不同血清型汉坦病毒基因重排的研究

目的探明汉坦病毒HTN型与SEO型、HTN型与PHV型之间基因重排的频率和特点。方法分别用相同滴度的汉坦病毒HTN型与SEO型(76-118株与SR-11株),及汉坦病毒HTN型与PHV型(76-118株与PHV株)毒株分别混合感染VeroE6细胞,用空斑形成试验挑选子代病毒克隆,挑得的单个病毒克隆在VeroE6细胞上扩增。分别用分型引物对子代病毒进行RT-PCR实验,以确定子代病毒L、M、S片段核酸的来源,鉴定子代病毒基因型。结果发现76-118株与SR-11株混种的子代病毒株基因30/44株来源于亲代病毒,9/44株子代病毒发生了基因片段重排。76-118株与PHV株混种的子代病毒株基因26/36株来源于亲代病毒,3/36株子代病毒发生了基因片段重排。76-118株与SR-11株混种的子代病毒株的基因重排率(20.45%)明显高于76-118株与PHV株混种的子代病毒株(8.33%)。结论这种重排率的差异可能是由于HTN型与SEO型HV基因核酸序列之间的同源性较高,而更容易发生基因重排。而HTN型与PHV型HV基因核酸序列之间的同源性较低,发生基因重排的机会显著降低。

Studies on Genetic Reassortment among different Serotype of Hantaviruses

Objective To determine the frequency and characteristics of reassortment between Hantaan and Seoul viruses,and between Hantaan and Prospect Hill viruses.Methods Mixed infections were initiated in Vero E6 cell by culturing with Hantaan(76-118 strains)and Seoul(SR-11 strain)viruses,or with Hantaan(76-118 strain)and Prospect Hill(PHV Strain)viruses.Potential reassortant virus plaques were picked for multiplex RT-PCR analysis,using primers specific for individual genome segments(L,M,and S)of each strain.Results The results showed that in the mixed infections using 76-118 and SR-11 strains,most of the progeny virus plaques are non-reassorted(68.19%,30/44),and with a few reassortant viruses plaques(9 out of the 44).Approximately 4.55% of the progeny virus plaques appeared to have two segments from one parental virus strains,and another one segment contain S or M segments originating from both parental virus strains.In the mixed infections using 76-118 and PHV strains,most of the progeny virus plaques(72.22%,26/36)had the parental genotype,while reassortant viruses were detected in 3 out of the 36 progeny plaques.Approximately 8.33% of the progeny virus plaques appeared to have two segments from one parental virus strains,and another one segment contain M segments originating from both parental virus strains.The reassortant ratio of virus progeny from mixed infections with 76-118 and SR-11 strains(20.45%)has been observed higher than that from mixed infections with 76-118 and PHV strains(8.33%).Conclusion The different frequency of reassortment might be explained by the difference of their genetic divergence of the three segments.