河北省汉坦病毒的分离、复苏及分型

The isolation, recovery and identification of Hantavirus in Hebei province

目的对2012年河北省鼠类标本进行汉坦病毒（HV）分离并复苏保存的病毒株,鉴定型别,了解河北省疫区HV的型别分布特点。方法利用间接免疫荧光法（IFA）对鼠肺标本进行初筛,采用细胞培养法分离阳性标本病毒,并复苏已保存的病毒株。提取病毒RNA后进行分型鉴定。结果 4份阳性鼠肺标本的组织悬液接种Vero-E6细胞2-3代后,经IFA检测,细胞内出现病毒特异性荧光颗粒,但未发现致细胞病变效应。4份标本全部分离成功。取已保存的病毒株接种Vero-E6细胞1-2代后,经IFA检测,细胞内出现较强的病毒特异性荧光颗粒,6株病毒全部复苏成功。通过分型鉴定,10株病毒均属于家鼠型汉城型汉坦病毒（SEOV）。结论河北省HV以SEO型为主。SEOV在Vero-E6细胞虽无明显的细胞病变,但可在该细胞中增殖并被检测。在-80℃条件下,HV至少保存20余年。我国东方田鼠为SEOV的宿主之一。

Objective To understand the distribution characteristics of Hantavirus （HV） genotype in Hebei province, the HV strains isolated from rodents in 2012 and recovered from formerly saved were typed. Methods Lung samples were screened by immunoinfluscent assay （IFA）. Antigen-positive lung samples were processed and inoculated onto cultures of confluent Veto-E6 cells. Formerly saved strains were recovered. Total RNA were extracted for genotype and identification. Results Four lung tissues were homogenized and inoculated onto confluent Vero-E6 cells. After the second-third passage, the characteristic HV antigen particle could be seen by IFA. At last, all four strains were isolated successfully. The formerly saved strains were used to infect fresh Vero-E6 cell cultures. After the first-second passage, the characteristic HV antigen particle can be seen by IFA. At last, all six strains were recovered successfully. All ten strains were typed by PCR and they belonged to SEO type. Conclusion SEO type was the predominant HV genotype in Hebei province. SEOV didn＇ t produce cytopathic effects, but the virus can proliferate in the cell. HV can be saved for at least more than 20 years at -80 ℃. In China, Microtusfortis was an alternative host for SEOV.