利用荧光定量PCR检测方法对柞蚕不同生产阶段柞蚕微孢子虫发生情况的初步调查  

Preliminary Investigation on Occurrence of Nosema pernyi at Different Production Stages of Antheraea pernyi by Fluorescence Quantitative PCR

作　　者：米锐 李青峰[3] 李佩佩[1,2] 马淑慧 叶博[1,2] 赵振军 孙永欣[1,2] 李亚洁 李学军[1,2] 孟楠 温志新[1,2] 朱有敏 都兴范[1,2] 范琦 Mi Rui;Li Qingfeng;Li Peipei;Ma Shuhui;Ye Bo;Zhao Zhenjun;Sun Yongxin;Li Yajie;Li Xuejun;Meng Nan;Wen Zhixin;Zhu Youmin;Du Xingfan;Fan Qi(Liaoning Ocean and Fisheries Science Research Institute,Dalian Liaoning 116023,China;Dalian Institute of Biotechnology,Liaoning Academy of Agriculture Sciences,Dalian Liaoning 116023,China;Sericultural Research Institute of Liaoning Province,Fengcheng Liaoning 118100,China)

机构地区：[1]辽宁省海洋水产科学研究院,辽宁大连116023 [2]辽宁省农业科学院大连生物技术研究所,辽宁大连116023 [3]辽宁省蚕业科学研究所,辽宁凤城118100

出　　处：《蚕业科学》2019年第6期837-843,共7页ACTA SERICOLOGICA SINICA

基　　金：辽宁省自然科学基金指导计划项目(No.201602418);辽宁省科学事业公益研究基金项目(No.20180013);大连市高层次人才创新支持计划项目(No.2018RQ59)。

摘　　要：柞蚕微粒子病是柞蚕生产中重点检测的病害,目前主要采用镜检淘汰带毒母蛾及子代卵的方法进行病害防控,而荧光定量PCR检测方法具有较高的灵敏度,能够有效提高柞蚕生产中对微孢子虫的阳性检出率。应用荧光定量PCR检测方法对来自2个蚕区柞蚕蛹、蛾的微孢子虫发生情况进行初步调查,同时考察蛹期不同积温发育时期微孢子虫的发生情况以及蛾期亲本带毒对子代卵带毒的影响。结果表明,在蛹期积温中后期(150℃以后)柞蚕微孢子虫的阳性检出率提高,可以考虑在这一发育阶段利用荧光定量PCR方法进行蛹期提前预检;在检验柞蚕微孢子虫垂直传播过程中,发现亲本蛾带毒情况对子代卵带毒存在不同程度的影响,有必要用荧光定量PCR检测方法在卵期对柞蚕微孢子虫进行辅助检测,可以弥补常规蛾期检测的漏检问题。研究结果为应用荧光定量PCR检测技术,在蛹期提前预检以提高种茧品质,在蛾期检测以及卵期辅助检测以减少柞蚕微孢子虫的垂直传播,最终有效控制柞蚕微粒子病的发生提供了参考依据。Microsporidiosis caused by Nosema pernyi is a major control disease in the production of Chinese oak silkworm,Antheraea pernyi.Microscopic examination was usually applied for N.pernyi detection in mother moth and their offspring eggs for disease control.Fluorescence quantitative PCR(qPCR)detection method has higher sensitivity and can improve the positive detection rate of N.pernyi in A.pernyi production.In this experiment,qPCR was used to investigate the occurrence of N.pernyi in pupae and moth from 2 different rearing regions and in pupae at different accumulated temperatures.Besides,effect of parent moth with N.pernyi on occurrence of N.pernyi in offspring eggs was also inspected.The results showed that higher positive detection rate of N.pernyi was found in the middle and late stage of pupae with accumulated temperature over 150℃,so N.pernyi in pupae could be prechecked by qPCR detection.Furthermore,parents with N.pernyi were found to infect the offspring eggs to some extent according to the experiment of detecting vertical transmission of N.pernyi.Thus,it was necessary to detect silkworm eggs in egg stage as a revise for the missing detection of N.pernyi during routine detection in moth stage.The study provides a reference for the application of qPCR throughout early detection in pupae stage to improve the quality of seed cocoons,routine detection in moth stage and auxiliary detection in egg stage to prevent vertical transmission of N.pernyi and effectively control the occurrence of microsporidiosis.

关 键 词：柞蚕 柞蚕微孢子虫 荧光定量PCR 发育时期 

分 类 号：Q959.115.92[生物学—动物学] S885.1[农业科学—特种经济动物饲养]