Hostname: page-component-586b7cd67f-dlnhk Total loading time: 0 Render date: 2024-11-26T11:15:08.844Z Has data issue: false hasContentIssue false
  • English
  • Français

Stability of Metarhizium anisopliae (Hypocreales: Clavicipitaceae) isolates during repeated in vitro subculture and evaluation of an oil-in-water mycoinsecticide

Published online by Cambridge University Press:  24 May 2022

Emine Sönmez ,
Hülya Uzunoğlu ,
Ardahan Eski ,
Zihni Demirbağ  and
İsmail Demir Open the ORCID record for İsmail Demir [Opens in a new window]
Emine Sönmez
Affiliation:
1Düzce University, Environment and Health Centre, 81620, Düzce, Turkey
Hülya Uzunoğlu
Affiliation:
2Karadeniz Technical University, Faculty of Science, Department of Biology, 61080, Trabzon, Turkey
Ardahan Eski
Affiliation:
3Bilecik Şeyh Edebali University, Vocational School, Program of Biomedical Equipment Technology, 11100, Bilecik, Turkey
Zihni Demirbağ
Affiliation:
2Karadeniz Technical University, Faculty of Science, Department of Biology, 61080, Trabzon, Turkey
İsmail Demir*
Affiliation:
2Karadeniz Technical University, Faculty of Science, Department of Biology, 61080, Trabzon, Turkey
*
*Corresponding author. Email: [email protected]
Get access Rights & Permissions [Opens in a new window]

Abstract

Nine Metarhizium anisopliae (Hypocreales: Clavicipitaceae) isolates were evaluated for efficacy against Melolontha melolontha (Coleoptera: Scarabaeidae) larvae, stability in culture, and the superior isolate used to produce an oil-in-water mycoinsecticide. The phenotypic and genotypic characters of four isolates with high virulence were evaluated for their stability after repeating 12 cycles of in vitro subculture. Repeated subculture did not affect the germination of conidia; however, the morphology of some isolates changed significantly. Three isolates lost their virulence, whereas the KTU-2 isolate remained highly pathogenic. Therefore, KTU-2 was selected as the superior isolate for mycoinsecticide production. After the conidia of KTU-2 were produced by solid-state fermentation using cracked rice as the substrate, the conidia were formulated as an oil-in-water emulsion and its efficacy was assessed. The formulation caused 80% mortality on Me. melolontha larvae even at the lowest application rate (1 × 105 conidia/mL) in pot experiments, and complete mortality was obtained with the concentration of 1 × 107 conidia/mL. Lethal concentrations that kill 50% and 95% of Me. melolontha larvae present were estimated as 9.29 × 103 and 2.1 × 106 conidia/mL, respectively. Oil-in-water mycoinsecticide could be a potential candidate for the commercial control of Me. melolontha and other white grubs.

Type
Research Paper
Information
The Canadian Entomologist , Volume 154 , Issue 1 , 2022 , e26
Check for updates
Copyright
© The Author(s), 2022. Published by Cambridge University Press on behalf of the Entomological Society of Canada

Access options

Get access to the full version of this content by using one of the access options below. (Log in options will check for institutional or personal access. Content may require purchase if you do not have access.)

Footnotes

Subject editor: Zhen Zhou

References

Abbott, W.S. 1925. A method of computing the effectiveness of an insecticide. Journal of Economic Entomology, 18: 265–267. https://doi.org/10.1093/jee/18.2.265a.CrossRefGoogle Scholar
Acheampong, M.A., Hill, M.P., Moore, S.D., and Coombes, C.A. 2020. UV sensitivity of Beauveria bassiana and Metarhizium anisopliae isolates under investigation as potential biological control agents in South African citrus orchards. Fungal Biology, 124: 304–310. https://doi.org/10.1016/j.funbio.2019.08.009.CrossRefGoogle ScholarPubMed
Ansari, M.A. and Butt, T.M. 2011. Effects of successive subculturing on stability, virulence, conidial yield, germination, and shelf-life of entomopathogenic fungi. Journal of Applied Microbiology, 110: 1460–1469. https://doi.org/10.1111/j.1365-2672.2011.04994.x.CrossRefGoogle ScholarPubMed
Arthurs, S. and Thomas, M.B. 2001. Effects of temperature and relative humidity on sporulation of Metarhizium anisopliae var. acridum in mycosed cadavers of Schistocerca gregaria . Journal of Invertebrate Pathology, 78: 59–65. https://doi.org/10.1006/jipa.2001.5050.CrossRefGoogle ScholarPubMed
Arzumanov, T., Jenkins, N., and Roussos, S. 2005. Effect of aeration and substrate moisture content on sporulation of Metarhizium anisopliae var. acridum. Process Biochemistry, 40: 1037–1042. https://doi.org/10.1016/j.procbio.2004.03.013.CrossRefGoogle Scholar
Athanassiou, C.G., Kavallieratos, N.G., Rumbos, C.I., and Kontodimas, D.C. 2017. Influence of temperature and relative humidity on the insecticidal efficacy of Metarhizium anisopliae against larvae of Ephestia kuehniella (Lepidoptera: Pyralidae) on wheat. Journal of Insect Science, 17: 22. https://doi.org/10.1093/jisesa/iew107.CrossRefGoogle Scholar
Barreto, L.P., Luz, C., Mascarin, G.M., Roberts, D.W., Arruda, W., and Fernandes, E.K.K. 2016. Effect of heat stress and oil formulation on conidial germination of Metarhizium anisopliae s.s. on tick cuticle and artificial medium. Journal of Invertebrate Pathology, 138: 94–103. https://doi.org/10.1016/j.jip.2016.06.007.CrossRefGoogle ScholarPubMed
Biryol, S., Efe, D., Eski, A., Demirbag, Z., and Demir, I. 2020. Fungal pathogens of Amphimallon solstitiale Linnaeus, 1758 (Coleoptera: Scarabaeidae). Turkish Journal of Entomology, 44: 375–384. https://doi.org/10.16970/entoted.663690.CrossRefGoogle Scholar
Bradford, M. 1976. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Analytical Biochemistry, 72: 248–254. https://doi.org/10.1016/0003-2697(76)90527-3.CrossRefGoogle ScholarPubMed
Braga, G.U.L., Flint, S.D., Messias, C.L., Anderson, A.J., and Roberts, D.W. 2001. Effect of UV-B on conidia and germlings of the entomopathogenic hyphomycete Metarhizium anisopliae . Mycological Research, 105: 874–882. https://doi.org/10.1017/S0953756201004270.CrossRefGoogle Scholar
Butt, T.M. and Goettel, M.S. 2000. Bioassays of entomopathogenic fungi. In Bioassays of entomopathogenic microbes and nematodes. Edited by Navon, A. and Ascher, K.R.S.. Centre for Agriculture and Bioscience International, Wallingford, United Kingdom. Pp. 141–195.CrossRefGoogle Scholar
Butt, T.M., Wang, C.S., Shah, F.A., and Hall, R. 2006. Degeneration of entomogenous fungi. In An ecological and societal approach to biological control. Edited by Eilenberg, J. and Hokkanen, H.M.T.. Kluwer Academic Press, Dordrecht, The Netherlands. Pp. 213–226.CrossRefGoogle Scholar
Chelvi, C.T., Thilagaraj, W.R., and Nalini, R. 2011. Field efficacy of formulations of microbial insecticide Metarhizium anisopliae (Hyphocreales: Clavicipitaceae) for the control of sugarcane white grub Holotrichia serrata F. (Coleoptera: Scarabidae). Journal of Biopesticide, 4: 186–189.Google Scholar
Cobb, B.D. and Clarkson, J.M. 1993. Detection of molecular variation in the insect pathogenic fungus Metarhizium anisopliae using RAPD-PCR. FEMS Microbiology Letters, 112: 319–324. https://doi.org/10.1111/j.1574-6968.1993.tb06469.x.CrossRefGoogle ScholarPubMed
Dimbi, S., Maniania, N.K., Lux, S.A., and Mueke, J.M. 2004. Effect of constant temperatures on germination, radial growth and virulence of Metarhizium anisopliae to three species of African tephritid fruit flies. BioControl, 49: 83–94. https://doi.org/10.1023/B:BICO.0000009397.84153.79.CrossRefGoogle Scholar
Erler, F. and Ates, A.O. 2015. Potential of two entomopathogenic fungi, Beauveria bassiana and Metarhizium anisopliae (Coleoptera: Scarabaeidae), as biological control agents against the June beetle. Journal of Insect Science, 5: 44. https://doi.org/10.1093/jisesa/iev044.CrossRefGoogle Scholar
Faria, M.R. and Wraight, S.P. 2007. Mycoinsecticides and mycoacaricides: a comprehensive list with worldwide coverage and international classification of formulation types. Biological Control, 43: 237–256. https://doi.org/10.1016/j.biocontrol.2007.08.001.CrossRefGoogle Scholar
Fernandes, E.K.K., Rangel, D.E.N., Braga, G.U.L., and Roberts, D.W. 2015. Tolerance of entomopathogenic fungi to ultraviolet radiation: a review on screening of strains and their formulation. Current Genetics, 61: 427–440. https://doi.org/10.1007/s00294-015-0492-z.CrossRefGoogle ScholarPubMed
Finney, D.J. 1971. Probit analysis. Cambridge University Press, Cambridge, United Kingdom.Google Scholar
Goffre, D., Jensen, A.B., Lopez Lastra, C.C., Humber, R., and Folgarait, P.J. 2021. Conidiobolus lunulus, a new entomophthoralean species isolated from leafcutter ants. Mycologia, 113: 56–64.CrossRefGoogle ScholarPubMed
Guijarro, B., Melgarejo, P., and Cal, A.D. 2007. Effect of stabilizers on the shelf-life of Penicillium frequentans conidia and their efficacy as a biological agent against peach brown rot. International Journal of Food Microbiology, 113: 117–124. https://doi.org/10.1016/j.ijfoodmicro.2006.06.024.CrossRefGoogle ScholarPubMed
Hall, R.A. 1980. Effect of repeated subculturing on agar and passaging through an insect host on pathogenicity, morphology, and growth rate of Verticillium lecanii . Journal of Invertebrate Pathology, 36: 216–222. https://doi.org/10.1016/0022-2011(80)90027-0.CrossRefGoogle Scholar
Hernandez-Velazquez, V.M., Berlanga-Padilla, A.M., and Barrientos-Lozano, L. 2000. Vegetable and mineral oil formulations of Metarhizium anisopliae var. acridum to control the central American locust (Schistocerca piceifrons piceifrons Walker) (Orthoptera: Acrididae). Journal of Orthoptera Research, 9: 223–227. https://doi.org/10.2307/3503650.CrossRefGoogle Scholar
Hutwimmer, S., Wagner, S., Affenzeller, M., Burgstaller, W., and Strasser, H. 2008. Algorithm-based design of synthetic growth media stimulating virulence properties of Metarhizium anisopliae conidia. Journal of Applied Microbiology, 105: 2026–2034. https://doi.org/10.1111/j.1365-2672.2008.03872.x.CrossRefGoogle ScholarPubMed
Ibrahim, R.A. 2017. Laboratory evaluation of entomopathogenic fungi, commercial formulations, against the rhinoceros beetles, Oryctes agamemnon arabicus (Coleoptera: Scarabaeidae). Egyptian Journal of Biological Pest Control, 27: 49–55.Google Scholar
Kocacevik, S., Sevim, A., Eroglu, M., Demirbag, Z., and Demir, I. 2015. Molecular characterization, virulence and horizontal transmission of Beauveria pseudobassiana from Dendroctonus micans (Kug.) (Coleoptera: Curculionidae). Journal of Applied Entomology, 139: 381–389. https://doi.org/10.1111/jen.12181.CrossRefGoogle Scholar
Laengle, T., Pernfuss, B., Seger, C., and Strasser, H. 2005. Field efficacy evaluation of Beauveria brongniartii against Melolontha melolontha in potato cultures. Sydowia, 57: 54–93.Google Scholar
Leal, S.C.M., Bertioli, D.J., Butt, T.M., Carder, J.H., Burrows, P.R., and Peberdy, J.F. 1997. Amplification and restriction endonuclease digestion of the pr1 gene for the detection and characterization of Metarhizium strains. Mycological Research, 101: 257–265. https://doi.org/10.1017/S0953756296002560.CrossRefGoogle Scholar
Leal, S.C.M., Bertioli, D.J., Butt, T.M., and Peberdy, J.F. 1994. Characterization of isolates of the entomopathogenic fungus Metarhizium anisopliae by RAPD-PCR. Mycological Research, 98: 1077–1081. https://doi.org/10.1016/S0953-7562(09)80436-X.CrossRefGoogle Scholar
Leal, S.C.M., Butt, T.M., Peberdy, J.F., and Bertioli, D.J. 2000. Genetic exchange in Metarhizium anisopliae strains coinfecting Phaedon cochleariae is revealed by molecular markers. Mycological Research, 104: 409–414. https://doi.org/10.1017/S0953756299001549.CrossRefGoogle Scholar
Lee, H., Song, M., and Hwang, S. 2003. Optimizing bioconversion of deproteinated cheese whey to mycelia of Ganoderma lucidum. Process Biochemistry, 38: 1685–1693. https://doi.org/10.1016/S0032-9592(02)00259-5.CrossRefGoogle Scholar
Leger, R.J.S., Charnley, A.K., and Cooper, R.M. 1986. Cuticle-degrading enzymes of entomopathogenic fungi: synthesis in culture on cuticle. Journal of Invertebrate Pathology, 48: 85–95. https://doi.org/10.1016/0022-2011(86)90146-1.CrossRefGoogle Scholar
Loesch, A., Hutwimmer, S., and Strasser, H. 2010. Carbon utilization pattern as a potential quality control criterion for virulence of Beauveria brongniartii . Journal of Invertebrate Pathology, 104: 58–65. https://doi.org/10.1016/j.jip.2010.01.007.CrossRefGoogle ScholarPubMed
Marcelino, D., Giordano, R., Gouli, S., Gouli, V., Parker, B.L., Skinner, M., TeBeest, D., and Cesnik, R. 2008. Colletotrichum acutatum var. fioriniae (telemorf: Glomerella acutata var. fioriniae var nov.) infection of a large scale insect. Mycologia, 100: 353–374.CrossRefGoogle Scholar
Ment, D., Gindin, G., Glazer, I., Perl, S., Elad, D., and Samish, M. 2010. The effect of temperature and relative humidity on the formation of Metarhizium anisopliae chlamydospores in tick eggs. Fungal Biology, 114: 49–56. https://doi.org/10.1016/j.mycres.2009.10.005.CrossRefGoogle ScholarPubMed
Muniz, E.R., Paixão, F.R.S., Barreto, L.P., Luz, C., Arruda, W., Angelo, I.C., and Fernandes, E.K.K. 2020. Efficacy of Metarhizium anisopliae conidia in oil-in-water emulsion against the tick Rhipicephalus microplus under heat and dry conditions. BioControl, 65: 339–351. https://doi.org/10.1007/s10526-020-10002-5.CrossRefGoogle Scholar
Nahar, P.B., Kulkarni, S.A., Kulye, M.S., Chavan, S.B., Kulkarni, G., Rajendran, A., et al. 2008. Effect of repeated in vitro sub-culturing on the virulence of Metarhizium anisopliae against Helicoverpa armigera (Lepidoptera: Noctuidae). Biocontrol Science and Technology, 18: 337–355. https://doi.org/10.1080/09583150801935650.CrossRefGoogle Scholar
Oliveira, D.G.P., Lopes, R.B., Rezende, J.M., and Delalibera, I. 2018. Increased tolerance of Beauveria bassiana and Metarhizium anisopliae conidia to high temperature provided by oil-based formulations. Journal of Invertebrate Pathology, 151: 151–157. https://doi.org/10.1016/j.jip.2017.11.012.CrossRefGoogle ScholarPubMed
Paixao, F.R.S., Muniz, E.R., Barreto, L.P., Bernardo, C.C., Mascarin, G.M., Luz, C., and Fernandes, E.K.K. 2017. Increased heat tolerance afforded by oil-based conidial formulations of Metarhizium anisopliae and Metarhizium robertsii . Biocontrol Science and Technology, 27: 324–337. https://doi.org/10.1080/09583157.2017.1281380.CrossRefGoogle Scholar
Perinotto, W.M.S., Angelo, I.C., Golo, P.S., Camargo, M.G., Quinelato, S., Santi, L., and Bittencourt, V.R.E.P. 2014. Metarhizium anisopliae (Deuteromycetes: Moniliaceae) Pr1 activity: biochemical marker of fungal virulence in Rhipicephalus microplus (Acari: Ixodidae). Biocontrol Science and Technology, 24: 123–132. https://doi.org/10.1080/09583157.2013.847903.CrossRefGoogle Scholar
Putnoky-Csicsó, B., Tonk, S., Szabó, A., Márton, Z., Bogdányi, F.T., Tóth, F., et al. 2020. Effectiveness of the entomopathogenic fungal species Metarhizium anisopliae strain NCAIM 362 treatments against soil inhabiting Melolontha melolontha larvae in sweet potato (Ipomoea batatas L.). Journal of Fungi, 6: 116. https://doi.org/10.3390/jof6030116.CrossRefGoogle Scholar
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: 883–891. https://doi.org/10.1080/09583157.2011.588317.CrossRefGoogle Scholar
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. https://doi.org/10.1111/j.1574-6968.2007.00666.x.CrossRefGoogle ScholarPubMed
Santoro, P.H., Zorzetti, J., Constanski, K., and Neves, P.M.O.J. 2014. Conidial production, virulence, and stress tolerance of Beauveria bassiana conidia after successive in vitro subculturing. Revista Colombiana de Entomologia, 40: 85–90.Google Scholar
Sevim, A., Demir, I., Hofte, M., Humber, R.A., and Demirbag, Z. 2010. Isolation and characterization of entomopathogenic fungi from hazelnut-growing region of Turkey. BioControl, 55: 279–297. https://doi.org/10.1007/s10526-009-9235-8.CrossRefGoogle Scholar
Shah, F.A. and Butt, T.M. 2005. Influence of nutrition on the production and physiology of sectors produced by the insect pathogenic fungus Metarhizium anisopliae. FEMS Microbiology Letters, 250: 201–207. https://doi.org/10.1016/j.femsle.2005.07.011.CrossRefGoogle ScholarPubMed
Shah, F.A., Wang, C.S., and Butt, T.M. 2005. Nutrition influences growth and virulence of the insect pathogenic fungus Metarhizium anisopliae . FEMS Microbiology Letters, 251: 259–266. https://doi.org/10.1016/j.femsle.2005.08.010.CrossRefGoogle ScholarPubMed
Small, C.L.N. and Bidochka, M.J. 2005. Up-regulation of Pr1, a subtilisin-like protease, during conidiation in the insect pathogen Metarhizium anisopliae . Mycological Research, 109: 307–313. https://doi.org/10.1017/S0953756204001856.CrossRefGoogle ScholarPubMed
Sonmez, E., Sevim, A., Demirbağ, Z., and Demir, I. 2016. Isolation, characterization and virulence of entomopathogenic fungi from Gryllotalpa gryllotalpa (Orthoptera: Gryllotalpidae). Applied Entomology and Zoology, 51: 213–223. https://doi.org/10.1007/s13355-015-0390-3.CrossRefGoogle Scholar
Tanyeli, E., Sevim, A., Demirbag, Z., Eroglu, M., and Demir, I. 2010. Isolation and virulence of entomopathogenic fungi against the great spruce bark beetle, Dendroctonus micans (Kugelann) (Coleoptera: Scolytidae). Biocontrol Science and Technology, 20: 695–701. https://doi.org/10.1080/09583151003717219.CrossRefGoogle Scholar
Tigano-Milani, M.S., Gomes, A.C.M.M., and Sobral, B.W.S. 1995. Genetic variability among Brazilian isolates of the entomopathogenic fungus Metarhizium anisopliae . Journal of Invertebrate Pathology, 65: 206–210. https://doi.org/10.1006/jipa.1995.1031.CrossRefGoogle Scholar
Ummidi, V.R.S. and Vadlamani, P. 2014. Preparation and use of oil formulations of Beauveria bassiana and Metarhizium anisopliae against Spodoptera litura larvae. African Journal of Microbiology Research, 8: 1638–1644. https://doi.org/10.5897/AJMR2013.6593.Google Scholar
Vandenberg, J.D. and Cantone, F.A. 2004. Effect of serial transfer of three strains of Paecilomyces fumosoroseus on growth in vitro, virulence, and host specificity. Journal of Invertebrate Pathology, 85: 40–45. https://doi.org/10.1016/j.jip.2003.12.004.CrossRefGoogle ScholarPubMed
Verma, M., Brar, S.K., Tyagi, R.D., Surampalli, R.Y., and Valero, J.R. 2007. Starch industry wastewater as a substrate for antagonist, Trichoderma viride production. Bioresource Technology, 98: 2154–2162. https://doi.org/10.1016/j.biortech.2006.08.032.CrossRefGoogle ScholarPubMed
Wang, C., Types, M.A., and Butt, T.M. 2002. Detection and characterisation of pr1 virulent gene deficiencies in the insect pathogenic fungus Metarhizium anisopliae . FEMS Microbiology Letters, 213: 251–255. https://doi.org/10.1111/j.1574-6968.2002.tb11314.x.CrossRefGoogle ScholarPubMed
Xavier-Santos, S., Lopes, R.B., and Faria, M. 2011. Emulsifiable oils protect Metarhizium robertsii and Metarhizium pingshaense conidia from imbibitional damage. Biological Control, 59: 261–267. https://doi.org/10.1016/j.biocontrol.2011.08.003.CrossRefGoogle Scholar
Yucel, B., Gozuacik, C., Gencer, D., Demir, I., and Demirbag, Z. 2018. Determination of fungal pathogens of Hypera postica (Gyllenhall) (Coleoptera: Curculionidae): isolation, characterization, and susceptibility. Egyptian Journal of Biological Pest Control, 28: 39. https://doi.org/10.1186/s41938-018-0043-2.CrossRefGoogle Scholar
Zimmermann, G. 1982. Effect of high temperatures and artificial sunlight on the viability of conidia of Metarhizium anisopliae . Journal of Invertebrate Pathology, 40: 36–40. https://doi.org/10.1016/0022-2011(82)90034-9.CrossRefGoogle Scholar
Zimmermann, G. 2007. Review on safety of the entomopathogenic fungus Metarhizium anisopliae. Biocontrol Science and Technology, 17: 879–920. https://doi.org/10.1080/09583150701593963.CrossRefGoogle Scholar