球孢白僵菌XJBB-01株Pr1丝氨酸蛋白酶高效表达菌株的构建及其对拉合尔钝缘蜱毒力测定OA

This study utilized Agrobacterium tumefaciens-mediated transformation and hygromycin re-sistance screening to introduce the Pr1 serine protease CDEP2 gene into Beauveria bassiana strain XJBB-01.Enzyme activity assays were conducted to obtain a high-efficiency expression strain of Pr1 serine protease in Beauveria bassiana.The virulence of strain Pr1 serine protease activity before and after transformation,as well as the spore suspension concentration of 1×107 CFU/mL,were measured at different stages of Or-nithodoros lahorensis.The results demonstrated that the engineered Beauveria bassiana strains Pr1-1 and Pr1-2 were successfully obtained through hygromycin resistance screening and PCR identification.Pheno-typic analysis of the two engineered strains revealed significantly higher colony growth rates and spore pro-duction compared to the XJBB-01 strain(P<0.05).Among them,the colony growth rates of Pr1-1 and Pr1-2 were 9.27 mm/d and 8.87 mm/d,respectively,which were 0.77 mm/d and 1.17 mm/d higher than those of strain XJBB-01.The spore production rates were 4.58×106 spores/cm2 and 5.22×106 spores/cm2,respectively,representing increases of 0.51×106 spores/cm2 and 1.15×106 spores/cm2 compared to strain XJBB-01.The enzyme activities of XJBB-01 strain and Pr1-1,Pr1-2 strains were meas-ured,with Pr1-1 and Pr1-2 showing enzyme activities of 34.13 U/mL and 35.58 U/mL,respectively,repre-senting significant increases of 5.46 U/mL and 6.91 U/mL compared to XJBB-01 strain(P<0.05).At 25℃and a spore suspension concentration of 1×107 CFU/mL,strains Pr1-1 and Pr1-2 exhibited signifi-cantly lower hatching rates of Ornithodoros lahorensis eggs compared to strain XJBB-01,with reductions of 23.3%and 26.6%,respectively.The mortality rates of nymphs increased by 17.4%and 20.7%,while the mortality rates of adult ticks rose by 16.7%and 23.3%.The median lethal time of larval ticks was accel-erated by 2.3 d and 2.5 d,respectively,while that of adult ticks was accelerated by 2.6 d and 3.3 d,show-ing significant differences(P<0.05).