Mycobiology 33(4): 235-239 (2005) Copyright © 2005 by The Korean Society of Mycology Notes on Cordyceps species Collected from the Central Region of Nepal 1 Bhushan Shrestha and Jae-Mo Sung * Entomopathogenic Fungal Culture Collection (EFCC), Department of Applied Biology, Kangwon National University, Chuncheon 200701, Korea (Received August 24, 2005) The present study was carried out to explore the Cordyceps species and other entomopathogenic fungal flora around Kathmandu Valley and a few high altitude locations of Nepal. In this paper, we report eight Cordyceps species as new to Nepal: C. gracilis, C. ishikariensis, C. liangshanensis, C. martialis, C. militaris, C. pruinosa, C. sphecocephala and C. tricentri. We also mention a few allied genera such as Beauveria, Hirsutella and Paecilomyces from Nepal. Further collections from different ecological regions of Nepal will show the richness of entomopathogenic fungal floral diversity of Nepal. KEYWORDS: Cordyceps, Entomopathogenic fungi, Kathmandu valley, Langtang area, Manang village Cordyceps species have been reported from many parts of the world. The genus Cordyceps belongs to the family 1939a, b, 1941; Kobayasi and Shimizu, 1983), Congo (Moureau, 1962), Norway (Eckblad, 1967), Ghana (Samson et al., 1982), Taiwan (Tzean et al., 1997), Amazonia (Evans and Samson, 1982, 1984; Samson and Evans, 1985), Thailand (Hywel-Jones 1994, 1995a, b, c, 1996; Hywel-Jones and Sivichai, 1995), Korea (Sung, 1996), China (Zang and Kinjo, 1998), and Mexico (Guzman et al., 2001). Apart from the work of Petch, very little work has been carried out on Cordyceps in South Asian region. To date, a few species of Cordyceps have been reported from Nepal. C. sinensis is one of them, which has been reported from high altitude areas (Balfour-Browne, 1955; Kobayasi, 1981; Adhikari and Durrieu, 1996). This species is the most popular species in Nepal due to its high medicinal values, thus regarded as a natural gift for humans as in other East Asian countries. Another Cordyceps species, C. nutans has been reported in Manang Region (Shrestha, 1985). The third Cordyceps species, C. nepalensis was reported as a new Cordyceps species from Mt. Kangchenjunga of Nepal (Zang and Kinjo, 1998). Recently, a glimpse of Cordyceps diversity of Nepal was presented (Shrestha and Sung, 2005). A Few Cordyceps exploration trips were carried out in Nepal during last several years to explore entomopathogenic fungal diversity of Nepal. Detailed taxonomic studies of Cordyceps species collected from Korea and other parts of the world, including Nepal, are undergoing at Entomopathogenic Fungal Culture Collection (EFCC), Kangwon National University, Korea (Shrestha et al., 2004; Sung et al., 2005). This paper intends to provide information on Cordyceps species collected from Nepal during last few years in order to highlight the entomopathogenic fungal diversity of Nepal (Table 1). Collection trips were made around Kathmandu Valley (Fig. 1) from 1997 to 2001 and in 2003. One trip was Clavicipitaceae in the order Hypocreales of Ascomycota. Cordyceps species mostly infect different stages of their host insects from larva to adult, kill them and eventually grow out of dead bodies of insects, except for a few species which grow on hypogeal Elaphomyces species. Very few Cordyceps species, including C. sinensis, have been reported from Nepal. However, scientific research on these species has been lacking although knowledge on their medicinal properties such as tonic and aphrodisiac values are common in the Nepalese society. Scientific studies on Cordyceps species started about 300 years ago, when Cordyceps militaris was described under the generic name, Clavaria, due to its Clavaria-like stromata (Linnaeus, 1753). Linnaeus followed the same generic name Clavaria and mentioned few Cordyceps species in his great work Species Plantarum (Linnaeus, 1753). Since then, it has attracted attention of great mycologists such as Persoon (1799), Fries (1823), Link (1833), Berkeley (1843), Tulasne Brothers (1865), Saccardo (1883) and Massee (1895) and was described under different generic names. Old literature proposed different generic names for Cordyceps such as Clavaria, Sphaeria and Torrubia, before Link (1833) finally erected Cordyceps as a new generic name. During last hundred years or more, regional exploration of Cordyceps species continued in many parts of the world such as Australia (Olliff, 1895; Willis, 1959), North America (Seaver, 1911; Mains, 1958), New Zealand (Dingley, 1953), Ceylon (present day Sri Lanka) (Petch, 1924), Great Britain (Petch, 1932, 1948), Japan (Kobayasi, *Corresponding author <E-mail: jmsung@kangwon.ac.kr> Present Address: Green Energy Mission/Nepal, Anam Nagar, Kathmandu P.O. Box 10647, Nepal 1 235 236 Shrestha and Sung Table 1. Cordyceps species collected from Nepal Cordyceps species C. gracilis C. C. C. C. C. C. ishikariensis liangshanensis martialis militaris nutans pruinosa C. sinensis C. sphecocephala C. tricentri Allied species Beauveria sp. Hirsutella sp. EFCC number EFCC 10528, 10529 Location Nagarkot Hill Collection date July 2003 Host Lepidopteran larva, usually Hepialus sp. EFCC 10407 Godawari Hill July 2003 Larva of cicadae EFCC 1520, 1521, 1522 Shivapuri Hill July 1997. Lepidopteran larva EFCC 3193, 3194 Shivapuri Hill July 1999 Coleopteran larva EFCC 7338, 7339 Nagarkot Hill July 2001 Lepidopteran pupae and larva EFCC 7206, 7207 Nagarjun Hill and Gokarna June 2001 Hemipteran bugs EFCC 10468, 10909 Shivapuri Hill July 2003 Limacodidae cocoon (Lepidoptera) EFCC 7282, 7283, 7284 Manang and Langtang Villages May 1999, May 2001 Larva of Hepialus sp. EFCC 7236, 7237, 7238 Nagarjun Hill June 2001 Bees and wasps EFCC 7251, 7252 Gokarna June 2001 Aadults of Vespidae (Homoptera) EFCC 7270 EFCC 3240, 3241 EFCC 7170, 7171 EFCC10570, 10571 Shivapuri Hill Shivapuri Hill Nagarjun Hill Shivapuri Hill July 2001 July 2001 June 2001 July 2003 1 Hemipteran bug Lepidopteran larva Larva of cicadae Lepidopteran larva Paecilomyces cicadae Paecilomyces sp. EFCC; Entomopathogenic Fungal Culture Collection, Kangwon National University, Chuncheon 200-701, Korea. 1 Fig. 1. Geographical map of Nepal. made to Langtang National Park in 2001, from where the specimens of were purchased from local collectors. specimens collected from local people of Manang Area were also observed. Specimens were airdried and have been preserved in Entomopathogenic Fungal Culture Collection (EFCC), Kangwon National UniverC. sinensis C. sinensis sity, Chuncheon, Korea. Morphological characters of species given by Mains (1958), Kobayasi (1941, 1982), Kobayasi and Shimizu (1983), Shimizu (1997), Tzean (1997) and Zang and Kinjo (1998) were referred to for the identification of species (Fig. 2). Cordyceps et al. Cordyceps Notes on Cordyceps species Collected from the Central Region of Nepal Fig. 2. 237 Cordyceps species collected from Nepal. a, C. gracilis; b, C. ishikariensis; c, C. liangshanensis; d, C. martialis; e, C. militaris; f, C. nutans; g, C. pruinosa; h, C. sinensis; i, C. sphecocephala; j, C. tricentri. Cordyceps gracilis Durieu & Montagne. Stroma consists of a stout cylindrical stipe, with a white to light brown, and ovoid to subglobose, brown-colored head. Stipe ranges from 30~49 mm in length and 1~2 mm in diameter. Head is 3~6 mm long and 3~4 mm wide. Stromata are produced in solitary per host, usually from head or thorax region. Both stalk and head are smooth in surface. Head is dotted with ostioles of perithecia. Perithecia are completely immersed in head and range in size from 500~600 × 150~200 µm. Perithecia are ovoid with a slightly long neck. Size of ascus is 325~350 × 5 µm. Size of ascus cap is 6~6.5 × 4~4.5 µm. Cordyceps ishikariensis Kobayasi & Shimizu. It produces five to six brown-yellow colored stromata per host. Stromata arise from the head region of the host. Stipe is 30~35 mm long, whereas head is 15~20 mm long. Stromata are 1.5~2 mm wide, stipe being more slender than head. Stipe is rough due to torn outer epidermal layer. Perithecia are semi-immersed and 500~570 × 240~300 µm in size. Perithecia are ovoid to broadly ovoid. Size of ascus is 250~360 × 4 µm. Size of ascus cap is 3~4 × 2 µm. Cordyceps liangshanensis M. Zang, D. Liu & R. Hu. Stromata are light to dark brown and are produced in solitary. Stromata arise from the head region of the host. Surface of stipe is longitudinally striate. Stipe ranges from 40~60 mm in length and 1.5~2 mm in diameter. Head is about 15 mm long and slightly wider than stipe. Perithecia are semi-immersed and 400~450 × 200~250 µm in size. Perithecia are ovoid to oval in shape. Size of ascus is 175~210 × 6~7 µm. Size of ascus cap is 4~5 × 3 µm. Cordyceps martialis Speg. It produces brownish orange stromata, one to a few per host. Stipe varies in length from 25 to 45 mm, while head ranges from 10 to 20 mm. Stipe and head are 0.5~1 and 1~2 mm wide, respectively. Surface of the head is rough due to presence of neck of immersed perithecia. Surface of stipe is sometimes covered by short spine-like structures. Upper parts of head are sometimes branched. Perithecia are ovoid or flaskshaped with long conical neck and almost immersed in head. They range in size from 500~530 × 150 µm. Size of ascus is 225~260 × 5~6 µm. Size of ascus cap is 6~7 × 4~5 µm. Cordyceps militaris. (L. ex Fr.) Link. Stromata are usually orange, club or clavate shaped. Stipe and head are 25~40 mm and 20~30 mm long, respectively. Stipe is 1~1.5 mm wide, head being slightly broader than stipe. Stromata are usually solitary, but sometimes a few per host. Surface of head is rough due to apices of perithecia. Stipe is sometimes slightly twisted and the surface is irregularly furrowed. Perithecia are semi-immersed in head and are 650~770 × 300~500 µm in size. Perithecia are broadly ovoid in shape. Size of ascus is 350~450 × 3~4 µm. Size of ascus cap is 3~4 × 3~3.5 µm. Size of partspore is 3~3.5 × 1 µm. Cordyceps nutans Patouillard. It produces black stipe with yellow to orange or red head. The uppermost part of stipe is usually similar to head in color. Stipe and head are 238 Shrestha and Sung 42~60 × 1 mm and 5~10 × 1~1.5 mm in size, respectively. The head is oval, obtuse or cylindrical and is erect. Surface of head is smooth and that of stipe is smooth, but longitudinally furrowed. Stipe is sometimes twisted. Stromata vary from solitary to a few per host. Perithecia are completely immersed and are obliquely vertical in head. They are 800~900 × 300 µm in size and elongated flask shaped. Size of ascus is 525~550 × 4~7 µm. Size of ascus cap is 10~12 × 6~8 µm. Size of part-spore is 7~8 × 1~1.4 µm. size. Perithecia are ovoid in shape. Ascus size is 300~320 × 5 µm. Ascus cap is 5~6 × 3~4 µm. The present study has explored Cordyceps species in Nepal. Additionally, this study has also explored allied species of Cordyceps, such as Beauveria, Hirsutella, Paecilomyces, etc. Among Paecilomyces species, P. cicadae (Miquel) Samson was found growing on larva of cicadae in Nagarjun Area of Kathmandu valley. It produces red colored stromata, single or few per host. Stipe is 10~15 × 1 mm in size and head is 5~10 mm long and 1.5 mm wide. The head is narrow-clavate. Surface of head is rough due to ostioles of semi-immersed perithecia. Stipe has irregular but short furrows on its surface. Perithecia are semiimmersed in head and are 350~500 × 240~290 µm in size. Perithecia are ovoid, with ostiole sometimes directed to one side. Size of ascus is 185~200 × 2 µm. Size of ascus cap is 3 × 1.5~2 µm. Part-spores are joined at both ends in a thin thread-like structure. The authors wish to acknowledge the financial support from Korea Science and Engineering Foundation (KOSEF) to Entomopathogenic Fungal Culture Collection (EFCC), Kangwon National University, Korea. Institute for Cordyceps Research of Kangwon National University is also thanked for providing research facilities to carry out this study. We also wishes acknowledge the permission for exploration of Cordyceps species in their respective areas from Department of Forests and Department of National Parks and Wildlife Conservation of the Government of Nepal. Cordyceps pruinosa Petch. Stromata are black with cylindrical stipe and slightly swollen head, usually with sterile apex. Stipe ranges from 40~45 mm in lengths and 1.5 mm wide. Head is slightly wider than stipe, usually 2 mm wide and 20~25 mm long. Perithecia are slightly projecting from the surface of the head. Surface of stipe is smooth with irregularly furrowed. Stipe is slightly twisted. Perithecia are 330~370 µm long and 170~270 µm wide. Perithecia are ovoid and have thick perithecial wall. Size of ascus is 200~210 × 7~8 µm. Cordyceps sinensis. (Berk.) Sacc. It produces pale-yellow to brownish-yellow slender stipe with ovoid to cylindrical yellow head. Stipe is 30~55 mm long and 1 mm wide. Head is 5~10 mm long and wider than stipe, about 1.5~2 mm wide. Each host produces single stroma. Stipe is smooth and longitudinally striate. Head is dotted with ostioles of perithecia on apex of ridges. Perithecia are immersed in head and are 620~780 × 200~230 µm in size. Perithecia are obliquely vertical in head, elongated flask-shaped or conoid in shape. Size of ascus is 350 × 6~7. Size of ascus cap is 8~9 × 5~6 µm. Size of partspore is 9~10.5 × 1~1.3 µm and is fusiform in shape with pointed ends. Cordyceps sphecocephala (Berk.) Sacc. Stromata are produced in solitary per host and are yellow in color. Stipe is 33~43 mm long and very slender, 0.25~0.3 mm wide. 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