鼠癣螨(Myocoptes musculinus)分子鉴定和感染调查

Prevalence and Molecular Identification of Myocoptes musculinus in Laboratory Animals in China

目的建立鼠癣螨(Myocoptes musculinus)快速鉴定方法,并对50个生产厂家843批5109只SPF实验动物鼠癣螨感染的检查结果进行分析,为制定鼠癣螨的净化策略提供科学数据和技术支持。方法直接镜检实时动态显微视屏摄录技术结合形态学鉴定方法,进行鼠癣螨感染筛查。多重PCR方法鉴定分离的鼠癣螨cytb(细胞色素b)、cox2(细胞色素C过氧化物酶亚基2)和ATP6(ATP合成酶F0亚基6)基因,从分子水平上进一步确认鼠癣螨感染。结果基于直接镜检实时动态显微视屏摄录技术,检出鼠癣螨阳性样本39份,阳性率为0.76%,检出形态为鼠癣螨虫卵、幼虫、若虫和成虫。根据鼠癣螨的虫卵、幼虫、若虫、雌雄成虫的大小和形态初步确定虫种。从阳性样本中分离的单个鼠癣螨的虫卵、幼虫、若虫和成虫扩增其cytb、cox2和ATP6基因,结果表明,39份形态学阳性样本,分子鉴定均为阳性,且与其他不同种属的寄生虫无交叉反应。序列比对结果显示,不同个体间鼠癣螨的cytb、cox2和ATP6部分基因序列核苷酸相似性达100%。结论直接镜检实时动态显微视屏摄录技术联合多重PCR技术检测鼠癣螨具有高的灵敏性和特异性。检测的实验动物样本中确有鼠癣螨阳性,提示我国实验动物病原学检查需进一步强化。本研究首次对中国实验动物的鼠癣螨进行了分子鉴定和感染调查,为净化鼠癣螨策略提供数据和技术支持。

Objective The aim of this study is to establish a method of rapid identification of Myocoptes musculinus infection, and analyze the test result of 843 batches specific pathogen free （SPF） laboratory animal from 50 different manufacturers in China, and to provide data and technical support for the strategy of Myocoptes musculinus purification. Method Direct microscopy real-time dynamic video recording techniques combined with morphological identification method were applied to screen Myocoptes musculinus infestation. Multiple PCR method were used to identify the isolated Myocoptes musculinus cytb （cytochrome b）, cox2 （cytochrome c oxidase subunit 2） and ATP6 （ ATP synthase F0 subunit 6） genes, and the Myocoptes musculinus infection was further confirmed at the molecular level. Result Based on the direct microscopy real-time dynamic microscopic video recording technology, 39 Myocoptes musculinus positive samples were detected, the positive rate was 0.76%, and the morphology was the egg, larva, nymph and adult. The parasite species were preliminarily determined according to the size and morphology of Myocoptes musculinus eggs, larvae, nymphs, and adults. The cytb, cox2 and ATP6 genes of Myocoptes musculinus were amplified from the single egg, larva, nymph and adult of Myocoptes musculinus separated from the positive samples, and the result showed that 39 positive samples of morphology, molecular identification are all positive, and no cross reaction with other different species of parasites. Sequence comparison result showed that the nucleotide similarity of cytb, cox2 and ATP6 gene sequence of Myocoptes musculinus in different individual animal was 100%. Conclusion Direct microscopy real-time dynamic microscopic video recording technology combined with multiple PCR technique has high sensitivity and specificity and can be used to detect Myocoptes musculinus. The test result showed that Myocoptes musculinus was positive in laboratory animal samples, suggesting that laboratory animal pathogen examination in our country should be further strengthened. This research was the first molecular identification and infection investigation of Myocoptes musculinus in laboratory animals in China.