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Characterization of Bacillus thuringiensis Strain DOR4 Toxic to Castor Semilooper Achaea janata: Proteolytic Processing and Binding of Toxins to Receptors

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Abstract

We have isolated a strain of Bacillus thuringiensis (Bt) from Indian soil samples that was shown to be toxic to Achaea janata larvae. The isolate, named B. thuringiensis DOR4, serotypically identified with the standard subspecies kurstaki (H3a3b3c) and produced bipyramidal inclusions along with an amorphous type. Although the plasmid pattern of DOR4 was different from that of the reference strain, a crystal protein profile showed the presence of two major bands (130 and 65 kDa) similar to those of Bt subsp. kurstaki HD-1. To verify the cry gene content of DOR4, triplex PCR analysis was performed; it showed amplification of the cry1C gene in addition to cry1Aa, cry1Ac, cry2A, and cry2B genes, but not the cry1Ab gene. RT-PCR analysis showed the expression of cry1Aa and cry1Ac genes. In vitro proteolysis of DOR4 protoxin with midgut extract generated products of different sizes. Zymogram analysis of DOR4 protoxin as substrate pointed to a number of distinct proteases that were responsible for activation of protoxins. Furthermore, toxin overlay analysis revealed the presence of multiple toxin-binding proteins in midgut epithelium. Based on all these characterizations, we suggest that the Bt DOR4 strain can be exploited for an A. janata control program.

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Acknowledgments

This work was partly supported by a research grant from the Department of Biotechnology (DBT), the Government of India (Sanction No. BT/PR2745/AGR/02/141/2001). MB and GM are recipients of a fellowship from the Council for Scientific and Industrial Research (CSIR), India.

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Authors and Affiliations

  1. School of Life Sciences, University of Hyderabad, Hyderabad, 500046, India

    Madhusudhan Budatha, P. B. Kirti & Aparna Dutta-Gupta

  2. Department of Pharmacology, University of Cambridge, Tennis Court Road, Cambridge, CB2 1PD, UK

    Gargi Meur

  3. Directorate of Oil Seeds Research, Rajendra Nagar, Hyderabad, 500030, India

    P. S. Vimala Devi

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  1. Madhusudhan Budatha
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  2. Gargi Meur
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  3. P. S. Vimala Devi
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  4. P. B. Kirti
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  5. Aparna Dutta-Gupta
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Correspondence to Aparna Dutta-Gupta.

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Budatha, M., Meur, G., Vimala Devi, P.S. et al. Characterization of Bacillus thuringiensis Strain DOR4 Toxic to Castor Semilooper Achaea janata: Proteolytic Processing and Binding of Toxins to Receptors. Curr Microbiol 57, 72–77 (2008). https://doi.org/10.1007/s00284-008-9155-3

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  • DOI: https://doi.org/10.1007/s00284-008-9155-3

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