Dataset Information

Selection and characterization of Bacillus thuringiensis strains from northwestern Himalayas toxic against Helicoverpa armigera.

ABSTRACT: In this study, we present the selection and the characterization of Bacillus thuringiensis (Bt) strains with respect to their cry/cyt gene content and toxicity evaluation toward one of the most important polyphagous lepidopteran pest, Helicoverpa armigera. Fifty-six Bt isolates were obtained from 10 different regions of northwestern Himalayas, recording a total B. thuringiensis index of 0.62. Scanning electron microscopy revealed presence of bipyramidal, spherical, flat and irregular crystal shapes; SDS-PAGE analysis of spore-crystal mixtures showed the prominence of 130, 70, and 100 kDa protein bands in majority of the isolates; PCR analysis with primers for eight cry and cyt gene families and 13 cry gene subfamilies resulted in isolates showing different combinations of insecticidal genes. Strains containing cry1 were the most abundant (57.1%) followed by cyt2 (46.42%), cry11 (37.5%), cry2 (28.57%), cry4 (21.42%), cyt1 (19.64%), cry3 (8.9%), and cry7, 8 (7.14%). A total of 30.35% of the strains did not amplify with any of the primers used in this study. Median lethal concentration 50 (LC₅₀ ) estimates of spore-crystal mixtures of Bt-JK12, 17, 22, 48, and 72 against second instar larvae of H. armigera was observed to be 184.62, 275.39, 256.29, 259.93 ?g ml^-1 , respectively. B. thuringiensis presents great diversity with respect to the presence of crystal protein encoding genes and insecticidal activity. Four putative toxic isolates identified in this study have potential application in insect pest control. B. thuringiensis isolate JK12 exhibited higher toxicity against H. armigera than that of B. thuringiensis HD1, hence can be commercially exploited to control insect pest for sustainable crop production. The results of this study confirm the significance of continuous exploration of new Bt stains from different ecological regions of the world.

SUBMITTER: Lone SA

PROVIDER: S-EPMC5727364 | biostudies-literature | 2017 Dec

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Selection and characterization of Bacillus thuringiensis strains from northwestern Himalayas toxic against Helicoverpa armigera.

Lone Showkat A SA Malik Abdul A Padaria Jasdeep C JC

MicrobiologyOpen 20171018 6

In this study, we present the selection and the characterization of Bacillus thuringiensis (Bt) strains with respect to their cry/cyt gene content and toxicity evaluation toward one of the most important polyphagous lepidopteran pest, Helicoverpa armigera. Fifty-six Bt isolates were obtained from 10 different regions of northwestern Himalayas, recording a total B. thuringiensis index of 0.62. Scanning electron microscopy revealed presence of bipyramidal, spherical, flat and irregular crystal sha ...[more]

PMID: 29047221

Similar Datasets

Project description:Helicoverpa armigera is a major insect pest of several crops worldwide. This insect is susceptible to some Bacillus thuringiensis (Bt) Cry insecticidal proteins expressed in transgenic crops or used in biopesticides. Previously, we identified H. armigera prohibitin (HaPHB) as a Cry1Ac-binding protein. Here, we further analyzed the potential role of PHB as a Cry toxin receptor in comparison to cadherin (CAD), well recognized as a Cry1Ac receptor. HaPHB-2 midgut protein and HaCAD toxin-binding region (TBR) fragment from H. armigera were expressed in Escherichia coli cells, and binding assays with different Cry1 toxins were performed. We demonstrated that Cry1Ab, Cry1Ac, and Cry1Fa toxins bound to HaPHB-2 in a manner similar to that seen with HaCAD-TBR. Different Cry1Ab mutant toxins located in domain II (Cry1AbF371A and Cry1AbG439D) or domain III (Cry1AbL511A and Cry1AbN514A), which were previously characterized and found to be affected in receptor binding, were analyzed regarding their binding interaction with HaPHB-2 and toxicity against H. armigera One β-16 mutant (Cry1AbN514A) showed increased binding to HaPHB-2 that correlated with 6-fold-higher toxicity against H. armigera, whereas the other β-16 mutant (Cry1AbL511A) was affected in binding to HaPHB-2 and lost toxicity against H. armigera Our data indicate that β-16 from domain III of Cry1Ab is involved in interactions with HaPHB-2 and in toxicity. This report identifies a region of Cry1Ab involved in binding to HaPHB-2 from a Lepidoptera insect, suggesting that this protein may participate as a novel receptor in the mechanism of action of the Cry1 toxins in H. armigeraIMPORTANCEHelicoverpa armigera is a polyphagous pest that feeds on important crops worldwide. This insect pest is sensitive to different Cry1 toxins from Bacillus thuringiensis In this study, we analyzed the potential role of PHB-2 as a Cry1 toxin receptor in comparison to CAD. We show that different Cry1 toxins bound to HaPHB-2 and HaCAD-TBR similarly and identify β-16 from domain III of Cry1Ab as a binding region involved in the interaction with HaPHB-2 and in toxicity. This report characterized HaPHB-Cry1 binding interaction, providing novel insights into potential target sites for improving Cry1 toxicity against H. armigera.

Project description:Midgut receptors play a critical role in the specificity of Cry toxins for individual insect species. Cadherin proteins are essential putative receptors of Cry1A toxins in lepidopteran larvae. Cry2A family members share common binding sites in Helicoverpa armigera, and one of them, Cry2Aa, has been widely reported to interact with midgut cadherin. Here, we studied the binding interaction and functional role of H. armigera cadherin in the mechanism of Cry2Ab toxicity. A region spanning from cadherin repeat 6 (CR6) to the membrane-proximal region (MPR) of cadherin protein was produced as six overlapping peptides to identify the specific binding regions of Cry2Ab. Binding assays showed that Cry2Ab binds nonspecifically to peptides containing CR7 and CR11 regions in a denatured state but binds specifically only to CR7-containing peptides in the native state. The peptides CR6-11 and CR6-8 were transiently expressed in Sf9 cells to assess the functional role of cadherin. Cytotoxicity assays showed that Cry2Ab is not toxic to the cells expressing any of the cadherin peptides. However, ABCA2-expressing cells showed high sensitivity to Cry2Ab toxin. Neither increased nor decreased sensitivity to Cry2Ab was observed when the peptide CR6-11 was coexpressed with the ABCA2 gene in Sf9 cells. Instead, treating ABCA2-expressing cells with a mixture of Cry2Ab and CR6-8 peptides resulted in significantly reduced cell death compared with treatment with Cry2Ab alone. Moreover, silencing of the cadherin gene in H. armigera larvae showed no significant effect on Cry2Ab toxicity, in contrast to the reduced mortality in ABCA2-silenced larvae. IMPORTANCE To improve the efficiency of production of a single toxin in crops and to delay the evolution of insect resistance to the toxin, the second generation of Bt cotton, expressing Cry1Ac and Cry2Ab, was introduced. Understanding the mode action of the Cry proteins in the insect midgut and the mechanisms insects use to overcome these toxins plays a crucial role in developing measures to counter them. Extensive studies have been conducted on the receptors of Cry1A toxins, but relatively little has been done about those of Cry2Ab. By showing the nonfunctional binding of cadherin protein with Cry2Ab, we have furthered the understanding of Cry2Ab receptors.

Dataset Information

Selection and characterization of Bacillus thuringiensis strains from northwestern Himalayas toxic against Helicoverpa armigera.

Publications

Selection and characterization of Bacillus thuringiensis strains from northwestern Himalayas toxic against Helicoverpa armigera.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets