H7N9亚型流感病毒的遗传进化分析及对鸡的致病性研究

发布时间：2018-03-25 06:07

本文选题：禽流感病毒　切入点：H7N9　出处：《华南农业大学》2015年硕士论文

【摘要】：H7N9亚型流感感染病例于2013年2月19日在中国首次出现,至今为止人感染H7N9亚型流感病毒出现了三波疫情,不仅给人们健康带来威胁,也为我国养禽业造成了的巨大经济损失(WHO,2015),因此对H7N9亚型流感病毒的遗传进化关系及致病性进行研究具有重要的现实意义。为了解家禽H7N9亚型流感病毒的流行情况,本研究于2013年至2015年采集了8770个家禽拭子和环境样品,共分离鉴定出28株H7N9亚型流感病毒,总分离率为0.32%。其中鸭源2株,环境2株,鸡源24株;来自江苏省的毒株2株,广西的毒株1株,其它26株均分离自广东。活禽交易市场24株,养殖场4株。为研究H7N9亚型流感病毒的遗传进化关系,本研究以禽源的28株病毒和人源的1株病毒作为研究对象,分析了毒株的遗传演化规律。研究发现,第三波疫情中的14株H7N9亚型流感病毒的HA和NA基因均由第二波疫情中的病毒进化而来,且形成了两个进化分支,划分为W3-a和W3-b分支,W3-a中的毒株主要分离自广东东莞、广州和惠州,该分支中的毒株和广东、香港和广西人源毒株位于同一进化分支,具有较近的亲缘关系,推测由第二波疫情中的本地毒株进化而来。W3-b中的毒株主要分离自潮州和汕尾,该分支中的毒株与新疆、福建、贵州、江苏人源或环境中的毒株在相同分支,具有较近的亲缘关系,推测本分支的毒株完成了通过野鸟或人员流动或家禽跨省调运的传播过程。第三波疫情中14株病毒的内部基因分析结果表明,H7N9亚型流感病毒的内部基因发生着进化和重组,个别毒株发生了多元重组。对29株H7N9病毒基因主要氨基酸位点的分析结果表明,人源毒株GH074/2013在PB2基因编码蛋白的第627个氨基酸位点为K,禽源毒株G1/2013在HA基因编码蛋白的第227个氨基酸位点为Q,其它毒株均具有氨基酸位点PB2-627E、PB2-701D、HA-226L(H3 numbering)、NA-289R(N9 numbering)、M2-31N和HA裂解位点PEIPKGRG。三波疫情中H7N9亚型流感病毒的主要氨基酸位点并无显著差异。为了了解H7N9病毒对鸡的致病性情况,本研究选取了4株H7N9病毒做对SPF鸡的致病性实验。研究结果表明,4株H7N9病毒对鸡的致病力都很弱,感染4周龄SPF鸡后,所有鸡均未呈现明显的临床症状。仅在攻毒后第4d,G1组出现轻微采食量减少,其中有2只鸡精神沉郁,但第6d又恢复正常。H7N9亚型流感病毒可以通过同居方式感染其他健康鸡。选取的4株H7N9亚型流感病毒表现出不同的组织嗜性和在鸡体内的复制能力,G1毒株的组织嗜性和在鸡体内的复制能力稍强,所有毒株在肺脏组织的复制能力强。病理切片结果表明,4株H7N9亚型流感病毒对鸡的致病力弱,内脏组织仅出现轻微病变,病变产生的严重程度依次为肺脏肾脏盲肠扁桃体肝脏脾脏。
[Abstract]:The H7N9 subtype influenza infection first appeared in China on February 19, 2013. So far, there have been three waves of human infection with H7N9 subtype influenza virus, which not only poses a threat to people's health. Therefore, it is of great practical significance to study the genetic and evolutionary relationship and pathogenicity of H7N9 subtype influenza virus. In order to understand the epidemic situation of H7N9 subtype influenza virus in poultry, In this study, 8770 poultry swabs and environmental samples were collected from 2013 to 2015, 28 strains of H7N9 subtype influenza virus were isolated and identified, the total isolation rate was 0.322.There were 2 strains of duck, 2 strains of environment, 24 strains of chicken and 2 strains of virus from Jiangsu Province. One strain was isolated from Guangxi, the other 26 strains were isolated from Guangdong, 24 strains in live poultry market and 4 strains in livestock farm. To study the genetic and evolutionary relationship of H7N9 subtype influenza virus, In this study, 28 strains of avian viruses and 1 strain of human viruses were used as research objects, and the genetic evolution of the virus strains was analyzed. The HA and na genes of 14 H7N9 subtype influenza viruses in the third wave were evolved from the viruses in the second wave and formed two evolutionary branches, W3-a and W3-b. The strains of W3-a and W3-b were mainly isolated from Dongguan, Guangdong Province. In Guangzhou and Huizhou, the virulent strains in this branch are closely related to those of Guangdong, Hong Kong and Guangxi, which are located in the same evolutionary branch. It is inferred that the strains from the local strains in the second wave of the epidemic are mainly isolated from Chaozhou and Shanwei, and the strains in this branch are in the same branch as those in Xinjiang, Fujian, Guizhou and Jiangsu. Have close kinship, The results of internal gene analysis of 14 strains of influenza virus in the third wave showed that the internal genes of H7N9 subtype influenza virus had evolved and recombined. The main amino acid loci of 29 strains of H7N9 virus were analyzed. The 627th amino acid locus of PB2 gene encoding protein in human strain GH074/2013 was K. the 227th amino acid locus of HA gene protein of avian strain G1 / 2013 was Q.Amino acid site PB2-627EN PB2-701D HA-226LH3 numbering NA-289RN9 numbering M2-31N and HA cleavage site PEIPKGRGG were found in all other strains. In order to understand the pathogenicity of H7N9 virus to chicken, there is no significant difference in the main amino acid loci of H7N9 subtype influenza virus in the three waves. In this study, four strains of H7N9 virus were selected to test the pathogenicity of SPF chicken. The results showed that the pathogenicity of H7N9 virus was very weak, and 4 weeks old SPF chickens were infected. No obvious clinical symptoms were found in all chickens. There was only a slight decrease in intake in group G 1 on the 4th day after drug attack, and 2 chickens were depressed in spirit. On the 6th day, however, the influenza virus returned to normal. H7N9 subtype influenza virus could infect other healthy chickens through cohabitation. The four strains of H7N9 subtype influenza virus showed different tissue tropism and replication ability in chicken. Sex and the ability to replicate in chickens is slightly stronger, The pathogenicity of the four strains of H7N9 subtype influenza virus to chicken was weak, and the visceral tissue was only slightly damaged. The severity of the lesion was followed by lung, kidney, cecum, tonsil, liver, spleen.
【学位授予单位】：华南农业大学
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：S852.65

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