诺如病毒GⅡ.4型变异株感染流行的研究进展

诺如病毒是-种在全球范围广泛分布并引起急性感染性胃肠炎的病原体^[1].美国每年约有1.79亿例急性胃肠炎感染者,其中超过50％(36％-59％)为感染诺如病毒所致;每年因诺如病毒感染导致住院病例可达70 000例,其中死亡800例,带来医疗负担高达2.84亿美元^[2,3].包括美国在内,许多发达国家已建立了诺如病毒胃肠炎监测系统.本文主要对诺如病毒病原学和流行病学特征以及GⅡ.4型变异株在多国的流行概况进行综述.     

江苏省近期急性胃肠炎暴发中诺如病毒分子特征分析

目的 了解江苏省胃肠炎暴发疫情中诺如病毒(NoV)感染状况,并对其病原分子流行病学特征进行初步研究.方法 收集江苏省2012年10月至2013年3月7起胃肠炎暴发疫情患者肛拭子标本共67份,采用实时荧光RT-PCR定性检测,NoV阳性标本用RT-PCR扩增其聚合酶区及VP1区基因片段进-步测序分型.结果 7起疫情均为NoV感染引起的暴发,检测阳性率为67.2％(45/67).全部45株阳性毒株序列分析表明7起疫情中除-起是由GⅡ.3型感染引起外,其余6起均由新GⅡ.4感染引起,其中38株毒株与2012年澳大利亚参考株GⅡ.4/Sydney株同源性均在99％以上.进-步对6株新型Sydney变异株的衣壳蛋白VP1区扩增测序,并与9株近年流行株的VP1区氨基酸序列比对,发现该6株毒株的VP1区部分氨基酸已出现突变,而氨基酸位点的突变是与病毒抗原性密切相关.结论 江苏省7起疫情暴发聚集且感染毒株型别单-,说明已出现新NoV变异株,其原型株GⅡ.4/Sydney已在国外多地引起暴发和流行,提示该毒株将是今后防控监测的重点.     

南宁市朝阳溪污水中诺如病毒GⅡ.4型变异株基因分析

目的了解南宁市朝阳溪污水中诺如病毒(Norovirus,NV)GⅡ.4变异株的基因特征。方法 2011年1~12月采集72份南宁市朝阳溪污水,经实时荧光逆转录-聚合酶反应(Realtime RT-PCR)方法进行NV核糖核酸(RNA)检测,阳性标本进行基因克隆测序和遗传进化分析。结果 72份污水中均能检测到NV RNA,基因Ⅰ组(GenogroupⅠ,GⅠ)NV RNA阳性率为70.83%,基因Ⅱ组(GenogroupⅡ,GⅡ)阳性率则达100.00%,共为16个基因型,有18株为GⅡ.4型,其中2株为2006b变异株,其余16株均为2010变异株。检测到的2006b变异株和2010变异株在衣壳蛋白N/S区第94位氨基酸均发生变异,而在第6位、第15位仅2010变异株发生了变异,2006b变异株(除SW2011041-2株外)均无变异。这些变异株与同年南宁市腹泻病例中NV GⅡ.4型变异株核苷酸型内同源性分别为96.70%~98.93%和97.42%~99.64%。结论南宁市朝阳溪污水中NV存在很高的遗传多样性,其来源可能是周边居民中的感染者;GⅡ.4型NV中2010变异株为优势株,其在人群中可能存在隐性感染;污水监测是人间NV感染监测的重要补充。     

广州市大学城诺如病毒GⅡ.4 Sydney 2012变异株感染疫情再次暴发的调查

2013 年1 月广州市大学城ZD 大学暴发国内首起诺如病毒GⅡ.4 Sydney 2012 变异株感染引起的急性胃肠炎疫情。2 个月后该地区其他4 所高校（HS、HG、GD、GG）和1所中学（GDFZ）相继又出现暴发。     

GⅡ-4型诺如病毒ORF2基因克隆及序列分析

目的 获得中国流行株GⅡ-4型诺如病毒(NoV)ORF2基因序列,为进一步研究诊断、疫苗提供基础依据.方法 从GⅡ-4型NoV粪便中提取病毒RNA,经逆转录后PCR并克隆到simple PMD18-T载体获得完整开放读码柜架(ORF2)阅读框序列.结果 获得了正确序列的基因Ⅱ-4,亚型(genogroup Ⅱ-4,GⅡ-4)NOV ORF2基因序列,经软件分析,3株NOV ORF2序列编码的氨基酸大小约为54KD,与VA387(GⅡ-4)氨基酸同源性为92%～93%.ORF2编码的氨基酸序列存在3个相对保守序列.结论 获得GⅡ-4型NoV ORF2基因序列,为进一步表达病毒衣壳蛋白及Nov感染的诊断试剂及防治疫苗药物研制提供基础依据.     

广州市2017年某高校诺如病毒GⅡ.4 Sydney 2012变异株感染暴发调查

目的 分析2017年广州市GⅡ.4 Sydney 2012变异株引起的诺如病毒感染暴发疫情的流行特征和分子生物学特征,为诺如病毒感染疫情的防控提供依据。方法 设计调查表收集病例发病资料和疫情数据,采集现症病例和厨工的肛拭子以及环境标本进行病原学检测,阳性标本进行基因测序和同源性分析。采用病例对照研究方法分析疫情的相关危险因素。结果 2017年9月17-21日,广州市大学城某高校累计发生诺如病毒感染病例226例,包括223例在校学生和3例厨工,学生罹患率为0.73%（223/30 711）;宿舍A区学生罹患率最高（1.73%,164/9 459）,无院系和班级聚集性;主要危险因素为18-20日在A区食堂就餐（OR=10.75,95% CI：5.56~20.79）,发病风险最高的是18日晚餐,另一危险因素是同宿舍有病例（OR=3.65,95% CI：1.92~6.94）;诺如病毒核酸阳性的厨工全部来自A区食堂,阳性率为26.67%（12/45）,病毒基因测序结果为GⅡ.4 Sydney 2012变异株,与学生病例的病毒基因序列相似度为100%。结论 本次疫情是继2013年之后诺如病毒GⅡ.4 Sydney 2012变异株再次在学生人群引起的较大规模暴发疫情,由食源性传播的可能性较大,同时不排除接触传播。     

成都市成华区诺如病毒GII.P16-GII.4 Sydney＿2012分子生物学特征

目的 研究2021年引发成都市急性胃肠炎聚集性疫情的诺如病毒GII.P16-GII.4 Sydney＿2012株的遗传进化和分子病原学特征。方法 采集2021年1—12月成都市65起急性胃肠炎聚集性疫情病例粪便及肛拭子样本825份,采用实时荧光定量PCR检测。结果为阳性的样本经由常规RT-PCR对RdRp区和VP1区扩增后测序,以BioEdit、MEGA-X将所得序列与全球各参考株序列比对并构建进化树,对其同源性、氨基酸变异、蛋白质三维结构等基因特征进行分析研究。结果 自上述65起疫情检出7种诺如病毒基因型,其中13起由Sydney＿2012株引起。比对分析结果显示,其同一起Sydney＿2012疫情中标本序列相似性为98.6%～100%,疫情之间序列相似性为96.2%～100%。结论 Sydney＿2012株可能成为成都地区流行的优势株,甚至有引起爆发疫情的可能性。持续加强诺如病毒疫情监测,有助于提升防控预警能力。     

诺如病毒GⅡ．4型Sydney 2012变异株与GⅡ．3型混合感染事件调查

目的 调查某大学一起急性胃肠炎暴发事件的感染来源和流行病学特征。方法 开展病例搜索,用描述性流行病学方法分析;用RT PCR法检测诺如病毒核酸,进行基因测序和核酸分型。结果 事件共发生感染性腹泻疑似病例473例,罹患率7.7%,确诊97例（20.5%）;临床症状以腹泻（100%）和呕吐（67%）为主,病例分布无聚集性,无性别差异;采集病例和厨工肛拭子、食堂剩余食物、环境涂抹样和生活饮用水样本共53份,细菌分离培养阴性,肛拭子诺如病毒RT PCR阳性率60.0%（9/15）,经基因测序分型5份为诺如病毒GⅡ.4型Sydney 2012变异株,4份为GⅡ.3型。结论 该起事件由诺如病毒GⅡ.4型Sydney 2012变异株和GⅡ.3型混合感染所致,传播途径可能为食源性传播和接触传播,卫生监管部门应加强对集体单位食堂的监管。     

一起诺如病毒GⅡ.17引起的成人腹泻病原学基因诊断研究

目的针对一起北京市丰台区2014年10月暴发的急性胃肠炎进行病原基因研究,鉴定其病原体,并对其基因分型做进一步分析。方法应用实时荧光定量PCR对疫情中采集的50份样品进行诺如病毒检测,检测后的阳性样本经逆转录RT-PCR扩增,对其产物测序,进行基因数据库比对,应用Mega 5.0软件构建序列进化树分析。结果 50份疫情样品中,诺如病毒阳性18份。阳性样本基因扩增后测序所得序列与基因数据库比对发现,18份阳性样本均为GⅡ.17型,阳性率为36%。结论本次北京市丰台区暴发的急性胃肠炎疫情是由罕见的诺如病毒GⅡ.17引起。     

安徽省诺如病毒G II.4/Sydney 2012变异株分子特征分析

目的了解安徽省某高校一起急性胃肠炎暴发疫情的病原体基因分型和分子特征。方法采用实时荧光定量PCR法(Real-time PCR)对6份粪便/肛拭子标本检测诺如病毒核酸,检出的阳性标本经传统RT-PCR扩增衣壳蛋白区(ORF2)部分序列、基因测序和型别鉴定,利用Clustal X 2.0和Mega 6.0软件对测定序列进行核苷酸同源性比对和构建基因进化和亲缘性关系树。结果检出4份标本诺如病毒核酸阳性,阳性率为66.67%(4/6);基因序列比对和进化亲缘性关系显示:诺如病毒基因型为GII.4型;2毒株序列之间核苷酸同源性为100%;与2014~2015年上海株、香港株核酸同源性最高,均为100%;与2012澳大利亚Sydney株(JX459908)同源性为99.6%;与上海和香港两地区毒株亲缘性关系最近,聚为独立一簇,同属GII.4/Sydney 2012分支。结论引发该起急性胃肠炎暴发的病原体为GII.4型诺如病毒,且属于GII.4 Sydney 2012变异株。     

Molecular epidemiology of norovirus associated with gastroenteritis and emergence of norovirus GII.4 variant 2012 in Japanese pediatric patients

In late 2012, an outbreak of acute gastroenteritis due to norovirus variant Sydney_2012 occurred and have been reported from many counties. In this study, we described surveillance study of the incidence of norovirus infections among Japanese pediatric patients in association with gastroenteritis and investigated the antigenic change of the new variant Sydney_2012 circulated in Japanese populations. A total of 2381 fecal specimens collected from children with acute gastroenteritis in Hokkaido, Tokyo, Shizuoka, Kyoto, Osaka, and Saga from 2009 to 2013 were examined for norovirus and further analyzed molecularly. A high proportion (39.3%) of norovirus positive samples and several genotypes were detected. Norovirus GII.4 dominated over other genotypes (71.4%). The Den_Haag_2006b (43.2%) was detected as the predominant variant and co-circulated with New_Orleans_2009 (17.8%) until March 2012. Subsequently, they were displaced by Sydney_2012. The Sydney_2012 variant has been responsible for the majority of norovirus infections in 2012–2013 (85.7%). Although Sydney_2012 variant has a common ancestor with New_Orleans_2009 variant, analysis of P2 sub-domain showed a high level of diversity in comparison with other variants in four amino acid changes at the antigenic sites. The change in particular residue 393 of new variant may affect HBGA recognition. Analysis of noroviruses circulating in the past 4 years revealed a change of predominant variant of norovirus GII.4 in each epidemic season. The change of amino acid in putative epitopes may have led the virus escape from the existing herd immunity and explain the increase of new variant outbreaks.     

2012-2014年广东省哨点医院诺如病毒GII.4 Sydney变异株流行状况及暴发疫情特征分析

目的：分析2012年1月至2014年6月广东省哨点医院诺如病毒GII.4 Sydney变异株流行状况,以及由诺如病毒GII.4 Sydney变异株感染引起的暴发疫情特征。方法2012年1月至2014年6月在广东省选取22家医院的门诊科室为监测哨点,将采集的腹泻病例粪便样本（共10750份）送至市级CDC,进行病毒核酸提取及诺如病毒核酸检测,所有阳性样本递送至广东省CDC,并按照随机数字法抽取了855份进行诺如病毒基因分型,共成功测序样本690份。采用χ2检验比较不同年龄组、不同时期内腹泻病例诺如病毒感染情况。通过广东省突发公共卫生事件管理信息系统,收集2012年1月至2014年6月广东省13起由诺如病毒GII.4 Sydney变异株感染引起的社区暴发疫情数据,进行社区流行病学分析。结果2012年8月首次检出诺如病毒GII.4 Sydney变异株,2012年11月检出比例为13/15。2012年11月至2013年1月（T1时期）,各月份中腹泻病例诺如病毒感染阳性率分别为23.8%（100/421）、15.9%（61/383）、19.2%（95/495）,2013年11月至2014年1月（T2时期）,各月份中腹泻病例诺如病毒感染阳性率分别为17.0%（90/529）、8.7%（37/426）、11.2%（46/409）,均低于T1时期（χ2值分别为6.65、9.93、10.74,P值分别为0.010、0.002、0.001）。T1时期内≥15、<15岁组腹泻病例诺如病毒感染阳性率分别为26.3%（143/543）、14.9%（113/756）（χ2=25.90,P<0.001）;T2时期≥15、<15岁组阳性率分别为10.1%（52/516）、14.3%（121/848）（χ2=5.09,P=0.024）。13起暴发疫情中,食源性传播占10/13。结论广东省2012年8月首次检出诺如病毒GII.4 Sydney变异株,2012年11月起呈现社区流行,流行1年后,强度降低;由诺如病毒GII.4 Sydney变异株引起的暴发疫情主要以食源性传播为主。     

Emergence of GII.e as a major ORF 1 norovirus genotype and its associated ORF 2 GII.4 variant forms

The noroviruses are a major cause of outbreaks of gastroenteritis. The norovirus genotype “GII.e”, identified by ORF (Open Reading Frame) 1 nucleotide sequencing, appears to be an obligatory recombinant, in that no unique GII.e ORF 2 genotype has been identified. In 2012 GII.e norovirus became the predominant ORF 1 genotype in norovirus outbreaks in Victoria, Australia, and the current study documents changes in the ORF 1 region of GII.e norovirus since it first emerged in 2008, as well as in the ORF 2 genotypes associated with GII.e norovirus. GII.e norovirus underwent significant genetic change in ORF 1 between 2010 and 2012 and this genetic change corresponded to a significant increase in the prevalence of the virus. Nucleotide sequencing of the ORF 2 region of GII.e specimens showed that in 2008–2009, all the ORF 2 sequences corresponded to the GII.4 (2007) variant, in 2010 all the ORF 2 sequences corresponded to the GII.4 (2012-like) variant and in 2012 all the ORF 2 sequences corresponded to the GII.4 (2012) variant, the GII.4 (2012-like) variant, or the GII.4 (2009-like) variant. The evidence indicated that the development of the 2012 GII.e epidemic strains was due to evolutionary change rather than a novel recombination event. The results also support the notion that ORF 1 is critical in determining the virulence of a norovirus strain.     

2015年-2017年湖州诺如病毒感染的流行特征研究

目的研究湖州地区诺如病毒(NoV)的流行病学模式和遗传特征。方法2015年1月-2017年12月,收集急性胃肠炎患者的粪便标本,进行实时RT-PCR(qPCR)检测NoV。RT-PCR用于基因组扩增和序列测定。病毒基因型别使用在线NoV分型工具(http://www.rivm.nl/mpf/norovirus/typingtool)分型。结果NoV感染的总体流行率为23.7%(414/1757)。在414个NoV阳性标本中,393个被鉴定为NoV GⅡ,11个属于NoV GⅠ,10个是NoV GⅠ和GⅡ共同感染。共235份标本成功测序,这3年的主要流行型别是GⅡ.Pe-GⅡ.4 Sydney2012和GⅡ.P17-GⅡ.17,2017年还出现新的流行优势株GⅡ.P16-GⅡ.2。全年均检测到NoV感染,流行高峰发生在每年的冬春期间。结论在2014年10月GⅡ.17首次在湖州出现后,稳步取代了以前流行的GⅡ.4 Sydney 2012菌株。但在2016年GⅡ.4 Sydney 2012又回到主导地位,2017年出现新的流行优势株GⅡ.P16-GⅡ.2。     

Recombinant norovirus GII.g/GII.12 gastroenteritis in children

Recombinant GII.g/GII.12 norovirus (NoV) strains emerged in 2008 in Australia and subsequently have been associated with gastroenteritis outbreaks worldwide. In the winter season 2009-2010 GII.12 strains caused 16% of the NoV outbreaks in the United States. During 2009-2010 we also identified GII.g/GII.12 strains during surveillance of sporadic cases of gastroenteritis in Italian children. Severity scores were calculated for the GII.g/GII.12 NoV infections using the Vesikari scale and in two out of three paediatric cases they exceeded the median value calculated for concomitant GII.4 infections. Upon sequence analysis, the Italian strains were found to be recombinant viruses and displayed different patterns of nucleotide polymorphisms. Phylodynamic analysis with other GII.g/GII.12 recombinants showed a high rate of evolution, comparable to the rates observed for GII.4 viruses. The mechanisms leading to worldwide emergence of GII.12 NoV strains in 2008-2010 are not clear. Monitoring of GII.12 NoV circulation is necessary to understand these mechanisms of evolution.     

Novel intergenotype human norovirus recombinant GII.16/GII.3 in Bangladesh

Noroviruses (NoVs) are one of the major etiological agents of acute gastroenteritis in all age groups. In this study, we identified an intergenotype NoV recombinant strain in the fecal specimens of two male infants with acute diarrhea in Bangladesh. Phylogenetic analysis showed that the identified strains were recombinant NoV strains with a GII.3 capsid and a GII.16 polymerase gene. The recombination breakpoint was located in the ORF1/ORF2 overlap region. To the best of our knowledge this is the first report of a NoV recombinant GII.16/GII.3 strain worldwide.     

Efficacy of an intramuscular bivalent norovirus GI.1/GII.4 virus-like particle vaccine candidate in healthy US adults

BackgroundWe performed this first-in-human efficacy trial of Takeda’s bivalent norovirus vaccine candidate (TAK-214) against moderate or severe acute gastroenteritis (AGE) in healthy adults.MethodsThis double-blind, randomized, placebo-controlled phase 2b trial was conducted over two winter seasons in 18–49 year-old US Navy recruits. Participants were randomized (1:1) to receive intramuscular injections of saline placebo (N = 2,357) or TAK-214 [15 μg GI.1 and 50 μg GII.4c VLPs, 0.5 mg Al(OH)₃] (N = 2,355), and monitored for 45 days post-vaccination for AGE. Norovirus genotypes were identified by RT-PCR and sequencing of stool/vomitus samples. Sera from AGE cases were used to assess immune responses as genotype-specific histo-blood group antigen (HBGA)-blocking antibodies.FindingsWith low rates of homotypic norovirus AGE detected the statistical analysis was proactively modified to account for AGE due to any norovirus genotype. Of the 48 norovirus AGE cases of “any severity”, 29 in placebo and 19 in vaccinees, causative genotypes were GI.1 (n = 1), G1.7a (n = 1), GII.2 (n = 39) and GII.4 (n = 7). Applying predefined definitions of moderate or severe AGE gave 26 vs. 10 cases due to any norovirus genotype in placebo vs. vaccine groups, a vaccine efficacy (VE) of 61.8% (95.01% CI, 20.8 to 81.6; p = 0.0097). Five vs. one moderate or severe cases due to vaccine GI.1/GII.4 homotypic genotypes in placebo vs. vaccine arms gave a primary endpoint vaccine efficacy of 80.0% (99.99% CI, −1318.1 to 99.7; p = 0.142). Levels of GI.1 and GII.4 HBGA-blocking antibodies were increased in vaccinees and in some placebo AGE cases infected with GII.2, indicating cross-reactivity in the immune responses to different genotypes.InterpretationDespite limited cases of homotypic norovirus AGE meaning the primary endpoint was not fully evaluable, we showed TAK-214 provided statistically significant efficacy against “any moderate/severe norovirus AGE” principally caused by the heterotypic GII.2 genotype, demonstrating induction of cross-genotype protection.