Dong Huijin, Sun Yu, Zhao Linqing, Deng Li, Zhu Runan, Chen Dongmei, Jia Liping, Liu Liying, Zhang You, Qian Yuan
Bing Du Xue Bao. 2016 Jul;32(4):436-44.
The first reported group A rotavirus(RVA)G9 strain T203 in mainland China detected from an infant in Beijing in 1994 clustered within VP7 lineage Ⅵ(G9-Ⅵ),whereas the common RVA G9 belong to VP7 lineage Ⅲ(G9-Ⅲ) worldwide. Interestingly, since it was first reported in 1994 there was no G9-Ⅵ circulating in Beijing, until an unexpected G9-Ⅵ strain was identified by sequencing in 2010 RVA surveillance. This present study was to develop a convenient and effective dot-blot hybridization method for differentiating G9-Ⅲ and G9-Ⅵ rotaviruses, to investigate the re-emerging circulating distribution of G9-Ⅵ RVAs in outpatient children with diarrhea from 2011 to 2012in Beijing. By multiple-sequence aligning and analyzing the collected VP7 gene nucleotide sequences of G9 RVAs worldwide from GenBank database using Clustal W software, a region within VP7 gene which is highly divergent between G9-Ⅲ and G9-Ⅵ rotaviruses and conserved within themselves was selected as probe. One pair of common primers at both side of this probe region was designed, and used for synthesizing DIG labeled G9-Ⅲ and G9-Ⅵ probes with PCR, respectively. Subsequently, RVA G and P genotypes were identified as previously described. Then G9-Ⅲ and G9-Ⅵ were further differentiated from G9 RVAs using dot-blot hybridization method established in this study. It was showed that G9 was the most prevalent genotype(43.5%),followed by G3(30.5%)、G1(12.2%)and G2(11.5%),and no G4 genotype was detected.Interestingly,G9-Ⅵ was the most predominant(96.5%) type, while only 3.5% was G9-Ⅲ among these G9 rotaviruses.Phylogenetically,G9-Ⅵ rotaviruses in this study clustered closely with human G9-Ⅵ rotaviruses which were more recently re-emerging in several countries including China around the world as well as porcine G9-Ⅵ rotavirus strain F7P4 in Canada, whereas they clustered distantly to worldwide common G9-Ⅲ strains.G9-Ⅵ rotaviruses were re-emerging in the world, whether it gained stronger spreading ability or virulence than ever common G9-Ⅲ during evolution with porcine G9-Ⅵ rotaviruses need further analysis.
1994年在中国北京一名婴儿中首次检测到的中国大陆第一株A组轮状病毒（RVA）G9毒株T203属于VP7谱系Ⅵ（G9-Ⅵ），而全球常见的RVA G9属于VP7谱系Ⅲ（G9-Ⅲ）。有趣的是，自1994年首次报道以来，G9-Ⅵ在北京一直没有传播，直到2010年RVA监测中通过测序意外鉴定出一株G9-Ⅵ毒株。本研究旨在开发一种简便有效的斑点杂交方法，用于区分G9-Ⅲ和G9-Ⅵ轮状病毒，并调查2011年至2012年北京门诊腹泻儿童中重新出现的G9-Ⅵ RVA的流行分布情况。通过使用Clustal W软件对从GenBank数据库中收集的全球G9 RVA的VP7基因核苷酸序列进行多序列比对和分析，选择VP7基因中G9-Ⅲ和G9-Ⅵ轮状病毒之间高度不同但自身保守的区域作为探针。在该探针区域两侧设计一对通用引物，分别用于通过PCR合成地高辛标记的G9-Ⅲ和G9-Ⅵ探针。随后，按照先前描述的方法鉴定RVA G和P基因型。然后使用本研究建立的斑点杂交方法从G9 RVA中进一步区分G9-Ⅲ和G9-Ⅵ。结果显示，G9是最流行的基因型（43.5%），其次是G3（30.5%）、G1（12.2%）和G2（11.5%），未检测到G4基因型。有趣的是，在这些G9轮状病毒中，G9-Ⅵ是最主要的类型（96.5%），而G9-Ⅲ仅占3.5%。在系统发育上，本研究中的G9-Ⅵ轮状病毒与最近在包括中国在内的几个国家重新出现的人类G9-Ⅵ轮状病毒以及加拿大的猪G9-Ⅵ轮状病毒株F7P4密切聚类，而它们与全球常见的G9-Ⅲ毒株聚类较远。G9-Ⅵ轮状病毒正在全球重新出现，其在与猪G9-Ⅵ轮状病毒共同进化过程中是否获得了比以往常见的G9-Ⅲ更强的传播能力或毒力，需要进一步分析。
