Variable induction of cuticle-degrading enzymes of Beauveria bassiana isolates in the presence of different insect cuticles

Volume 9, Issue 4
December 2020
Pages 563-576

Document Type : Original Research

Authors

1 Department of Plant Protection, Faculty of Agriculture, Urmia University, Urmia, Iran.

2 Department of Plant Protection, College of Agriculture and Natural Resources, University of Tehran, Karaj, Iran.

Abstract
In the present study, the cuticle-degrading enzymes production potential of five native Beauveria bassiana (TV, OZ, UN, DV and DE) isolates was investigated in the presence of cuticles from Eurygaster integriceps, Ephestia kuehniella and Zophobas morio. Furthermore, histopathology of infected insects by B. bassiana was studied. The level of cuticle degrading enzymes was the highest and lowest for TV (as the most virulent isolate) and DE (as the weakest isolate), respectively. E. integriceps nymphs as the most sensitive host produced the highest level of cuticle degrading enzymes (Pr2, exochitinase, and lipase) while Z. morio as the most resistant host, produced the lowest level of hydrolytic enzymes. According to histopathological study, the fungal isolate could not penetrate into Z. morio cuticle, as no mycelia or hyphae were observed in its tissues after inoculation, while fungal bodies were detected in microscopic slides of the other two insects. Overall, the chemical and topographical structure of insect cuticle had a substantial effect on the virulence of entomopathogenic fungus. Production of enzymes including proteases (especially Pr2), chitinase (N-acetyl-glucosaminidase), and lipases was positively related to virulence of fungus isolates. It can be concluded that not only the hydrolytic activity of B. bassiana isolates, but also host cuticle composition determine the pathogenesis and virulence cascade in fungus-insect interactions.

Keywords

Altre, J. A., Vandenberg, J. D. and Cantone, F. A. 1999. Pathogenicity of Paecilomyces fumosoroseus isolates to diamondback moth, Plutella xylostella: correllation with spore size, germination speed, and attachment to cuticle. Journal of Invertebrate Pathology, 73: 332–338.
Andersen, S. O., Hojrup, P. and Roepstorff, P. 1995. Insect cuticular proteins. Insect Biochemistry and Molecular Biology, 25: 153–176.
Barra, P., Etcheverry, M. and Nesci, A. 2015. Improvement of the insecticidal capacity of two
Purpureocillium Lilacinum strains against Tribolium Confusum. Insects, 6: 206-223.
Boldo, J. T., Junges, A., Amaral, K. B., Staats, C., Vainstein, M. H. and Schrank, A. 2009. Endochitinase CHI2 of the biocontrol fungus Metarhizium anisopliae affects its virulence toward the cotton stainer bug Dysdercus peruvianus. Current Genetics, 55: 551-560.
Boucias, D. G. and Pendland, J. C. 1991. Attachment of mycopathogens to cuticle. The initial event of mycoses in arthropod hosts. In: Cole, G. T. and Hoch, H.C. (Eds.) The fungal Spore and Disease Initiation in Plants and Animals. Plenum Press, New York, pp. 101-127.
Bradford, M. M. 1976. Rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Analytical Biochemistry, 72: 248–254.
Charnley, A. K. 2003. Fungal pathogens of insects: cuticle degrading enzymes and toxins. Advances in Botanical Research, 40: 241-321.
Chui-Chai, N., Krutmuang, P., Nalumpang, S., Mekchay, S., Khanongnuch, C. and Chanbang, Y. 2012. Insecticidal activity and cuticle degrading enzymes of entomopathogenic fungi against Plutella xylostella (Lepidoptera: Plutellidae). Journal of Nature and Science, 11(1): 147-155.
Clarkson, J. M. and Charnely, A. K. 1996. New insights into mechanisms of fungal pathogenesis in insects. Trends in Microbiology, 4: 197–204.
Crespo, R., Juárez, M. P., dal Bello, G. M., Padín, S., Calderón Fernández, G. and Pedrini, N. 2002. Increased mortality of Acanthoscelides obtectus by alkane-grown Beauveria bassiana. BioControl, 47: 685–696.
Dhar, P. and Kaur, G. 2010. Cuticle-degrading proteases produced by Metarhizium anisopliae and their induction in different media. Indian Journal of Microbiology, 50: 449-455.
Dhawan, M. and Joshi, N. 2017. Enzymatic comparison and mortality of Beauveria bassiana against cabbage caterpillar Pieris brassicae LINN. Brazilian Journal of Microbiology, 48: 522-529.
Duo-Chuan, L. 2006. Review of fungal chitinases. Mycopathologia, 161: 345-360.
Dombrovsky, A., Huet, H., Zhang, H., Chejanovsky, N. and Raccah, B. 2003. Comparison of newly isolated cuticular protein genes from six aphid species. Insect Biochemistry and Molecular Biology, 33: 709–715.
El-Sayed, G. N., Ignoffo, C. M., Leathers, T. D. and Gupta, S. C. 1993. Effects of cuticle source and concentration on expression of hydrolytic enzymes by an entomopathogenic fungus, Nomuraea rileyi. Mycopathologia, 122: 149-152.
Erlacher, A., Sousa, F., Schroeder, M., Jus, S., Kokol, V., Cavaco-Paulo, A. and Guebitz, G. M. 2006. A new cuticle scale hydrolysing protease from Beauveria brongniartii. Biotechnology Letters, 28: 703–710.
Fang, W., Leng, B., Xiao, Y., Jin, K., Ma, J., Fan, Y., Feng, J., Yang, X., Zhang, Y. and Pei, Y. 2005. Cloning of Beauveria bassiana chitinase gene Bbchit1 and its application to improve fungal strain virulence. Applied and Environmental Microbiology, 71: 363–370.
Faria, M., Lopes, R. B., Souza, D. A. and Wraight, S. P. 2015. Conidial vigor vs. viability as predictors of virulence of entomopathogenic fungi. Journal of Invertebrate Pathology, 125: 68-72.
Feng, M. 1998. Reliability of extracellular protease and lipase activities of Beauveria bassiana isolates used as their virulence indices. Acta Microbiologica Sinica, 38: 461-467.
Gillespie, J. P., Bateman, R. and Charnley, A. K. 1998. Role of cuticle-degrading protease in the virulence of Metarhizium spp. for the desert locust, Schistocerca gregaria. Journal of Invertebrate Pathology, 71: 128-137.
Goettel, M.S. and Inglis, G. D. 1997. Fungi: hyphomycetes. In: Lacey, L. A. (Ed) Manual of Techniques in Insect Pathology, Academic Press, San Diego, pp. 213–249.
Goettel, M. S., St. Leger, R. J., Rizzo, N. W., Staples, R. C. and Roberts, D. W. 1989. Ultrastructural localization of a cuticle degrading protease produced by the entomopathogenic fungus, Metarhizium anisopliae during penetration of host cuticle. Journal of General Microbiology, 135: 2223–2239.
Gupta, S. C., Leathers, T. D., El-Sayed, G. N. and Ignoffo, C. M. 1994. Relationships among enzyme activities and virulence parameters in Beauveria basiana infections of Galleria mellonella and Trichoplusia ni. Journal of Invertebrate Pathology, 64:13-17.
Kaur, G. and Padmaja, V. 2009. Relationships among activities of extracellular enzyme production and virulence against Helicoverpa armigera in Beauveria bassiana. Journal of Basic Microbiology, 49: 264-274.
Khachatorians, G. G. and Qazi, S. S. 2008. Entomopathogeneic fungi: biochemistry and molecular biology. In: Brakhage, A. A. and Zipfei, P. F. (Eds.), The Mycota, VI: Human and Animal Relationships, 2nd ed. Springer-Verlag, Berlin, pp. 33-61.
Khan, S., Guo, L., Shi, H., Mijit, M. and Qiu, D. 2012. Bioassay and enzymatic comparison of six entomopathogenic fungal isolates for virulence or toxicity against green peach aphids Myzus persicae. African Journal of Biotechnology, 11(77): 14193-14203.
Lacey, L. A., Horton, D. R., Chauvin, R. L. and Stocker, J. M. 1999. Comparative efficacy of Beauveria bassiana, Bacillus thuringiensis, and aldicarb for control of Colorado potato beetle in an irrigated desert agroecosystem and their effects on biodiversity. Entomologia Experimentalis et Applicata, 93: 189-200.
Lu, H.-L. and St. Leger, R. J. 2016. Insect immunity to entomopathogenic fungi. Advances in Genetics, 94: 251-285.
Mayoral, F., Benuzzi, M. and Ladurner, E. 2006. Efficacy of the Beauveria bassiana strain ATCC 74040 (Naturalis®) against whiteflies on protected crops. Integrated Control in Protected Crops, Mediterranean Climate, IOBC/wprs Bulletin, 29: 83-88.
Mohanty, S. S., Raghavendra, K. and Dash, A. P. 2008. Induction of chymoelastase (Pr1) of Metarhizium anisopliae and its role in causing mortality to mosquito larvae. World Journal of Microbiology and Biotechnology, 24(10): 2283–2288.
Montesinos-Matias, R., Viniegra-González, G., Alatorre-Rosas, R. and Loera, O. 2011. Relationship between virulence and enzymatic profiles in the cuticle of Tenebrio molitor by 2-deoxy-D-glucose-resistant mutants of Beauveria bassiana (Bals.) Vuill. World Journal of Microbiology and Biotechnology, 27: 2095–2102.
Mustafa, U. and Kaur, G. 2009. Extracellular enzyme production in Metarhizium anisopliae isolates. Folia Microbiologica, 54: 499-504.
Padmini Palem, P. C. and Padmaja, V. 2013. Functional interplay of certain pathogenicity determinant factors governing virulence of the entomopatogenic fungal species Beauveria. International Journal of Research in Biosciences, 2 (2): 83-92.
Pelizza, S. A., Eliades, L. A., Scorsetti, A. C., Cabello, M. N. and Lange, C. E. 2012. Entomopathogenic fungi from Argentina for the control of Schistocerca cancellata (Orthoptera: Acrididae) nymphs: fungal pathogenicity and enzyme activity. Biocontrol Science and Technology, 22(10): 1119-1129.
Petrisor, C. and Stoian, G. 2017. The role of hydrolytic enzymes produced by entomopathogenic fungi in pathogenesis of insects. Romanian Journal for Plant Protection, 10: 66-72.
Pinto, F. G. S., Fungaro, M. H. P., Ferreira, J. M., Valadares-Inglis, M. C. and Furlaneto, M. C. 2002. Genetic variation in the cuticle degrading protease activity of the entomopathogen Metarhizium flavoviride. Genetics and Molecular Biology, 25: 231-234.
Qazi, S. and Khachatourians, G. 2008. Addition of exogenous carbon and nitrogen sources to aphid exuviae modulates synthesis of proteases and chitinase by germinating conidia of Beauveria bassiana. Archives of Microbiology, 189: 589–596.
Ramzi, S. and Zibaee, A. 2014. Biochemical properties of different entomopathogenic fungi and their virulence against Chilo suppressalis (Lepidoptera: Crambidae) larvae. Biocontrol Science and Technology, 24(5): 597-610.
Revathi, N., Ravikumar, G., Kalaiselvi, M., Gomathi, D. and Uma, C. 2011. Pathogenicity of three entomopathogenic fungi against Helicoverpa armigera. Plant Pathology and Microbiology, 2: 1-4.
Richard, J. S., Neal, T. D., Karl, J. K. and Michael, R. K. 2010. Model reactions for insect cuticle sclerotization: participation of amino groups in the cross-linking of Manduca sexta cuticle protein MsCP36. Insect Biochemistry and Molecular Biology, 40: 252–258.
Robledo-Monterrubio, M., Alatorre-Rosas, R., Viniegra-Gonzalez, G. and Loera, O. 2009. Selection of improved Beauveria bassiana (Bals.) Vuill. strains based on 2-deoxy-D-glucose resistance and physiological analysis. Journal of Invertebrate Pathology, 101: 222-227.
Rosato, Y. B., Messias, C. L. and Azevedo, J. L. 1981. Production of extracellular enzymes by isolates of Metarhizium anisopliae. Journal of Invertebrate Pathology, 38: 1–3.
Safavi, S. A. 2011. Successive subculturing alters spore-bound Pr1 activity, germination and virulence of the entomopathogenic fungus, Beauveria bassiana. Biocontrol Science and Technology, 21(8): 883-890.
Safavi, S. A. 2012. Attenuation of the entomopathogenic fungus Beauveria bassiana following serial in vitro transfers. Biologia, 67(6): 1062-1068.
Safavi, S. A., Shah, F. A., Pakdel, A. K., Rasoulian, G. R., Bandani, A. R. and Butt, T. M. 2007. Effect of nutrition on growth and virulence of the entomopathogenic fungus Beauveria bassiana. FEMS Microbiology Letters, 270: 116-123.
SAS Institute. 2002. SAS for Windows, Release 9.1, Cary, NC, USA.
Schofield, D. A., Westwater, C., Warner, T. and Balish, E. 2005. Differential Candida albicans lipase gene expression during alimentary tract colonization and infection. FEMS Microbiology Letters, 244(2): 359-65.
Seyedtalebi, F. S., Safavi, S. A., Talaei-Hassanloui, R. and Bandani, A. R. 2017. Quantitative comparison for some immune responses among Eurygaster integriceps, Ephestia kuehniella and Zophobas morio against the entomopathogenic fungus Beauveria bassiana. Invertebrate Survival Journal, 14: 174-181.
Seyed talebi, F. S., Safavi, S. A., Talaei-Hassanloui, R. and Bandani, A. R. 2018. Study of the virulence and conidial germination types for some Beauveria bassiana isolates. Biological Control of Pests & Plant Diseases, 7(1): 65-73.
St. Leger, R. J. 1993. Biology and mechanisms of invasion of deuteromycete fungal pathogens. In: Beckage, N. C., Thompson, S. N. and Federici, B. A. (Eds.), Parasites and Pathogens of Insects. Academic Press, San Diego, pp. 211–229.
St. Leger, R. J., Charnley, A. K. and Cooper R. M. 1987. Characterization of cuticle-degrading proteases produced by the entomopathogen Metarhizium anisopliae. Archives of Biochemistry and Biophysics, 253:221–232.
St. Leger, R. J., Durrands, P. K., Charnley, A. K. and Cooper, R. M. 1988. Role of extracellular chymoelastase in the virulence of Metarhizium anisopliae for Manduca sexta. Journal of Invertebrate Pathology, 52:285–293.
St. Leger, R. J., Joshi, L., Bidochka, M. J., Rizzo, N., Roberts, D. W. 1996. Biochemical characterization and ultrastructural localization of two extracellular trypsins produced by Metarhizium anisopliae in infected insect cuticles. Applied and Environmental Microbiology, 62: 1257 1264.
St. Leger, R., Joshi, L. and Roberts, D. W. 1998. Ambient pH is a major determinant in the expression of cuticle-degrading enzymes and hydrophobin by Metarhizium anisopliae. Applied and Environmental Microbiology, 64(2): 709–713.
Svedese, V., Vieira, P., Pereira, J., Mesquita, L., Luna, E. and Figueiredo, A. 2013. Pathogenicity of Beauveria bassiana and production of cuticle-degrading enzymes in the presence of Diatraea saccharalis cuticle. African Journal of Biotechnology, 12(46): 6492 – 6497.
Talaei-Hassanloui, R., Kharazi-Pakdel, A., Goettel, M. S., Little, S. and Mozaffari, J. 2007. Germination polarity of Beauveria bassiana conidia and its possible correlation with virulence. Journal of Invertebrate Pathology, 94: 102-107.
Toledo, A. V., Remes Lenicov, A. M. M. and Lopez Lastra, C. C. 2010. Histopathology caused by the entomopathogenic fungi, Beauveria bassiana and Metarhizium anisopliae, in the adult planthopper, Peregrinus maidis, a maize virus vector. Journal of Insect Science, 10:35.
Tsujita, T., Ninomiya, H. and Okuda, H. 1989. P-nitrophenyl butyrate hydrolyzing activity of hormone sensitive lipase from bovine adipose tissue. Journal of Lipid Research, 30: 997–1004.
van Aalten, D. M. F., Komander, D., Systand, B., Gaseidnes, S., Peter, M. G. and Eijsink, V. G. H. 2011. Structural Insights into the Catalytic Mechanism of a Family 18 Exochitinase. Proceedings of the National Academy of Sciences, 98(16): 8979-8984.
Vega, F. E., Meyling, N. V., Luangsa-Ard, J. J. and Blackwell, M. 2012. Fungal entomopathogens. In: Vega, F. E. and Kaya, H. K. (Eds.) Insect Pathology. Elsevier, London, pp. 171-220.