Microsoft Word - 10. 57 BJPT 16 - 57 -editKA.doc Bangladesh J. Plant Taxon. 23(2): 181-188, 2016 (December) © 2016 Bangladesh Association of Plant Taxonomists MYCOFLORA ASSOCIATED WITH INFECTED FRUITS OF MOMORDICA COCHINCHINENSIS (LOUR.) SPRENG. SHAMIM SHAMSI1, SAROWAR HOSEN, MD. AL-MAMUN AND MOMTAZ BEGUM Department of Botany, University of Dhaka, Dhaka-1000, Bangladesh. Keywords: Mycoflora; Fruit rot; Anthracnose; Momordica cochinchinensis. Abstract Eleven species of anamorphic fungi associated with anthracnose and fruit rot symptoms of Momordica cochinchinensis (Lour.) Spreng. (Ban Kakrol) have been described. The associated fungi were Aspergillus niger Van Tieghem, Colletotrichum gloeosporioides (Penz.) Sacc., C. orbiculare (Berk. & Mont.) Arx., Corynespora cassiicola (Berk. & Curt.) Wei, Curvularia clavata Jain, Dendryphiella vinosa (Berk. & Curt.) Reisinger, Fusarium moniliforme J. Sheld., Lasidiodiplodia theobromae (Pat.) Griff. & Maubl., Pestalotiopsis guepinii (Desm.) Stey., Penicillium digitatum Sacc. and Xylohypha pinicola D. Hawksw. Xylohypha pinicola is a new record for Bangladesh. Introduction Momordica cochinchinensis (Lour.) Spreng. (Ban Kakrol) belongs to the family Cucurbi- taceae (Ahmed et al., 2008). Its roots, leaves and fruits are important for its nutritional and medicinal properties. The plant is native to South East Asia. In Bangladesh, it grows mainly in the Chittagong Hill Tracts in wild condition. It is known as Ban Kakrol in Bangladesh, Gac in Vietnam and variously red melon, babyjackfruit, spiny bitter gourd or cochinchin gourd in English. Momordica cochinchinensis fruit is traditionally used in Asia to provide red colour for cuisines and enhance visional health. Recently, M. cochinchinensis fruit has emerged as a potential source of carotenoids, especially lycopene and β-carotene. Carotenoids and other identified bioactives from this fruit including phenolics, flavonoids and trypsin inhibitors are associated with many beneficial bioactivities such as antioxidant, anticancer and provitamin A activities. In addition to the traditional utilization, commercial products like M. cochinchinensis powder and oil is manufactured as natural colourants and medicinal supplements (Chuyenet al., 2015). In Vietnam, gac is prized by natives for promoting longevity and vitality. In a supplementation trial among Vietnamese children, gac increased serum vitamin A levels more than synthetic beta- carotene (Burke et al., 2005). From the above discussion it is clear that diseases free M. cochinchinensis fruits are carrying need for nutritional and medicinal point of view. From India Mukerji and Bhasin (1986) reported eight fungal diseases of two species of Momordica L. viz., M. charantia L. and M. dioica Roxb. ex Willd. They exclusively reported only fruit rot disease on M. cochinchinensis species. So far there is no report is available regarding fungal diseases of Momordica spp. in Bangladesh. Recently fruit rot and anthracnose symptoms showing severe damage of M. cochinchinensis fruits was recorded on the plants grown in Botanical garden, Curzon Hall, Dhaka University. Present investigation was conducted to find out the fungi associated with infected fruits of M. cochinchinensis. 1Corresponding author. Email: prof.shamsi@gmail.com 182 SHAMSI et al. Materials and Methods Infected fruits of Momordica cochinchinensis (Lour.) Spreng. having characteristic symptoms were collected from selected fields of the Botanical Garden of Dhaka University to record the prevalence of the diseases and association of fungi during the tenure of December 2015 to January 2016. Samples were collected in separate sterile polyethylene bags, labeled properly and then brought to the laboratory for isolating associated pathogenic fungi following “Tissue planting method” on PDA medium. From infected fruits of Ban Kakrol showing anthracnose and soft rot symptoms, each measuring 2 mm2 sized were cut separately with a pair of sterilize scissors and kept in a separate sterilized Petri plate. The inocula were washed with sterile water and then surface sterilized by dipping in 10% chlorox solution for three minutes. The inocula were again washed with sterile water. For fruit rot symptoms a total of 100 inocula were placed separately on 30 sterilized Petri plates containing 15 ml of PDA medium with an addition of 1 drop (ca 0.03 ml) of lactic acid to check the bacterial growth and incubated in an incubator (25 ± 2˚C) for 7 days. Detail morphological studies of the fungal isolates were made in order to determine their identification. The microscopic structural characters of the isolated fungi were recorded under a digital camera. Species identification was done by Camera Lucida drawing. All specimens, included in the present study were preserved in Mycology and Plant Pathology section and Salar Khan Herbarium, Department of Botany, University of Dhaka, Bangladesh. Identities of the isolates were determined following the standard literature (Booth, 1971; Ellis, 1971; Barnet and Hunter, 1972; Sutton, 1980). Results and Discussion A total of eleven species of fungi were associated with two types of symptoms viz., anthracnose and fruit rot were recorded on infected fruits of Momordica cochinchinensis (Lour.) Spreng. (Fig. 1). The fungi associated with anthracnose symptoms were Colletotrichum gloeosporioides, C. orbiculare, Curvularia clavata, Fusarium moniliforme and Penicillium digitatum. Fruit rot infected samples showed the association of Aspergillus niger, Colletotrichum gloeosporioides, Corynespora cassiicola, Curvularia clavata, Dendryphiella vinosa, Fusarium moniliforme, Lasidiodiplodia theobromae, Pestalotiopsis guepinii and Xylohypha pinicola. Of these X. pinicola is a new record for Bangladesh. Taxonomic treatment of fungal taxa 1. Aspergillus niger Van Tieghem, Ann. Sci. Nat. Bot., Ser. 5, 8: 240 (1867). (Fig. 2A). Colonies effuse, black. Mycelium well-developed, septate, profusely branched and brownish. Cells are multinucleate. Conidiophores brown 200−400 × 7−10 µm. Vesicles globose or subglobose, thick walled, commonly 20−50 µm, occasionally up to 100 µm in diameter. Sterigmata 20−30 × 6−8 µm. Conidia dark brown, one celled globose, spinose 2−4 (5) µm in diameter. Cattenulate. Specimen examined: Isolated from infected fruits of M. cochinchinensis, Botanical Garden, University of Dhaka, Dhaka, S. Hosen 12, 6 December 2015. 2. Colletotrichum gloeosporioides (Penz.) Sacc., Fung. Agrum. 2: 6 (1882). (Fig. 2B). Colonies greyish. Acervuli black, sub-epidermal but later the epidermis are ruptured and expose them. Hyphae septate, hyaline, both inter and intra-cellular. Conidia hyaline, straight, obtuse at the apex, 11.2−25.2 3.6−5.0 µm. Specimen examined: Isolated from infected fruits of M. cochinchinensis, Botanical Garden, University of Dhaka, Dhaka, S. Shamsi 3077, 17 December 2015. MYCOFLORA ASSOCIATED WITH MOMORDICA COCHINCHINENSIS 183 Fig. 1. Momordica cochinchinensis. A. Healthy fruit, B. Rotten fruit, C−F. Fruit showing different stages of anthracnose symptom. 184 SHAMSI et al. 3. Colletotrichum orbiculare (Berk. & Mont.) Arx.,Verh. Akad. Wet. Amst. 51(3): 112(1957). (Fig. 2C, D). Colonies with abundant white greyish aerial mycelium, reverse dark brown, conidial masses salmon pink. Setae present, conidia straight, cylindrical obtuse at the apices, 11.7−14.4 × 4.5−6 µm. Specimen examined: Isolated from infected fruits of M. cochinchinensis, Botanical Garden, University of Dhaka, Dhaka, S. Shamsi 3078, 1January 2016. 4. Corynespora cassiicola (Berk. & Curt.) Wei, Mycol. Pap. 34: 5 (1950). (Fig. 2E). Colonies effuse, grey, thinly hairy. Mycelium mostly immersed; no stroma. Conidiophores pale to mid brown, with up to 9 successive cylindrical proliferations. Conidia solitary or in chains of 2−6, very variable in shape. Obclavate to cylindrical, straight or curved, subhyaline to rather pale olivaceous brown or brown, smooth, with 4−20 pseudosepta, 48.6−131.4 ×12−23 µm. Specimen examined: Isolated from infected fruits of M. cochinchinensis, Botanical Garden, University of Dhaka, Dhaka, S. Hosen 13, 6 December 2015. 5. Curvularia clavata Jain, Trans. Brit. Mycol. Soc. 45(4): 542 (1962). (Fig. 2F). Colonies effuse, brown, grey or black, hairy, cottony or velvety. Mycelium brown, septate, branched. Conidiophores dark brown, paler towards the apex, septate, mostly geniculate. Conidia solitary, simple, often curved, clavate, ellipsoidal, broadly fusiform with 3 septate, dark brown, end cells paler than the others, smooth, 16.5−25 × 7−11 µm. Specimen examined: Isolated from infected fruits of M. cochinchinensis, Botanical Garden, University of Dhaka, Dhaka, S. Hosen 14, 17 December 2015. 6. Dendryphiella vinosa (Berk. & Curt) Reisinger, Bull.trimest. Soc. Mycol. Fr. 84: 27−39 (1968). (Fig. 3A). Colonies effuse, rust-colored, brown or black, hairy or velvety. Mycelium mostly immersed. Stroma none. Setae and hyphopodia absent. Conidiophores reddish brown, verruculose. Conidia 3septate, distinctly verruculose when mature, pale clear brown to burnt sienna, darker at the hilum, 16.2−27 ×3.6−7.2µm. Specimen examined: Isolated from infected fruits of M. cochinchinensis, Botanical Garden, University of Dhaka, Dhaka, S. Hosen 15, 6 December 2015. 7. Fusarium moniliforme J. Sheld., Annual Rep. Neb. Agric. Exp. Sta. 17: 23−32 (1904). (Fig. 3B). Colony white, cottony, reverse violet in PDA medium. Microconidia are formed in chains under optimum growing conditions and these can readily be observed in situ under the low power of the microscope. Microconidia measure 8−11 ×1.8−2.25 µm and are fusiform to clavate with a slightly flattened base; they occasionally become 1 septate. Macroconidial formation is rare in many strains, where present they develop from conidiophores formed as lateral branches on the hyphae. The conidiophore consists of a single basal cell bearing 2−3 apical phialides or it may form 2−3 metulae which in turn bear simple doliform to obclavate phialides, 17−23.4 ×1.8−2.7µm. Specimen examined: Isolated from infected fruits of M. cochinchinensis, Botanical Garden, University of Dhaka, Dhaka, S. Hosen 16, 6 December 2015. MYCOFLORA ASSOCIATED WITH MOMORDICA COCHINCHINENSIS 185 Fig. 2. A. Aspergillus niger, B. Colletotrichum gloeosporioides, C−D. C. orbiculare, E. Corynespora cassiicola and F. Curvularia clavata. (Bar = 50 µm). 186 SHAMSI et al. Fig. 3. A. Dendryphiella vinosa, B. Fusarium moniliforme, C. Lasidiodiplodia theobromae, D. Penicillium digitatum, E. Pestalotiopsis guepinii, F. Xylohypha pinicola. (Bar = 50 µm). MYCOFLORA ASSOCIATED WITH MOMORDICA COCHINCHINENSIS 187 8. Lasidiodiplodia theobromae (Pat.) Griff. & Maubl, Bull. Trimest. Soc. Mycol. Fr. 8:136 (1892). (Fig. 3C). Colonies greyish brown, cottony, reverse brownish black. Hyphae septate, branched, dark chocolate brown. Pycnidia globose, dark brown, ostiolate. Conidiophore short, hyaline. Conidia dark brown, two-celled, ellipsoidal, 16−22 × 8−12 µm. Specimen examined: Isolated from infected fruits of M. cochinchinensis, Botanical Garden, University of Dhaka, Dhaka, S. Hosen 17, 1 January 2016. 9. Penicillium digitatum Sacc., Bur. Anim. Ind., Bul. 118: 31−33 (1910). (Fig. 3D). Colony small, cottony, greenish, reverse creamy. Hyphae septate, branched, hyaline. Conidiophores hyaline, septate. Sterigmata equally variable, 15−28 µm long and 3.5−5.0 µm width. Conidia elliptical to subglobose, smooth aseptate with greenish, tinge commonly 3.5−5.0 µm and occasionally up to 10−12 µm in diameter. Cattenulate. Specimen examined: Isolated from infected fruits of M. cochinchinensis, Botanical Garden, University of Dhaka, Dhaka, S. Hosen 18, 17 December 2015. 10. Pestalotiopsis guepinii (Desm.) Stey., Bull. Jard. Bot. État Brux. 19(3): 312 (1949). (Fig. 3E). Colonies white, cottony, reverse white. Hyphae septate, branched, hyaline. Acervuli black, small, shining. Conidiophores septate, branched, dark brown, cylindrical or lageniform, formed from the upper cells of the pseudoparenchymata. Conidia fusiform, straight or slightly curved, mostly 3 euseptate: basal cells hyaline, truncate, with an endogenous, cellular, appendage: apical cell conic, hyaline, with 2 or more apica, simple or branched, spathulate or espathulate appendages: median cells brown, sometimes versicoloured, thicker-walled, smooth, 14−23 × 5−7.5 µm. Specimen examined: Isolated from infected fruits of M. cochinchinensis, Botanical Garden, University of Dhaka, Dhaka, S. Hosen 19, 6 December 2015. 11. Xylohypha pinicola D. Hawksw., Trans. Brit. Mycol. Soc. 64: 89−99 (1975). (Fig. 3F). Colonies effuse, dark brown to black. Conidiophores septate, simple or branched, straight or flexuous, mid to dark brown, thick-walled, smooth, up to 160 µm long, 4−7 µm thick. Conidia in simple or branched acropetal chains ellipsoidal, pale to mid brown, smooth, 4−10 × 3.5−5 µm. Specimen examined: Isolated from infected fruits of M. cochinchinenesis, Botanical Garden, University of Dhaka, Dhaka, S. Shamsi 3079, 6 December 2015. Acknowledgements The authors wish to thank Professor Dr. Mohammad Zashim Uddin, Department of Botany, University of Dhaka for collecting Momordica cochinchinensis plants from Fasiakhali union, Chakaria Upazilla, Cox’s Bazar district and rearing it in the botanical garden of Dhaka University. References Ahmed, Z.U., Hassan, M.A., Begum, Z.N.T., Khondker, M., Kabir, S.M.H., Ahmed, M., Ahmed, A.T.A., Rahman, A.K.A. and Haque, E.U. (Eds.) 2008. Encyclopedia of Flora and Fauna of Bangladesh, Vol. 7. Angiosperms: Dicotyledons (Balsaminaceae−Euphorbiaceae). Bangladesh Asiat. Soc., Dhaka, pp. 1−546. Barnett, H.L. and Hunter, B.B. 1972. Illustrated Genera of Imperfect Fungi. Burgess Pub. Co. U. S. A. pp. III +241. 188 SHAMSI et al. Booth, C. 1971. The genus Fusarium. The Commonwealth Mycological Institute, Kew, Surrey, England, pp. 237. Burke, D.S., Smidt, C.R. and Vuong, L.T. 2005. Momordica cochinchinensis, Rosa roxburghii, Wolfberry, and Sea Buckthorn—highly nutritional fruits supported by tradition and science . Current Top. Nutraceut. Res. 3(4): 259−266. Chuyen, H.V., Nguyen, M.H., Roach, P.D., Golding, J.B. and Parks, S.E. 2015. Gac fruit (Momordica cochinchinensis Spreng.): a rich source of bioactive compounds and its potential health benefits. Int. J. Food Sci. & Technol. 50: 567−577. Ellis, M.B. 1971. Dematiaceous Hyphomycetes. The Commonwealth Mycological Institute, England, pp. 608. Mukerji, K.G. and Bhasin, J. 1986. Plant diseases of India. A source Book. Tatta McGrew-Hill Publishing Company Ltd. New Delhi, pp. 468. Sutton, B.C. 1980. The Coelomycetes. Fungi Imperfecti with Pycnidia, Acervuli and Stromata. Common- wealth Mycological Institute, England, pp. 696. (Manuscript received on 10 May 2016; revised on 23 August 2016)