【作者】 张烨；

【导师】 舒跃龙；

【作者基本信息】 中国疾病预防控制中心 ， 公共卫生， 2013， 硕士

【摘要】 目的：A型流感病毒可以感染野鸟和家禽,在人、猪、马、老虎等哺乳动物以及水生哺乳动物都能分离到。野生水禽是A型流感病毒的天然储存库,外表健康的鸭、鹅、鸟类,可以携带病毒并排除体外,污染环境。人感染H5N1禽流感病毒的病例中接触城乡活禽市场及禽类制品是危险因素之一。为了了解我国禽类相关环境中禽流感病毒的分布情况,2009-2012年我们对我国31个省市的城乡活禽市场、家禽养殖场,禽类屠宰加工厂等禽类相关环境中开展了禽流感病毒分布情况的调查研究。方法：2010年-2012年在全国范围内,采集城乡活禽市场、禽类养殖场、禽类屠宰加工厂环境环境标本共43944份,其中城乡活禽市场16711份,家禽养殖场23976份,禽类屠宰加工厂3257份。共采集禽类粪便12229份、笼具表面15141份、禽类饮用水7535份、清洗禽类污水4435份、宰杀处理禽类制品案板2534份,其他类型环境标本2070份。对采集的标本采用Real-time RT-PCR进行A型禽流感病毒核酸检测,核酸阳性的标本采用9日龄SPF鸡胚进行病毒分离,血凝阳性的分离物采用one-step RT-PCR鉴定HA和NA的亚型。结果：一年四季均能检测到A型禽流感病毒核酸阳性的环境样本并能分离到病毒,冬春季节高于其他季节。2010-2012南方省份环境样本A型核酸阳性率及病毒分离率分别为6.11%和1.13%,北方省份环境样本A型核酸阳性率及病毒分离率分别为0.93%和0.10%。南方省份和北方省份环境样本的A型核酸阳性率及病毒分离率之间存在显著性差异(P<0.01)。南方省份城乡活禽市场、家禽饲养场、家禽屠宰加工厂环境样本A型核酸阳性率及病毒分离率分别为10.02%、1.72%及5.74%。南方省份不同监测场所环境样本的A型核酸阳性率存在显著性差异(P<0.01)。南方省份城乡活禽市场、家禽饲养场、家禽屠宰加工厂环境样本病毒分离率分别为1.89%、0.23%及0.92%。南方省份不同监测场所环境样本的病毒分离率率存在显著性差异(P<0.01)北方省份城乡活禽市场、家禽饲养场、家禽屠宰加工厂环境样本A型核酸阳性率及病毒分离率分别为2.64%、0.30%及1.59%。北方省份不同监测场所环境样本的A型核酸阳性率存在显著性差异(P<0.01)。北方省份城乡活禽市场、家禽饲养场、家禽屠宰加工厂环境样本病毒分离率分别为0.35%、0.03%及0.13%。北方省份不同监测场所环境样本的病毒分离率率存在显著性差异(P<0.01)2010-2012年环境样本中共分离到302份禽流感病毒,H5N1亚型占57.96%,H9N2亚型占26.49%,H3、H4、H6共占12.91%,混合病毒共占2.64%。城乡活禽市场、禽类饲养场、家禽屠宰加工厂H5N1亚型禽流感病毒分离率分别为0.99%、0.02%及0.03%,不同监测场所H5N1亚型病毒分离率存在显著性差异(P<0.01)；H9N2亚型禽流感病毒的分离率分别为0.17%、0.32%及0.09%。不同监测场所H9N2亚型病毒分离率存在显著性差异(P<0.01)。经过统计学分析,城乡活禽市场和禽类饲养场不同环境样本A型核酸阳性率存在显著性差异(P<0.01)。家禽屠宰加工厂不同样本A型核酸阳性率无显著性差异(P>0.05)。结论1.我国家禽相关环境中一年四季都能检测到并分离到禽流感病毒,冬春季节是病毒分离高峰。2.南方省份环境中A型禽流感病毒核酸阳性率和病毒分离率高于北方省份。3.南北方省份中不同监测场所A型核酸阳性率及病毒分离率存在显著性差异。城乡活禽市场高于其他监测场所。4.环境样本分离病毒以H5N1及H9N2亚型为主,不同监测场所中H5N1和H9N2病毒分离率存在显著性差异,城乡活禽市场环境样本H5N1亚型病毒分离率高于其他监测场所,禽类饲养场环境样本H9N2亚型病毒分离率高于其他监测场所。5.城乡活禽市场和禽类饲养场中不同环境样本A型核酸阳性率存在显著性差异。更多还原

【Abstract】 Objectives:Influenza A virus can be isolated from birds and mammalian animals, such as humans, swine, horses, tigers and marine mammalian. Wild aquatic birds are the reservoir of influenza A virus, the apparent healthy ducks, geese and birds may shed and excrete viruses to the environments around them. One of risk factors for infection with H5N1cases are visiting live poultry markets. To investigate the prevalence of avian influenza viruses in the related environments, we performed a project to test the avian influenza virus distribution in live poultry markets, poultry farms, poultry packing factories in31Provinces in China from2009to2012.MethodsA total48706samples from poultry related environments were collected from live poultry markets, poultry farms, poultry packing factories from2009to2012. A total187911samples from live poultry markets,26269samples from poultry farms,3536samples from poultry packing factories.A total of12928feces,17268swabs from cages,8259drinking water of birds,5003water of sewage,2779swabs of chopping boards were collected. Samples were tested for influenza A nucleotide by real-time reverse transcription polymerase chain reaction (rRT-PCR) and rRT-PCR positive samples were isolated in embryonated eggs. Subtypes of the isolates with hemagglutination positive were determined by one-step rRT-PCR using specific HA and NA primers.Results:Avian influenza viruses can be isolated from the environments of live poultry markets, poultry farms, poultry packing factories in the whole year from2010to2012, with a seasonal peak during winter and spring season.For analysis the differences of variety of sampling places and sampling types for virus detection using both rRT-PCR and egg culture test, the samples were divided2parts according provinces from South or North of China.During2012-2012, In South Provinces environmental sample the nucleotide and isolation positive rate of influenza A for rRT-PCR were6.11%and1.13%respectively,in northern province environmental sample the nucleotide and isolation positive rate of influenza A were0.93%and0.10%respectively, environmental sample the nucleotide and isolation positive rate of influenza A separation rate was significant difference between southern province and northern provinces (P<0.01).In the southern province of live poultry market, poultry farms, poultry packing factories environment sample the virus nucleic acid positive rate type A influenza were10.02%,1.72%and5.74%respectively. Southern provinces different monitoring sites environmental samples of type A nucleic acid positive rate exists significant difference (P<0.01).The isolation positive rate of influenza of environment sample of live poultry markets, virus samples from poultry, poultry packing factories were1.89%,0.23%and0.92%respectively. Southern provinces different monitoring sites environmental samples of the isolation positive rate exists significant difference (P<0.01)。In the northern province of live poultry market, poultry farms, poultry packing factories environment sample the virus nucleic acid positive rate type A influenza were10.02%,1.72%and5.74%respectively. Southern provinces different monitoring sites environmental samples of type A nucleic acid positive rate exists significant difference (P<0.01)。The isolation positive rate of influenza of environment sample of live poultry markets, virus samples from poultry, poultry packing factories were1.89%,0.23%and0.92%respectively, northern provinces different monitoring sites environmental samples of the isolation positive rate exists significant difference (P<0.01)。During2012-2012, influenza302was isolated in the environmental samples. mainly H5N1subtype (57.94%) and H9N2subtype (26.49%).H3, H4, H6, H11subtypes and N6, N8, N9subtypes also has been isolated. Live poultry markets and poultry farms,H5Nl subtype of avian influenza virus separation rate is0.99%,0.02%and0.03%respectively, different monitoring sites subtype H5N1viruses isolated rate exists significant difference (P<0.01); H9N2subtype avian influenza virus separation rate were0.17%,0.32%and0.09%respectively. Different monitoring sites subtype H9N2virus separation rate exists significant difference (P<0.01).After statistical analysis, urban and rural live poultry markets and poultry farms in different environmental samples of type A nucleic acid positive rate exists significant difference (P<0.01). Poultry slaughter plant samples of different type A nucleic acid positive rate there was no significant difference (P>0.05)Conclusions1.Avian influenza viruses can be isolated from the environments of live poultry markets, poultry farms, poultry packing factories in the whole year from2010to2012, with a seasonal peak during winter and spring season.2. The rRT-PCR positive rate of influenza A virus nucleic acid in the southern province environment positive rate and virus separation rate is higher than the northern provinces.3. The different monitoring sites in north and south provinces the positive rate of influenza A virus nucleic acid and the virus isolation rate exist significant differences. Live poultry markets between urban and rural areas is higher than other places of surveillance.4. Environmental sample separation is mainly H5N1and H9N2subtype virus, H5N1and H9N2viruses in different monitoring sites.Virus separation rate exist significant differences, urban and rural live poultry market environment sample subtype H5N1virus separation rate is higher than other monitoring places, the poultry farm environment sample subtype H9N2virus separation rate is higher than other places of surveillance.5. The live poultry markets and poultry farms in different environmental samples of type A nucleic acid positive rate exists significant differences.更多还原