与HIV-1共感染的HHV-6毒株的分离、传代培养与鉴定

目的：对一株从中国河南省一名艾滋病患者体内分离的、可在MT4细胞上稳定传代的未知病毒29A进行鉴定，确定该病毒种类及型别。方法：用从该患者外周血单核细胞中分离的病毒株感染MT4细胞，获得可在该细胞上稳定传代的病毒株29A。通过细胞病变特征、电镜观察结果、HIV-1 p24抗原检测以及HIV-1 pol区基因扩增结果，对该毒株是否为HIV-1进行鉴别。在电镜下对病毒感染细胞的超薄切片进行观察，发现该病毒呈现疱疹病毒形态特征，因此设计人类疱疹病毒6型（HHV-6）U69、U16／U17、U60／U66三个不同基因片段的特异性引物并进行巢式PCR扩增，对扩增产物进行序列测定和结果分析。结果：该毒株HIV-1 p24抗原检测为阴性；用HIV-1 pol区特异性引物未扩增出目的片段；细胞病变形态不同于HIV-1引起的病变；电镜观察该病毒直径为160～200nm，明显大于HIV—1，表明该传代株不是HIV-1。用人类疱疹病毒6型（HHV-6）U69、U16／U17、U60／U66等基因的特异性引物均扩增出了目的片段，序列分析的结果表明该毒株为HHV-6病毒B亚型。结论：在国内首次从艾滋病患者的外周血淋巴细胞中分离出HHV-6，为进一步研究HHV-6和HIV-1的相互作用奠定了基础。

Isolation,passage and identification of HHV-6 co-infected with HIV-1

Objective: To identify and characterize an unknown viral strain 29A,which was isolated from the peripheral blood lymphocytes of an AIDS patient from Henan Province of China and was able to passage in MT4.Methods:The virus strain was isolated from the peripheral blood lymphocytes of an AIDS patient.The primary isolated strain was used to infect human transformed T-cell lines MT4,and a stable passage strain 29A was gained.The CPE(cytopathic effect)observation,electron microscopic examination,HIV-1p24 antigen test and nested-PCR(nPCR) with HIV-1 pol gene primers were performed to determine whether 29A was HIV-1.These viruses exhibited a characteristic herpesvirus morphology,so nPCR was performed to amplify HHV-6 U69,U16/U17,U60/U66 genes.Results: p24 Antigen was negative in the strain 29A,pol gene of HIV-1 was not amplified,and the characteristics of CPE were different from those of HIV-1.The diameter of this virus particle was about 160-200 nm,apparently bigger than that of HIV-1.So the strain 29A was not considered as HIV-1.nPCR was performed with HHV-6 primer,and the U69,U16/U17,U60/U66 genes of HHV-6 could be amplified.Based on the result of sequence analysis, the strain 29A was HHV-6 subtype B.Conclusion: The HHV-6 strain was isolated firstly from the peripheral blood lymphocytes of a patient with AIDS in China,which could be propagated stably on MT4 cells.It offers a useful platform for further studying the interaction between HIV and HHV-6.