Native Bacillus Strains Crude Culture Product Exhibits High Biocidal Activities Against Larvae and Adult Anopheles...

Anopheles mosquitoes are major vectors of malaria, a disease that continues to cause substantial morbidity and mortality worldwide. Bacillus-based insecticides are widely recognized as effective, environmentally friendly, and sustainable tools for mosquito control. Therefore, this study aimed to isolate native Bacillus strains from diverse environmental sources and evaluate their potential for mosquito control. Bacillus strains with high biocidal activity were isolated, identified, genetically characterized, and physiologically characterized. The larvicidal and adulticidal activities of culture supernatants and cell pellets were evaluated separately. Eight Bacillus strains exhibited strong biocidal activity in their culture supernatants at a concentration equivalent to 8 McFarland units (approximately 2.4 × 10⁹ CFU/mL) within 24 h. The culture supernatants showed significant larvicidal activity, with LC₅₀ and LC₉₅ values of 0.24 (95% CI: 0.01-0.69) CFU/mL and 1.57 (95% CI: 0.55-4.34) CFU/mL, respectively. The most active strains were identified by biochemical characterization and 16S rRNA gene sequencing. Laboratory bioassays demonstrated that the culture supernatants were effective against both larval and adult Anopheles mosquitoes. SDS-PAGE analysis revealed protein bands ranging from 15 to 130 kDa, including bands within molecular weight ranges commonly associated with putative Cry/Cyt-like proteins. These findings provide preliminary evidence of proteins that may be associated with the observed mosquitocidal activity. The most active Bacillus preparations also showed greater activity than clothianidin under the laboratory conditions tested. Overall, these findings highlight the potential of native Bacillus culture supernatants as environmentally friendly candidates for the development of biological mosquito control agents.