北京地区2007—2008年G9型A组人轮状病毒VP7和VP4基因分析

Analysis on VP7 and VP4 genes of human rotavirus G9 identified from children with diarrhea in Beijing, from 2007 to 2008

目的了解北京地区2007—2008年检测到的G9型A组人轮状病毒外壳蛋白VP7和VP4的基因特征。方法选取经过轮状病毒核酸杂交方法检测为G9型轮状病毒的12份儿童腹泻患儿的粪便标本，应用针对VP7全长基因的特异引物对进行RT-PCR扩增，对所获得的VP7全长基因进行克隆和测序，将所获得的序列与GenBank中的G9型原型病毒株和近期流行株的VP7基因进行序列和种系进化分析；经巢式PCR对G9型的VP4进行P基因分型。结果12株G9型轮状病毒经VP7基因的序列比较分析得到确认。P基因分型结果显示北京地区近年来存在G9P[8]和G9P[6]型两种组合的轮状病毒感染。序列和种系进化分析发现北京G9型株VP7基因与世界范围内近期流行的G9型株一样都属于进化分支Ⅲ，彼此间的核苷酸和氨基酸同源性较高，而与国内最早报道的G9型T203进化关系较远，且北京G9P[8]和G9P[6]型株分别与国内近期报道的新疆G9P[8]和G9P[6]型株及相应的武汉G9型株VP7基因，在氨基酸位点上存在一些共同的氨基酸残基取代。结论北京地区近年存在G9P[8]和G9P[6]两种不同基因组合的G9型轮状病毒感染，需要进一步加强对G9型轮状病毒的分子流行病学监测。

Objective To characterize the outer capsid protein VP7 and VP4 encoding genes of human rotavirus G9 strains detected in Beijing, from 2007 to 2008. Methods Full length of VP7 genes of G9 rotaviruses from 12 fecal specimens previously detected by dot-blot hybridization assay were amplified by RT-PCR and sequenced after being cloned into T vector. The sequences of these VP7s were compared to VP7 genes of rotavirus G9 prototype strains and recently circulating strains around the world. VP4 genes of these 12 G9 strains were amplified by nested-PCR for P genotyping. Results Sequence analysis for the full length of VP7 genes from these 12 specimens confirmed that they were G9 rotaviruses. P genotyping for VP4 genes revealed that both P [ 8 ] G9 and P [ 6 ] G9 were circulating in Beijing in the last 2 years. Sequence and phylogenetic analysis demonstrated that VP7 genes of G9 strains from Beijing in this study were clustered in the lineage Ⅲ which resembled the G9 strains circulating in other places around the world, indicated by high identities of nucleotide and deduced amino acid sequences and were distant with the first reported G9 strain T203 identified in China in 1994. It was found that there were some consistent amino acid substitutes at the corresponding positions among VPTs from these 12 specimens and from Xinjiang and Wuhan, both in G9P [ 8 ] and G9P [ 6 ] strains. Conclusion The rotavirus G9 strains both in combination of G9P [ 8 ]and G9P [6] were circulating in Beijing in the past years. It seemed that rotavirus G9 should be included in the list of surveillance for rotavirus in China.