Effect And Toxic Constituents Of Bacillus Thuringiensis 010 With Nematicidal Activity

Application of chemicals against insect pests, although efficacious in most cases, has severe drawbacks related to the environmental pollution. Bacillus thuringiensis is spore-forming bacterium which had been used as an insecticide around the world. Recently, it had been found to have effect on plant disease as well. Therefore, it is necessary for researchers to screen new Bt strains with higher toxicity, and clone its novel action genes for GMOs.The free-living nematode Caenorhabditis elegans is an important pathogen in fungus and straw mushrooms, and being used increasingly as a model system for it could be bred laboratory. In this study, C. elegance was raised with Escherichia coli and Bt 010 stored in our laboratory was identified with significant activity in inhibiting the development of C. elegans. Crystal protein and chitinase genes were detected and the latter was confirmed by mass spectrum. Both sequences had been submitted to GenBank at accession numbers EF065179 and EU030625.Bioassay for 118 Bt isolates on this free-living nematode, we demonstrate that six of them are active against C. elegance. Within 36 hours, the LC₅₀ of the nematicidal toxins of Bt 010 is 0.498μg/ml. In addition, PCR was used to analyse genes of this insecticidal bacterium. Except cry 1Ia, we did not find any reported nematocidal cry genes. The changes of C. elegans its both on body wall and in intestine were observed through transmission electron microscope (TEM). These observations imply the significance of crystal protein from Bt 010 as a toxin against nematodes, and the degeneration of body wall might be caused by chitinase from Bt 010. Bioassays were also carried out with E. coli TB1 containing Bt crystal and chitinase genes. The results indicated that the chitinase gene from Bt 010 might be functional gene against C.elegans.Bt 010 could also target plant parasitical nematode Meloidogyne spp.and DNA ladder survey showed that the toxic protein did not damage the DNA of nematodes as chemical pesticides.In summary, our research work was focused on the following:Established a new technique for nematocidal bioassay;Confirmed the existence of the chitinase in Bt strain by MALDITOF-TOF-MS ;Firstly found the biological activity of chitinase from Bt strain against C. elegans .