Screening Of Bacillus Thuringiensis Strains Of Strong Virulence Against Lepidopterous Pest

Bt as a microbial insecticide, harmless to environment, showing a great prospect, plays an important role for the green plant protection. Pest control is an important task in the agricultural production. Besides as pesticides, the Bt toxin genes were also applied to transfer into crops to develop insect-resistant crops. Now, transgenic cotton with crylA gene has been commercialized widely in China, resulting in solution to the hazards of cotton bollworm. However, with the large-scale cultivation of transgenic insect-resistant crops, the risk of resistace from insects is more and more serious, and the challenges are severe for insect pests control.To screen high active and no cross-resistance strains with crylA class of genes for the development and application of insecticide, based on the sequences of cry gene which has no cross-resistance to crylA class,8primers were designed for192laboratory-save strain resources for lepidopteran pests and efficient screening. Comparison of plasmid profiles of its diversity. The insecticidal activity of Bt strains of the beet armyworm, diamondback moth larvae were further determined,15high-virulence strains were filtered out.Fifteen high-virulence strains were divided into8types according to large plasmid profile analysis; they contained several gene types:cry1B, cry1C, cry1D, cry11, cry2A and cry9genes according to gene classical identification, and shaped biconical square and spherical crystal by electronic scanning. Most of the fifteen expressed protein with the molecular weight of130ku, except the protein with the molecular weight of both60ku and130ku expressed by one strain, another expressed protein with the molecular weight of both70ku and130ku were analyzed by SDS-PAGE. The concentration of crystal protein in the mixtures of spores and crystals was determined.The bioassays of eight types strains against Spodoptera exigua, Plutella xylostella, Helicoverpa armigera Cry1Ac-susceptible and resistant strain with the production strains KN11as control showed that PS4-E12strain was nearly4times higher toxicity against P. xylostella than control strain KN11; three strains of the eight showed strong toxicity against S. exigua, and the LC50of PS3-C2was1.87μg·mL-1; PS4-E12, PS1-F11, SCL-B3and PS8-B6strain exhibited high insecticidal activity against H. armigera Cry1Ac-susceptible strain, while PS3-C2, PS1-F11, SCL-B3, PS8-B6, PS4-E12and MO3-E5were no cross-resistance strains against H. armigera Cry1Ac-resistant strain, but the results need further validation.All of eight types Bt strains contained chitinase genes identified by PCR with primers were designed according to the dates on GenBank.The growing status of each strain at same time were determined, and the results showed that there was no significant difference on growth speed among8strains. Bt strains were evaluated in strain feature, growth speed, active spore number, insecticidal activity and so on in this study. The results will show great importance on isolation and exploration of new Bt strains.