Ohyama T, Yoshizumi S, Sawada H, Uchiyama Y, Katoh Y, Hamaoka N, Utagawa E
Hokkaido Institute of Public Health, Sapporo, Japan.
Microbiol Immunol. 1999;43(6):543-50. doi: 10.1111/j.1348-0421.1999.tb02440.x.
Samples of feces and vomit collected from patients during 13 non-bacterial gastroenteritis outbreaks which occurred in Hokkaido between 1995 and 1998 were examined by electron microscopy (EM) and reverse-transcription polymerase chain reaction (RT-PCR) for evidence of infection with human caliciviruses (HuCVs). In 6 food-borne outbreaks, oysters were the probable source of infection, while the origin of HuCVs was not found out for the other 7 outbreaks. One-hundred-eleven of 214 stool, vomit and oyster specimens examined gave positive results by RT-PCR, while HuCVs were detected by EM in 36 of 121 stool specimens examined. We determined the nucleotide sequences of 470-bp or 373-bp PCR products amplified from the RNA polymerase region of the HuCV genomes with primer sets MR3/4 and Yuri22F/R, respectively. The sequences of different strains revealed great heterogenicity, with a range of 60 to 100% homology among strains. In a few cases, a mixed genotype was found in the same patient or same outbreak by means of nested PCR and cloning of PCR products into an appropriate vector. Of the 19 different strains found, 4 strains could be classified as Norwalk virus (genogroup 1) and the other 15 strains as Snow Mountain agent (genogroup 2) based on genotyping with homology analysis. Furthermore, the strains belonging to genogroup 2 could be classified into 4 subgroups with more than 93% homology in amino acids among strains in the subgroup.
对1995年至1998年期间在北海道发生的13起非细菌性肠胃炎疫情中从患者采集的粪便和呕吐物样本进行了电子显微镜（EM）检查和逆转录聚合酶链反应（RT-PCR），以寻找人类杯状病毒（HuCVs）感染的证据。在6起食源性疫情中，牡蛎可能是感染源，而在其他7起疫情中未查明HuCVs的来源。在214份检查的粪便、呕吐物和牡蛎样本中，111份通过RT-PCR检测呈阳性，而在121份检查的粪便样本中，36份通过EM检测到HuCVs。我们分别用引物组MR3/4和Yuri22F/R从HuCV基因组的RNA聚合酶区域扩增出470 bp或373 bp的PCR产物，并测定了其核苷酸序列。不同毒株的序列显示出很大的异质性，毒株之间的同源性范围为60%至100%。在少数情况下，通过巢式PCR和将PCR产物克隆到合适载体中，在同一患者或同一疫情中发现了混合基因型。在发现的19种不同毒株中，根据同源性分析进行基因分型，4种毒株可归类为诺如病毒（基因组1），其他15种毒株可归类为雪山病毒（基因组2）。此外，属于基因组2的毒株可分为4个亚组，亚组内毒株的氨基酸同源性超过93%。
