Sains Malaysiana 43(5)(2014): 697–705

In vitro and In vivo Evaluation of Streptomyces Suppressions against Anthracnose in

Chili Caused by Colletotrichum

(Penilaian Penindasan Streptomyces Secara In vitro dan In vivo terhadap Antraknos dalam Cili Disebabkan oleh Colletotrichum)

PEDRAM SHAHBAZI*, MD. YUSOFF MUSA, GEOK YUAN ANNIE TAN, FARHAT AHMADI AVIN, WEE FEI AARON TEO & VIKINESWARY SABARATNAM

Institute of Biological Sciences, Faculty of Science, University of Malaya, 50603 Kuala Lumpur

Malaysia

Diserahkan: 8 Januari 2013/Diterima: 4 September 2013

ABSTRACT

The isolation of 66 streptomycetes from rhizosphere soil of chili plants was done for their inhibitory activities against three different dominant species of Colletotrichum namely C. acutatum, C. gloeosporioides and C. capsici. Twenty one streptomycetes strains were active against at least one of the Colletotrichum species. In addition, ten strains that inhibited the in vitro growth of Colletotrichum species showed chitinase activity. Strain P42, which displayed the highest inhibitory activity against all three anthracnose fungi species and high chitinase activity was tested as biological control agent in a greenhouse study. The strain successfully controlled chili anthracnose disease by significantly reducing the disease severity. Phylogenetic analysis of the 16S rRNA gene sequences showed that strain P42 belongs to the Streptomyces rochei clade. The results of the current study showed that rhizosphere-derived soil of chili plants is an important source of bioactive streptomycetes which are antagonistic against Colletotrichum.

Keywords: Biological control; chili pepper; Colletotrichum; inhibitory activity; Streptomyces

ABSTRAK

Enam puluh enam pencilan streptomiset daripada tanah rizosfera tumbuhan cili telah dilakukan saringan aktiviti antimikrob terhadap tiga spesies kulat Colletotrichum yang berbeza iaitu C. acutatum, C. gloeosporioides dan C. capsici. Dua puluh satu strain streptomiset didapati aktif terhadap sekurang-kurangnya salah satu spesies Colletotrichum. Tambahan pula, sepuluh terikan streptomiset yang mampu merencat pertumbuhan Colletotrichum secara in vitro juga menunjukkan aktiviti kitinase. Strain P42 yang memaparkan aktiviti perencatan tertinggi terhadap ketiga-tiga spesies kulat kajian dan juga mempunyai aktiviti kitinase tinggi diuji sebagai agen kawalan biologi dalam kajian rumah hijau. Strain P42 ini berjaya mengawal penyakit antraknos cili. Analisis filogenetik gen jujukan 16S rRNA menunjukkan bahawa strain P42 kepunyaan klad Streptomyces rochei. Keputusan kajian ini memberikan gambaran bahawa tanah rizosfera tumbuhan cili adalah sumber penting streptomiset bioaktif yang berupaya mengawal pertumbuhan Colletotrichum.

Kata kunci: Cili padi; Colletotrichum; kawalan biologi; saringan aktiviti; Streptomyces

RUJUKAN

Bell, A.A. & Mace, M.E. 1981. Biochemistry and physiology of resistance. In Fungal Wilt Diseases of Plants, edited by Mace, M. New York: Academic Press.

Berdy, J. 2005. Bioactive microbial metabolites. The Journal of Antibiotics 58(1): 1-26.

Cho, S.J., Lee, S.K., Cha, B.J., Kim, Y.H. & Shin, K.S. 2003. Detection and characterization of the Gloeosporium gloeosporioides growth inhibitory compound iturin A from Bacillus subtilis strain KS03. FEMS Microbiology Letters 223(1): 47-51.

Cook, R.J. & Baker, K.F. 1983. The Nature and Practice of Biological Control of Plant Pathogens. USA: American Phytopathological Society.

Crawford, D.L., Lynch, J.M., Whipps, J.M. & Ousley, M.A. 1993. Isolation and characterization of actinomycete antagonists of a fungal root pathogen. Applied and Environmental Microbiology 59(11): 3899-3905.

De Boer, W., Klein Gunnewiek, P.J.A., Lafeber, P., Janse, J.D., Spit, B.E. & Woldendorp, J.W. 1998. Anti-fungal properties of chitinolytic dune soil bacteria. Soil Biology and Biochemistry 30(2): 193-203.

De, S. & Chandra, A.L. 1983. A-19, a new antifungal antibiotic. Indian Journal of Experimental Biology 21: 31-33.

Demain, A.L. & Fang, A. 1995. Emerging concepts of secondary metabolism in actinomycetes. Actinomycetologica 9(2): 98-117.

El-Tarabily, K., Soliman, M., Nassar, A., Al-Hassani, H., Sivasithamparam, K., McKenna, F. & Hardy, G.E.S.T. 2000. Biological control of Sclerotinia minor using a chitinolytic bacterium and actinomycetes. Plant Pathology 49(5): 573- 583.

Ezziyyani, M., Requena, M.E., Egea-Gilabert, C. & Candela, M.E. 2007. Biological control of Phytophthora root rot of pepper using Trichoderma harzianum and Streptomyces rochei in combination. Journal of phytopathology 155(6): 342-349.

Freeman, S., Katan, T. & Shabi, E. 1998. Characterization of Colletotrichum species responsible for anthracnose diseases of various fruits. Plant Disease 82(6): 596-605.

Getha, K. & Vikineswary, S. 2002. Antagonistic effects of Streptomyces violaceusniger strain G10 on Fusarium oxysporum f. cubense race 4: Indirect evidence for the role of antibiosis in the antagonistic process. Journal of Industrial Microbiology & Biotechnology 28(6): 303-310.

Gupta, R., Saxena, R., Chaturvedi, P. & Virdi, J. 1995. Chitinase production by Streptomyces viridificans: Its potential in fungal cell wall lysis. Journal of Applied Microbiology 78(4): 378-383.

Hermosa, M., Grondona, I., Iturriaga, E.A., Diaz-Minguez, J., Castro, C., Monte, E. & Garcia-Acha, I. 2000. Molecular characterization and identification of biocontrol isolates of Trichoderma spp. Applied and Environmental Microbiology 66(5): 1890-1898.

Hsu, S. & Lockwood, J. 1975. Powdered chitin agar as a selective medium for enumeration of actinomycetes in water and soil. Applied and Environmental Microbiology 29(3): 422.

Ilic, S., Konstantinovic, S., Todorovic, Z., Lazic, M., Veljkovic, V., Jokovic, N. & Radovanovic, B. 2007. Characterization and antimicrobial activity of the bioactive metabolites in streptomycete isolates. Microbiology 76(4): 421-428.

Ishikawa, J., Tsuchizaki, N., Yoshida, M., Ishiyama, D. & Hotta, K. 2000. Colony PCR for detection of specific DNA sequences in actinomycetes. Actinomycetologica 14(1): 1-5.

Jetiyanon, K. & Kloepper, J.W. 2002. Mixtures of plant growth-promoting rhizobacteria for induction of systemic resistance against multiple plant diseases. Biological Control 24(3): 285-291.

Jiang, C.L. & Xu, L.H. 1996. Diversity of aquatic actinomycetes in lakes of the middle plateau, Yunnan, China. Applied and Environmental Microbiology 62(1): 249-253.

Jones, C.R. & Samac, D.A. 1996. Biological control of fungi causing alfalfa seedling damping-off with a disease-suppressive strain of Streptomyces. Biological Control 7(2): 196-204.

Jukes, T. & Cantor, C. 1969. Evolution of protein molecules. In Mammalian Protein Metabolism, edited by Munro, H. New York: Academic Press.

Kim, O.S., Cho, Y.J., Lee, K., Yoon, S.H., Kim, M., Na, H., Park, S.C., Jeon, Y.S., Lee, J.H. & Yi, H. 2012. Introducing EzTaxon-e: A prokaryotic 16S rRNA gene sequence database with phylotypes that represent uncultured species. International Journal of Systematic and Evolutionary Microbiology 62(3): 716-721.

Kim, S.B., Falconer, C., Williams, E. & Goodfellow, M. 1998. Streptomyces thermocarboxydovorans nov. and Streptomyces thermocarboxydus nov., two moderately thermophilic carboxydotrophic species from soil. International Journal of Systematic Bacteriology 1(4): 59-68.

Labeda, D., Goodfellow, M., Brown, R., Ward, A., Lanoot, B., Vanncanneyt, M., Swings, J., Kim, S.B., Liu, Z. & Chun, J. 2012. Phylogenetic study of the species within the family Streptomycetaceae. Antonie van Leeuwenhoek 101(1): 73-104.

Labeda, D. & Shearer, M. 1990. Isolation of actinomycetes for biotechnological applications. In Isolation of Biotechnology Organisms from Nature, edited by Labeda, D., New York: McGraw Hill.

Lane, D.J. 1991. 16S/23S rRNA sequencing. In Nucleic Acid Techniques in Bacterial Systematics, edited by Stackebrandt,  E. & Goodfellow, M. Chichester, United Kingdom: John Wiley & Sons.

Links, J., Rombouts, J. & Keulen, P. 1957. The ‘Bulging Factor’, a fungistatic antibiotic produced by a Streptomyces strain, with evidence of an active water-excreting mechanism in fungi. Journal of General Microbiology 17(3): 596-601.

Locci, R. 1989. Streptomycetes and related genera. In Bergey’s Manual of Systematic Bacteriology, edited by Williams, S.T., Sharpe, M.E., Holt, J.G., Murray, R.G.E., Brenner, D.J., Krieg, N.R., Moulder, J.W., Pfennig, N., Sneath, P.H.A. & Staley, J.T., Baltimore: Williams and Wilkins.

Marois, J. 1990. Biological control of diseases caused by Fusarium oxysporum. In Fusarium Wilt of Banana, edited by Ploetz, R. California: APS Press.

Pakdeevaraporn, P., Wasee, S., Taylor, P. & Mongkolporn, O. 2005. Inheritance of resistance to anthracnose caused by Colletotrichum capsici in Capsicum. Plant Breeding 124(2): 206-208.

Peres, N.A.R., Kuramae, E.E., Dias, M.S.C. & De Souza, N.L. 2002. Identification and characterization of Colletotrichum spp. affecting fruit after harvest in Brazil. Journal of Phytopathology 150(3): 128-134.

Saadoun, I., Hameed, K., Al-Momani, F., Malkawi, H., Meqdam, M. & Mohammad, M. 2000. Characterization and analysis of antifungal activity of soil streptomycetes isolated from North Jordan. Egyptian Journal of Microbiology 35(4): 463-471.

Saitou, N. & Nei, M. 1987. The neighbor-joining method: A new method for reconstructing phylogenetic trees. Molecular Biology and Evolution 4(4): 406-425.

Satomi, T., Kusakabe, H., Nakamura, G., Nishio, T., Uramoto, M. & Isono, K. 1982. Neopeptins A and B, new antifungal antibiotics. Agricultural and Biological Chemistry 46(10): 2621-2623.

Shirling, E.B. & Gottlieb, D. 1966. Methods for characterization of Streptomyces species. International Journal of Systematic Bacteriology 16(3): 313-340.

Skujins, J., Potgieter, H. & Alexander, M. 1965. Dissolution of fungal cell walls by a streptomycete chitinase and β-(1→ 3) glucanase. Archives of Biochemistry and Biophysics 111(2): 358-364.

Taddei, A., Rodriguez, M.J., Marquez-Vilchez, E. & Castelli, C. 2006. Isolation and identification of Streptomyces spp. from Venezuelan soils: Morphological and biochemical studies. Microbiological Research 161(3): 222-231.

Tamura, K., Dudley, J., Nei, M. & Kumar, S. 2007. MEGA4: Molecular evolutionary genetics analysis (MEGA) software version 4.0. Molecular Biology and Evolution 24(8): 1596- 1599.

Than, P.P., Prihastuti, H., Phoulivong, S., Taylor, P.W.J. & Hyde, K.D. 2008. Chilli anthracnose disease caused by Colletotrichum species. Journal of Zhejiang University- Science B 9(10): 764-778.

Vikineswary, S., Nadaraj, P., Wong, W. & Balabaskaran, S. 1997. Actinomycetes from a tropical mangrove ecosystem- Antifungal activity of selected strains. Asia-Pacific Journal of Molecular Biology and Biotechnology 5(2): 81-86.

Wang, Y., Zhang, Z., Ruan, J., Wang, Y. & Ali, S. 1999. Investigation of actinomycete diversity in the tropical rainforests of Singapore. Journal of Industrial Microbiology & Biotechnology 23(3): 178-187.

Xu, L., Li, Q. & Jiang, C. 1996. Diversity of soil actinomycetes in Yunnan, China. Applied and Environmental Microbiology 62(1): 244-248.

Yakoby, N., Zhou, R., Kobiler, I., Dinoor, A. & Prusky, D. 2001. Development of Colletotrichum gloeosporioides restriction enzyme-mediated integration mutants as biocontrol agents against anthracnose disease in avocado fruits. Phytopathology 91(2): 143-148.

Yeo, W.H., Kim, S., Kim, S.S., Yu, S. & Park, E. 1994. Isolation, physico-chemical properties, and biological activity of new thiopeptide antibiotics, kimorexins. Journal of Microbiology and Biotechnology 4: 349-353.

Yoshida, S., Shirata, A. & Hiradate, S. 2002. Ecological characteristics and biological control of mulberry anthracnose. Japan Agricultural Research Quarterly 36(2): 89-96.

Yun, H.K., Ahmad, A.H., Muid, S. & Seelan, J.S.S. 2009. First report of Colletotrichum spp. causing diseases on Capsicum spp. in Sabah, Borneo, Malaysia. Journal of Threatened Taxa 1(8): 419-424.

*Pengarang untuk surat-menyurat; email: ped.shahbazi@gmail.com