基于组学的寄生蜂抗菌肽基因鉴定及其对寄主小菜蛾免疫反应的作用研究

We use the diamondback moth, Plutella xylostella, and its two solitary parasitoids, Cotesia vestalis (Braconidae) and Diadegma semiclausum (Ichneumonidae), as a model system to examine the innate immunity of these two endoparasitoids against foreign pathgens and identify antimicrobial peptide (APM) genes that involved in parasitoid-host interaction. Firstly we will characterize the diversity of antimicrobial peptide (APM) genes (defensin，abaecin, hymenoptaecin and so on) in these two types of parasitoids and gene transcripts of these APMs at different developmental stages and different tissues. Secondly we will examine the gene expression in the host larval and impact on the immune response of the host larval. Lastly we will verify the function of AMPs genes that derived from different parasitism factors, i.e, venom, PDV or teratocytes which reveal the molecular basis of immune evolutionary in these two parasitoids and the host. The results will not only reveal the consistent and dissimilar between these two parasitoids, but also provide us a profound base for understanding the regulation mechanism of parasitoid on the host immune responses and for developing new techniques for sustainable management of insect pests..

以携带不同寄生因子的2种内寄生蜂菜蛾盘绒茧蜂（毒液，PDV和畸形细胞）和半闭弯尾 姬蜂（毒液和PDV）寄生同一寄主（小菜蛾）为研究体系，在分析和鉴定寄生蜂defensin，abaecin和hymenoptaecin等抗菌肽基因基础上，研究两种寄生蜂自身应对外源刺激的天然免疫反应，鉴定与寄生蜂-寄主免疫互作相关的抗菌肽分子，检测其在寄主小菜蛾体内转录表达，分析其在小菜蛾体内免疫反应的作用，从而揭示寄生蜂抗菌肽在寄生蜂与寄主免疫互作中的功能及其规律，解析携带不同寄生因子的寄生蜂与寄主免疫进化的分子基础,为最终阐寄生蜂调控寄主免疫反应的机制及发展持续控制害虫的新技术奠定扎实的基础。

抗菌肽，是一类具有免疫效应的小分子多肽，已经被发现具有抵御包括细菌、真菌、病毒、原生生物在内的多种病原体的活力，部分抗菌肽还具有帮助伤口愈合、抗氧化、抑制酶活的作用，是昆虫免疫系统的重要效应因子。本项目基于菜蛾盘绒茧蜂基因组和转录组，筛选其中的抗菌肽，通过荧光定量PCR明确了龄期组织表达特性；获取了其中膜翅肽基因CvHym的开放阅读框序列，构建载体，体外表达获得了活性蛋白，对其抗菌活性进行了检测；通过qPCR检测了寄生对寄主小菜蛾抗菌肽表达的影响，诱导寄主小菜蛾后畸形细胞和幼蜂抗菌肽表达水平变化；通过免疫组织化学和Western Blot技术对CvHym的进行了检测，明确表达和分泌情况。本研究表明，菜蛾盘绒茧蜂含有丰富的抗菌肽资源，对多种病原物有抗菌活力。蜂畸形细胞能表达并分泌抗菌肽。寄生蜂与寄主是一个有机整体，寄生会影响寄主抗菌肽的表达，畸形细胞和幼蜂液会为寄主抗菌免疫提供补偿。