北京市市售花蛤诺如病毒和轮状病毒检出情况及基因特征分析

目的了解北京市市售花蛤诺如病毒和轮状病毒检出情况,分析其基因特征。方法2014年11月-2015年10月,采集北京市三个水产品销售市场新鲜花蛤各24件。用两种方法对花蛤鳃和肠内容物进行病毒浓缩,提取核酸后用实时荧光RT-PCR法检测诺如病毒GI/GII组和A组轮状病毒核酸。用半巢式RT-PCR方法,扩增诺如病毒阳性样本衣壳蛋白VP1区基因,对PCR阳性产物进行测序,用诺如病毒分型工具进行基因亚型鉴定,用MEGA软件最大似然法构建进化树。结果72件花蛤中未检出A组轮状病毒,诺如病毒检出率为34.7%(25/72),GII组、GI组和GI/GII组混合检出率分别为27.8%(20/72)、4.2%(3/72)和2.8%(2/72)。11月诺如病毒检出率最高,为100.0%(6/6);三个水产销售市场诺如病毒检出率分别为25.0%(6/24)、50.0%(12/24)和29.2%(7/24)。10件测序成功,其中6株为GII.17,GII.6、GII.2、GI.3a和GI.4各1株。这些毒株与腹泻患者和外环境参考毒株相似性为97.9%~100.0%。结论北京市市售花蛤存在诺如病毒污染,而且与腹泻患者和外环境参考毒株高度同源,应加强花蛤及销售人群和环境的诺如病毒监测。

Detection of norovirus and rotavirus and analysis of the genetic characteristics on clams sold in Beijing

Objective To investigate the detection of norovirus and rotavirus and to analyze norovirus genetic characteristics on clams sold in Beijing.Methods Each 24 pieces of fresh clams were collected from A,B and C aquatic products sales markets in Beijing during November 2014 and October 2015.Two method were used to concentrate the virus in gills and intestinal contents of clams.The virus RNA were extracted,real-time RT-PCR was tested for norovirus GI/GII and Rotavirus A.Norovirus capsid protein region was amplified by semi-nested RT-PCR from positive specimens.PCR products were sequenced directly.The subtypes were identified using the norovirus genotyping tool,and the phylogenetic tree was built using MEGA software with maximum likelihood.Results Among 72 sets of scallops,rotavirus was detected negative,and the detection rate of norovirus was 34.7%(25/72).The detection rates in genogroup GII,GI and GI/G II were 27.8%(20/72),4.2%(3/72)and 2.8%(2/72),respectively.The highest detection rate was 100.00%(6/6)in November、The detection rates of norovirus from A,B and C aquatic products sales markets were 25.0%(6/24),50.0%(12/24)and 29.2%(7/24)respectively.6 Strains GII.17 and 4 Strains(GII.6,GII.2,GI.3a and GI.4)were sequenced successfully.The similarity of these strains to diarrhea patients and environmental reference strains were 97.9%-100.0%.Conclusions There is norovirus pollution in clams sold in Beijing,the strains were highly homologous with diarrhea patients and environmental reference strains.The surveillance of norovirus in clams and the population and environment in which they are sold should be strengthened.