浙江省5例输入性卵形疟原虫wallikeri亚种感染病例的鉴定

Iditification of Five Imported Cases of Plasmodium ovale wallikeri Infection in Zhejiang Province

目的对浙江省5例镜检阳性、常规巢式PCR检测阴性的输入性疟疾病例进行卵形疟原虫wallikeri亚种(Plasmodium ovale wallikeri)的鉴定与分析。方法收集5例待检病例的流行病学资料和血样,进行镜检、快速疟疾诊断试纸条检测(RDT)和巢式PCR检测(采用疟原虫属特异性引物、种特异性引物和卵形疟原虫wallikeri亚种特异性引物),将PCR扩增片段测序后,与Gen Bank中相关序列进行Blast比对分析。结果 5例疟疾病例均为自非洲归国人员,并有疟疾发病史。镜检结果显示,5例患者均疟原虫阳性。RDT检测结果为泛疟原虫阳性2例,阴性3例。应用常规巢式PCR检测5份血样DNA,4种疟原虫均为阴性。改用卵形疟原虫wallikeri亚种特异性引物进行扩增,结果均为阳性。将5份血样的阳性扩增产物进行测序,所得序列经Blast分析,与Gen Bank中编号为KF219561.1的卵形疟原虫wallikeri亚种clone RSH10 18S核糖体RNA基因的序列一致性均达100%。结论综合镜检和PCR检测结果,证实该5例镜检疟原虫阳性、常规PCR检测阴性的病例均为卵形疟原虫wallikeri亚种感染。

Objective To identify and analyze Plasmodium ovale wallikeri in 5 imported malaria cases, who were detected positive by microscopy and negative by conventional PCR. Methods Epidemiological information and blood samples were collected from the five patients. The detection was conducted by microscopy, Rapid Diagnostic Test （RDT） and nested PCR with Plasmodium genus-specific, species-specific and Plasmodium ovale wallikeri-specific primers. The amplified products were sequenced and Blast analysis was performed on line in NCBI. Results The five patients returned from Africa, and all had a history of malaria. They were microscopically positive for Plasmodiurn sp., and two cases showed Pan positive RDT result. All blood samples were negative for four Plasmodium spp. by conventional nested PCR, but positive by nested PCR with Plasmodium ovale wallikeri-specific primers. Blast analysis showed that the amplified sequences of the five cases had complete homology with P. ovale wallikeri clone RSH10 18S ribosomal RNA gene （Accession No. KF219561.1）. Conclusion The five cases which classified as positive by microscopy while negative by conventional PCR have been confirmed as Plasmodium ovale wallikeri infection by nested PCR with P. ovale wallikeri-specific primers.