关键词: 玫烟色棒束孢, 昆虫病原真菌, 遗传工程, 生物测定, 毒力, 马尾松毛虫

Abstract: Using SMART RACE RT-PCR, the complete full-length cDNA encoding a chitinase was cloned from the biocontrol agent Isaria fumosorosea. The gene, designated as Ifuchi1, contains an open reading frame (ORF) with 1 275 bp encoding 424 amino acids, which contains a N-terminal 24 amino acid residues displaying the characteristics of signal peptide. The mature chitinase has a molecular mass of 47.6 kDa and a calculated pI of 4.89. The protein sequence contains two highly conserved regions including a putative enzymatic active site and a potential chitin-binding domain. The chinitase belongs to the class V of family 18 of glycosyl hydrolases. The Ifuchi1 gene from I. fumosorosea was engineered into Beauveria bassiana by blastospore transformation. Compared to the wild-type strain Bb13, the chitinase activity of a selected transformant was increased by 86.2% after induction for 36 h. Bioassayed against the larvae of Masson’s pine caterpillar Dendrolimus punctatus, the median lethal time (LT₅₀) of transgenic isolates was reduced by 29%, and the mortality rate was increased by 52.9%, as compared to the wild-type at a concentration of 1×10⁷ conidia / mL. The median lethal concentration (LC₅₀) values of the wild-type and transgenic isolates were 4.71×10⁶ conidia / mL, and 1.74×10⁶ conidia / mL (approximately 1.7-fold decrease), respectively. The transgenic strain thus showed significantly improved virulence against D. punctatus.

Key words: Isaria fumosorosea, entomopathogenic fungi, genetic engineering, bioassay, virulence, Dendrolimus punctatus