蜱传人兽共患无形体和立克次体的分子生物学鉴定

Molecular Identifi cation of Tick-borne Zoonotic Anaplasma and Rickettsia in Ticks

目的调查承德市周边农村蜱传立克次氏体目细菌的流行现状,分析承德地区人兽共患无形体和立克次体的种类及其遗传特征,为蜱传立克次氏体病的防控提供理论依据。方法本研究从承德市三沟镇和六沟镇采集蜱并提取总DNA,使用巢氏PCR技术扩增16S rRNA基因进行立克次氏体目细菌的鉴定。扩增无形体(Anaplasma)的热休克蛋白(groEL)基因和立克次体(Rickettsia)的外膜蛋白A(ompA)基因,测序后对序列进行同源性与系统发生分析,对蜱传人兽共患无形体和立克次体病原体进行鉴定。结果在三沟镇和六沟镇共采集到336只蜱,经鉴定均为长角血蜱。蜱感染无形体的阳性率为25.3%(85/336),立克次体的阳性率为17.3%(58/336),蜱感染无形体的阳性率显著高于立克次体(P=0.01)。无形体包括山羊无形体(Anaplasma capra)和羊无形体(Anaplasma ovis),阳性率分别为8.3%(28/336)和17.0%(57/336);立克次中西伯利亚立克次体(Rickettsia sibirica)的阳性率为3.6%(12/58),饶氏立克次体(Rickettsia raoultii)的阳性率为6.3%(21/58),Candidatus Rickettsia jingxinensis的阳性率7.4%(25/58)。结论承德市长角血蜱携带有2种人兽共患无形体与3种立克次氏体,表明承德周边农村地区流行立克次氏体目细菌具有较高的遗传多样性,需加强承德地区蜱与蜱传病原体的监测与防控。

Objective To investigate the prevalence and species of tick-borne Rickettsiales in rural areas of Chengde City,and to provide a theoretical basis for the prevention and control of diseases caused by tick-borne Rickettsiales.Methods Total DNA was extracted from ticks collected from Sangou Town and Liugou Town of Chengde City,and the 16S rRNA gene was amplifi ed by nested PCR to identify Rickettsiae bacteria.The heat shock protein(groEL)gene of Anaplasma and outer membrane protein A(ompA)gene of Rickettsia were amplifi ed and sequenced for homology and phylogenetic analysis to identify the pathogens of tick-borne zoonotic Anaplasma and Rickettsia.Results A total of 336 ticks were collected from Sangou Town and Liugou Town,and all of them were identifi ed as Haemaphysalis longicornis.The positive rate of Anaplasma was 25.3%(85/336)in ticks and 17.3%(58/336)in Rickettsia.The positive rate of Anaplasma in ticks was signifi cantly higher than that in Rickettsia(P=0.01).The positive rates of Anaplasma capra and Anaplasma ovis were 8.3%(28/336)and 17.0%(57/336),respectively.The positive rate of Rickettsia sibirica and Rickettsia raoultii was 3.6%(12/58)and 6.3%(21/58),respectively.The positive rate of Candidatus Rickettsia jingxinensis was 7.4%(25/58).Conclusion Two zoonotic Anaplasma and three Rickettsia species were identifi ed in H.longicornis in Chengde,indicating that Rickettsiae bacteria prevalent in rural areas around Chengde present high genetic diversity.And it is necessary to strengthen the monitoring and prevention and control of ticks and tick-borne pathogens in Chengde.