Cathepsin L-like Cysteine Proteinase Genes Are Associated with the Development and Pathogenicity of Pine Wood Nematode, Bursaphelenchus xylophilus

摘要

The pine wood nematode (PWN), Bursaphelenchus xylophilus , is the pathogen of pine wilt disease (PWD), resulting in huge losses in pine forests. However, its pathogenic mechanism remains unclear. The cathepsin L-like cysteine proteinase (CPL) genes are multifunctional genes related to the parasitic abilities of plant-parasitic nematodes, but their functions in PWN remain unclear. We cloned three cpl genes of PWN ( Bx-cpls ) by rapid amplification of cDNA ends (RACE) and analyzed their characteristics using bioinformatic methods. The tissue specificity of cpl gene of PWN ( Bx-cpl ) was studied using in situ mRNA hybridization (ISH). The functions of Bx-cpl s in development and pathogenicity were investigated using real-time quantitative PCR (qPCR) and RNA interference (RNAi). The results showed that the full-length cDNAs of Bx-cpl-1 , Bx-cpl-2 , and Bx-cpl-3 were 1163 bp, 1305 bp, and 1302 bp, respectively. Bx-cpl s could accumulate specifically in the egg, intestine, and genital system of PWN. During different developmental stages of PWN, the expression of Bx-cpl s in the egg stage was highest. After infection, the expression levels of Bx-cpl s increased and reached their highest at the initial stage of PWD, then declined gradually. The silencing of Bx-cpl could reduce the feeding, reproduction, and pathogenicity of PWN. These results revealed that Bx-cpl s play multiple roles in the development and pathogenic processes of PWN.