Isolation, identification and evaluation of mosquito entomopathogenic Bacillus species and related genera from randomly selected sites in Kenya

This study was carried out to identify specific mosquitocidal Bacillus species and related genera for future development of biopesticides in local mosquito control program in Kenya. Bacterial isolation was conducted from 100 soil samples through pasteurization method and preliminary identification conducted through phenotypical analysis. Toxicity analysis was performed through bioassays and lethal concentrations (LC) were determined using probit analysis. Toxic isolates were further identified through analysis of the 16s rRNA and screening of toxin genes through PCR. Expression of toxin proteins was performed using SDS-PAGE. Out of 453 isolates, 7 of them were found to yield highly potent toxicity (>50% mortality) against Culex quinquefasciatus during the initial toxicity assays. Among them, two isolates KDHa3 and SKDHb5, with LC50 values of 0.007mg/L and 0.008mg/L, respectively, were the most toxic against the target. Phylogenetic analysis based on 16s rRNA showed high homology to Lysinibacillus sphaericus (six isolates) and Bacillus thuringiensis (one isolate). Various toxin genes encoding BinA, BinB, Mtx (1, 2 and 3), Cry48A, Cry49A, Cry4A, Cry11A and Cyt1A were detected among the isolates. The protein profiles using SDS-PAGE were consistent with the standard strains Lysinibacillus sphaericus C3-41 and B. thuringiensis var. israelensis. Native toxic Bacillus species and related genera were identified with this study being the first to report highly toxic strains of L. sphaericus and B. thuringiensis strains from Kenyan soil samples. Key words: Bacillus species, Bacillus thuringiensis, Culex quinquefasciatus, Entomopathogenic, Lysinibacillus sphaericus, toxin.