应用PCR技术检测媒介恙螨体内恙虫病立克次体(英语)

本文PCR技术的引物是参考恙虫病立克次体Karp株Sta 58序列设计合成的一对DNA引物.以恙虫病立克次体DNA为模板,成功扩增长约1kb的DNA片段.该对引物首次应用于人工成虫腹内接种羔虫病立克次体(人工接种法)和幼虫叮咬恙虫病鼠(叮咬病鼠法)的我国主要恙虫病媒介地里纤羔螨的研究.PCR检测人工接种法成虫的不同时期和经卵传递的子代立克次体DNA均获满意结果,立克次体在恙螨体内生长持续360天及产卵孵出的第4代幼虫均呈阳性.叮咬病鼠法立克次体在恙螨亲代饱食蚴、若蛹、若虫、成虫及第2代子产代幼虫,PCR检测均呈阳性.结果显示PCR技术检测恙螨体内恙虫病立克次体的特异性和敏感性高;体外基因扩增检测恙螨体内及鼠宿主体内立克次体是流行病学调查的新的敏感方法.本法为恙虫病分子体流行病学调查提供了新的应用技术,亦为临床病人诊断提供了敏感方法.

DETECTION OF RICKETTSIA TSVTSUGAMUSHI FROM VECTOR TROMBICULID MITE BY DNA AMPLIFICATION USING POLYMERASE CHAIN REACTION TECHNIQUE

With reference to the Sta 58 major antigen gene of Rickettsia tsutsugamushi (Karp strain), we had designed and synthesized a pair of DNA primers; and a kilobase fragment of Sta 58 is amplified for using in polymerase chain reaction (PCR) with the primers.This is the first report on detection of R. tsutsugamushi in trombiculids by using PCR technique. The experimental results indicated that the time for which R. tsutsugamushi could exist in the adult of Leptotrombidium deliense when inoculated into its abdomenal cavity was 360 days at least and R. tsutsugamushi could be transovarially transmitted to the offsprings for 4 generations; and the time for which R.tsutsugamushi could exist in L. deliense after biting the infective mouse was 270 days at least and R. tsutsugamushi could be transmitted transovarially to the offsprings for 2 generations. The results also showed that the PCR technique was highly specific and sensitive for detection of R. tsutsugamushi; and the amplification of gene outside body to detect the rickettsiae in the body of trombiculid and mouse can be used as a new method for investigation of scrub typhus epidemiology.