Characterization of entomotoxic and nematotoxic genes from indigenous Bacillus thuringiensis strains and their biocontrol potential

Abstract

Abstract Background The gram pod borer (Helicoverpa armigera Hubner), Bihar hairy caterpillar (Spilosoma obliqua Walker), tobacco caterpillar (Spodoptera litura Fabricius) and root knot nematode (Meloidogyne incognita (Kofoid and White) Chitwood) are the major insect pest and nematodes infesting mungbean, urdbean and other legumes. Bacillus thuringiensis (Bt) is a promising biocontrol agent that helps in managing the above pest. Deploying Bt biopesticides instead of chemical pesticides in legume cultivation enhances the soil health substantially. The present study envisages morphological, molecular cum biochemical characterization, entomotoxicity and nematotoxicity potential of four indigenous Bt strains (Ak2.IIPR, F8.IIPR, F5.IIPR and F6.IIPR) along with HD1 (standard check from BGSC). Results The SEM micrograph analysis of above four Bt strains along with F1.IIPR revealed the presence of different combinations of insecticidal crystal proteins, viz. cuboidal crystal (CC), bipyramidal crystal (BC), spherical crystal (SC) and irregularly pointed crystal (IC). This was further confirmed by PCR amplification of Cry1, Cry2Aa, Cry2Ac, VIP3A and chi36 gene-specific primers and their phylogenetic analysis. F8.IIPR and Ak2.IIPR recorded the highest toxicity index against second-instar S. obliqua larvae (292.56 and 174.55), H. armigera (150.00 and 113.65) and S. litura (210.71 and 114.19) based on probit analysis. F5.IIPR (91.67%) and F8.IIPR (89.00%) recorded the highest juvenile mortality against M. incognita, followed by Ak2.IIPR (70.33%). The protein profiling of supernatant of terrific broth inoculated with F8.IIPR indicated the presence of VIP3A and Cry1I. Conclusion The present study concluded that F8.IIPR and Ak2.IIPR were potential Bt strains isolated from Indo-Gangetic plains which is having both entomocidal and nematicidal properties. Further, these strains were deposited at national repository, ICAR-NBAIM with the following accession no.: NAIMCC-SB-065 and NAIMCC-SB-066, respectively. These strains can be formulated as microbial biopesticide.

Keywords

• Insecticidal genes
• Exochitinase gene
• Toxicity index
• Nematicidal genes
• Bt DNA gyrase
• SEM analysis