粤北山区域恙虫病自然疫源地的研究

Study on Tsutsugamushi Disease in YueBei area

目的了解粤北山区域是否存在恙虫病自然疫源地。方法采用分子生物学诊断技术结合间接免疫荧光方法调查粤北山区发热人群、宿主动物感染状况;采用荧光定量PCR技术对粤北山区域恙虫病病人的标本检测Ot-Sta56kDa基因片段,对阳性标本进行基因分型和序列分析,并与巢式PCR检测结果相比较。结果在660份发热人群血清标本中有224份检测到恙虫病东方体,阳性率为33.94%;在55份鼠类脾脏组织标本中有10份黄毛鼠(R.lossea Swinboe)检测到恙虫病东方体,阳性率为18.18%,有2份褐家鼠(R.norvegicus Berkenhout)检测恙虫病东方体,阳性率为3.64%,各调查点之间无显著性差异(P>0.05)。经荧光定量PCR和巢式PCR鉴定为Gilliam型和Kawasaki型,间接免疫荧光法也诊断为恙虫病东方体Gilliam型。从病人、动物宿主、地里纤恙螨选取4株粤北代表株,其同源性为100%,与Kawasaki型的同源性为96.25%;另有3份阳性黄毛鼠标本的核苷酸序列与TA686型的同源性为78.5%。结论从宿主动物、媒介、分子水平上证实粤北山区域存在恙虫病自然疫源地。

Objective To explore the existence of Orientia Tsutsugamushi （Ot） disease in YueBei area of the Guangdong province. Methods Molecular biology techniques and indirect immunofluoresence antibody method were used to investigate the fever patients and host animals. The Ot-Sta56kDa gene of the fever patients in YueBei area was tested by quantitative real-time PCR （qPCR） assay. Nested-polymerase chain reaction（nPCR） was used to identify the genetypes and analyze sequence. Results 660 fever patients＇ and 55 mice samples were determined by qPCR. The positive rate of patients was 33.94% （224/660）, the positive rate of R.lossea Swinboe were 18.18% （10/50） and the positive rate of R. norvegicus Berkenhout was 3.64% （2/50）. There was no significant difference among five suggestive areas （P〉0.05）. Gilliarm genetype and Kawasaki strain of the patients and mice were identified by qPCR and nPCR. Gilliarm genetype was identified by indirect immunofluorescence antibody method （IFA）. The rates of homology of nucleotide sequences among 4 representative strains in the patients and animals were 100%. The homology of Japan Kawasaki strain were 96.25%. The homology rate of the TA686 starin in the nucleotide sequences among mice were 78.5%. Conclusion It has been proved that epidemic area of Tsutsuganushi disease did exist in YueBei area of the Guangdong peovince confirmed on patients, transmission vectors and molecular biology levels.