Formulation of Metarhizium pinghaense and Metarhizium robertsii and the infection potential of the formulations against Pseudococcus viburni (Hemiptera: Pseudococcidae) after storage




conidial viability, pathogenicity, Pseudococcus viburni, storage, mineral carriers, oil carriers


Formulation of entomopathogens refers to the mixing of various inert ingredients, like clays and mineral oils, with the active ingredients which are the entomopathogens. Successful formulation enhances the survival of the entomopathogen and also eases their transportation, storage, preparation and application. The aim of this study was to develop a formulation to maintain the longevity and pathogenicity of the mass-produced conidia of local Metarhizium pinghaense and M. robertsii, for above-ground future commercial field application against Pseudococcus viburni. The objectives were to develop a cost-effective protocol for formulation of infective propagules and to test their effectiveness under laboratory conditions. The conidia of both isolates were formulated using four different oils (liquid paraffin, coconut, canola and olive oils) as liquid carriers, and diatomaceous earth as a mineral carrier. Conidial viability and pathogenicity were assessed over a period of eight weeks. In the study, it was observed that the conidia formulated in oil carriers maintained a high conidial viability and survival rate of >95 % over a period of eight weeks for both isolates, relative to when formulated in mineral carriers, or when stored as dry conidial powder. The conidia in all the oil formulations were also observed to induce high mortality, ranging between 60 % and 90 % for M. pinghaense, and between 70 % and 90 % for M. robertsii, when used against P. viburni. The ability of conidia of both isolates to maintain viability and pathogenicity, following storage in the oil formulations, increased the likelihood of the local isolates being successfully integrated as biological control agents for management of P. viburni  under field conditions.


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Alves RT, Bateman RP, Gunn J, Prior C, Leather SR. 2002. Effects of different formulations on viability and medium-term storage of Metarhizium anisopliae conidia. Neotropical Entomology 31(1): 91–99.

Bateman RP, Carey M, Moore DE, Prior C. 1993. The enhanced infectivity of Metarhizium flavoviride in oil formulations to desert locusts at low humidities. Annals of Applied Biology 122(1): 145–152.

Batta YA. 2003. Production and testing of novel formulations of the entomopathogenic fungus Metarhizium anisopliae (Metschinkoff) Sorokin (Deuteromycotina: hyphomycetes). Crop Protection 22(2): 415–422.

Blumberg D, Van Driesche RG. 2001. Encapsulation rates of three encyrtid parasitoids by three mealybug species (Homoptera: Pseudococcidae) found commonly as pests in commercial greenhouses. Biological Control 22(2): 191–199.

Bukhari T, Takken W, Koenraadt CJ. 2011. Development of Metarhizium anisopliae and Beauveria bassiana formulations for control of malaria mosquito larvae. Parasites & Vectors 22;4(1): 23.

Burges HD. 2012. Formulation of Microbial Biopesticides: Beneficial Microorganisms, Nematodes and Seed Treatments. Berlin, Germany: Springer Science & Business Media.

Charles JG. 1993. A survey of mealybugs and their natural enemies in horticultural crops in North Island, New Zealand, with implications for biological control. Biocontrol Science and Technology 3(4): 405–418.

Charles JG. 2011. Using parasitoids to infer a native range for the obscure mealybug, Pseudococcus viburni, in South America. BioControl 56(2): 155–161.

Couch TL, Ignoffo CM. 1981. Formulation of insect pathogens. In: Burges HD, editor. Microbial Control of Pests and Plant Diseases 1970–1980. London, U.K.: Academic Press.

Da Silva VCP, Nondillo A, Galzer ECW, Garcia MS, Botton M. 2017a. Effect of host plants on the development, survivorship, and reproduction of Pseudococcus viburni (Hemiptera: pseudococcidae). Florida Entomologist 100(4): 718–724.

Da Silva VP, Garcia M, Botton M. 2017b. Biology of Blepyrus clavicornis (Compere) (Hymenoptera: Encyrtidae), a parasitoid of Pseudococcus viburni (Signoret) (Hemiptera: Pseudococcidae). Revista Brasileira de Entomologia 61(3): 257–261.

Daane KM, Almeida RPP, Bell VA, Walker JTS, Botton M, Fallahzadeh M, Mani M, Miano JL, Sforza R, Walton VM, et al. 2012. Biology and management of mealybugs in vineyards. In: Bostanian NJ, Vincent C, Isaacs, R, editors. Arthropod Management in Vineyards. Dordrecht, Netherlands: Springer. p. 271–307.

Daoust RA, Ward MG, Roberts DW. 1983. Effect of formulation on the viability of Metarhizium anisopliae conidia. Journal of Invertebrate Pathology 41(2): 151–160.

Ekesi S, Maniania NK, Lux SA. 2002. Mortality in three African tephritid fruit fly puparia and adults caused by the entomopathogenic fungi, Metarhizium anisopliae and Beauveria bassiana. Biocontrol Science and Technology 12(1): 7–17.

Franco JC, Zada A, Mendel Z. 2009. Novel approaches for the management of mealybug pests: application and resistance management. In: Ishaaya I, Horowitz AR, editors. Biorational Control of Arthropod Pests. Dordrecht, Netherlands: Springer. p. 233–278.

Franco JC, Zada A, Mendel Z. 2009. Novel approaches for the management of mealybug pests. In: Ishaaya I, Horowitz A, editors. Biorational Control of Arthropod Pests. Dordrecht, Netherlands: Springer. p. 233–278.

Fravel DR, Connick WJ, Lewis JA. 1998. Formulation of microorganisms to control plant diseases. In: Burges HD, editor. Formulation of Microbial Biopesticides: Beneficial Microorganisms, Nematodes and Seed Treatments. Dordrecht, Netherlands: Springer. p. 187–202.

Hedgecock S, Moore D, Higgins PM, Prior C. 1995. Influence of moisture content on temperature tolerance and storage of Metarhizium flavoviride conidia in an oil formulation. Biocontrol Science and Technology 5(3): 371–378.

Humber RA. 1997. Fungi: preservation of cultures. In: Lacey LA, editor. Manual of Techniques in Insect Pathology. Cambridge, Massachusetts: Academic Press. p. 269–279.

Inglis GD, Enkerli JUERG, Goettel MS. 2012. Laboratory techniques used for entomopathogenic fungi: Hypocreales. In: Lacey LA, editor. Manual of Techniques in Invertebrate Pathology. London, U.K.: Academic Press. p. 189–253.

Inyang EN, Mccartney HA, Oyejola B, Ibrahim L, Pye BJ, Archer SA, Butt TM. 2000. Effect of formulation, application and rain on the persistence of the entomogenous fungus Metarhizium anisopliae on oilseed rape. Mycological Research 104(6): 653–661.

Jaronski ST, Mascarin GM. Chapter 9, Mass production of fungal entomopathogens. Microbial Control of Insect and Mite Pests: From Theory to Practice. In: Lacey LA, editor. Microbial Control of Insect and Mite Pests: From Theory to Practice. Amsterdam, Netherlands: Elsevier; 2017. p. 141–155.

Langewald J. 1999. LUBILOSA: Green Muscle User Handbook. Version 4. Biological Locust and Grasshopper Control Project. CABI.

Mathulwe LL, Malan AP, Stokwe NF. Laboratory screening of entomopathogenic fungi and nematodes for pathogenicity against the obscure mealybug, Pseudococcus viburni (Hemiptera: pseudococcidae). Biocontrol Science and Technology 2022;32(4): 397–417.

Mathulwe LL, Malan AP, Stokwe NF. Mass Production of entomopathogenic fungi, Metarhizium robertsii and Metarhizium pinghaense, for commercial application against Insect Pests. Journal of Visual Experiment 2022 Mar 31;181(181): e63246. PMID:35435892

Mibey RK. 1.2. 2.2 Scooty moulds. World Crop Pests. 1997;7:275–290.

Mkiga AM, Mohamed SA, du Plessis H, Khamis FM, Akutse KS, Nderitu PW, Niassy S, Muriithi BW, Ekesi S. Compatibility and efficacy of Metarhizium anisopliae and sex pheromone for controlling Thaumatotibia leucotreta. Journal of Pest Science 2021;94(2): 393–407.

Moore D, Higgins PM. Viability of stored conidia of Metarhizium flavoviride Gams and Rozsypal, produced under differing culture regimes and stored with clays. Biocontrol Science and Technology 1997;7(3): 335–344.

Moore D, Bateman RP, Carey M, Prior C. 1995. Long term storage of Metarhizium flavoviride conidia in oil formulations for the control of locusts and grasshoppers. Biocontrol Science and Technology 5(2): 193–200.

Moore D, Lord JC, Smith SM. 2000 Pathogens. In: Subramanyam B, Hagstrum DW, editors. Alternatives to Pesticides in Stored-Product IPM. Dordrecht, Netherlands: Kluwer Academic Publishers. p. 193–227.

Mudavanhu P. 2009. An investigation into the integrated pest management of the obscure mealybug, Pseudococcus viburni (Signoret) (Hemiptera: Pseudococcidae), in pome fruit orchards in the Western Cape Province, South Africa. Doctoral dissertation, Stellenbosch University, Stellenbosch, South Africa.

Mugonza J, Otim MH, Egonyu JP. 2020. The comparative virulence of an atoxigenic strain of Aspergillus flavus (Eurotiales: Trichocomaceae) and the commercial ICIPE 69 Metarhizium anisopliae (Hypocreales: Clavicipitaceae) to the bean leaf beetle Ootheca mutabilis (Coleoptera: Chrysomelidae). International Journal of Tropical Insect Science 40(2): 403–411.

Nel PJ. 1983. Decidous Fruits and Vines: Pests and Diseases and their Control. David Philip, Cape Town, South Africa.

Oerke EC, Dehne HW. 2004. Safeguarding production losses in major crops and the role of crop protection. Crop Protection. 23(4): 275–285.

Park DS, Leem YJ, Hahn KW, Suh SJ, Hong KJ, Oh HW. 2010. Molecular identification of mealybugs (Hemiptera: Pseudococcidae) found on Korean pears. J Econ Entomol. 103(1): 25–33.

Polar P, Kairo MT, Moore D, Pegram R, John SA. 2005. Comparison of water, oils and emulsifiable adjuvant oils as formulating agents for Metarhizium anisopliae for use in control of Boophilus microplus. Mycopathologia. 160(2): 151–157.

Prior C, Jollands P, Le Patourel G. 1988 Infectivity of oil and water formulations of Beauveria bassiana (Deuteromycotina: Hyphomycetes) to the cocoa weevil pest Pantorhytes plutus (Coleoptera: Curculionidae). Journal of Invertebrate Pathology 52(1): 66–72.

Roberts DW, St Leger RJ. 2004. Metarhizium spp., cosmopolitan insect-pathogenic fungi: mycological aspects. Adv Appl Microbiol. 54:1–70.

Sedighi N, Abbasipour H, Askary H, Sheikhi Gorjan A. 2013. Effect of oil suspended conidia of Metarhizium anisopliae var. major on mortality of the sunn pest, Eurygaster integriceps Puton (Hemiptera: Scutelleridae). Arch Phytopathol Pflanzenschutz. 46(2): 128–140.

Shah PA, Pell JK. 2003. Entomopathogenic fungi as biological control agents. Appl Microbiol Biotechnol. 61(5-6): 413–423.

Stathers TE, Moore D, Prior C. 1993. The effect of different temperatures on the viability of Metarhizium flavoviride conidia stored in vegetable and mineral oils. Journal of Invertebrate Pathology 62(2): 111–115.

TIBCO Software Inc. 2018. STATISTICA (data analysis software system), version Palo Alto, CA, USA: TIBCO Software Inc.

Tupe SG, Pathan EK, Deshpande MV. 2017. Development of Metarhizium anisopliae as a mycoinsecticide: from isolation to field performance. Journal of Visual Experiment 125(125): e55272.

Wakgari WM, Giliomee JH. 2004. Description of adult and immature female instars of Pseudococcus viburni (Hemiptera: Pseudococcidae) found on apple in South Africa. African Entomology 12:29–38.

Wood BW, Tedders WL, Reilly CC. 1988. Sooty mold fungus on pecan foliage suppresses light penetration and net photosynthesis. HortScience. 24:231–256.

Wraight SP, Jackson MA, De Kock SL. 2001. Production, stabilization and formulation of fungal biocontrol agents. In: Butt TM, Jackson C, Magan N, editors. Fungi as Biocontrol Agents: Progress, Problems and Potential. Wallingford, U.K.: CABI. p. 253–287.

Xavier-Santos S, Lopes RB, Faria M. 2011. Emulsifiable oils protect Metarhizium robertsii and Metarhizium pingshaense conidia from imbibitional damage. Biological Control 59(2): 261–267.

Zimmermann G. 2007. Review on safety of the entomopathogenic fungi Beauveria bassiana and Beauveria brongniartii. Biocontrol Science and Technology 17(6): 553–596.




How to Cite

Mathulwe LL, Malan AP, Stokwe NF. Formulation of Metarhizium pinghaense and Metarhizium robertsii and the infection potential of the formulations against Pseudococcus viburni (Hemiptera: Pseudococcidae) after storage. Afr. Entomol. [Internet]. 2023 Feb. 10 [cited 2024 Apr. 14];31. Available from: