Abstract
Cladode brown spot (CBS) is an important disease of Nopalea cochenillifera in the semiarid region of Northeastern Brazil. It has been reported in several countries, but its etiology is controversial, attributed to a complex of pathogens. Fifty fungal isolates were obtained and identified based on morphology and phylogeny, through analysis of the ribosomal DNA internal transcribed spacers (ITS), translation elongation factor 1-alpha (TEF1-alpha), β-tubulin (TUB-2), second largest subunit of RNA polymerase (RPB2), glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and the mating type locus MAT1–2 (ApMAT) partial gene sequences. Seven fungal general and twenty species were identified – Alternaria sp., A. longipes and A. tenuissima; Colletotrichum fructicola and C. siamense; Fusarium lunatum, F. incarnatum, and F. verticillioides; Lasiodiplodia euphorbicola, L. iraniensis, L. pseutheobromae, L. theobromae and Lasiodiplodia sp.; Neofusicoccum batangarum; Neopestalotiopsis australis, N. protearum and Neopestalotiopsis sp.; and Nigrospora sphaerica, N. hainanensis and Nigrospora sp. Four of these species had already been reported on Cactaceae of the genus Opuntia, but only N. batangarum, C. fructicola and C. siamense have been reported to date on the genus Nopalea. All isolates were pathogenic to detached cladodes of N. cochenillifera ‘Miúda’. Differences in aggressiveness were observed among the species, with L. iraniensis and F. lunatum characterized as the most aggressive species, whilst F. verticillioides as the least aggressive. This study provide important information on the fungi associated with cladode brown spot and improve the strategies for the management of disease on N. cochenillifera.
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References
Alves, A., Crous, P. W., Correia, A., & Phillips, A. J. L. (2008). Morphological and molecular data reveal cryptic speciation in Lasiodiplodia theobromae. Fungal Diversity, 28, 1–13.
Ammar, M. I., Shltout, A. B., & Kamhawy, M. A. (2004). Cladode and fruit rots of prickly pear (Opuntia ficus-indica L. Mill.) in Egypt. Egyptian Journal of Phytopathology, 32, 119–128.
Barbosa, R. S., Cavalganti, V. A. L., Lopes, E. B., & Araujo, E. (2007) Doenças da Palma in E. B. Lopes (Ed.), Palma forrageira: cultivo, uso atual e perspectivas de utilização no semiárido nordestino (pp. 47–56). João Pessoa, Brazil: EMEPA/FAEPA.
Barbosa, R. S., Calvalcanti, V. A. L., Lopes, E. B, & Araújo, E. (2012). Doenças da Palma forrageira. In E. B. Lopes (Ed.), Palma forrageira: Cultivo, uso atual e perspectivas de utilização no semiárido nordestino (pp. 81–98). João Pessoa: EMEPA/FAEPA.
Barnet, H. B., & Hunter, B. B. (1998). Illustrated genera of imperfect fungi (4th ed.). St. Paul: APS Press.
Begoude, B. A. D., Slippers, B., Wingfield, M. J., & Roux, J. (2010). Botryosphaeriaceae associated with Terminalia catappa in Cameroon, South Africa and Madagascar. Mycological Progress, 9, 101–123.
Berbee, M. L., Pirseyedi, M., & Hubbard, S. (1999). Cochliobolus phylogenetics and the origin of known, highly virulent pathogens, inferred from ITS and glyceraldehyde-3-phosphate dehydrogenase gene sequences. Mycologia, 91, 964–977.
Bezerra, J. D., Santos, M. G., Svedese, V. M., Lima, D. M., Fernandes, M. J., Paiva, L. M., & Souza-Motta, C. M. (2012). Richness of endophytic fungi isolated from Opuntia ficus-indica Mill. (Cactaceae) and preliminary screening for enzyme production. World Journal of Microbiology and Biotechnology, 28, 1989–1995.
Bragança, C. A. D., Silva, L. L., Haddad, F., & Oliveira, S. A. S. (2016). First report of Colletotrichum fructicola causing anthracnose in cassava (Manihot esculenta) in Brazil. Plant Disease, 100, 857–858.
Capobiango, N. P., Pinho, D. B., Zambolim, L., Pereira, O. L., & Lopes, U. P. (2016). Anthracnose on strawberry fruits caused by Colletotrichum siamense in Brazil. Plant Disease, 100, 859.
Carbone, I., & Kohn, L. M. (1999). A method for designing primer sets for speciation studies in filamentous ascomycetes. Mycologia, 91, 553–556.
Conforto, C., Lima, N. B., Garcete-Gómez, J. M., Câmara, M. P. S., & Michereff, S. J. (2016). First report of cladode brown spot in cactus prickly pear caused by Neofusicoccum batangarum in Brazil. Plant Disease, 100, 1238.
Conforto, C., Lima, N. B., Garcete-Gómez, J. M., Câmara, M. P. S., & Michereff, S. J. (2017). First report of Colletotrichum siamense and C. fructicola causing cladode brown spot in Nopalea cochenillifera in Brazil. Journal of Plant Pathology, 99, 812.
Correia, K. C., Silva, M. A., Morais, M. A., Jr., Armengol, J., Phillips, A. J. L., Câmara, M. P. S., & Michereff, S. J. (2016). Phylogeny, distribution and pathogenicity of Lasiodiplodia species associated with dieback of table grape in the main Brazilian exporting region. Plant Pathology, 65, 92–103.
Costa, J. F. O., Ramos-Sobrinho, R., Chaves, T. P., Silva, J. R. A., Pinho, D. B., Assunção, I. P., & Lima, G. S. A. (2017). First report of Colletotrichum fructicola causing anthracnose on Annona leaves in Brazil. Plant Disease, 101, 386–387.
Dodge, B. O. (1938). A further study of the dry-rot disease of Opuntia. Mycologia, 30, 82–96.
Doyle, J. J., & Doyle, J. L. (1987). A rapid DNA isolation procedure for small quantities of fresh leaf tissue. Phytochemical Bulletin, 19, 11–15.
Farr, D. F., & Rossman, A. Y. (2019). Fungal database: Fungus-host distributions. http://nt.ars-grin.gov/fungaldatabases/fungushost/fungushost.cfm. Accessed 23 Mar 2019.
Ferreira, M. A., Bispo, S. V., Rocha Filho, R. R., Urbano, S. A., & Costa, C. T. F. (2012). The use of cactus as forage for dairy cows in semi-arid regions of Brazil. In P. Konvalina (Ed.), Organic farming and food production (pp. 169–189). Rijeka: InTech.
Fisher, P. J., Sutton, B. C., Petrini, L. E., & Petrini, O. (1994). Fungal endophytes from Opuntia stricta: a first report. Nova Hedwigia, 59, 195–200.
Flores-Flores, R., Velázquez del Valle, M. G., León-Rodriguez, R., Flores-Moctezuma, H. E., & Hernández-Lauzardo, A. N. (2013). Identification of fungal species associated with cladode spot of prickly pear and their sensitivity to chitosan. Journal of Phytopathology, 161, 544–552.
Glass, N. L., & Donaldson, G. (1995). Development of primer sets designed for use with PCR to amplifyconserved genes from filamentous ascomycetes. Applied and Environmental Microbiology, 61, 1323–1330.
Goh, T. K. (1999). Single-spore isolation using a handmade glass needle. Fungal Diversity, 2, 47–63.
Granata, G. (2001). Doenças bióticas e abióticas. In G. Barbera, P. Inglese, & E. Pimienta-Barrios (Eds.), Agroecologia, cultivo e usos da palma forrageira (pp. 112–122). Rome: FAO/Sebrae.
Hyde, K. D., Nilsson, R. H., Alias, S. A., Ariyawansa, H. A., Blair, J. E., Cai, L., De Cock, A. W. A. M., Dissanayake, A. J., Glockling, S. L., Goonasekara, I. D., Gorczak, M., Hahn, M., Jayawardena, R. S., Al Van Kan, J. A. L., Laurence, M. H., Lévesque, C. A., Li, X., Liu, J. K., Maharachchikumbura, S. S. N., Manamgoda, D. S., Martin, F. N., Mckenzie, E. H. C., Mctaggart, A. R., Mortimer, P. E., Nair, P. V. R., Pawłowska, J., Rintoul, T. L., Shivas, R. G., Spies, C. F. J., Summerell, B. A., Taylor, P. W. J., Terhem, R. B., Udayanga, D., Vaghefi, N., Walther, G., Wilk, M., Wrzosek, M., Xu, J.-C., Yan, J.-Y., & Zhou, N. (2014). One stop shop: backbones trees for important pytopathogenic genera: I. Fungal Diversity, 67, 21–125.
Katoh, K., & Standley, D. M. (2013). MAFFT multiple sequence alignment software version 7: Improvements in performance and usability. Molecular Biology and Evolution, 30, 772–780.
Lima, G. S. A., Assunção, I. P., Martins, R. B., Santos, H. V., & Michereff, S. J. (2011). Development and validation of a standard area diagram set for assessment of Alternaria spot on the cladodes of the prickly pear cactus. Journal of Plant Pathology, 93, 691–695.
Lima, N. B., Batista, M. V. D. A., Morais, M. A., Jr., Barbosa, M. A. G., Michereff, S. J., Hyde, K. D., & Câmara, M. P. S. (2013). Five Colletotrichum species are responsible for mango anthracnose in northeastern Brazil. Fungal Diversity, 61, 75–88.
Liu, Y. J., Whelen, S., & Hall, B. D. (1999). Phylogenetic relationships among ascomycetes: Evidence from an RNA polymerse II subunit. Molecular Biology and Evolution, 16, 1799–1808.
Lopes, E. B., Santos, D. C. E., & Vasconcelos, M. F. (2012). Cultivo da palma forrageira. In E. B. Lopes (Ed.), Palma forrageira: Cultivo, uso atual e perspectivas de utilização no semiárido nordestino (pp. 21–60). João Pessoa: EMEPA/FAEPA.
Maharachchikumbura, S. S. N., Guo, L. D., Cai, L., Chukeatirote, E., Wu, W. P., Sun, X., Crous, P. W., Bhat, D. J., McKenzie, E. H. C., Bahkali, A. H., & Hyde, K. D. (2012). A multi-locus backbone tree for Pestalotiopsis, with a polyphasic characterization of 14 new species. Fungal Diversity, 56, 95–129.
Maharachchikumbura, S. S. N., Hyde, K. D., Groenewald, J. Z., Xu, J., & Crous, P. W. (2014). Pestalotiopsis revisited. Studies in Mycology, 79, 121–186.
Marques, W. M., Lima, N. B., Morais, M. A., Jr., Barbosa, M. A. G., Souza, B. O., Michereff, S. J., Phillips, A. J. L., & Câmara, M. P. S. (2013a). Species of Lasiodiplodia associated with mango in Brazil. Fungal Diversity, 61, 181–193.
Marques, W. M., Lima, N. B., Morais, M. A., Jr., Michereff, S. J., Phillips, A. J. L., & Câmara, M. P. S. (2013b). Botryosphaeria, Neofusicoccum, Neoscytalidium and Pseudofusicoccum species associated with mango in Brazil. Fungal Diversity, 61, 195–208.
Netto, M. S. B., Assunção, I. P., Lima, G. S. A., Marques, M. W., Lima, W. G., Monteiro, J. H. A., Balbino, V. Q., Michereff, S. J., Phillips, A. J. L., & Câmara, M. P. S. (2014). Species of Lasiodiplodia associated with papaya stem-end rot in Brazil. Fungal Diversity, 67, 127–141.
Netto, M. S. B., Lima, W. G., Correia, K. C., da Silva, C. F. B., Thon, M., Martins, R. B., Miller, R. N. G., Michereff, S. J., & Câmara, M. P. S. (2017). Analysis of phylogeny, distribution, and pathogenicity of Botryosphaeriaceae species associated with gummosis of Anacardium in Brazil, with a new species of Lasiodiplodia. Fungal Biology, 121, 437–451.
Nuno Silva, D., Talhinhas, P., Varzea, V. M. P., Cai, L., Salgueiro Paulo, O., & Batista, D. (2012). Application of the Apn2/MAT locus to improve the systematics of the Colletotrichum gloeosporioides complex: an example from coffee (Coffea spp.) hosts. Mycologia, 104, 396–409.
Nylander, J. A. A. (2004). MrModeltest v2. Program distributed by the author. Uppsala: Uppsala University, Evolutionary Biology Centre.
Page, R. D. M. (1996). TreeView: an application to display phylogenetic trees on personal computers. Computer Applications in the Biosciences, 12, 357–358.
Rehner, S. A. (2001). Primers for elongation factor 1-alpha (EF1-alpha). http://www.aftol.org/pdfs/EF1primer.pdf. Accessed 15 Feb 2015.
Rojas, E. I., Rehner, S. A., Samuels, G. J., Van Bael, S. A., Herre, E. A., Cannon, P., Chen, R., Pang, J., Wang, R., Zhang, I., Peng, I. Q., & Sha, T. (2010). Colletotrichum gloeosporioides s.l. associated with Theobroma cacao and other plants in Panama: multilocus phylogenies distinguish host associated pathogens from asymptomatic endophytes. Mycologia, 102, 1318–1338.
Ronquist, F., & Huelsenbeck, J. P. (2003). MrBayes3: Bayesian phylogenetic inference under mixed models. Bioinformatics, 19, 1572–1574.
Sharma, G., Pinnaka, A. K., & Shenoy, N. D. (2014). Resolving the Colletotrichum siamense species complex using ApMat marker. Fungal Diversity, 71, 247–264.
Silva-Hughes, A. F., Wedge, D. E., Cantrell, C. L., Carvalho, C. R., Pan, Z., Moraes, R. M., Madoxx, V. L., & Rosa, L. H. (2015). Diversity and antifungal activity of the endophytic fungi associated with the native medicinal cactus Opuntia humifusa (Cactaceae) from the United States. Microbiological Research, 175, 67–77.
Souza, A. E. F., Nascimento, L. C., Araújo, E., Lopes, E. B., & Souto, F. M. (2010). Ocorrência e identificação dos agentes etiológicos de doenças em palma forrageira (Opuntiaficus-indica Mill.) no semiárido paraibano. Biotemas, 23, 11–20.
Staden, R., Beal, K. F., & Bonfield, J. K. (1998). The Staden package, 1998. In S. Misener & A. S. Krawetz (Eds.), Bioinformatics methods and protocols (pp. 115–130). New York: Academic Press.
Sung, G. H., Sung, J. M., Hywel-Jones, N. L., & Spatafora, J. W. (2007). A multi-gene phylogeny of Clavicipitaceae (Ascomycota, Fungi): identification of localized incongruence using a combinational bootstrap approach. Molecular Phylogenetics and Evolution, 44, 1204–1223.
Suryanarayanan, T., Wittlinger, S. K., & Faeth, S. H. (2005). Endophytic fungi associated with cactiin Arizona. Mycological Research, 109, 635–639.
Swart, W. J., & Kriel, W. M. (2002). Pathogens associated with necrosis of cactus pear cladodes in South Africa. Plant Disease, 86, 693.
Tuite, J. (1969). Plant pathological methods - Fungi and bacteria. Minneapolis: Burgess Publishing.
Wang, M., Liu, F., Crous, P. W., & Cai, L. (2017). Phylogenetic reassessment of Nigrospora: ubiquitous endophytes, plant and human pathogens. Persoonia, 39, 118–142.
Weir, B. S., Johnston, P. R., & Damm, U. (2012). The Colletotrichum gloeosporioides species complex. Studies in Mycology, 73, 115–180.
White, T. J., Bruns, T., Lee, S., & Taylor, J. (1990). Amplification and direct sequencing of fungal ribosomal RNA genes for phylogenetics. In M. A. Innis, D. H. Gelfand, J. J. Sninsky, & T. J. White (Eds.), PCR protocols: A guide to methods and applications (pp. 315–322). New York: Academic Press.
Woudenberg, J. H. C., Groenewald, J. Z., Binder, M., & Crous, P. W. (2013). Alternaria redefined. Studies in Mycology, 75, 171–212.
Acknowledgments
This study was financed by Instituto Nacional de Tecnología Agropecuária (INTA, Argentina) and by Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES, Brasil). The first author was partially supported by a doctor scholarship of the INTA. M. P. S. Câmara and S. J. Michereff also acknowledge the Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq, Brasil) research fellowship.
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Conforto, C., Lima, N.B., Silva, F.J.A. et al. Characterization of fungal species associated with cladode brown spot on Nopalea cochenillifera in Brazil. Eur J Plant Pathol 155, 1179–1194 (2019). https://doi.org/10.1007/s10658-019-01847-3
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DOI: https://doi.org/10.1007/s10658-019-01847-3