Skip to main content
SpringerLink
Log in

Transformation and attachment of Beauveria bassiana conidia on the cuticle of Tribolium confusum and Sitophilus oryzae in conjunction with diatomaceous earth

Journal of Pest Science volume 85, pages 387–394 (2012)Cite this article

Abstract

Agrobacterium tumefaciens-mediated transformation (AMT) was used in order to create fluorescent conidia of Beauveria bassiana (Balsamo) Vuillemin (Ascomycota: Hypocreales). From the fungal transformants, one Enhanced Green Fluorescent Protein B. bassiana strain indicated stability (maintained the GFP characteristics) in fluorescent signal, while its pathogenicity did not differ significantly from the wild B. bassiana type. After exposure on wheat treated with the transformed conidia, fluorescence was evident on all body parts but especially on metasternon of adults of Tribolium confusum Jacquelin du Val (Coleoptera: Tenebrionidae) and on elytra in of Sitophilus oryzae (L.) (Coleoptera: Curculionidae). No significant differences were noted in fluorescence levels in the case of T. confusum between the transformed and the wild strain. However, the addition of diatomaceous earth (DE) increased attachment, especially at increased intervals after exposure. For S. oryzae, fluorescence of the transformed strain was decreased with the increased post-exposure time, but the simultaneous presence of DE increased fluorescence. The results of this study demonstrate that fluorescent conidia of B. bassiana can be utilized as a tool for the evaluation of parameters that affect the attachment of entomopathogenic fungi in insects’ cuticle.

This is a preview of subscription content, access via your institution.

Access options

Buy single article

Instant access to the full article PDF.

USD 39.95

Price includes VAT (Canada)

Fig. 1
Fig. 2
Fig. 3
Fig. 4

References

  • Akbar W, Lord JC, Nechols JR, Howard RW (2004) Diatomaceous earth increases the efficacy of Beauveria bassiana against Tribolium castaneum larvae and increases conidia attachment. J Econ Entomol 97:273–280

    Article  PubMed  Google Scholar 

  • Amey RC, Athey-Pollard A, Mills PR, Bailey A, Foster GD (2002) PEG-mediated and Agrobacterium-mediated transformation in the mycopathogen Verticillium fungicola. Mycol Res 106:4–11

    Article  Google Scholar 

  • Anonymous (2000) Biopesticide technical document Beauveria bassiana strain GHA. Biol Control. http://www.epa.gov/pesticides/biopesticides/factsheets/fs128924t.htm

  • Athanassiou CG, Steenberg T (2007) Insecticidal effect of Beauveria bassiana (Balsamo) Vuillemin (Ascomycota: Hypocreales) in combination with three diatomaceous earth formulations against Sitophilus granarius (L.) (Coleoptera: Curculionidae). Biol Control 40:411–416

    Article  Google Scholar 

  • Bidochka MJ, Khachatourians GG (1992) Growth of the entomopathogenic fungus Beauveria bassiana on cuticular components from the migratory grasshopper, Melanoplus sanguinipes. J Invertebr Pathol 59:165–173

    Article  Google Scholar 

  • Boucias DG, Pendland JC, Latge JP (1988) Nonspecific factors involved in attachment of entomopathogenic Deuteromycetes to host insect cuticle. Appl Environ Microbiol 54:1795–1805

    PubMed  CAS  Google Scholar 

  • Cantone FA, Vanderberg JD (1999) Use of the green fluorescent protein for investigations of Paecilomyces fumosoroseus in insect hosts. J Invertebr Pathol 74:193–197

    Article  PubMed  CAS  Google Scholar 

  • Cao Y, Peng G, He Z, Wang Z, Yin Y, Xia Y (2007) Tranformation of Metarhizium anisopliae with benomyl resistance and green fluorescent protein genes provides a tag for genetically engineered strains. Biotechnol Lett 29:907–911

    Article  PubMed  CAS  Google Scholar 

  • Chen N, Hosing T, Goodwin PH (2003) Use of green fluorescent protein to quantify the growth of Colletotrichum during infection of tobacco. J Microbiol Methods 53:113–122

    Article  PubMed  CAS  Google Scholar 

  • Cody CW, Prasher DC, Wrestler WM, Prendergast FG, Ward WW (1993) Chemical structure of the hexapeptide chromophore of the Aequorea green-fluorescent protein. Biochemistry 32:1212–1218

    Article  PubMed  CAS  Google Scholar 

  • Ebeling E (1971) Sorptive dusts for pest control. Annu Rev Entomol 16:123–158

    Article  PubMed  CAS  Google Scholar 

  • Fernandez S, Groden E, Vandenberg JD, Furlong MJ (2001) The effect of mode of exposure to Beauveria bassiana on conidia acquisition and host mortality of Colorado potato beetle, Leptinotarsa decemlineata. J Invertebr Pathol 77:217–226

    Article  PubMed  CAS  Google Scholar 

  • Greenspan L (1977) Humidity fixed points of binary saturated aqueous solutions. J Res Natl Bur Stand A 81:89–96

    Article  Google Scholar 

  • Hallsworth JE, Magan N (1999) Water and temperature relations of growth of the entomogenous fungi Beauveria bassiana, Metarhizium anisopliae and Paecilomyces farinosus. J Invertebr Pathol 74:261–266

    Article  PubMed  Google Scholar 

  • Hansen LS, Steenberg T (2007) Combining larval parasitoids and an entomopathogenic fungus for biological control of Sitophilus granarius (Coleoptera: Curculionidae) in stored grain. Biol Control 40:237–242

    Article  Google Scholar 

  • Inglis PW, Aragao FJL, Frazao H, Magalhaes BP, Valadares-Inglis MC (2000) Biolistic co-transformation of Metarhizium anisopliae var. acridum strain CG423 with green fluorescent protein and resistance to glufosinate ammonium. FEMS Microbiol Lett 191:249–254

    Article  PubMed  CAS  Google Scholar 

  • Jin S, Xu R, Wei Y, Goodwin PH (1999) Increased expression of a plant actin gene during a biotrophic interaction between round-leaved mallow, Malva pusilla, and Colletotrichum gloeosporioides f. sp. malvae. Planta 209:487–494

    Article  PubMed  CAS  Google Scholar 

  • Kavallieratos NG, Athanassiou CG, Michalaki MP, Batta YA, Rigatos HA, Pashalidou FG, Balotis GN, Tomanovic Z, Vayias BJ (2006) Effect of the combined use of Metarhizium anisopliae (Metschinkoff) Sorokin and diatomaceous earth for the control of three stored-product beetle species. Crop Prot 25:1087–1094

    Article  Google Scholar 

  • Kavallieratos NG, Wakefield ME, Athanassiou CG (2008) Factors affecting the attachment of conidia of Metarhizium anisopliae (Metschnikoff) Sorokin (Deuteromycotina: Hyphomycetes) to different body parts of Tribolium confusum Jacquelin du Val (Coleoptera: Tenebrionidae) adults. IOBC/WPRS Bull 40:181–190

    Google Scholar 

  • Korunic Z (1998) Diatomaceous earths, a group of natural insecticides. J Stored Prod Res 34:87–97

    Article  Google Scholar 

  • Lee SB, Taylor JW (1990) Isolation of DNA from fungal mycelia and single spores. In: Innis MA et al (eds) PCR protocols: a guide to methods and applications. Academic Press, San Diego, pp 282–287

    Google Scholar 

  • Lord JC (2001) Desiccant dusts synergize the effect of Beauveria bassiana (Hyphomycetes: Moniliales) on stored-grain beetles. J Econ Entomol 94:367–372

    Article  PubMed  CAS  Google Scholar 

  • Lord JC (2005) Low humidity, moderate temperature, and desiccant dust favour efficacy of Beauveria bassiana (Hyphomycetes: Moniliales) for the lesser grain borer, Rhyzopertha dominica (Coleoptera: Bruchidae). Biol Control 34:180–186

    Article  Google Scholar 

  • Mahe A, Grisvard J, Dron M (1992) Fungal- and plant-specific gene markers to follow the bean anthracnose infection process and normalize a bean chitinase mRNA induction. Mol Plant Microb Interact 5:242–248

    Article  CAS  Google Scholar 

  • Mewis I, Ulrichs Ch (2001) Action of amorphous diatomaceous earth against different stages of the stored product pests Tribolium confusum, Tenebrio molitor, Sitophilus granarius and Plodia interpunctella. J Stored Prod Res 37:153–164

    Article  PubMed  CAS  Google Scholar 

  • Michalaki MP, Athanassiou CG, Kavallieratos NG, Batta YA, Balotis GN (2006) Effectiveness of Metarhizium anisopliae (Metschinkoff) Sorokin applied alone or in combination with diatomaceous earth against Tribolium confusum Du Val larvae: Influence of temperature, relative humidity and type of commodity. Crop Prot 25:418–425

    Article  Google Scholar 

  • Moore D, Higgins PM (1997) Viability of stored conidia of Metarhizium flavoviride Gams and Rozsypal, produced under different culture regimes and stored with clays. Biocontrol Sci Technol 7:335–343

    Article  Google Scholar 

  • Moore D, Lord JC, Smith SM (2000) Pathogens. In: Subramanyam Bh, Hagstrum DW (eds) Alternatives to pesticides in stored-product IPM. Kluwer Academic Publishers, Dordrecht, pp 193–227

    Chapter  Google Scholar 

  • Mullins ED, Chen X, Romaine P, Rain R, Geiser DM, Kang S (2001) Agrobacterium-mediated transformation of Fusarium oxysporum: an efficient tool for insertional mutagenesis and gene transfer. Phytopathology 91:173–180

    Article  PubMed  CAS  Google Scholar 

  • SAS Institute (1995) SAS user’s guide: statistics. SAS Institute, Gary

    Google Scholar 

  • Smith RJ, Grula EA (1981) Nutritional requirements for conidial germination and hyphal growth of Beauveria bassiana. J Invertebr Pathol 37:222–230

    Article  CAS  Google Scholar 

  • Sokal RR, Rohlf FJ (1995) Biometry, 3rd edn. Freedman and Company, New York

    Google Scholar 

  • Sosa-Gomez DR, Boucias DG, Nation JL (1997) Attachment of Metarhizium anisopliae to the southern green stinkbug Nezara viridula cuticle and fungistatic effect of cuticular lipids and aldehydes. J Invertebr Pathol 69:31–39

    Article  PubMed  CAS  Google Scholar 

  • Spellig T, Bottin A, Kahmann R (1996) Green fluorescent protein (GFP) as a new vital marker in the phytopathogenic fungus Ustilago maydis. Mol Gen Genet 252:503–509

    PubMed  CAS  Google Scholar 

  • Vanden Wymelenberg AJ, Cullen D, Spear RN, Schoenike B, Andrews JH (1997) Expression of green fluorescent protein in Aureobasidium pullulans and quantification of the fungus on leaf surfaces. Biotechniques 23:686–690

    PubMed  CAS  Google Scholar 

  • Vassilakos TN, Athanassiou CG, Kavallieratos NG, Vayias BJ (2006) Influence of temperature on the insecticidal effect of Beauveria bassiana in combination with diatomaceous earth against Rhyzopertha dominica and Sitophilus oryzae on stored wheat. Biol Control 38:270–281

    Article  Google Scholar 

  • Vayias BJ, Athanassiou CG (2004) Factors affecting efficacy of the diatomaceous earth formulation SilicoSec against adults and larvae of the confused beetle Tribolium confusum Du Val (Coleoptera: Tenebrionidae). Crop Prot 23:565–573

    Article  CAS  Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Laboratory of Agricultural Zoology and Entomology, Agricultural University of Athens, Iera Odos 75, 11855, Athens, Greece

    V. K. Stephou & C. G. Athanassiou

  2. Laboratory of Plant Pathology, Agricultural University of Athens, Iera Odos 75, 11855, Athens, Greece

    S. E. Tjamos & E. J. Paplomatas

  3. Department of Agriculture, Plant Production and Rural Environment, Laboratory of Entomology and Agricultural Zoology, University of Thessaly, Phytokou Str, 38443, Nea Ionia, Magnesia, Greece

    C. G. Athanassiou

Authors
  1. V. K. Stephou
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. S. E. Tjamos
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. E. J. Paplomatas
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. C. G. Athanassiou
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to C. G. Athanassiou.

Additional information

A contribution to the Special Issue on Recent Advances in Stored-Product Protection.

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Stephou, V.K., Tjamos, S.E., Paplomatas, E.J. et al. Transformation and attachment of Beauveria bassiana conidia on the cuticle of Tribolium confusum and Sitophilus oryzae in conjunction with diatomaceous earth. J Pest Sci 85, 387–394 (2012). https://doi.org/10.1007/s10340-012-0428-z

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s10340-012-0428-z

Keywords

  • Beauveria bassiana
  • Tribolium confusum
  • Sitophilus oryzae
  • Diatomaceous earth
  • Entomopathogenic fungi
  • Conidial transformation
  • GFP conidia

Search

Navigation