Unraveling the abundance of vip3-type genes in Indian Bacillus thuringiensis across the agroclimatic landscape and impact of amino acid substitutions for safer agriculture

Vegetative insecticidal protein (vip) genes of Bacillus thuringiensis (Bt) are candidates for gene pyramiding in the resistance management of pests. The prevalence of vip genes in Bt isolates is relatively under-explored. Bt isolates recovered from 29 diverse sources in nine agro-climatic zones of India were screened for the presence of vip3-type genes by PCR with 4 sets of oligonucleotide primers. Out of 155 Bt isolates, 70.32 % (109) and 44.52 % (69) isolates were positive for amplification of partial vip3-type genes with primer sets 1 and 4, respectively. The primer set-2 was found to be more efficient for amplifying full-length genes (29.03 % /45 isolates) as compared with primer set-3 (3.23 %/ 5 isolates), also corroborated in the amplification of full-length vip3 genes in ten Bt BGSC strains used as reference. Frequency analysis revealed presence of vip3 genes in Bt isolates across all agro-climatic zones. Thus, Indian Bt isolates from diverse sources have a rich repertoire of vip3-type genes. Our study reports the highest number (45) of full-length vip3-type genes detected in a native Bt isolates collection, demonstrating enrichment of Indian Bt isolates for vip3 genes. Twelve of these genes have been cloned, sequenced, and out of these, six were found to be effective against Helicoverpa armigera in our laboratory previously. Comparison of substitutions in deduced amino acids sequence of these genes and expression of Vip3 proteins in SDS-PAGE analysis of selected native Bt isolates positive for full-length vip3-type genes indicated their biopesticidal potential.