2019年河南省一例输入性基孔肯雅热病例的发现与处置

目的诊断输入性疑似基孔肯雅热(CHIK)病例,处置输入性CHIK疫情,防止出现本地感染病例。方法2019年8月河南省新乡市辉县发生一起疑似输入性CHIK疫情。采用荧光定量反转录聚合酶链式反应、病毒分离方法检测病例、共同暴露者、接触者血清样本和蚊虫媒介样本,采用Clustal X和MEGA 7.0软件分析基孔肯雅病毒(CHIKV)E1基因序列;根据《基孔肯雅热预防控制技术指南(2012版)》开展流行病学调查、采取相应的预防控制措施。结果病例血清样本CHIKV核酸阳性、CHIKV分离阳性。分离株属于ECSA基因型,在E1基因区段与2017年中国浙江省输入株MG912993核苷酸/氨基酸同源性为100%,与河南省既往输入株MG925665核苷酸和氨基酸同源性分别为99.4%和99.8%。流行病学调查显示,病例共同暴露者1人、接触者3人,均无相应临床表现;以病例居住地为疫点进行媒介控制和蚊媒应急监测,疫点布雷图指数由首次监测的9.5降至第3次监测的3.2并一直维持在较低水平,蚊虫媒介CHIKV核酸阴性。结论河南省新乡市辉县输入性CHIK病例由ECSA基因型CHIKV导致,未引起本地疫情扩散。     

杭州市首例本地感染登革热病例病原溯源调查

目的 对杭州市首例本地感染登革热病例开展调查及病原分子学溯源。方法 对首例本地感染登革热病例开展流行病学调查,采集患者血清进行登革病毒核酸和抗体检测。提取病毒核酸后扩增E基因并测序,利用生物信息学软件进行多序列比对排列及构建进化树。结果 该病例的登革病毒核酸及IgM抗体均为阳性,基因序列比对及进化分析,病毒株为登革病毒1型GⅣ亚型,与韩国2007年1例从菲律宾旅游输入病例分离的病毒株亲缘关系最近,核苷酸和氨基酸同源性为98.9%和99.6%,与菲律宾2010年的2株病毒核苷酸和氨基酸同源性为98.6%和99.6%,病毒株来源于菲律宾的可能性较大。结论 杭州市首例本地感染登革热病例可能是由菲律宾旅游归国的输入性病例引发的二代病例。     

2009年江西省新分离流行性乙型脑炎病毒株全基因组分子生物学特性研究

目的对2009年分离自江西省蚊虫标本的流行性乙型脑炎(乙脑)病毒JX0939株进行全基因组序列测定,分析其基因组特征。方法使用针对乙脑病毒全基因组测序引物进行PCR扩增基因组片段,PCR产物直接测序,拼接后获得全基因组序列。利用生物信息学软件进行核苷酸序列及氨基酸序列分析和病毒的系统进化分析。结果新分离乙脑病毒JX0939株基因组全长10965个核苷酸,从97位到10392位,共10296个核苷酸编码一个开放阅读框,编码3432个氨基酸。与GenBank中登录的所有59株乙脑病毒全基因组序列比较发现,其核苷酸总体差异率为1%～17%,氨基酸总体差异率为0.1～5%。与目前使用的减毒活疫苗株SA-14-14-2株相比较,只有88%的核苷酸同源性,全基因组共存在1237个核苷酸差异,83个氨基酸差异。提取GenBank登录的及本实验室测定的不同省份的乙脑病毒全基因组序列,进行全基因组序列系统进化分析显示JX0939株属于基因1型乙脑病毒。结论新分离的乙脑病毒JX0939株属于基因1型,与2007年中国分离株SH17M-07进化关系最接近,与疫苗株SA-14-14-2相比关键位点氨基酸未见变异。     

云南新分离四株流行性乙型脑炎病毒全基因组序列特征研究

目的 对云南省2009年和2010年分离的4株流行性乙型脑炎(乙脑)病毒进行全基因组序列测定和分析,阐明近期乙脑病毒流行株的分子生物学特征及基因型。 方法 通过反转录-聚合酶链反应和核苷酸序列测定方法获得病毒全基因组序列,采用Clustal X、DNAstar和Mega等生物学软件进行核苷酸和推导氨基酸序列分析及系统进化分析。 结果 2009年和2010年在云南省中部和西部地区三带喙库蚊(Culex tritaeriorhynchus)中分离到4株乙脑病毒,其中YN09M57株基因组全长10 967 bp,DH10M865、DH10M978和DHL10M62株基因组全长均为10 965 bp,均编码3432个氨基酸。这4株病毒核苷酸和氨基酸同源性分别为98.3%～99.9%和99.5%～99.9%,与来自GenBank的7株基因Ⅰ型乙脑病毒核苷酸和氨基酸同源性分别为98.3%～99.9%和99.7%～99.9%;与基因Ⅱ、Ⅲ、Ⅳ和Ⅴ型参考株核苷酸和氨基酸同源性分别在78.7%～89.7%和91.4%～98.4%;与乙脑减毒活疫苗SA14-14-2株在E蛋白编码区有15个氨基酸位点差异,均位于抗原关键位点之外。基于E基因、全基因组系统进化分析显示云南4株乙脑病毒均为基因Ⅰ型,并形成2个进化分支,分别与相邻省份和东南亚流行株进化关系较近。 结论 云南新分离4株乙脑病毒属基因Ⅰ型,虽然它们之间及其与该型参考株核苷酸和氨基酸位点存在某些差异,但决定病毒毒力和抗原性的关键位点未见明显变化。本研究提示这些乙脑病毒流行株具有稳定的遗传特性和地域特征。     

一起甲型病毒性肝炎暴发疫情的病原学诊断与基因分析

目的 查明引起2004年宁波局部地区甲型病毒性肝炎（甲肝）暴发疫情的病原并分析其基因特征。 方法 用酶联免疫吸附试验检测血清中的甲肝病毒IgM抗体（HAV IgM）和戊型肝炎病毒IgM抗体（HEV IgM），利用逆转录-聚合酶链反应（RT PCR）检测粪便样品中的HAV核酸（RNA）并扩增VP1基因，测序结果利用DNA star软件进行比对分析。 结果　172份血清样品中HAV IgM均为阳性，HEV IgM均为阴性。172份粪便样品中共检测到HAV RNA阳性样品74份，占43.0%。选择5份样品进行HAV VP1基因扩增并测序，测序结果经过比对后发现4株宁波株(NB2004-10、NB2004-11、 NB2004-51和 NB2004-71)的VP1基因是完全一致的；NB2004-75与以上4株的核苷酸的同源性为99.9％，氨基酸的同源性为99.7％；宁波株与HAV各基因型标准株比较， 与IA亚型毒株AH2的核苷酸和氨基酸的同源性最高， 分别为98.9％和99.7%～100.0％；与中国其他地区流行株比较，核苷酸和氨基酸的同源性波动于89.4%～96.7%和97.0％~100.0％。结论　引起这次肝炎暴发疫情的病原是甲肝病毒，其基因型为IA亚型，与日本株AH2、AH3亲缘关系最近，与中国其他地区流行株处于不同的进化簇上。     

双重荧光定量PCR检测西尼罗病毒和基孔肯雅病毒

摘要：目的：建立同时检测西尼罗病毒（WNV）、基孔肯雅病毒（CHIKV）的双重荧光定量PCR法,为临床疑似病例的诊断提供依据。 方法：分别针对WNV CAP基因、CHIKV E1基因保守区设计特异性引物和TaqMan探针,建立并优化双重荧光定量PCR反应体系,评价方法的特异性和灵敏度。 结果：建立的双重荧光定量PCR可同时检测WNV、CHIKV核酸,标准曲线相关系数（r）分别达0.999、0.998,灵敏度达10 copies/μL,具有良好的特异性。 结论：建立了同时检测WNV、CHIKV的双重荧光定量PCR法,但尚需临床进一步验证。     

浙江省台州市两株狂犬病病毒流行株的基因序列分析

目的分析浙江省台州市2株狂犬病病毒核蛋白及糖蛋白的基因序列,了解狂犬病病毒野毒株与人用及兽用狂犬病疫苗株间的差异。方法以免疫荧光法检测2008年采自台州市的144只犬脑标本,以RT-(nested)PCR法扩增病毒核蛋白及糖蛋白全基因,克隆测序后以生物信息学软件进行遗传特征分析。结果检测到2株狂犬病病毒阳性样品,序列分析表明两样品所携病毒均为基因1型狂犬病病毒,所携病毒的N基因核苷酸及氨基酸同源性均为100%,G基因核苷酸和氨基酸同源性分别为99.8%和99.4%。所携病毒与CTN疫苗株核苷酸同源性较高,N基因与G基因分别为88.8%和85.9%~86.1%,两样品所携病毒与Hep-Flury疫苗株的核蛋白氨基酸同源性最高,为97.6%,CTN次之,为97.1%,与CTN糖蛋白氨基酸同源性较高,为92.3%~92.5%。与诸基因1型狂犬病病毒参考株相比,两样品所携病毒与浙江温州Wz0(H)、衢州株ZJ-QZ及印度尼西亚株病毒同源性最高。系统发育分析表明XY20及JJ22与浙江温州Wz0(H)、衢州株ZJ-QZ以及印度尼西亚株、疫苗株CTN,以及泰国与马来西亚株进化关系最近,而与疫苗株aG、PV、ERA,及标准攻击毒CVS株等进化关系较远。结论两份阳性犬脑所携狂犬病病毒是基因1型狂犬病病毒,但无论是N基因还是G基因的核苷酸序列,以及推导出来的氨基酸序列与已知的1型狂犬病病毒株及疫苗株存在差异。     

2015年浙江省洞头区病毒性脑炎暴发疫情病原学研究

目的 了解2015年11月浙江省温州市洞头区鹿西乡疑似病毒性脑炎暴发疫情的病原及基因特征,分析其基因变异情况并进行病原溯源。方法 开展流行病学调查,采集32例患者和75例密切接触者的咽拭子和粪便样本共140份,检测人类肠道病毒（HEV）核酸和分离病毒,对HEV分离株VP1基因测序,进行同源性与进化分析。结果 疫情波及鹿西乡小学和幼儿园共计396名学生,其中发病32例,罹患率8.1%（32/396）。临床症状为发热、头痛、呕吐等。从32例患者和75例密切接触者样本中,分别检测到30例和5例HEV核酸阳性,阳性率分别为93.8%和6.7%,共分离到23株肠道埃可病毒30型（ECHO30）。ECHO30分离株之间VP1区核苷酸和氨基酸同源性分别为99.9%~100.0%和99.7%~100.0%,新分离株与ECHO30原型株之间VP1区核苷酸和氨基酸同源性分别为81.8%和92.1%。系统进化分析显示新分离株位于ECHO30进化支Ⅰ的基因亚型分支上,与2013年浙江省CHN/ZJ/RA-72/2013株,2014年ZJ/JH Chen/2014株亲缘关系最近。结论 引起2015年11月浙江省温州市洞头区鹿西小学及幼儿园聚集性病毒性脑炎暴发疫情的病原为ECHO30 GⅠ进化支病毒。     

广州市境外输入性基孔肯雅热病例的流行病学和病原学特征分析

目的 了解2017-2019年广州市输入性基孔肯雅热(CHIKF)病例流行特征和基孔肯雅病毒(CHIKV)E1基因特征,为疫情防治工作提供科学依据.方法 使用SPSS 19.0软件,采用描述性流行病学方法分析2017-2019年广州市输入性CHIKF病例流行特征,并以CHIKV的E1基因序列进行同源分析和构建系统发育树...     

天津市输入性基孔肯亚病毒全基因组序列分析

目的:分析天津市1例输入性基孔肯亚病毒（CHIKV）的全基因组序列特征与进化,为CHIKV的监测与防控提供科学依据。方法:收集2019年11月4日于天津市第二人民医院采集的血清标本200 μl提取核酸,设计叠瓦式引物扩增CHIKV基因组全长,然后利用Illumina Miniseq平台进行高通量测序。结果:本研究的CH...     

Chikungunya virus and the safety of plasma products

BACKGROUND: Chikungunya virus (CHIKV) outbreaks were previously restricted to parts of Africa, Indian Ocean Islands, South Asia, and Southeast Asia. In 2007, however, the first autochthonous CHIKV transmission was reported in Europe. High‐level viremia, a mosquito vector that is also present in large urban areas of Europe and America, and uncertainty around the resistance of this Alphavirus toward physiochemical inactivation processes raised concerns about the safety of plasma derivatives. To verify the safety margins of plasma products with respect to CHIKV, commonly used virus inactivation steps were investigated for their effectiveness to inactivate this newly emerging virus. STUDY DESIGN AND METHODS: Pasteurization for human serum albumin (HSA), vapor heating for Factor VIII inhibitor bypassing activity, solvent/detergent (S/D) treatment for intravenous immunoglobulin (IVIG), and incubation at low pH for IVIG were investigated for their capacity to inactivate CHIKV and the closely related Sindbis virus (SINV). The obtained results were compared to previous studies with West Nile virus and the commonly used model virus bovine viral diarrhea virus. RESULTS: The data generated demonstrate the effective inactivation of CHIKV as well as SINV by the inactivation steps investigated and thereby support results from earlier validation studies in which model viruses were used. CONCLUSION: High inactivation capacities with respect to CHIKV were demonstrated. This provides solid reassurance for the safety of plasma products and the results verify that the use of model viruses is appropriate to predict the inactivation characteristics of newly emerging viruses when their physicochemical properties are well characterized.     

The Potential Impact of Chikungunya Virus Outbreaks on Blood Transfusion

Chikungunya virus (CHIKV) is responsible for large periodic epidemics in both endemic and nonendemic areas where competent mosquitoes are present. Transmission of CHIKV by transfusion during explosive outbreaks has never been documented, and the true impact of CHIKV infection on blood transfusion during an outbreak is unknown. Considerations include not only transfusions in the active outbreak areas but also returning travelers to nonendemic areas. Because there are no documented cases of transfusion-transmitted CHIKV, there are no standard guidelines regarding transfusion policies during a chikungunya fever outbreak. We review current information from studies during outbreaks with the goal of estimating the potential effect of different blood safety interventions (eg, querying donors for possible CHIKV exposure, chikungunya fever–related symptoms, screening for CHIKV RNA).     

Chikungunya disease in nonhuman primates involves long-term viral persistence in macrophages

Chikungunya virus (CHIKV) is a mosquito-borne alphavirus that induces in humans a disease characterized by fever, rash, and pain in muscles and joints. The recent emergence or reemergence of CHIKV in the Indian Ocean Islands and India has stressed the need to better understand the pathogenesis of this disease. Previous CHIKV disease models have used young or immunodeficient mice, but these do not recapitulate human disease patterns and are unsuitable for testing immune-based therapies. Herein, we describe what we believe to be a new model for CHIKV infection in adult, immunocompetent cynomolgus macaques. CHIKV infection in these animals recapitulated the viral, clinical, and pathological features observed in human disease. In the macaques, long-term CHIKV infection was observed in joints, muscles, lymphoid organs, and liver, which could explain the long-lasting CHIKV disease symptoms observed in humans. In addition, the study identified macrophages as the main cellular reservoirs during the late stages of CHIKV infection in vivo. This model of CHIKV physiopathology should allow the development of new therapeutic and/or prophylactic strategies.     

浙江省温州市首例输入性基孔肯雅热病例流行病学调查

2017年9月9日1名由孟加拉国入境的商人经广州出入境检验检疫局白云机场口岸旅检科确认为基孔肯雅热病毒核酸阳性;当天该商人由广州机场转机到达温州市入住市区酒店,经调查证实为1例输入性基孔肯雅热病例且为温州市首例,温州市疾病预防控制中心及时采取一系列防控措施后未发生二代病例。应对此类疾病加强监测,积极提高应对能力,预防可能出现的暴发与流行。     

Estimated risk of Chikungunya viremic blood donation during an epidemic on Reunion Island in the Indian Ocean, 2005 to 2007

BACKGROUND: Between 2005 and 2007, Chikungunya virus (CHIKV) caused a massive epidemic on Reunion Island with a major peak in the number of cases in February 2006. Blood donation was interrupted on the island in January 2006. STUDY DESIGN AND METHODS: Estimates of the mean risk of viremic blood donation on Reunion Island were computed for different phases of the epidemic. Calculations used CHIKV incidence estimates derived from sentinel surveillance, duration of viremia, and frequency of asymptomatic infection. Data on these two last parameters were initially based on hypotheses and subsequently obtained from studies carried out during the outbreak. The estimated risk was compared to the results of CHIKV nucleic acid testing (NAT) implemented for platelet (PLT) donations screening. RESULTS: Over the course of the outbreak, the mean risk was estimated at 132 per 100,000 donations. The risk peaked at 1,500 per 100,000 donations at the height of the outbreak in February 2006. In total, 47 blood donations would have been potentially viremic if blood collection had not been interrupted. During this period, an estimated 312,500 of 757,000 inhabitants had been infected by mosquito-borne transmission. From January to May 2006, the estimated mean risk (0.7%) and observed risk on PLT donations (0.4%) were of the same order of magnitude. CONCLUSION: During this large outbreak, the estimated risk of viremic blood donation was high, but low compared to the risk of mosquito-borne CHIKV transmission. The estimated risk was corroborated by the concordant results with the observed risk.     

Clinical and radiological features of imported chikungunya fever in Japan: a study of six cases at the National Center for Global Health and Medicine

Chikungunya fever (CHIKF) is currently distributed in Africa and in South and Southeast Asia; outbreaks have occurred periodically in the region over the past 50 years. After a large outbreak had occurred in countries in the western Indian Ocean region in 2005, several countries reported cases of CHIKF from travelers who had visited affected areas. In Japan, there have been only 15 cases of CHIKF patients so far, according to the National Institute of Infectious Diseases. Therefore, to evaluate the clinical and radiological features associated with the disease, we describe 6 imported cases of CHIKF. All of the patients had had prolonged arthralgia on admission to our hospital, and diagnosis was confirmed with specific antibodies by using an IgM-capture enzyme-linked immunoassay and a plaque reduction neutralizing antibody assay. Magnetic resonance imaging (MRI) of one patient revealed erosive arthritis and tenosynovitis during the convalescence stage. Clinicians should be aware of the late consequences of infection by the chikungunya virus (CHIKV) and recognize the possible association of subacute and chronic arthritis features. In addition, competent vectors of CHIKV, Aedes aegypti, can now be found in many temperate areas of the eastern and western hemispheres, including Japan. This fact raises concern that the virus could be introduced and become established in these areas. This necessitates an increased awareness of the disease, because imported cases are likely to contribute to the spread of CHIKV infection wherever the competent mosquito vectors are distributed.