2011-2012年深圳市呼吸道腺病毒分子分型及其流行特征

目的了解2011—2012年深圳地区致病性呼吸道腺病毒（AdV）的病原体基因亚型和遗传进化及其流行特征．分析深圳地区呼吸道腺病毒的流行规律和传播途径。方法收集2011年1月至2012年12月深圳地区31个流感样病例监测哨点2822份样本,先用荧光定量PCR方法筛查流感样病例,筛查后的阴性样本进一步进行呼吸道腺病毒等13种常见呼吸道病毒的核酸检测。对检测出的109份腺病毒阳性标本进行病毒分离,感染Hep-2受体细胞后共获得71份AdV稳定细胞毒株,用腺病毒六邻体基因作为靶基因,扩增其中的50份腺病毒阳性标本,纯化扩增产物后直接用于序列分析,在GenBank上进行序列比较,确定其病毒亚型并进行系统进化分析。结果2011—2012年深圳市腺病毒感染患者的高发年龄段人群为小于7岁的儿童,1～2岁婴幼儿发病率最高;4-6月和10—12月为每年发病高峰期。在2822份流感样病例咽拭子标本中,经定量PCR方法确定为AdV核酸阳性共109份,核酸阳性率为3．86％,从109份核酸阳性样本中分离病毒7l份,病毒分离率为68．93％。50份分离的呼吸道腺病毒分子分型结果为1型AdV6例,占12％;2型AdV4例,占8％;3型AdV27例,占54％;4型AdV3例,占6％;5型AdV4例,占8％;7型AdV5例,占10％;另外有复合型AdV（HAdV—un）1例,占2％。结论近两年来深圳地区呼吸道Adv感染以3型为主,腺病毒引起的呼吸道疾病主要发生在每年的4—6月,感染人群主要是1~2岁婴幼儿。     

西安市流感样病例中人腺病毒感染特点分析CSCD

目的 分析2013-2015年西安市流感样病例中非流感患者腺病毒（HAdV）感染的流行病学特点和病毒分型特征.方法 收集2013年1月到2015年12月西安市两所国家级流感监测哨点医院的流感样病例样本,经荧光PCR筛检流感病毒阴性而腺病毒阳性的标本,进行病毒分离后对HAdV六邻体（Hexon）基因扩增、测序及分型鉴定,与HAdV 1-7型的参考序列构建基因进化树.结果 2 367份标本中检测出88例腺病毒阳性,阳性率为3.72％.共检出7种亚型的HAdV,其中HAdV-1型占9.09％,HAdV-2型占22.73％,HAdV-3型占23.86％,HAdV-4型占5.68％,HAdV-5型占7.95％,HAdV-6型占3.41％,HAdV-7型占1.14％.在腺病毒阳性病例中,男性检出率高于女性,但差异不具有统计学意义（P＞0.05）;按年龄分6个组,1岁以下3例阳性,1～3岁36例阳性,4～6岁26例阳性,7 ～18岁16例阳性,19 ～59岁5例阳性,60以上2例阳性.7岁以下儿童高发,年龄组间差异具有统计学意义（Х^2=0.10,P ＜0.05）.结论 2013-2015年西安市上呼吸道感染中HAdV的主要的流行型别为3型和2型,7岁以下儿童高发;HAdV的感染全年散发,无明显季节性.     

兰州地区呼吸道感染患儿中人类博卡病毒1～3型检测与临床研究

目的了解兰州地区急性呼吸道感染患儿中人博卡病毒1—3型（HBoV1～3）感染的临床及分子流行病学特征。方法收集兰州大学第一医院2009年12月至2010年11月急性呼吸道感染患儿鼻咽分泌物及咽拭子标本524份,用：巢氏PCR扩增人博卡病毒（HBoV）NS1片段,检测HBoV1～3;同时PCR检测常见呼吸道病毒。结果524份标本中检出HBoV43例,检出率为8．2％,仅次于鼻病毒、呼吸道合胞病毒、副流感病毒3型;混合感染率为69．8％。其中人博卡病毒1型（HBoV1）在下呼吸道感染中检出率显著高于上呼吸道感染的检出率;2例人博卡病毒2型（HBoV2）患儿都出现胃肠道症状,与标准株GU048662．1的核苷酸同源性分别为99％和100％;1例人博卡病毒3型（HBoV3）与标准株HM132056．1的核苷酸同源性为99％。结论本地区儿童急性呼吸道感染中博卡病毒感染以HBoV1为主,首次检出HBoV3;人博卡病毒与其他病毒有较高的合并感染。人博卡病毒是本地区儿童急性呼吸道感染的重要病原之一。     

南京地区儿童人偏肺病毒感染的流行病学及临床特征分析

目的了解南京地区儿童人偏肺病毒（hMPV）感染的流行病学特点及临床特征。方法收集2009年8月至2010年7月南京医科大学附属南京儿童医院住院及门诊呼吸道感染患儿的鼻咽抽吸物（NPA）及咽拭子（NPS）共642例,采用逆转录聚合酶链反应法（RT—PCR）检测hMPVM基因,将阳性PCR扩增产物进行测序、同源性和进化分析。结果642例标本中共检出hMPV阳性扩增产物35份,检出率为5．5％。系统进化分析显示南京地区hMPVB1型占51．4％,A2b型占31．4％。hMPV的发病高峰在4月份。其致呼吸道感染以1岁以内多见（71．4％）。35例hMPV感染患儿中有15例（42．8％）存在混合感染,其中与HRV的混合感染检出率最高。临床诊断以肺炎（17例,48．6％）最为常见。结论人偏肺病毒是南京地区儿童急性呼吸道感染的重要病原之一,该年度其优势流行型别为B1型,南京地区A、B两型hMPV感染患儿临床特征无明显差异。     

重症急性呼吸道感染儿童中人博卡病毒的分型检测与基因进化分析

目的 了解重症急性呼吸道感染住院儿童中人博卡病毒（HBoV）的感染状况,流行病学特征及其进化特征.方法 采用巢式PCR的方法,对来自北京儿童医院重症急性呼吸道感染住院儿童的259份鼻咽抽吸物,进行人博卡病毒（HBoV）分型检测与测序,同时进行了合并感染检测、流行病学、临床特点及基因多态性分析.结果 共检出56份人博卡病毒感染阳性标本,阳性率为21.6％,[95％ CI（16.0％～27.3％）,P＜0.0001],其中2岁以下儿童感染率较高.与其他呼吸道常见病毒的合并感染率为94.6％.HBoV阳性产物测序分析发现,人博卡病毒1型占96.4％ （54/56）,2、3型各1份.HBoV阳性株分型区（VP1/VP2）序列变异不明显.结论 人博卡病毒是儿童急性呼吸道感染常见的病原体,以I型最为常见,分型区（VP1/VP2）序列较保守.HBoV在重症急性呼吸道感染儿童中是否起到真正的致病作用还需进一步的研究.     

长沙地区呼吸道感染儿童Saffold心肌炎病毒的调查

目的了解长沙地区Saffold病毒（以下简称为SAFV）在儿童呼吸道感染中的流行情况,探讨其与儿童呼吸道感染的相关性。方法选取湖南省人民医院儿科医学中心2007年11月至2008年10月间643名因呼吸道感染住院儿童的鼻咽抽吸物。采用实时荧光定量聚合酶链式反应（Realtime．PCR）方法扩增SAFV的5UTR的基因片段,并且统计分析临床资料。结果643份样本中共检测出SAFV阳性67份,阳性检出率为10．42％（67／643）,5岁以上未检测出该病毒。31例患迁延性肺炎和慢性肺炎的患儿标本检出SAFV8例（25．81％）,差异有统计学意义。结论本研究表明长沙地区下呼吸道感染住院儿童存在SAFV感染;SAFV可能与下呼吸道感染及病程迁延相关。     

兰州地区5岁以下婴幼儿病毒性腹泻的分子流行病学研究

目的调查兰州地区5岁以下婴幼儿病毒性腹泻的流行情况,了解四种主要腹泻病毒在儿童中的分布情况。方法采集2009年7月至2010年6月兰州大学第一医院儿科5岁以下腹泻患儿粪便标本290份及儿童保健中心健康婴幼儿正常粪便标本114份,采用酶联免疫吸附试验（ELISA）检测轮状病毒抗原,采用巢式聚合酶链反应对轮状病毒阳性标本进行分型;采用反转录．聚合酶链反应（RT—PCR）检测杯状病毒和星状病毒,聚合酶链反应（PCR）检测腺病毒。结果290份腹泻标本中四种病毒的阳性率分别为：轮状病毒39．31％,杯状病毒11．38％,腺病毒10．69％,星状病毒4．83％;对114份轮状病毒阳性标本G、P分型,G3型及P[8]型为优势株;114份正常标本轮状病毒检出率为0,杯状病毒检出7例,星状病毒检出1例,腺病毒检出5例。结论病毒性病原在兰州地区婴幼儿腹泻中占有重要地位,长期系统的监测具有重要意义。     

2010—2011年兰州地区5岁以下腹泻患儿杯状病毒和腺病毒感染的流行特征

目的了解兰州地区腹泻患儿中杯状病毒和腺病毒感染的分子流行病学及临床特点。方法收集兰州大学第一医院2010年7月至2011年6月腹泻患儿粪便标本295份,采用RT-PCR或PCR的方法检测杯状病毒及腺病毒,腺病毒阳性标本利用多重PCR及巢式PCR的方法分型,并对序列进行分析。结果295份粪便标本中杯状病毒的检出率为13．2％（39／295）,腺病毒的检出率是5．1％（15／295）。分型结果显示：杯状病毒中69．2％为诺如病毒,其余是札如病毒,诺如病毒中以GII-3（13例）为主,其次为GII-4（12例）,GII-6（2例）;腺病毒主要以F组的41型（10／15）为主,同时还检测到1例A组的31型,2例B组的3型及C组的1例5型和1例6型,两种病毒均主要感染2岁以下儿童,无明显的季节高峰。结论杯状病毒和腺病毒是2010—2011年兰州地区病毒性腹泻患儿的重要病原,长期监测具有重要意义。     

2010—2012年韶关市儿童呼吸道感染病毒病原学分析

目的探讨韶关市儿童常见呼吸道感染的病原学特点和分布特征。方法采集2010年7月至2012年7月因呼吸道感染于粤北人民医院的住院患者呼吸道标本171份,采用荧光定量PCR方法,对呼吸道标本同时进行甲型流感病毒（FluA）,乙型流感病毒（FluB）,腺病毒（ADV）,博卡病毒（BoV）,副流感病毒1型（PIV1）、2型（PIV2）、3型（PIV3）,鼻病毒（HRV）,呼吸道合胞病毒（RSV）,冠状病毒229E、0C43、HKUl、NL63,偏肺病毒（MPV）等14种常见呼吸道病毒核酸检测。结果171份标本中检出阳性标本93份,核酸阳性率为54．4％（93／171）,其中FluA占首位,阳性率为8．2％（14／171）,其他依次为ADV7．6％（13／171）,HRV7．6％（13／171）,PIVI／II／III 7．0％（12／171）,RSV 6.4％（11／171）,FluB5．8％（10／171）,BoV5．3％（9／171）,MPV3．5％（6／171）,冠状病毒（HCoV—OC43／HKUl）2．9％（5／171）。不同性别问患儿呼吸道病毒阳性率差异无统计学意义（P〉0．05）。≥6月龄组阳性率最低（37．5％）,1—3岁年龄组阳性率最高（62．1％）。结论韶关地区儿童发热呼吸道病毒感染病例的病原体以甲型流感病毒、腺病毒和鼻病毒为主。     

2007—2011年长沙地区急性下呼吸道感染住院儿童人偏肺病毒的流行状况

目的了解人偏肺病毒（hMPV）在长沙地区急性下呼吸道感染住院患儿中的流行病学特点。方法收集2007年9月至2011年2月因急性下呼吸道感染在湖南省人民医院儿科医学中心住院儿童的鼻咽抽吸物（nasopharyngealaspirates,NPA）样本2613份,采用逆转录聚合酶链反应法（RT—PCR）检测hMPVM基因,将阳性PCR扩增产物测序并与GenBank中已知的hMPV参考株进行比对、分析。结果2613份标本中hMPV阳性检出数135例,检出率为5．2％,男女之间的检出率比较有统计学差异（x2=8．007,P=0．003）,hMPV阳性检出患儿的年龄以1岁以内多见（63．2％）。hMPV阳性检出率在春季呈现高峰,从检出季节分布显示A2b型主要在冬春季节流行,而B2型主要在春夏季流行。135例hMPV长沙株分为A型和B型两个主要的基因型,其中A2b亚型在2007--2008年为优势流行型别,2009--2010年A2b和B2型共同流行,B2亚型在2011年呈优势流行型别。135例hMPV检出阳性患儿中有66例（48．9％）存在混合病毒感染,其中与HBoV混合检出率最高。结论长沙地区部分儿童的急性下呼吸道感染与hMPV有关,且阳性检出患儿年龄主要集中在1岁以下,男多于女,主要流行季节在春季,A2b型和B2型优势基因型在长沙地区共同流行,与其他病毒混合检出率较高。     

Adenovirus types associated with severe respiratory diseases: A retrospective 4‐year study in Kuwait

Human adenovirus (HAdV) infection can result in a severe respiratory disease. The aim of this study was to identify HAdV types detected in patients hospitalized for severe respiratory illness. The study population consisted of 743 patients with severe respiratory disease admitted to four major hospitals in Kuwait between January 2013 and December 2016. Respiratory specimens were retrospectively screened for 20 respiratory viruses by real‐time PCR. The HAdV hexon gene was amplified and directly sequenced, and HAdV types were identified by performing Bayesian phylogenetic analysis. HAdV DNA was detected in 27 (3.6%) patients, with peaks in November and March. Most patients were infants and young children suffering from pneumonia or acute bronchiolitis. The detected HAdV types were C1, C2, C5, B3, and B7. Clusters of HAdV C1, C2, and C5 were observed with high posterior probability. All patients infected with HAdV C5 and 50% of patients infected with HAdV C2 or B7 were admitted to the intensive care unit (ICU). Co‐infection with other viruses was detected in 44.4% of patients. The most common co‐infecting virus was rhinovirus (HRV). HAdV/HRV co‐infection was detected in two children who presumably developed disseminated HAdV infection and died. This is the first report describing the circulation of HAdV types associated with severe outcomes in Kuwait. These findings highlight the need for a national surveillance system to monitor changes in predominant HAdV types and increased numbers of severe respiratory infections.

     

Molecular characterization of human adenoviruses associated with respiratory infection in Turkey

Human adenoviruses (HAdVs) are responsible for various clinical diseases. Molecular epidemiological studies of respiratory HAdVs are limited in Turkey. To determine the main genotypes and epidemiological characteristics of HAdVs in patients with respiratory symptoms. HAdV PCR‐positive extracts of nasal/nasopharyngeal specimens sent to the Turkish Public Health Institution from various cities of Turkey in 2015–2016 were investigated by seminested PCR. Partial sequence analysis of the hexon gene of HAdVs was performed. SPSSv.24.0 was used. A total of 23/68 (33.82%) HAdV‐positive samples were amplified. Mastadenovirus B, C, D, and F were detected and mastadenovirus B (10/23; 43.5%) and C (10/23; 43.5%) were predominant strains. Interestingly, HAdV‐F known to have gastrointestinal system tropism was detected in two patients with respiratory symptoms. HAdV‐B3 was the most prevalent genotype (9/23; 39.1%). Also, HAdV‐B7 is defined as a reemerging pathogen. It is noteworthy that there is a cluster of four HAdV‐C strains showing a close paraphyletic relationship with HAdV‐2/6 intertypic recombination. To our knowledge, this is the first study showing that HAdV‐B7 reemerging pathogen circulating in patients with respiratory infections in our country. It is also necessary to emphasize that HAdV‐2/6 recombinant strains were detected in this study for the first time in Turkey.     

Molecular typing of clinical adenovirus specimens by an algorithm which permits detection of adenovirus coinfections and intermediate adenovirus strains

BackgroundEpidemiological data suggest that clinical outcomes of human adenovirus (HAdV) infection may be influenced by virus serotype, coinfection with multiple strains, or infection with novel intermediate strains. In this report, we propose a clinical algorithm for detecting HAdV coinfection and intermediate strains.Study designWe PCR amplified and sequenced subregions of the hexon and fiber genes of 342 HAdV-positive clinical specimens obtained from 14 surveillance laboratories. Sequences were then compared with those from 52 HAdV prototypic strains. HAdV-positive specimens that showed nucleotide sequence identity with a corresponding prototype strain were designated as being of that strain. When hexon and fiber gene sequences disagreed, or sequence identity was low, the specimens were further characterized by viral culture, plaque purification, repeat PCR with sequencing, and genome restriction enzyme digest analysis.ResultsOf the 342 HAdV-positive clinical specimens, 328 (95.9%) were single HAdV strain infections, 12 (3.5%) were coinfections, and 2 (0.6%) had intermediate strains. Coinfected specimens and intermediate HAdV strains considered together were more likely to be associated with severe illness compared to other HAdV-positive specimens (OR = 3.8; 95% CI = 1.2–11.9).ConclusionsThe majority of severe cases of HAdV illness cases occurred among immunocompromised patients. The analytic algorithm we describe here can be used to screen clinical specimens for evidence of HAdV coinfection and novel intermediate HAdV strains. This algorithm may be especially useful in investigating HAdV outbreaks and clusters of unusually severe HAdV disease.     

广州流动儿童病毒性腹泻监测

目的了解广州流动儿童病毒性腹泻的感染情况。方法收集2010年5月至2011年4月广州市白云区某社区卫生服务站241例流动儿童腹泻患者的人口资料学以及粪便标本,利用酶联免疫技术（ELISA）检测轮状病毒（RV）,采用逆转录聚合酶链反应（RT-PCR）法检测标本中诺如病毒（NVs）、星状病毒（AstV）,聚合酶链反应（PCR）法检测腺病毒（AdV）。结果 241例患者粪便标本中RV、NVs、AdV、AstV4种病毒总检出率为56.0%（135/241）,各病毒检出率分别为43.6%（105/241）、11.2%（27/241）、7.9%（19/241）、6.6%（16/241）。27株NVs阳性毒株均为GII-4;19株AdV中14株为AdV-41型,并有AdV1、2、3、31型的散在感染;AstV感染15株,均为AstV-1型。结论病毒为流动儿童腹泻的重要病因,其中轮状病毒是最主要的病原体,其他依次为NVs、AdV和AstV。     

Detection of three human adenovirus species in adults with acute respiratory infection in China

Human adenoviruses (HAdVs) are recognized as causal agents in a wide range of human diseases. However, researchers lack sufficient data on the exact HAdV species and serotypes associated with adult acute respiratory tract infections (ARTIs). To detect and characterize HAdV infections in adults in China, clinical specimens were collected from 10,310 adults with ARTIs from May 2005 to July 2010. The partial HAdV hexon gene was amplified by polymerase chain reaction (PCR), sequenced, and phylogenetically analyzed. HAdVs were detected in 86 samples (0.8%), of which 67 (77.9%) were species B (HAdV-3, -7, -11, and -14), 7 (8.1%) were species C (HAdV-1, -2, and -6), and 12 (14%) were species E (HAdV-4). HAdV-3 was the most frequently detected serotype (41/86, 47.7%), followed by HAdV-7 (13/86, 15.1%), HAdV-4 (12/86, 14.0%), and HAdV-11 (11/86, 12.8%). Patients 14–25 years old (60.5%) exhibited a higher rate of adenovirus detection than older patients. Co-infections with other respiratory viruses were observed in samples positive for HAdV species B and E. Human rhinovirus was the most commonly found virus in patients with HAdV infection. These findings provide baseline data for the surveillance and control of HAdV infection in China.     

Detection and sequence-based typing of human adenoviruses using sensitive universal primer sets for the hexon gene

The universal primer sets for identification of human adenovirus (HAdV) targeting hexon gene were designed and applied to 121 clinical samples suspected of HAdV infection. The primer sets amplified at least 20 HAdV reference strains of six species. Of these clinical samples, 81 (66.9%) samples were positive for HAdV. They were classified into 11 serotypes belonging to 5 HAdV species (B-F). The primer sets described here are sensitive and reactive to the broad spectrum of HAdV and are useful for rapid diagnosis of various HAdV infections.     

Quantitative Real-Time PCR Assay Panel for Detection and Type-Specific Identification of Epidemic Respiratory Human Adenoviruses

Outbreaks of human adenovirus (HAdV) acute respiratory illness (ARI) have been well documented among civilians and unvaccinated military recruits. Among the 7 recognized HAdV species (A to G), species B (particularly serotypes 3, 7, 11, 14, and 21) and E (serotype 4) have more often been associated with epidemic ARI. Rapid detection and type-specific identification of these viruses would enhance outbreak response and help guide prevention and control measures. To this end, we developed type-specific real-time quantitative PCR (qPCR) assays for HAdV types 3, 4, 7, 11, 14, 16, and 21 targeting the HAdV hexon gene. All type-specific qPCR assays reproducibly detected as few as 10 copies/reaction of quantified hexon recombinant plasmids with a linear dynamic range of 8 log units (10¹ to 10⁸ copies); in contrast, a generic qPCR assay that detects all HAdV types run concurrently detected between 10 and 100 copies/reaction, depending on the virus type. No nonspecific amplifications were observed with concentrated nucleic acid from 51 HAdV prototype strains or other common respiratory pathogens. All members of a panel of 137 previously typed HAdV field isolates and positive clinical specimens were correctly characterized by the type-specific qPCR assays; two different HAdV types were detected in three of the clinical specimens and confirmed by amplicon sequencing. The qPCR assays permit sensitive, specific, and quantitative detection and identification of seven clinically important respiratory HAdVs and should provide a convenient adjunct to classical typing methods for a rapid response to HAdV outbreaks.     

南京地区婴幼儿杯状病毒感染的分子流行病学研究

目的了解南京地区婴幼儿杯状病毒腹泻的感染状况、临床表现以及分子流行病学特征。方法采集2010年7月至2011年6月南京医科大学附属南京儿童医院5岁以下腹泻患儿粪便标本及儿童保健中心健康婴幼儿粪便标本各428份。采用反转录-聚合酶链反应（RT—PCR）检测杯状病毒,测序确定其基因型别。结果428份腹泻样本中有63份为杯状病毒阳性,检出率为14．72％。其中诺如病毒GⅡ型58例,未检出诺如病毒GI型,札如病毒5例,以诺如病毒GⅡ-42006b型为主要流行株。428份健康对照组标本杯状病毒检出19例,诺如病毒6例,札如病毒11例,2例为诺如病毒GⅡ型和札如病毒混合感染。结论南京地区婴幼儿中存在不同基因型杯状病毒感染,流行毒株以GⅡ-2006b为主。