Variable region of 16s rRNA is essential for the identification of Group 1 mosquito-pathogenic strains of Lysinibacillus

Abstract

A study was carried out to characterize and identify the potential mosquito-pathogenic bacilli strains isolated from the soil samples obtained from various locations in Goa, India. Six isolates were characterized morphologically, biochemically and phylogentically. Phylogenetic relationships of these strains were determined based on comparison of the 16S rRNA sequences with that of the closest Lysinibacillus species obtained from nucleotide database. Their mosquitocidal abilities were confirmed by bioassay and presence of genes responsible for pathogenicity and toxin production. Molecular characterization by partial sequencing of 16S rRNA gene has confirmed their identity as Lysinibacillus sp. Group 1. Comparison of the 16S rRNA sequences with that of the five groups of “Bacillus sphaericus like organisms”, re-designated off lately as Lysinibacillus, revealed that a variable region of 695 bp and hypervariable region of 22 bp were highly conserved among these five groups. These isolates were found effective against Culex quinquefasciatus larvae with LC50 values ranging from 0.018 to 0.58 ppm. The PCR amplification also showed the presence of binA, binB binary genes and mtx1, mtx2, mtx3 mosquitocidal genes. Larvicidal activity against Culex quinquefasciatus appeared related with expression of binary toxins. These strains were found more potent than the commercial strain and hence could be formulated for controlling Culex species transmitters of Japanese encephalitis and filariasis.