Isolation and Characterization of Indigenous Endotoxin-Forming Bacillus sp. with Insecticidal Activity from Northern Iranian Soil

The main goal of the current study was to isolate and characterize endospore-forming soil bacteria that produce parasporal crystalline proteins against larvae of Lepidoptera insects during stationary phase of growth. Two different methods were applied for the isolation of spore-forming strains. Polymerase Chain Reaction (PCR) was used for the characterization of cry gene content of the isolated strains and 16S rRNA sequencing was carried out to identify bacterial strains. Efficiency of the isolates as insecticide was evaluated in a bioassay experiment using insect's larvae. Regarding presence of parasporal inclusion crystals and SDS-PAGE patterns, 10 bacterial strains were isolated from about 200 soil samples. Analysis of crystal-spore mixtures with SDS-PAGE showed a broad range of proteins with molecular weight between 11-230 kDa. Data from PCR analysis indicated that only two isolates (RIPI6 and RIPI18) may contain cry3 gene. Isolate RIPI21 and reference strain (Bacillus thuringiensis subsp. kurstaki) were positive for cry1 gene. 16S rRNA gene sequences of all isolates showed at least 96% sequence match with B. thuringiensis strains deposited in the GenBank. The results of bioassay experiments showed the efficacy of strains RIPI7, 10 and 22 on killing of larvae of both Anagasta kuehniella Zeller and Plutella xylostella Curt. It is concluded that there are some bacterial candidates for biological control of major agricultural pests in north of Iran.