Genetically Modified Beauveria bassiana Strains Harboring the Isaria fumosorosea Protease Gene Exhibit Increased Virulence against Dendrolimus punctatus

摘要

In this study, we cloned the complete full-length cDNA of Ifupr1 from the biocontrol agent Isaria fumosorosea using SMART rapid amplification of cDNA ends reverse transcription polymerase chain reaction. Our results showed that the Ifpr1 gene contained an open reading frame (ORF) with 1287 bp encoding 428 amino acids; the N-terminal 16 amino acid residues displayed the characteristics of signal peptide. The mature protease had a molecular mass of 46.3 kDa and a calculated isoelectric point of 7.37. The protein sequence contained two highly conserved regions, a putative enzymatic active site, and a potential chitin-binding domain. The Ifupr1 gene from I. fumosorosea was engineered into Beauveria bassiana by blastospore transformation. Compared with the wild-type strain Bb13, the protease activity was increased by 2.65-fold after induction for 36 h. In bioassays against the larvae of Masson’s pine caterpillar Dendrolimus punctatus, the median lethal time of the transgenic strain was reduced by 33.7% (P< 0.05), and mortality rates were increased by 53.3% (P< 0.05) as compared with the wild-type at a concentration of 1 × 107 spores/mL. The median lethal concentrations of the wild-type and transgenic isolates were 4.55 × 106 and 0.869 × 106 spores/mL, respectively. Taken together, these findings showed that the transgenic strain exhibited significantly improved virulence against D. punctatus.