Current Microbiology

, Volume 57, Issue 3, pp 175–180 | Cite as

Use of a Cry1Ac-Resistant Line of Helicoverpa armigera (Lepidoptera: Noctuidae) to Detect Novel Insecticidal Toxin Genes in Bacillus thuringiensis

  • Cheryl E. Beard
  • Leon Court
  • Roslyn G. Mourant
  • Bill James
  • Jeroen Van Rie
  • Luke Masson
  • Raymond J. Akhurst


This paper describes a screening strategy incorporating resistant insect lines for discovery of new Bacillus thuringiensis toxins against a background of known genes that would normally mask the activity of additional genes and the application of that strategy. A line of Helicoverpa armigera with resistance to Cry1Ac (line ISOC) was used to screen Cry1Ac-expressing strains of B. thuringiensis for additional toxins with activity against H. armigera. Using this approach, a number of Cry1Ac-producing strains with significant toxicity toward Cry1Ac-resistant H. armigera were identified. When the insecticidal protein complement of one of these strains, C81, was examined in detail, a novel cry2 gene (cry2Af1) was detected.


Bacillus Thuringiensis Cry1Ab Protein cry2Ab Gene cry1Ac Gene Cry1Ac Protein 
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  1. 1.
    Akhurst RJ, James WJ (1999) Helicoverpa armigera resistance to transgenic cotton expressing the Cry1Ac δ-endotoxin of B. thuringiensis. In: Yu S, Sun M, Liu Z (eds) Biotechnology of Bacillus thuringiensis. Vol 3. Science Press, Beijing, p 200Google Scholar
  2. 2.
    Akhurst RJ, James W, Bird LJ, Beard C (2003) Resistance to the Cry1Ac δ-endotoxin of Bacillus thuringiensis in the cotton bollworm, Helicoverpa armigera (Lepidoptera: Noctuidae). J Econ Entomol 96:1290–1299PubMedCrossRefGoogle Scholar
  3. 3.
    Arantes O, Lereclus D (1991) Construction of cloning vectors for Bacillus thuringiensis. Gene 108:115–119PubMedCrossRefGoogle Scholar
  4. 4.
    Bah A, van Frankenhuyzen K, Brousseau R, Masson L (2004) The Bacillus thuringiensis Cry1Aa toxin: effects of trypsin and chymotrypsin site mutations on toxicity and stability. J Invert Pathol 85:120–127CrossRefGoogle Scholar
  5. 5.
    Bird LJ, Akhurst RJ (2004) Relative fitness of Cry1A-resistant and -susceptible Helicoverpa armigera (Lepidoptera: Noctuidae) on conventional and transgenic cotton. J Econ Entomol 97:1699–1709PubMedGoogle Scholar
  6. 6.
    Bird LJ, Akhurst RJ (2007) Variation in susceptibility of Helicoverpa armigera (Hübner) and Helicoverpa punctigera (Wallengren) (Lepidoptera: Noctuidae) in Australia to two Bacillus thuringiensis toxins. J Invertbr Pathol 94:84–94CrossRefGoogle Scholar
  7. 7.
    Carozzi NB, Kramer VC, Warren GW, Evola S, Koziel MG (1991) Prediction of insecticidal activity of Bacillus thuringiensis strains by polymerase chain reaction product profiles. Appl Environ Microbiol 57:3057–3061PubMedGoogle Scholar
  8. 8.
    Crickmore N, Bone EJ, Williams JA, Ellar DJ (1995) Contribution of the individual components of the δ-endotoxin crystal to the mosquitocidal activity of Bacillus thuringiensis subsp. israelensis. FEMS Microbiol Lett 131:249–254Google Scholar
  9. 9.
    Dankocsik C, Donovan WP, Jany CS (1990) Activation of a cryptic crystal protein gene of Bacillus thuringiensis subspecies kurstaki by gene fusion and determination of the crystal protein insecticidal specificity. Mol Microbiol 4:2087–2094PubMedCrossRefGoogle Scholar
  10. 10.
    Donovan WP, Donovan JC, Engleman JT (2001) Gene knockout demonstrates that vip3A contributes to the pathogenesis of Bacillus thuringiensis toward Agrotis ipsilon and Spodoptera exigua. J Invertebr Pathol 78:45–51PubMedCrossRefGoogle Scholar
  11. 11.
    Höfte H, Van Rie J, Jansens S, Van Houtven A, Vanderbruggen H, Vaeck M (1988) Monoclonal antibody analysis and insecticidal spectrum of three types of Leptidopteran-specific insecticidal crystal proteins of Bacillus thuringiensis. Appl Environ Microbiol 54:2010–2017PubMedGoogle Scholar
  12. 12.
    Höfte H, Whiteley HR (1989) Insecticidal crystal proteins of Bacillus thuringiensis. Microbiol Rev 53:242–255PubMedGoogle Scholar
  13. 13.
    Jain DS, Udayasuriyan V, Arulsevi PI, Dev SS, Sangeetha P (2006) Cloning, characterization, and expression of a new cry2Ab gene from Bacillus thuringiensis strain 14–1. Appl Biochem Biotechnol 128:185–194PubMedCrossRefGoogle Scholar
  14. 14.
    Johnson DE, McGaughey WH (1996) Contribution of Bacillus thuringiensis spores to toxicity of purified Cry proteins towards Indianmeal moth larvae. Curr Microbiol 33:54–60PubMedCrossRefGoogle Scholar
  15. 15.
    Kranthi KR, Kranthi S Ali S, Banerjee SK (2000) Resistance to Cry1Ac δ-endotoxin of Bacillus thuringiensis in a laboratory selected strain of Helicoverpa armigera (Hübner). Curr Sci 78:1001–1004Google Scholar
  16. 16.
    Liang G, Tan W, Guo Y (2000) Study on screening and inheritance mode of resistance to Bt transgenic cotton in H. armigera. Acta Entomol Sin 43(Suppl):57–62Google Scholar
  17. 17.
    Liao C, Heckel DG, Akhurst R (2002) Toxicity of Bacillus thuringiensis insecticidal proteins for Helicoverpa armigera and Helicoverpa punctigera (Lepidoptera: Noctuidae), major pests of cotton. J Invertebr Pathol 80:55–63PubMedCrossRefGoogle Scholar
  18. 18.
    Liu Y-B, Tabashnik BE, Moar WJ, Smith RA (1998) Synergism between Bacillus thuringiensis spores and toxins against resistant and susceptible diamondback moths (Plutella xylostella). Appl Environ Microbiol 64:1385–1389PubMedGoogle Scholar
  19. 19.
    Mahon RJ, Olsen KM, Downes S, Addison S (2007) Frequency of alleles conferring resistance to the Bt toxins Cry1Ac and Cry2Ab in Australian populations of Helicoverpa armigera (Hübner) (Lepidoptera: Noctuidae). J Econ Entomol 100:1844–1853Google Scholar
  20. 20.
    Masson L, Erlandson M, Puzstai-Carey M, Brousseau R, Juárez-Pérez V, Frutos R (1998) A holistic approach for determining the entomopathogenic potential of Bacillus thuringiensis strains. Appl Environ Microbiol 64:4782–4788PubMedGoogle Scholar
  21. 21.
    Poncet S, Delécluse A, Klier A, Rapoport G (1995) Evaluation of synergistic interactions among the CryIVA, CryIVB and CryIVD toxic components of B. thuringiensis subsp. israelensis crystals. J Invertebr Pathol 66:131–135CrossRefGoogle Scholar
  22. 22.
    Rajamohan F, Alzate O, Cotrill JA, Curtiss A, Dean DH (1996) Protein engineering of Bacillus thuringiensis δ-endotoxin: Mutations at domain II of Cry1Ab enhance receptor affinity and toxicity toward gypsy moth larvae. Proc Natl Acad Sci USA 93:14338–14343PubMedCrossRefGoogle Scholar
  23. 23.
    Sambrook J, Fritch EF, Maniatis T (1989) Molecular cloning: a laboratory manual. 2nd ed. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NYGoogle Scholar
  24. 24.
    Wu S-J, Koller CN, Miller DL, Bauer LS, Dean DH (2000) Enhanced toxicity of Bacillus thuringiensis Cry3A δ-endotoxin in coleopterans by mutagenesis in a receptor binding loop. FEBS Lett 473:227–232PubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC 2008

Authors and Affiliations

  • Cheryl E. Beard
    • 1
  • Leon Court
    • 1
  • Roslyn G. Mourant
    • 1
  • Bill James
    • 1
  • Jeroen Van Rie
    • 2
  • Luke Masson
    • 3
  • Raymond J. Akhurst
    • 1
  1. 1.CSIRO EntomologyCanberraAustralia
  2. 2.Bayer BioScience N.VGentBelgium
  3. 3.National Research Council of Canada, Biotechnology Research InstituteQuébecCanada

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