, respectively in both the 

years. Among the weed control treatments maximum weed 

control efficiency (83.83 and 82.37%) and higher herbicide 

efficiency index (2.82 and 2.39) was under this treatment 

compared with the other sequential application of herbicides. 

On other hand weed control efficiency of different herbicide 

combinations were like wise, Glyphosate@0.75kg a.i. ha -1 at 15 

days before crop establishment + Butachlor@1.5 kg a.i. ha -1 at 

0-5 DAT (69.07 and 80.60 %) and Butachlor@1.5 kg a.i. ha -1 at 

0-5 DAT (70.14 and 72.02%), followed by alone applications of 

Glyphosate@0.75kg a.i. ha -1 at 15 days before crop establishment 

(64.21 and 69.17) 

Key words 

Bio-fficacy, Weed flora, Herbicide combination,, 

Transplanted rice, Weed control 

In India rice is gown in an area of around 44 million ha 

annually with a production of 104 million tones contributing 

45 % of the total food grain production of the country. Weed 

competition is one of the prime yield limiting biotic constraints 

in rice. Transplanted rice, in particular, is infested by 

heterogeneous types of weed flora consisting of grassy, 

broadleaf weeds and sedges causing yield reduction up to 

76%. Effective control of weeds had increased the grain yield 

by 85.5% (Mukherjee and Singh 2004). About 60% of weeds 

emerge during 7-30 days after transplanting and strongly 

compete with rice plants up to maximum tillering stage. Weeds 

are most competitive when the crop is small. So, controlling 

weeds when preparing the land for a rice crop is essential. 

Weed seed should be allowed to germinate and then killed 

before flooding or drilling the crop. Ploughing is traditionally 

used to bury weeds, but this is laborious and costly. Nonselective 

herbicides which control a broad spectrum of weeds 

can be used. Herbicides like glyphosate have no residual 

activity in the soil and do not affect the rice crop. Glyphosate 

gives good control of perennial weeds. They found that notill 

could save the time and costs associated with ploughing 

and that the soil structure improved so water management 

was better. Therefore, timely weed control at early stage is 

imperative for realizing desired level of productivity from 

transplanted rice. The use of herbicides offers selective and 

economic control of weeds right from the beginning, giving 

crop an advantage of good start and competitive superiority. 

Moreover, continuous use of same herbicides leads to a shift 

of weed flora and annual sedges (Rajkhowa, et al., 2006). 

Therefore, the present study was undertaken to evaluate bioefficacy 

of herbicide combination on weed control and yield 

performance of transplanted rice. 


A field experiment was conducted during kharif season 

of 2008 and 2009 at the Agricultural Research Station, 

Ummedganj, Kota (Rajasthan). The soil was clayey in texture, 

slightly alkaline in reaction (pH 7.5), low in organic carbon 

(0.49%) and medium in available nitrogen (278 kg ha -1 ), medium 

in available phosphorus (14.3 kg ha -1 ) and high in available 

potassium (305 kg ha -1 ). The experiment was laid out in 

randomized block design with seven treatments comprises of 

Glyphosate @ 0.75kg a.i. ha -1 at 15 days before crop 

establishment (T1), Butachlor@1.5 kg a.i. ha -1 at 0-5 DAT 

(T2), Bensulfuron - methyl + pretilachlor (6.6GR) @ 0.06 + 0.60 

kg a.i.ha -1 at 8-15 DAT (T3), Glyphosate @ 0.75kg a.i. ha -1 at 

15 days before crop establishment + Butachlor@1.5 kg a.i. 

ha -1 at 0-5 DAT (T4 ), Glyphosate @ 0.75kg a.i. ha -1 at 15 days 

before crop establishment + Bensulfuron - methyl + pretilachlor 

(6.6GR) @ 0.06 + 0.60 kg a.i.ha -1 at 8-15 DAT (T5), Hand 

weeding twice (20 and 40 DAT) (T6) and Non weeded control 

(T7) replicated four times. Fertilizers were applied to the plots 

as N-P-K @ 120-60-40 kg/ha from Urea, SSP, MOP respectively. 

The whole amount of P and K was applied as basal dose 

during final land preparation. N was top-dressed in three equal 

installments at 20, 40 and 55 DAT, respectively The variety 

‘Jaya’ was used as the test crop Thirty-day old rice seedlings 

were transplanted 20 cm x 10 cm apart on 25 th July, 2008 and 

28 th July, 2009 at the seed rate of 25 kg in nursery for one 

hectare. Two sprays of Monocrotophos @ 1 litre ha -1 were 

applied as prophylactic measure against insects-pests. The 

crops were kept under constant observation from transplanting 

till harvesting. The data on weed infestation and weed density 

were collected from plot at 60 DAT. A quadrate of 0.25 m 2 was 

placed randomly at three different spots outside an area of 12 

m 2 in the middle of the plot. The infesting species of weeds


within each quadrate were identified and their number was 

counted species-wise. The average number of three samples 

was then multiplied by 4 to obtain the weed density per m 2 . 

The collected weeds were first dried in the sun and then in an 

electric oven for 72 hours maintaining a constant temperature 

of 80°C. After drying, weight of each species was taken and 

expressed in g/m. Weed control efficiency and Herbicidal 

efficiency index were calculated with the following formula: 

Weed control efficiency (WCE) = DMC –DMT X 

100 

DMC 

Where, DMC = Weed dry matter production in Unweeded 

treatment 

DMT = Weed dry matter production in weed control treatment 

Weed index = Yield in weed free plot- Grain yield in treated plot X 100 

Yield in weed free plot 

Herbicide Efficiency Index (HEI) = YT-YC X 100/YC divided 

by DMT x 100/DMC, 

Where, YT and YC stand for the yields of treated and 

weedy control, respectively, while DMT and DMC refer to 

respective weed dry matter. The Weed Persistency index and 

HEI denote the tolerance of weeds to a particular treatment 

and efficiency of various herbicide doses in eradicating weeds, 

respectively. A lower value of WPI and higher value of HEI 

depict satisfactory levels of weed control. Results of both the 

years were analyzed statistically and data which did not show 

the homogeneity hence were given individual year- wise Least 

Significant Differences (LSD), was used for means verification 

and for discussion of the results under probability level of 

0.05. 


Weed Flora 

The major weeds observed in the experimental plots 

included grasses Echinochloa colonum and Echinochloa 

crusgalli sedges like Cyperus rotundus, Cyperus difformis 

Table 1. 

and Cyperus iria and broad leaf weeds Eclipta alba and 

Ammania baccifera. 

Effect on Weed control 

Effect of different herbicide combination on weed 

density, dry weight of weeds, and weed control efficiency 

given in Table 1. The results revealed that all the herbicide 

combination gave significant control of weed population over 

non weeded control. However, the highest weed control 

efficiency was given by twice hand weeding (91.80 and 

89.57%), respectively in both the years. These findings are in 

agreement with Rekha, et. al., 2002 who reported that twice 

hand weeding resulted in lower weed density compared to 

weedicides and untreated control. Among the weedicides 

combinations tested, application of Glyphosate @ 0.75kg a.i. 

ha -1 at 15 days before crop establishment + Bensulfuron - 

methyl + pretilachlor (6.6GR) @ 0.06 + 0.60 kg a.i.ha -1 at 8-15 

DAT was most effective in lowering the weed density (9 and 

19) and their dry weight (12.35 and 15.63 g), weed competition 

index (5.78 and 4.33), higher weed control efficiency (83.83 

and 82.37%) and herbicide efficiency index (2.82 and 2.39) 

thus enhancing yield attributes and yield (6.06 and 5.76 t ha -1 ) 

of rice, respectively in both the years.. The results further 

revealed that weed density reduced significantly in all the 

treated plots over untreated (check) plots. These findings are 

in accordance with Singh, et. al., 2007 who reported 20 to 63% 

yield losses in uncontrolled weed fields. It means that in 

uncontrolled weed fields, their densities continuously 

remained increasing that may adversely affect the crop growth. 

These results are in close conformity with the findings of 

Mukherjee and Singh (2004). 

Effect on yield attributes and yield 

All the weed control treatment combinations 

significantly reduced the weeds as compare to weedy check 

and recorded higher yield attributes and grain yield of rice 

(Table 2). Significantly higher number of panicles m -2 (347 and 

363) and panicle weight (3.68 and 3.70 g), grain yield (6.06 and 

Effect of herbicide combination on weed density, dry weight of weeds, weed competition index, weed control efficiency 

and herbicidal efficiency index in rice. 

Treatments 

Weed 

density 

Dry weight of 

weeds (g) 

Weed 

competition index 

(%) 

Weed control 

efficiency (%) 

Herbicide 

efficiency index 

2008 2009 2008 2009 2008 2009 2008 2009 2008 2009 

Glyphosate@0.75kg a.i /ha -1 at 15 days before crop establishment 24 23 27.33 27.33 13.3 14.03 64.21 69.17 0.95 0.90 

Butachlor@1.5 kg a.i. ha -1 at 0-5 DAT 19 20 22.8 24.81 10.99 13.09 70.14 72.02 1.26 1.04 

Bensulfuron - methyl + pretilachlor (6.6GR) @ 0.06 +0.60 kg 

a.i.ha -1 at 8-15 DAT 

46 45 50.2 60.35 24.29 18.28 34.26 31.93 0.26 0.32 

Glyphosate@0.75kg a.i /ha -1 at 15 days before crop establishment + 

Butachlor@1.5 kg a.i ha -1 at 0-5 DAT 

21 13 23.6 17.2 12.15 5.79 69.09 80.60 1.16 2.06 

Glyphosate@0.75kg a.i. ha -1 at 15 days before crop establishment + 

Bensulfuron - methyl + pretilachlor (6.6GR) @ 0.06 +0.60 kg a.i 9 10 12.35 15.63 5.78 4.33 83.83 82.37 2.82 2.39 

ha -1 at 8-15 DAT 

Hand weeding twice (20 and 40 DAT) 5 6 6.26 9.25 - - 91.80 89.57 6.65 4.66 

Non weeded control 68 72 76.36 88.66 35.27 32.73 - - - - 

CD (P=0.05) 6.2 4 4.54 3.05

Table 2. 

CHANDA PAKASH et al., Bio-efficacy of Herbicide Combination on Weed Control and Yield Performance 117 

Effect of herbicide combination on yield attributes and yield of transplanted rice. 

Treatments 

No of panicles 

m -2 

Panicle weight 

(g) 

Grain yield 

( t/ha) 

2008 2009 2008 2009 2008 2009 

Glyphosate@0.75kg a.i. /ha at 15 days before crop establishment 322 334 3.38 3.38 5.58 5.18 

Butachlor@1.5 kg a.i. /ha at 0-5 DAT 330 342 3.47 3.42 5.72 5.23 

Bensulfuron - methyl + pretilachlor (6.6GR) @ 0.06 +0.60 kg a.i./ha at 8-15 303 310 3.20 3.16 4.83 4.92 

DAT 

Glyphosate@0.75kg a.i. /ha at 15 days before crop establishment + 

325 356 3.44 3.63 5.65 5.47 

Butachlor@1.5 kg a.i. /ha at 0-5 DAT 

Glyphosate@0.75kg a.i. /ha at 15 days before crop establishment + 

347 363 3.68 3.70 6.06 5.76 

Bensulfuron - methyl + pretilachlor (6.6GR) @ 0.06 +0.60 kg a.i./ha at 8-15 

DAT 

Hand weeding twice (20 and 40 DAT) 368 388 3.92 3.92 6.43 6.02 

Non weeded control 268 278 2.91 2.85 4.16 4.05 

CD (P=0.05) 15 16 0.18 0.15 0.29 0.22 

5.76 t ha -1 ) of rice, respectively in both the years. were observed 

in plots treated with Glyphosate @ 0.75kg a.i. ha -1 at 15 days 

before crop establishment + Bensulfuron - methyl + pretilachlor 

(6.6GR) @ 0.06 + 0.60 kg a.i.ha -1 at 8-15 DAT. The per cent 

increase in grain yield of rice was 45.67 and 42.22 over nonweeded 

check respectively in 2008 and 2009.. All the weedicides 

combination showed significantly higher number of panicles 

and panicle weight over the non-weeded check. This was due 

to the fact the less competition for moisture, light and nutrient 

uptake by the crop plants. The higher assimilation of 

photosynthates in weedicide treated plots may be the reason 

for higher yield attributes and ultimately higher yield. The 

results were in close conformity with those of Kumar and 

Sharma, 2005, Singh, et al., 2005 and Amarjit, et al., 2006. 

Thus, the combination of Glyphosate @ 0.75kg a.i. ha -1 

at 15 days before crop establishment + Bensulfuron - methyl 

+ pretilachlor (6.6GR) @ 0.06 + 0.60 kg a.i.ha -1 at 8-15 DAT 

may be recommended in transplanted rice for controlling 

predominant weeds of south east plain zone of Rajasthan. 


Amarjit, S., Bali, Singh, Mahinder Kachroo Dileep, Sharma, B.C. and 

Shivam, D.R. 2006. Efficacy of herbicides inntransplanted, medium- 

duration rice under sub-tropical conditions of Jammu. Indian Journal 

of Agronomy, 51(3): 128-30. 

Kumar, M. and Sharma, G. 2005. Effect of herbicides alone and in 

combination on direct seeded rice. Indian Journal of Weed Science, 

37(3/4): 197-201. 

Mukherjee, P.K. and Singh, R.P. 2004. Efficacy of low doses of herbicides 

on divergent weed flora in transplanted rice. Oryza, 41(1): 20-23. 

Rajkhowa, D.J., Borah, N., Barua, I.C. and Deka, N.C. 2006. Effect of 

pyrazosulfuron ethyl on weeds and productivity of transplanted 

rice during rainy season. Indian Journal of Weed Science, 38(1&2): 

25-8. 

Rekha, K.B., Raju, M.S. and Reddy, M.D. 2002. Effect of herbicides in 

transplanted rice. Indian Journal of Weed Science, 34(1-2): 123- 

125. 

Singh, Purshotam, Singh, Parmeet, Singh, Rekhi and Singh, K.N. 2007. 

Efficacy of new herbicides in transplanted rice under temperate 

conditions of Kashmir. Indian Journal of Weed Science, 39(3&4): 

167-71. 

Singh, V.P., Govindra, S., Singh, R.K., Singh, S.P., Abnishkumar, 

V.C.D., Kumar, M. and Sharma, G. 2005. Effect of herbicides 

alone and in combination on direct seeded rice. Indian Journal of 

Weed Science, 37(3/4): 197-201. 



SHORT COMMUNICATION 

Genetic Variability, Heritability and Correlation Studies in Wheat Genotypes 

(Triticum aestivum L.) 

EKTA SINGH 1 , RAMTEKE P.W. 2 , M. RAM 2 , B.A. WANI 2 AND SADHNA SINGH 3 

Department of Biological Sciences 1 , Genetics and Plant Breeding 2 , Plant Protection 3 

Sam Higginbottom Institute of Agriculture, Technology and Sciences, (Formerly Allahabad Agriculture Institute) 

Allahabad U.P. 211 007 

email: ekta1701@gmail.com 1 

Wheat (Triticum aestivum L.) is one of the most important 

crop of the world and contributes about 40% of India’s food 

grain pool. The present investigation was conducted for 

selected 24 genotypes to determine the extent of genetic 

variability, genetic coefficient, heritability, genetic advance 

and correlation of different characters in wheat. 

The field experiment was carried at the experimental field 

of Genetics and Plant Breeding/ Seed Science and Technology, 

Department of GPB, SHIATS, Allahabad. The experimental 

methods consist of 24 diverse wheat lines. The trail was laid 

down in RBD with three replications during two consecutive 

years (2009-2010 and 2010-2011) with average plot size of 3 

sq.cm. sowing was done during both the years at the row to 

row spacing of 20 cm apart and 5 cm plant to plant distances. 

The observation were recorded on 5 randomly selected plants 

from each line in each replicate for 17 characters Table 1. The 

phenotypic and genotypic coefficient of variability was 

calculated according to the method suggested by Burton, 

1953. For estimation of heritability (Broad sense), genetic 

advance and correlation were calculated according to the 

suggested by Johnson, et al., 1955. 

Analysis of variance of two year data revealed significant 

differences among the genotypes for all the traits indicating 

the presence of sufficient genetic variability in the genotypes 

and considerable scope for their improvement. Sufficient 

genetic variability for many of the quantitative traits studied 

in wheat. The extent of variability with respect to 17 characters 

in different genotypes is measured in terms of range, genotypic 

coefficient of variation (GCV), phenotypic coefficient of 

variation (PCV), along with the heritability expected genetic 

advance and genetic advance as percentage of mean (GAM) 

are given in (Table 1). The considerable amount of variation 

was observed for all the characters. The phenotypic 

coefficient of variability (PCV) was at par with the genotypic 

coefficient of variability in all the characters (Table 1). The 

estimates of GCV and PCV were high for leaf nitrogen (35.34 

and 35.44), gluten content (32.47 and 32.70), chlorophyll a 

(18.62 and 19.13), total chlorophyll (16.19 and 17.10), tillers per 

plant (13.58 and 15.66) and low for harvest index (3.08 and 

4.16). The yield per plot (11.51 and 11.70) and spike length 

(11.20 and 11.52) recorded higher GCV and PCV values for 

yield attributing traits in wheat which was also reported by 

Walia and Garg, 1996 and Gupta and Verma, 2000. The high 

heritability was noticed for protein content (%) (99.63%) and 

low for chlorophyll b (66.70%). This indicates that chlorophyll 

b is highly influenced by environmental factors and not much 

exploited as selection criteria while all those which 

demonstrated high heritability values could be exploited as 

selection criteria by integrating with conventional breeding 

methods. Our findings are supported by observation of Singh 

and Singh, 2001 and Dwivedi, et al., 2002. Similarly, among 

yield and yield traits , yield is a well known trait not directly 

under genetic control (Grafius, 1965). But among the yield 

component characters, the heritability for tillers/plant 

(75.15%), spike length (94.46), grains/spike (93.84%) and 1000 

grain weight (94.75%) were found of very high order. Therefore 

these characters are well under genetic control and selection 

can result high yielding lines. Shoran, 1995 also reported high 

heritability for 1000 grain weight, harvest index, tillers/plant 

and plant height. 

In the present experiment, the study of correlation among 

different characters revealed that, in general the genotypic 

correlation coefficients were positive and of high order which 

indicate high degree of association of physiologic traits with 

the yield and yield components. This indicates that the 

physiologic traits such as chlorophyll-a had significant 

positive correlation with grain yield/plot and yield components 

like 1000 grain weight, spike length and tillers/plant and plant 

height; chlorophyll-b with grain yield/plot and yield 

components like grains/spike; total chlorophyll content with 

yield/plot; 1000 grain weight and plant height; leaf nitrogen 

with grain yield/plot, tillers/plot and 1000 grain weight; Harvest 

Index with 1000 grain/weight and grain filling period with plant 

height; Days to 50% flowering was found negatively correlated 

with plant height and days to maturity positively correlated 

with plant height. 

As regards genotypic correlation coefficient of 

physiologic traits with quality characteristics, chlorophyll a 

showed significant positive correlation with protein; 

chlorophyll b, showed negative correlation with gluten; total 

chlorophyll content showed positive correlation with protein 

content; leaf nitrogen showed positive correlation with protein

Table 1. 

Traits 

SINGH et al., Genetic Variability, Heritability and Correlation Studies in Wheat Genotypes (Triticum aestivum L.) 119 

Range, mean, GCV, PCV, heritability and genetic advance of mean for 17 traits in wheat 

Range 

(pooled) 

Grand 

mean 

(Pooled) 

GCV= Genotypic coefficient of variance, PCV= Phenotypic coefficient of variance. 

GCV PCV Heritability 

(%) (Broad 

scence) 

Genetic 

advance 

Genetic advance as 

(%)of mean (Genetic 

gain) 

Chlorophyll a (mg/g) 0.62-1.49 1.29 18.62 19.13 94.70 0.46 37.32 

Chlorophyll b (mg/g) 0.25-0.45 0.36 15.61 19.11 66.70 0.09 26.26 

Total chlorophyll (mg/g) 1.02-1.92 1.60 16.19 17.10 89.66 0.50 31.59 

Leaf nitrogen (mg/g) 1.28-4.40 2.96 35.34 35.44 99.48 2.15 72.63 

Nitrate Reductase Activity 0.135-0.175 0.156 7.42 7.89 88.36 0.02 14.37 

Harvest Index(%) 40.08-46.03 43.27 3.80 4.16 83.46 3.09 7.15 

Grain Filling Period 33.83-44.00 39.56 5.91 6.16 91.78 4.61 11.66 

Days to 50% flowering 65.00-80.00 73.49 5.14 5.23 96.55 7.65 10.41 

Days to maturity 109.00-119.00 113.07 2.40 2.48 93.38 5.40 4.78 

Grains/spike 41.00-52.33 45.45 6.62 6.83 93.84 6.00 13.21 

1000 grain weight(gm) 32.04-47.64 39.76 10.26 10.55 94.75 8.17 20.55 

Spike length(cm) 9.58-14.34 11.73 11.20 11.52 94.46 2.63 22.43 

Gluten content (%) 4.12-11.98 7.45 32.47 32.70 98.60 4.95 66.43 

Protein content (%) 10.90-12.51 11.53 4.06 4.11 99.63 0.955 8.28 

Plant height (cm) 90.16-122.07 101.16 7.96 7.99 99.26 16.53 16.34 

Tillers/plant 5.50-9.83 8.06 13.58 15.66 75.15 1.95 24.25 

Grain yield/ plot 2.04-3.21 2.73 11.51 11.70 98.60 0.63 23.33 

content; NRA also showed negative correlation with both 

protein and gluten content; HI positive correlation with protein 

only. Grain filling period though non-significant but showed 

positive correlation with protein and gluten content. Days to 

50% flowering and maturity showed significantly negative 

correlation with gluten content and non-significant positive 

correlation with protein content. 

The physiologic traits like chlorophyll a, b, leaf nitrogen 

are synonym to enhance photosynthesis and higher NRA 

synonym to efficient in plant nutrient (Nitrogen) uptake from 

soil could be integrated with conventional breeding as 

selection criteria to enhance the yield without raising input 

level (nitrogen in particular). Not the least, integration of 

physiologic traits having higher correlation with yield and 

yield components can be tapped to achieve another 

breakthrough in the per se productivity of wheat. Our findings 

are supported by Collaku and Harrison, 2005, Mohammadi, et 

al., 2006, Limulwar, et al., 2003 who also independently 

reported significant positive correlation of chlorophyll with 

yield. Rafat and Malik, 2005 observed significant positive 

correlation of plant height with grain yield. Payal, et al., 2007 

observed significant positive correlation of Harvest Index and 

tillers/plant with grain yield. 

The present study showed the presence of considerable 

variations among wheat genotypes for all traits tested which 

gives an opportunity to plant breeders for the improvement 

of these traits. Genetic correlation coefficient analysis 

indicated that important agronomic traits are positively 

correlated with grain yield. This suggests a common genetic/ 

physiological basis among these traits. Hence, simultaneous 

improvement of these traits would be possible and can be 

considered as suitable selection criteria for the development 

of high yielding wheat varieties. 


Burton, G.W. 1952. Quantitative inheritance in grasses. Proc. 6 th Int. 

Grassland Cong., 1:227-283. 

Collaku, A. and Harrison, S.A. 2005. Heritability of water logging 

tolerance in wheat. Crop Sciences, 45:722-727. 

Dwivedi, A.N., Pawar, L.S. and Madan, S. 2002. Studies on variability 

parameters and characters association among yield and quality 

attributing traits in wheat. Haryana Agric Univ. J. Res., 32:77-80. 

Grafius, J.E. 1965. A geometry for plant breeding, Crop Science, 4:241-246. 

Gupta, S.K. and Verma, S.R. 2000. Variability ,heritability and genetic 

advance under normal and rainfed conditions in durum wheat. Indian 

J. Agric. Research, 34(2):122-125. 

Johnson, H.W., Robinson, H.R. and Comstock, R.E. 1955. Estimation 

of genetic and environmental variability in soybean. Agron. J., 

47:477-433. 

Limulwar, S.R., Hatmode, C.N., Deotale, R.D., Vandana, D.B., Mahalle, 

S.N. and Pawar, P.S. 2003. Correlation of biochemical and yield 

contributing parameters with seed yield in mustard. J. of Soils and 

crops., 13(1):158-161. 

Mohammadi, M., Ceccarelli, S. and Naghavi, M.R. 2006. Variability 

and genetic parameters for related traits to drought tolerance in 

double haploid population of barley (Hordium Vulgare) 

International J. of Agriculture and Biology, 8(5):694-697. 

Payal-Sexana, Rawat, R.S., Verma, J.S., and Meena, B.K. 2007. Variability 

and character association analysis for yield and quality traits in 

wheat. Pantnagar J. of Research, 5(2):58-60. 

Rafat-Sultan and Malik, S.K. 2005. Genetic variability and character 

association between yield and yield attributing traits in bread wheat 

(Triticum aestivum L.). Annals of Agriculture Research, 26(1): 

118-125. 

Raha, P. and Ramgiri, S.R. 1998. Genetic variability of metric traits in 

wheat and Triticales crosses over environment. Crop Research Hisar, 

16(3): 318-320. 

Shoran, J. 1995. Estimation of variability parameters and path 

coefficient for certain metric traits in winter wheat (Triticum 

aestivum L. em. Thell) Indian J. of Genet., pp.463-467. 

Singh, S.B. and Singh, T.B. 2001. Correlation and path analysis in common 

wheat under light texture soil. Research on crops, 2(1):99-101. 

Walia, D.P. and Garg, D.K. 1996. Evaluation of genetic divergence in 

wheat (Triticum aestivum L.) germplasm. Indian J. of Genet., 

56:452-457. 




Screening of Horsegram (Macrotyloma uniflorum) Genotypes against Horsegram 

Yellow Mosaic Virus (HgYMV) Disease in Karnataka 

PREMA, G.U. 1 , RUDRASWAMY, P. 2 , NAGARAJU 1 AND RANGASWAMY, K.T. 1 

1 

Department of Plant Pathology, University of Agricultural Sciences, GKVK, Bangalore 560 065 

2 

AICRP on Arid Legumes, University of Agricultural Sciences, GKVK, Bangalore 560 065 

email: prems.gu@gmail.com 1 

Horsegram [Macrotyloma uniflorum (Linn.)] is a hardy 

grain legume known for its easily digestible quality protein. 

The left over material after harvest has a great nutritional value 

and a relished feed for farm animals. The grains contain on an 

average of 22% protein and also a source of vitamins (Thakur, 

1979). 

Horsegram yellow mosaic virus disease transmitted by 

the whitefly Bemisia tabaci (Gennadius) is prevalent in most 

parts of south India (Muniyappa and Reddy, 1976., 

Muniyappa, et al., 1975 and Williams, et al., 1968). The disease 

incidence ranged from 50 to 100% in both summer and early 

rainy season crops (Muniyappa, et al., 1975) causing 

substantial loss in grain yield. The increasing spread of the 

yellow mosaic disease due to increase in Bemisia tabaci 

population which resulted in almost complete loss of the crop 

during summer (Muniyappa, et al., 1978). 

The chemical methods recommended for the 

management of HgYMV are not economical and practicable 

because of very low yield potential of the crop and the crop is 

mainly grown as alternate crop whenever early monsoon fails. 

Further, it is mainly grown in dry lands by marginal and poor 

farmers who have minimum resources to spend on chemicals. 

Therefore, the only feasible and economical method for control 

of HgYMV disease is the development of resistant varieties. 

Therefore, there is a need to screen larg number of germplasm 

lines to identify the resistant source to HgYMV. In the present 

study, 100 horsegram genotypes have been evaluated under 

field conditions with a view to identify resistant sources for 

HgYMV under field condition during 2011. Each genotype 

was sown in five meter long rows. A susceptible genotype 

was planted after every five tester lines to serve as a source 

for disease. The per cent disease severity in each genotype 

was recorded at fifteen days interval and they were grouped 

into different categories employing disease scoring scale 

suggested by Muniyappa, et al., 1987. 

Among the different genotypes screened, 38 genotypes 

viz., TCR 2S-A, TCR 1432, TCR 1177, TCR 392-C, TCR 365-A, 

TCR 425-A, TCR 886, TCR 171-A, TCR 1293, TCR 117-A, TCR 

1151, TCR 914, TCR 1135, TCR 450-A, TCR 1266, TCR 1449, 

TCR 244-A, TCR 1063, TCR 896, TCR 888, TCR 8-A, TCR 

Table 1. 

Grouping of horsegram genotypes/germplasm lines into different degree of resistance against Horsegram yellow 

mosaic virus (HgYMV) disease during 2011 under field condition 

Sl. 

Reaction 

No. 

1. Resistant 

(Small necrotic yellow specks with 

restricted spread covering 0.1-5% leaf 

area) 

2. Moderately resistant 

(Yellow mottling of leaves, covering 

10-15% leaf area) 

3. Moderately susceptible 

(Yellow mottling and discoloration of 

leaves covering 15.1-30% leaf area) 

4. Susceptible 

(Pronounced yellow mottling and 

discoloration of leaves, pods and 

reduction in leaf size and stunting of 

plants covering 50-75% of foliage) 

5. Highly susceptible 

(Sever yellowing of entire leaves, 

stunting of plants and no pod formation 

covering 85-100% of foliage) 

Genotypes 

TCR 2S-A, TCR 1432, TCR 1177, TCR 392-C, TCR 365-A, TCR 425-A, TCR 886, TCR 171-A, TCR 

1293, TCR 117-A, TCR 1151, TCR 914, TCR 1135, TCR 450-A, TCR 1266, TCR 1449, TCR 244-A, 

TCR 1063, TCR 896, TCR 888, TCR 8-A, TCR 1247, TCR 161-A, TCR 1295, TCR 872, TCR 1058, TCR 

307-A, TCR 66-A, TCR 151-A, TCR 1210, TCR 1303, TCR 275-A, TCR 873, TCR 1204, TCR 167-A, 

TCR 1050, TCR 116-A, TCR 1133 

TCR 1056, TCR 172-A, TCR 1291, TCR 1100, TCR 1433, TCR 2 

TCR 232-A, TCR 1290, TCR 95-A 

TCR 1016, TCR 74-A, TCR 275-A, TCR 883, TCR 1263, TCR 425-A, TCR 466-A, TCR 726, TCR 1076, 

TCR 288-A, TCR 1402, TCR 222-A, TCR 179-B, TCR 1450, TCR 1358, TCR 1240, TCR 215-A, TCR 

243-A, TCR 1113, TCR 848, TCR 1316, TCR 250-A, TCR 1214 

TCR 875, TCR 1330, TCR 731-A, TCR 193-A, TCR 206-A, TCR 189-A, TCR 1070, TCR 356-A, TCR 

557-A, TCR 432-A, TCR 182-A, TCR 434-A, TCR 282-A, TCR 453-A, TCR 489-A, TCR 1790-A, TCR 

1294, TCR 170-A, TCR 266-A, TCR 224-A, TCR 1292, TCR 122-A, TCR 61-A, TCR 134-A, TCR 29-A, 

TCR 85-A, TCR 904, TCR 874, TCR 59-A, TCR 1176

PREMA et al., Screening of horsegram (Macrotyloma uniflorum) genotypes against Horsegram Yellow Mosaic Virus 121 

1247, TCR 161-A, TCR 1295, TCR 872, TCR 1058, TCR 307-A, 

TCR 66-A, TCR 151-A, TCR 1210, TCR 1303, TCR 275-A, TCR 

873, TCR 1204, TCR 167-A, TCR 1050, TCR 116-A and TCR 

1133 showed resistant reaction. Six genotypes showed 

moderately resistant reaction. Three genotypes showed 

moderately susceptible reaction. 23 genotypes were found 

susceptible to the virus and remaining 30 genotypes showed 

highly susceptible reaction to HgYMV (Table 1). Muniyappa, 

et al., 1976 and 1978 screened horsegram genotypes for yellow 

mosaic virus disease under filed conditions during summer, 

1975 and the genotypes exhibiting mild as well as moderate 

symptoms for HgYMV were identified. The results are in 

conformity with the findings of Rajkumar, et al., 2009, 

Srinivasulu, et al., 1991 and Parimala, et al., 2011. The 

genotypes identified in the present study can be used in 

breeding programme for developing tolerant varieties as no 

other management methods are economical for the 

management of this disease. 


First author thankfully acknowledges Department of 

Science and Technology (DST), New Delhi for providing 

financial assistance through Inspire programme. 


Muniyappa, V., Chandrashekaraiah, S.C. and Shivashankar, G. 1978, 

Horsegram cultures tolerant to yellow mosaic. Indian J. Genet. Pl. 

Breed., 38: 148-149. 

Muniyappa, V., Rajeshweri, R., Bharathan, N., Reddy, D.V.R. and Nolt, 

B.L. 1987. Isolation and characterisation of a geminivirus causing 

yellow mosaic disease of horsegram (Macrotyloma uniflorum) in 

India. J. Phytopathol., 119: 81-87. 

Muniyappa, V. and Reddy, H.R. 1976. Studies on the yellow mosaic 

disease of horsegram (Dolichos biflorus L.)- Virus vector relationships. 

Mysore J. Agric. Sci., 10: 605-610. 

Muniyappa, V., Reddy, H.R. and Shivashankar, G. 1975. Yellow mosaic 

disease of Dolichos biflorus L. (horsegram). Curr. Res., 4: 176. 

Muniyappa, V., Reddy, H.R. and Shivashankar, G. 1976. Studies on the 

yellow mosaic disease on horsegram (Macrotyloma uniflorus Syn. 

Dolichos biflorus). III. Evaluation of germplasm for the disease. 

Curr. Res., 5: 52-53. 

Parimala, K., Meenakumari, K. V.S., Sudhakar, R. and Kanaka Durga, 

K. 2011. Screening of horsegram genotypes against yellow mosaic 

and powdery mildew diseases, Indian J. Pl. Protect., 39: 160-161. 

Rajkumar, S.G., Prameela, H.A., Rangaswamy, K.T., Divya, B.L., 

Shankarappa, K.S., Viswanatha, K.P. and Maruthi, M.N. 2009. 

Sources of resistance to Horsegram yellow mosaic virus disease. J. 

Pl. Protect. Environ., 6(1): 86-89. 

Srinivasulu, B., Rao, A.S., Basheeruddin, M. and Reddy, M.N. 1991. 

Reaction of some horsegram genotypes to yellow mosaic disease in 

Andhra Pradesh. Legume Res., 14(2): 101-102. 

Thakur, C. 1979. Horsegram. In: Scientific Crop Production, Vol. I, 

pp.319-320, Metropoliton Book Co. Pvt., New Delhi, pp.495. 

Williams, F.J., Grewal, J.S. and Amin, K.S. 1968. Serious and new diseases 

of pulse crops in India in 1966. Plant Dis. Reptr., 52: 300-304. 




Evaluation of Garlic Genotypes Under Temperate Conditions of Kashmir Valley 

MUSHTAQ, A. CHATOO, PARVAZE, A. SOFI 1 , KHALID, R. DAR, ANGREZ ALI AND TASADUK SHAFI 

Faculty of Agriculture, SKUAST-K, Wadura, Sopore, 193 201, J&K, India 

e-mail: parvazesofi@gmail.com 1 

Garlic (Allium sativum L.) is one of the most important 

bulb vegetables, which is used as spice and flavoring agent 

for foods. Globally the leading producers include China with 

an area of 6.9 lakh hectares followed by India (1.7 lakh hectares) 

and Russia (0.28 lakh hectares) FAOSTAT, 2010. 

For any successful breeding programme, presence of 

adequate genetic variability is an important prerequisite. 

Breeders employ various selection strategies to utilize the 

available genetic variability for seeking improvement in 

economically important traits such as yield. Therefore, it is 

imperative to characterize the available germplasm resources 

in terms of nature and extent of variability. A number of workers 

have reported tremendous variability, association and direct 

and indirect effect among bulb yield and yield traits (Figliuolo, 

et al., 2001; Shri Dhar, 2002; Naruka and Dhaka, 2004 and 

Tsega, et al., 2010 in their work on garlic).Therefore, the present 

study was aimed at characterizing the germplasm aqquired 

under AICRP programme for development of locally suitable 

cultivars of garlic to boost the production and productivity. 

Present study which was conducted during 2010-11 at 

the research farm of Faculty of Agriculture, SKUAST-K, 

Wadura (34 o 17’ North and 74 o 33 E at altitude of 1594 m amsl), 

20 diverse genotypes (19 procured from Directorate of Onion 

and Garlic under AICRP and one local check) were evaluated 

in an Randomised Block Design with four replications. Each 

genotype was represented by 20 rows of 2 meter length with 

row to row spacing of 15 cm and plant to plant spacing of 10 

cm. Standard agronomic practices were followed to ensure a 

good crop. Data was collected for 13 quantitative traits (Table 

1) from 10 randomly selected competitive plants from each 

replication and subjected to standard statistical analysis. 

Variability components were calculated as per the 

method of Johnson, et al., 1955 and the correlation coefficients 

were computed following Aljibouri, et al., 1958. 

The results pertaining to mean performance of the 

germplasm accessions for 13 quantitative traits are presented 

in Table 1. Perusal of the means table reveals wide variation in 

trait expression and superiority of several genotypes over 

local check in respect of maturity, morphological and yield 

traits. The wide range and the mean suggested existence of 

sufficient variability among the tested genotypes for majority 

of characters indicating that there is considerable potential in 

improvement of garlic. High genetic variability has also been 

reported in garlic by Fugliuolo, et al., 2001 and Tsega, et al., 

2010. 

Analysis of variance for the 13 traits (Table 2) revealed 

significant variance in all traits except for days to maturity, 

leaf length, leaf width, pseudo-stem length, equatorial diameter 

and marketable yield (%). This is also quite evident from small 

range and CV recorded for these traits. PCV estimates were 

invariably higher for all traits except days to maturity and 

marketable yield (%). Generally the PCV estimates were higher 

than their corresponding estimates of GCV possibly due to 

the influence of environment on these traits due to quantitative 

nature of the traits under study. PCV values ranged from 5.09 

% for Marketable bulb yield to 320.33 % for number of cloves, 

while as GCV values ranged from 3.46 % for Marketable bulb 

yield to 320.09 % for number of cloves. There was close 

correspondence in the values of GCV and PCV for number of 

leaves, average bulb weight, number of cloves, average clove 

weight and total yield, indicating greater correspondence 

between genotype and phenotype as is also depicted by high 

heritability estimates for these traits. Similar results have been 

reported by Tsega, et al., 2010 in garlic, Abayneh, 2001 in 

onion. Heritability (broad sense) estimates ranged from low 

(24%) in case of equatorial diameter to very high (99%) in 

case of number of cloves, but were invariably high for most of 

the traits, which indicates that there is scope for improvement 

for these traits through selection. Genetic gain was higher for 

all traits except days to maturity, leaf width and marketable 

yield (%). Generally in plant breeding, high mean, high 

heritability, high GCV and high genetic advance are taken as 

the indicators of the possible improvements in trait expression 

that could be achieved through appropriate selection 

strategies. In light of such proposition it is expected that traits 

like plant height, number of leaves, leaf length, pseudo-stem 

length, polar diameter, equatorial diameter, average bulb weight, 

number of cloves, average clove weight and total bulb yield 

would respond to selection as they posses high values of the 

important genetic parameters as outlined above. 

However, since most of the economically important traits 

are complex in inheritance especially the yield that is governed 

by a large number of traits through direct as well as indirect 

influences. Therefore, it is imperative to elucidate the nature 

and extent of trait interrelationships to arrive at an optimal 

selection index for improvement of yield.this is all ther more 

important in view of the general observation that direct 

selection for yield per se has been invariably less effective. In 

the present study, the degree of trait associations as measured 

by correlation coefficients revealed that the total bulb yield 

was significantly correlated with average bulb yield (r=0.898),

Table 1. 

Entry 

Table 2. 

Source of 

variation 

CHATOO et al., Evaluation of Garlic Genotypes under Temperate Conditions of Kashmir Valley 123 

Analysis of variance for different morphological and yield traits in Garlic 

d.f Days to 

maturity 

followed by average clove weight (r=0.842), leaf width 

(r=0.7980, equatorial diameter (r=0.796), leaf length (r=0.691), 

polar diameter (r=0.6880 and plant height (r=0.605). Yield was 

negatively correlated with pseudo-stem length (r=-0.249) even 

though the value was non significant. 


Plant 

height 

Number 

of 

leaves 

Leaf 

length 

Abayneh, M. 2001. Variability 1. and association among bulb yield, 

quality and related traits in onion (Allium cepa L.). M.Sc. Thesis 

submitted to School of Graduate Studies, Alemaya University, pp.51. 

Polar Equatorial Average 

diameter diameter bulb 

weight 

Number 

of cloves 

Average 

clove 

weight 

Marketable 

bulb yield 

(%) 

Total yield 

Replication 3 51.85 32.93 0.29 11.22 0.42 1.49 0.46 0.41 18.56 13.23 230.92 33.68 833.03 

Genotypes 19 336.17** 422.31** 4.65 100.35** 1.09** 12.18** 2.33** 1.17* 853.13** 7175.63** 11128.86** 42.63** 37938.85** 

Error 57 42.34 41.56 0.41 11.89 0.17 1.70 0.26 0.46 11.35 2.98 186.67 10.26 503.36 

Table 3. 

Parameter 

Mean performance of 20 garlic genotypes for 13 quantitative traits. 

Days to 

maturity 

Variability Parameters for different morphological and yield traits in Garlic 

Days to 

maturity 

Plant 

height 

(cm) 

Plant 

height 

Number 

of leaves 

Number 

of leaves 

Leaf 

length 

(cm) 

Leaf 

length 

Leaf 

width 

(cm) 

Leaf 

width 

Pseudostem 

length 

(cm) 

Leaf Pseudostem 

width 

length 

Pseudostem 

length 

Polar 

diameter 

(cm) 

Polar 

diameter 

Equatorial 

diameter 

(cm) 

Equatorial 

diameter 

Average 

bulb 

weight 

(g) 

Average 

bulb 

weight 

Number 

of cloves 

Number of 

cloves 

Average 

clove 

weight 

(g) 

Average 

clove 

weight 

Marketable 

bulb yield 

(%) 

Marketable 

bulb yield 

(%) 

Total 

yield (g) 

AG 102 204.50 50.00 6.35 27.55 0.70 4.80 3.72 3.35 30.00 15.00 146.50 93.60 199.99 

AG 103 212.50 58.25 7.25 30.00 1.90 6.00 4.12 4.36 60.00 14.00 243.75 92.25 399.99 

AG 104 213.00 58.75 7.00 30.00 0.72 5.70 4.04 3.50 45.00 12.00 178.75 93.90 299.99 

AG 105 216.25 59.75 7.95 27.65 0.62 3.75 3.39 4.01 38.75 12.00 131.50 92.99 255.88 

AG 120 213.50 35.75 4.35 20.00 0.40 3.00 2.37 2.34 14.01 10.00 48.00 91.92 93.41 

AG 121 215.25 49.75 6.35 20.30 0.40 8.00 3.77 3.20 19.85 20.00 64.00 91.41 132.33 

BG 106 189.50 60.00 4.75 31.00 0.35 8.59 3.72 3.24 20.12 14.92 68.67 86.72 134.16 

BG 107 206.00 45.00 5.00 23.00 0.26 7.47 2.89 3.10 11.75 7.85 52.90 86.30 78.33 

BG 108 203.75 43.00 7.06 24.00 0.27 8.70 2.75 2.82 10.00 8.10 46.00 86.42 65.16 

BG 109 209.50 60.00 4.90 24.20 0.32 12.20 3.82 3.33 16.43 10.15 55.25 88.56 109.53 

BG 110 187.00 42.75 5.80 23.00 0.36 11.30 3.41 3.00 14.00 9.40 49.97 87.64 93.33 

BG 111 219.00 52.75 8.00 26.27 0.33 11.50 3.55 3.23 16.00 11.65 57.87 88.91 106.66 

CG 112 184.00 48.75 5.00 28.00 0.31 8.20 4.50 3.45 30.12 23.00 66.74 80.67 200.82 

CG 113 196.25 55.30 5.90 27.90 0.40 9.00 3.85 3.10 18.02 17.95 50.24 83.10 120.20 

CG 114 196.25 34.25 5.05 21.07 0.21 13.00 2.80 2.75 10.00 10.65 47.80 75.79 66.79 

CG 115 203.25 46.75 4.42 24.90 0.30 9.90 3.35 3.00 15.05 16.02 47.07 71.41 100.37 

CG 116 194.00 68.00 4.95 38.15 1.20 11.00 5.70 4.50 59.94 16.30 185.53 81.78 399.61 

CG 117 197.50 65.00 5.00 39.20 1.30 10.90 5.00 4.15 46.00 14.95 154.04 80.90 306.66 

CG 118 216.50 32.75 4.50 23.15 0.32 9.20 2.75 3.20 11.00 7.40 78.94 77.56 73.47 

CG 119 196.00 55.05 5.00 25.00 0.30 10.00 3.30 3.30 13.55 14.60 44.50 84.67 90.37 

Local 210.00 60.00 10.00 28.00 1.00 10.00 3.82 5.00 35.00 12.00 140.00 90.00 233.33 

Check 

Mean 203.97 51.50 5.93 26.77 0.57 8.67 3.64 3.42 25.45 13.23 93.23 86.02 169.54 

CV (%) + 10.48 9.97 1.48 5.05 0.43 2.75 0.77 0.63 16.07 4.06 59.12 6.23 107.13 

Mean 203.97 51.50 5.93 26.77 0.57 8.67 3.64 3.42 25.45 13.23 93.23 86.02 169.54 

Range 187.00- 

219.00 

32.75- 

68.00 

4.35- 

10.00 

21.07- 

39.20 

0.21- 

1.90 

3.00- 

13.00 

2.37-5.70 2.34-5.00 10.00- 

60.00 

7.40-23.00 46.00- 

146.50 

71.41-93.60 65.16- 

399.99 

σ 2 G 73.45 95.18* 1.60** 22.11 0.23 2.62 0.51* 0.18 210.44** 1793.15** 2735.54** 8.92 9108.12** 

σ 2 E 42.34 41.56 0.41 11.89 0.17 1.70 0.26 0.46 11.35 2.98 186.67 10.26 503.36 

PCV 5.27 22.70 26.30 21.78 110.95 23.97 24.10 23.39 58.51 320.33 57.98 5.09 57.82 

GCV 4.20 18.94 21.33 17.56 84.13 18.66 19.61 12.40 57.00 320.07 56.10 3.46 56.29 

Heritability 0.63 0.69 0.79 0.65 0.57 0.60 0.66 0.28 0.94 0.99 0.93 0.46 0.94 

(bs) 

Genetic 

Advance 

6.83 32.26 42.80 23.51 1.29 29.54 32.76 13.49 113.27 653.21 111.07 4.73 111.92 

Total 

yield 

FAO. 2010. FOASTAT. 2010. 

Figliuolo, G., Candido, V., Logozzo, G. and Zeuli, P.L.S. 2001. Genetic 

evaluation of cultivated garlic germplasm. Euphytica, 121: 325- 

34. 

Naruka, I.S. and Dhaka, R.S. 2004. Correlation studies in garlic (Allium 

sativum L.). Prog. Hort. 36: 128-31. 

Shri Dhar. 2002. Genetic variability and character association in garlic. 

Prog. Hort., 34: 88-91. 

Tsega, K., Tiwari, A. and Woldetsadik, K. 2010. Genetic variability, 

correlation and path coefficient among bulb yield and yield traits in 

Ethiopian garlic germplasm. Indian J. Hort., 67: 489-499. 


Trends in Biosciences 5 (1-4): 124-127, 2012 

Author Index 

Vol. 5 (1-4) 2012 

A 

Achahbar, Sanaa 266 

Afroza, Baseerat 343 

Aftab, Khan Uzma 218 

Ahmad, I. 306 

Ahmad, Iffat Zareen 218 

Ahmad, Ishfaq 136 

Ahmad, Nabeel 214 

Akhtar, Md. Humayoon 17 

Akhtar, Shirin 299 

Ali, Gowhar 97, 119, 136, 306 

Ali, Hasmat 225 

Ali, S.S. 71, 352 

Alim, Md. A. 287 

Anand, Lalitha 8 

Ansari, M.S. 41 

Ansari, Mohammad Israil 95 

Ansari, Rizwan Ali 202 

Arora, Neeraj 191 

Arulkirubakaran, M. 274 

Arya, Arvind 168 

Ashoka, H. G. 163 

Aziz, Ozair 101 

B 

Bahar, Fayaz Ahmed 107 

Bal, R.S. 127, 244 

Balai, Laxman Prasad 147, 152 

Barooah, Madhumita 220 

Behura, A.K. 287 

Bharadwaj, Renu 61 

Bharathiraja, B. 274 

Bhareti, Priyanka 161 

Bhat, F.A. 301 

Bhat, M. Afzal 284 

Bhat, Mohd. Yaqub 51 

Bhavani, P. 272, 312 

Bind, Akhilesh 214 

Biswas, Bikash Kanti 212 

Bora, Sudipta Sankar 220 

Boro, Robin Ch. 220 

Burange, P.S. 199 

C 

Capoor, Ajay 45, 57 

Chami, Fouzia 266 

Chami, Najat 266 

Chandra, Kunj 261 

Chandra, Sourav 212 

Chandran, M. 274 

Chaturvedi, Rajesh 218 

Chaubey, B.K. 64 

Chaudhary, Puja 349 

Choudhary, Saumya 89 

Choudhury, D. Roy 129, 143 

D 

Dar, Manzoor Hussain 284 

Dar, S.A. 97, 119, 338 

Dar, Z.A. 306, 317, 338, 343 

Das, M. 129, 143 

Das, Mrs. R. 212 

Dash, Debiprasad 234 

Deka, Archana 220 

Deshpande, Smita 61 

Devasahayam, Mercy 114 

Dhiman, Anil Kumar 1 

Durande, G.B. 196 

G 

Gade, R.S. 196 

Ganai, M.A. 338


Gangaprasad, S. 184 

Gaur, Isha 176 

Gautam, K.M. 297 

Ghrmah, Hamid A. 166 

Gonde, A.D. 199 

Gowda, Byre 140 

Gulfishan, Mohd 14 

Gulzaffar 317 

Gupta, Dolly 301 

Gupta, Kamlesh 79 

Gupta, Nidhi 160 

Gupta, Praveen 1, 180 

Gupta, Suman 114 

Gupta, Vishal 301 

Gusain, Poonam 246 

H 

Habib, M. 317, 338, 343 

Hasan, Wajid 41 

Hussain, Barkat 284 

Hussain, K. 343 

I 

Iqbal Asif, M. 97, 306 

Ishfaq, A. 119 

J 

Jagtap, R.N. 196 

Jain, Piyush 22 

Janaiah, C. 205 

Jayamuthunagai, J. 274 

Jayaram, Neetha 140 

Jirli, Basvaprabhu 222 

Joshi, Anita 61 

Joshi, Sarita 79 

K 

Kalha, C.S. 301 

Kalita, Rituparna 220 

Kamaluddin 136 

Kanoujia, Budh Prakash 214 

Kant, Rama 225 

Kapoor, Shalini 191 

Karmakar, Prabir Kumar 212 

Karunakar, V. 205 

Katoch, Anu 54 

Kaur, Amandeep 176 

Kaur, Gurpreet 176 

Kaur, Harpreet 54, 133, 332 

Khan, Ainul Haq 14 

Khan, Khalil 346 

Khan, M. Luqman 111, 332 

Khan, M.H. 119 

Khan, Mohd Kamran 89 

Khan, S.H. 343 

Khan, Tabreiz Ahmad 202 

Khanna, Veena 133 

Krishna, Ram 225 

Krupakar, A. 208 

Kumar, A 199, 244 

Kumar, Anil 4, 47 

Kumar, Arvind 77, 310 

Kumar, Ashok 127 

Kumar, B. Ravi 74 

Kumar, Binod 261 

Kumar, G. N. Veera 140 

Kumar, H. B. Manoj 140 

Kumar, Jeewesh 154, 240 

Kumar, Jitendra 4 

Kumar, K. 64 

Kumar, Kshitij 171, 279 

Kumar, Rajesh 191, 246 

Kumar, Sandeep 168 

Kumar, Vinod 222 

Kumar, Vipul 68 

Kundagrami, S. 129, 143 

L 

Lahan, Jyoti Prasad 220

126 Trends in Biosciences 5 (1-4), 2012 

Lal, Gabrial M. 335 

Lal, Kanhaiya 225 

Lather, V.S. 315 

Lingaraju, S. 74 

Lodam, V. A. 122 

Lone, Aijaz A. 306 

M 

Madhusudhan, R 8, 184 

Madke, P.K. 310 

Mahesh, Y.S. 74 

Manjunath, B. 8, 74, 140 

Mankar, D.M. 349 

Manzar, Abu 119 

Markanday, Jagdish 326 

Marker, S. 208 

Masih, Sam A 114 

Masih, Sama 214 

Mastan, S.A. 35 

Mehandi, Suhel 261, 329 

Mishra, Abha 25 

Mishra, V.K. 64 

Mittal, Pallavi 176 

Mohiddin, F.A. 301 

N 

Nagaraja, T.G. 20 

Nagaraju, H. K. 140 

Naik, Abhishek 299 

Najeeb, S. 338 

Nanda, S.S. 287 

Nandni, Durgesh 31 

Nayeem, Md. Rashid 17 

Neha, Kumari 335 

Nehvi, F.A. 97 

O 

Ojha, M.D. 295 

P 

Pal, Rishi 38, 321 

Palakshappa, M.G. 74 

Pandey, Anamika 89 

Pandey, V. P. 11 

Panickar, B. 138, 149 

Panwar, R.K. 161 

Parida, D. 287 

Parray, G.A. 338 

Patel, I. S. 138, 149, 157 

Patel, J. R. 157 

Patel, P. S. 138, 149 

Patial, Pankaj 326 

Patil, P. P. 122 

Patil, P.D. 188 

Patil, S. S. 122 

Patil, V. A. 122 

Patro, H. 287 

Patro, Hrusikesh 234 

Pervez, Rashid 11, 71 

Pir, F.A. 119 

Pradhan, Bhavana 61 

Prakash, Chandra 104, 297 

Prasad, C.S. 38, 321 

Prasad, D. Theertha 272, 312 

Prasad, Keshav 47 

Prasad, P.S. 303 

Prashantha, G. M. 163 

R 

Raghuwanshi, Arun 31 

Rahman, S. M. A. 11 

Rai, Manju 176 

Rajput, M.M. 4 

Ramappa, M. 140 

Ramaswamy, G. R. 294 

Rangare, N.R. 329 

Ranjeet 225 

Ravi, B.A. 184 

Ravindrababu, Y. 138, 149


Rawal, R. D. 294 

Reddy, S.M. 205 

Remmal, Adnane 266 

Risbud, Shilpa 61 

S 

Sachan, B.S. 4, 47 

Sagwal, Pardeep Kumar 289 

Saifulla, Muhammad 303 

Sarkar, M. 129 

Saroj, Pushpendra 222 

Sen, Anand 346 

Shah, Naseer Hussain 51 

Shahid, Mohd 68 

Shaikh, S.S. 20 

Shamim, Mohammad 231 

Sharan, Abhishek 214 

Sharma, Arun 297 

Sharma, Dinesh 289 

Sharma, J.C. 297 

Sharma, Poonam 133 

Sharma, R.C. 341 

Shikari, Asif 338 

Shivran, R.K. 104, 297 

Shrivastava, Vinoy Kumar 31 

Singh, A. P. 240 

Singh, Abhishek Pratap 11 

Singh, Anuradha 68 

Singh, Chandra Mohan 261, 329 

Singh, Charandeep 176 

Singh, D. C. 240 

Singh, Guriqbal 133 

Singh, H. 295 

Singh, Hamveer 77, 310 

Singh, Ishwer 289 

Singh, Jai 147, 152 

Singh, K. N. 107, 171, 279 

Singh, Krishna 25 

Singh, R. B. 147, 152 

Singh, S.K. 154 

Singh, Sunil Kumar 329 

Singh, Vir 246 

Sofi, P.A. 136 

Srinivasulu, Ch. 205 

Srivastava, Mukesh 68 

Srivastava, Richa 114 

Subha, Jyotsna 208 

T 

Tanveer, Hasan 77, 310 

Tarafdar, S. 143 

Tayade, D.S. 196 

Thakur, Neelam 332 

Thomas, George 89 

Tiwari, G.N. 38 

Tripathi, Atul 208 

Tulankar, K.P. 199 

U 

Usmani, Mohd. Kamil 17 

V 

Vashi, R. D. 122 

Verma, Anil K. 111 

Vijaya Mahantesha, S. R. 294 

Vivek, B.C. 8 

W 

Wagh, S.S. 188 

Wani, A.H. 51 

Wani, Shafiq. A. 317 

Wargantiwar, R.K. 199 

Warsi, Zeenat 45, 57 

Y 

Yadav, A.S. 341 

Yadav, C.B. 64 

Yogeesh, L. N. 184 

Z 

Zaffar, Gul 136

Trends in Biosciences 5 (1-4): 128-132, 2012 

A 

2, 4-D 171 

2, 4-D, somaclonal variation 279 

Abelmoschus esculentus 199 

Abiotic factors 17 

Abortion 212 

Absolute frequency 202 

Absolute 202 

Abundance 25 

Acrididae 17 

Activity 205 

Advance as per cent of mean 335 

Aeromonas hydrophila 35 

Agrobacterium 279 

Agro-morphometric 208 

Aligarh fort 17 

Alternaria alternate 205 

Alternaria 294 

Analysis of variance 338 

Anthracnose 140 

Anti bacterial 20 

Antibacteria 312 

Antibiotic resistance 101 

Antibody 212 

Anticarcinogenic effect 97 

Antifungal 312 

Antigen 212 

Antiglycemic effect 97 

Antiinsecticidal 312 

Aspergillus clavatus 205 

Aspergillus niger 214 

Assay 321 

Autecology 31 

Azotobacter 107 

B 

Bacillariophyceae 220 

Bacillus megatarium 20 

Bacillus 57 

Banana 8 

Basmati rice 289 

Bio 154 

Biocontrol 68 

Subject Index 

Vol. 5 (1-4) 2012 

Bioefficacy 41 

Bioenergy crops 89 

Bioenergy 89 

Bioethanol 214 

Biofertilizers 274 

Biofuels 89 

Biohydrogen 114 

Biological control 321 

Biomass 89 

Biopharma 1 

Bioprocess 180 

Biotechnology 1 

Blood Groups 191 

Bombyx mori 196 

Brachistin ae 231 

Braconidae 166, 231 

Brassica juncea 225 

Breeding 326 

Broad sense heritability 335 

Broad 61 

Brood-stock 326 

Brown sarson 317 

Brucella 212 

Bt. cotton 341 

C 

Cabbage aphid 284 

Cabbage 284 

Canal water 47 

Carbofuran 310 

Carp 326 

Cartaphydrochloride 310 

Catantopidae 17 

cDNA library 8 

Cell Culture 180 

Cell membrane 143 

Cellular damage 218 

Cellulase enzyme 214 

Cercospora 127 

Channa striatus 35 

Chemicals 51 

Chickpea 104, 119, 129, 240 

Chlorella 220


Chlorophyceae 220 

Clusters 64 

Coccicidal activity 266 

Coccidiosis 266 

Cocophilus 332 

Colletotrichum falcatum 95 

Colony Forming Unit (CFU) 303 

Combining ability 225, 317 

Combiningability 338 

Compatibility 71 

Component analysis 343 

Component traits 329 

Consumptions 4 

Cormel 168 

Cotton cultivators 349 

Cotton 332 

Coulombic efficiency 114 

Cow urine decoctions 41 

Cowpea 149 

Crop growth parameters 289 

Crop improvement 208 

Cross sowing 22 

Curcuma 11 

Cyanophyceae 220 

D 

Demand 4 

Density 202 

Desynapsis 14 

Diacrisia obliqua 45, 57 

Diallel 343 

Dietary patterns 4 

Disease Incidence 301 

Disease Severity 301 

Disease 127 

DOE 180 

Dolichos bean 140 

Dominance 25 

Dry bean 97 

Dual purpose mungbean 315 

Dynamics 284 

E 

Earias vittella 199 

Early blight 294 

Ecological factors 332 

Economics 289 

Efficacy insecticide 57 

Efficacy 45, 244 

Efficiency 163 

Egg hatch 54 

Eimeria oocyst 266 

Enterobacter 114 

Entomopathogenic nematodes 71, 321 

Entomopathogens 38 

Environment 338 

EO components 266 

Epidemic 95 

Escherchia coli 20 

Euglenophyceae 220 

Expressed sequence tag 8 

Extension gap 295 

Extention contact 349 

F 

Fababean 64 

False smut 301 

Farm yard manure (FYM) 287 

Fenugreek 299 

Fertilizers 47, 301 

Field capacity 163 

Field pea 244 

Fipronil 310 

FIRBS 22 

Foliar 140 

Food 4 

Formulation 45 

Free radical 218 

Fungicidal 205 

Fungicides 244 

Fusarium udum 303 

G 

Gall index 51 

Garlic 54 

gca 306, 317 

GCV 119 

Gene action 306 

Genetic advance 119, 329 

Genetic divergence 64 

Genetic variability 11 

Genetic 335 

Genotypes 157 

Germplasm 184, 299, 301, 315


Gladiolus 168 

Grain mould 74 

Grain yield 234 

Green foddern 136 

Growth 4, 22, 51 

H 

H. armigera 154, 240 

Hardening 176 

Harvesting 163 

Heat tolerance 143 

Heavy metals 101 

Helminth parasites 25 

Hemicriconemoides 332 

Herbicides 133 

Heritability 11, 119, 136, 329 

Heterohabditis indica 38, 321 

Heteropteran bugs 205 

Heterosis 122 

Hoagland solution 129 

Homolobus 166 

Hymenop tera 231 

Hypertension 191 

I 

Identification 111 

Ideotype 315 

Imbibition rate 129 

Impact 222 

In vitro growth 133 

India 231 

Indian patent law 1 

Indica rice 171, 279 

Infection 140 

Infertility 212 

Inheritance 184 

Inorganic fertilizers 274 

Input levels 22 

Insecticide resistance 341 

Insecticides 149, 199 

Instars 31 

IPM modules 188 

IPM 341 

IR 64 171, 279 

Irrigated groundnut 287 

Irrigation regimes 297 

K 

K 234 

Kharif Onion 295 

L 

Laggards 14 

Land degradation 47 

Land races 315 

Larval duration 196 

Leafhopper 138 

Lepidopteron 321 

Life history 31 

Line sowing 22 

Line x tester 122 

Lipaphis erysimi 41 

Lipid per oxidation 218 

Lipid Profile 191 

Liriomyza trifolii 188 

Livestock 161 

Longa 11 

Lymphocyte transformation 35 

M 

Maize 107, 157 

Management 199, 240 

Manure 104 

Maruca vitrata 149 

Mass production 38 

Maturation 326 

MDA 218 

Mean intensity 25 

Medium Optimization 180 

Meloidogyne javanica 352 

Meloidogyne 54 

MFC 114 

MIC 61 

Micrococcus sp. 20 

Micronuclei 14 

Micropropagation 176 

Microshoots 176 

Mint 54 

MMS 14 

Modulus of elasticity 163 

Monoclonal Antibodies 180 

Morphological traits 261 

Mortalit 45 

Moth bean 184 

Moth emergence 196 

Mulch 107 

Multiplication 176


Mungbean 147, 152 

Mutagenesis 208 

Mycrobes 274 

MYMV 127 

N 

N 234 

Natural 161 

Nematodes 352 

Net returns 22 

New species 166, 231 

Newer 149 

Nitrogen 107, 289 

Nodulation-efficiency 133 

Nodules 51 

North Karnataka 74 

Novel antimicrobial 61 

Novel insecticides 154 

Nutrient yield maximization 287 

O 

Oats 136 

Oocyst destruction 266 

Organic sources 234 

Organic substrates 68 

Ornamental 168 

Oryza sativa 301 

P 

P 234 

Papilio demoleus 31 

Papilionidae 31 

Patenting 1 

Pathogenicity 38 

PCV 119 

Pesticides 71 

Petrochemical industry 101 

Phosphorous levels yield 289 

Physiological traits 261 

Pigeonpea 154, 303 

PIs 312 

Pituitary gland 326 

Plant extracts 54 

Plant products 199 

Plant-parasitic nematodes 332 

Plasmid 8 

Pod infection 140 

Pollen fertility 14 

Polyenzyme formulation 61 

Population dynamics 240, 303 

Population index 31 

Population 284 

Pratylenchus thornei 352 

Premature birth 212 

Prevalence 25 

Production 4, 222 

Productivity 161 

Protein 104 

Pseudomonas aeruginosa 20 

Pupal duration 196 

Pyrgomorphidae 17 

Q 

Quality traits 329 

R 

RBPT 212 

Reapers 163 

Red rot 95 

Red soil 346 

Regeneration 168, 171, 279 

Relative density 202 

Relative frequency 202 

Relative injury 143 

Renewable energy 89 

Resistance 127 

Resources 161 

Rhizobium 101, 133 

Rice 122, 171, 234, 261, 279, 329, 335, 338 

Root explant 168 

Root induction 176 

Root-knot nematode 51 

ROS 218 

Rotylenchulus reniformis 352 

Roving survey 140 

Rust 244 

S 

S content 234 

Saccharomyces cerevisiae 214 

Salinity tolerance 129 

Saudi Arabia 166 

sca 306, 317 

Scaly fishes 25 

Scent components 205 

Schizoprymnus 231 

Screening 157, 294 

Seed leaf 272


Seed 129, 138 

Seedling vigour 129 

Segregants 64 

Sequencing 8 

Shear stress 163 

Sodic soil 346 

Sodium azide 208 

Soil moisture 346 

Soil nitrogen fixation 161 

Soil 161 

Solid 214 

Sorghum 74 

Sowing dates 152 

Spacing 152 

Specific gravity 214 

Spectrum antimicrobial 61 

Spotted pod borer 149 

State fermentation 214 

Stembores 157 

STI 312 

Strawberry 111 

Subabul trypsin inh ibitor 312 

Subabul 272 

Substrate moisture levels 68 

Sucking pests 138, 341 

Sugarcane 95 

Surface sterilization 176 

Survey 74, 202, 352 

Survival 143 

Sustainable 161 

Swarna Transformation 279 

Swarna 171 

Sweet pepper 343 

Symbiosis 133 

Synergy 61 

T 

Technology dissemination 295 

Technology gap 295 

Technology index 295 

Test 35 

Thermostability 143 

Thrips 138 

Thuringiensis 57 

Tomato cultivars 294 

Tomato 188, 274 

Traits 208 

Treatment 129, 138 

Trypsin inhibitor 272 

Tubebell 47 

Turmeric 11 

U 

Urdbean 127 

V 

Variability 329, 335 

Variation 299 

Varietal effect 196 

Varities 284 

Vicia faba 352 

Vigna radiata L. 51 

W 

Wastewater 101 

Water stress 261 

Watershed development 222 

Web blight 147, 152 

Weed 107 

Wheat 208, 297 

Whitefly 138 

X 

X. bacaniboia 111 

X. insigne 111 

X. radicicola 111 

X. simillimum 111 

Xanthophyceae 220 

Xiphinema fragariae 111 

Y 

Yellow stem borer 310 

Yield component traits 261 

Yield components 122, 225 

Yield loss 147 

Yield 22, 122, 136, 143, 299, 338 

YMV 184 

Z 

Zea mays 306 

Zero tillage 297 

Zinc 346

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Fox, P.C. and Atkinson, H.J. 1984. Glucose phosphate isomerase polymorphism in field population of the potato cyst 

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