Comprehensive genomic analysis and functional characterization of a new entomopathogenic strain, Bacillus thuringiensis subsp. israelensis VCRC B646, antagonistic to mosquito vectors (Diptera: Culicidae), isolated from soil in Tamil Nadu, India

BACKGROUND Mosquito‐borne diseases such as malaria, dengue and filariasis remain major public health threats, demanding effective and environmentally‐safe control measures. Bacillus thuringiensis subsp. israelensis (Bti) is widely employed as a biopesticide owing to its Cry and Cyt δ‐endotoxins. Continuous isolation and genomic characterization of potent indigenous strains are crucial for strengthening vector control strategies. This study aimed to assess the larvicidal efficacy, genomic features, plasmid architecture and environmental safety of a novel Indian isolate, Bti VCRC B646. RESULTS Bti VCRC B646 (JBAMKE010000000) exhibited potent larvicidal activity against Aedes aegypti , Anopheles stephensi and Culex quinquefasciatus , with median lethal concentrations (LC₅₀) values of 0.0067, 0.0085 and 0.0072 μg mL −1 , respectively, and LC₉₀ values of 0.0112, 0.0146 and 0.0129 μg mL −1 . Whole‐genome sequencing revealed a 6.6‐Mbp genome (34.83% GC) comprising a chromosome and seven plasmids, including one harboring insecticidal toxin genes (Cry4A, Cry4B, Cry11A, Cry10Aa, Cry15A, Cyt1, Cyt2). Comparative genomics (ANI, dDDH, phylogeny) confirmed its identity as Bti , closely related to the WHO‐reference strain Bti AM65‐52. SDS‐PAGE and toxin gene mining validated δ‐endotoxins and accessory chitinases and metalloproteases. Secondary metabolite biosynthetic clusters, phage elements and mobile genetic elements were identified, whereas no major human pathogenic markers were detected. Nontarget aquatic organisms remained unaffected at ≤30‐fold LC₉₀ doses. CONCLUSIONS Bti VCRC B646 demonstrates exceptional larvicidal potency and genomic richness, with plasmid‐borne δ‐endotoxins and biosynthetic versatility. Its strong efficacy, safety profile and ecological adaptability underscore its potential as a next‐generation, sustainable microbial larvicide for integrated vector management.