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Combined analysis of supernatant-based feeding bioassays and PCR as a first-tier screening strategy for Vip -derived activities in Bacillus thuringiensis strains effective against tropical fall armyworm

Combined analysis of supernatant-based feeding bioassays and PCR as a first-tier screening strategy for Vip -derived activities in Bacillus thuringiensis strains effective against tropical fall armyworm

Journal of Applied Microbiology 93(2): 269-277

ISSN/ISBN: 1364-5072

PMID: 12147075

DOI: 10.1046/j.1365-2672.2002.01694.x

Aims: To assess whether feeding bioassays using culture-supernatant proteins could be combined with PCR into a first-tier screening strategy for Vip3A-like genes efficient against tropical Spodoptera frugiperda. Methods and Results: Out of 12 Bacillus thuringiensis strains studied, the total protein concentrated from the culture supernatant of only the strain HD125 yielded a significantly increased armyworm mortality and an intense band of the predicted size for VIP3A protein in SDS-PAGE. However, PCR and sequencing data indicated Vip-like genes are ubiquitous in tropical B. thuringiensis isolates. Interestingly, the HD125 strain was also the only one displaying a single-band amplification pattern and the highest sequence identity to the reported Vip3A(a) gene. Conclusions: Results suggest the insecticidal effectiveness of putative VIPs in B. thuringiensis isolates can be preliminarily estimated by the use of supernatant-derived total protein in feeding experiments, though only in a limited manner. Significance and Impact of the Study: A simple and cost-effective first-tier screening strategy for VIP-derived activities in B. thuringiensis collections can be developed by combining PCR and feeding bioassays. Moreover, the employed primers showed to be useful as a tool for strains differentiation at DNA level, and for characterization and isolation of Vip-like genes in tropical B. thuringiensis germplasm.

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Accession: 003683487

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