广西牛星状病毒分子流行病学调查与全基因克隆及序列分析
发布时间：2021-02-20 16:27
星状病毒是小的无囊膜的单股线状RNA病毒,1975年首次通过电镜从英格兰产科病房胃肠炎的婴儿中观察到该病毒。随后,在陆生的哺乳类和禽类及水生动物中都检测到星状病毒。目前,不断发现动物和人类的新型星状病毒。星状病毒的多型性和重组可能造成宿主间的交叉感染,和与其他病毒的或者非病毒的病原混合感染,星状病毒通过不断适应经典的和新的宿主,可能导致人畜共患的星状病毒发生的风险。此外,已经有报道人和不同动物的星状病毒能引起神经症状。本研究主要针对早期发现的30年来未受重视的牛星状病毒。（1）首先对广西奶牛和水牛的牛星状病毒进行流行病学调查。从5个规模化牛场（3个奶牛场和2个水牛场）收集0-6月龄有腹泻症状小牛的直肠拭子共211份。用RT-PCR检测结果显示,牛星状病毒的阳性率43.6%（92/211）,其中奶牛阳性率46.10%（71/154）,水牛阳性率36.84%（21/57）,说明牛星状病毒在广西奶牛和水牛场普遍存在。（2）为了了解牛星状病毒与其它常见牛肠道病毒如BEV, BVDV, BCoV, BRV,BToV等的混合感染情况,从211份的直肠拭子取32份进行这些病原的检测。结果发现牛星状病...
【文章来源】：广西大学广西壮族自治区 211工程院校
【文章页数】：102 页
【学位级别】：硕士
【文章目录】：
ABSTRACT
摘要
Abbreviations
Chapter Ⅰ. LITERATURE AND REVIEW
1.1 History of astrovirus
1.2 Mamastrovirus
1.2.1 Human astrovirus
1.2.2 Animal astrovirus
1.3 Avastrovirus
1.4 Astrovirus genome
1.4.1 Non structural proteins（nsp1a,nsp1ab）
1.4.2 Structural proteins（Capsid protein）
1.5 Astrovirus pathogenesis
1.5.1 Astrovirus pathogenesis in humans
1.5.2 Astrovirus pathogenesis in animals
1.6 Astrovirus hazard in the food-chain
1.7 Astrovirus antigenicity
1.8 Astrovirus diagnostic
1.8.1 Electron microscopy and virus isolation
1.8.2 Serological essay
1.8.3 Biotechnology
1.9 Astrovirus immunization and control
1.10 Bovine astrovirus（BAstV）
1.11 Brief history of water buffalo
1.12 Aims and importance of the research
1.12.1 Importance
1.12.2 Aims
Chapter Ⅱ.MOLECULAR EPIDEMIOLOGY OF BOVINE ASTROVIRUS IN CATTLEAND WATER BUFFALO IN GUANGXI
2.0 Experimental design and technical route
2.1 Materials
2.1.1 Samples
2.1.2 Samples preparation
2.1.3 Principal Reagents
2.1.4 Reagents and culture media preparation
2.1.5 Principal Instruments and Equipments
2.2 Methods
2.2.1 Primers design
2.2.2 Total RNA extraction
2.2.3 1st strand cDNA synthesis （RT）
2.2.4 PCR detection for Astrovirus and gene amplification
2.2.5 Agarose Gel Electrophoresis （AGE）
2.2.6 Gene cloning
2.2.7 Gene sequencing
2.2.8 Results analysis and computation
2.3 Results analysis
2.3.1 RT-PCR detection results
2.3.2 Partial ORF2 amplification results
2.3.3 Trial study on co-infection of Astrovirus and other known viral enteropathogens
2.3.4 Phylogenetic analysis based on partial ORF2 of the Guangxi BAstVs
2.4 Discussion
Chapter Ⅲ. COMPLETE GENOME CLONING AND SEQUENCE ANALYSIS OF THEGUANGXI BOVINE ASTROVIRUSES （BAstV-GX/CHN/2014）
3.0 Experimental design and technical route
3.1 Introduction
3.2 Materials
3.2.1 Samples
3.2.2 Principal Reagents, instruments and equipments
3.3 Methods
3.3.1 Primers design
3.3.2 Total RNA extraction
st strand cDNA bank synthesis"> 3.3.3 1st strand cDNA bank synthesis
st strand cDNA bank synthesis'> 3.3.4 3'-end amplification 1st strand cDNA bank synthesis
st strand cDNA bank synthesis'> 3.3.5 5'-end amplification 1st strand cDNA bank synthesis
3.3.6 The complete genome PCR amplification
3.3.7 Gene cloning and sequencing
3.3.8 Computation and analysis
3.4 Results analysis
3.4.1 ORF1a analysis
3.4.2 ORF1b analysis
3.4.3 ORF2 analysis
3.4.4 Analysis of the ORF1b/ORF2 junction and the overlap region
3.4.5 Complete genome sequences analysis
3.5 Discussion
Chapter Ⅳ. GENERAL CONCLUSION
REFERENCES
ANNEXES
ACKOWLEDMENTS
【参考文献】：
期刊论文
[1]牛病毒性腹泻病毒RT-PCR检测方法的建立及应用[J]. 张淑琴,郭利,张亭亭,吴永旺,武华. 中国畜牧兽医. 2011(06)
本文编号：3043027
【文章来源】：广西大学广西壮族自治区 211工程院校
【文章页数】：102 页
【学位级别】：硕士
【文章目录】：
ABSTRACT
摘要
Abbreviations
Chapter Ⅰ. LITERATURE AND REVIEW
1.1 History of astrovirus
1.2 Mamastrovirus
1.2.1 Human astrovirus
1.2.2 Animal astrovirus
1.3 Avastrovirus
1.4 Astrovirus genome
1.4.1 Non structural proteins（nsp1a,nsp1ab）
1.4.2 Structural proteins（Capsid protein）
1.5 Astrovirus pathogenesis
1.5.1 Astrovirus pathogenesis in humans
1.5.2 Astrovirus pathogenesis in animals
1.6 Astrovirus hazard in the food-chain
1.7 Astrovirus antigenicity
1.8 Astrovirus diagnostic
1.8.1 Electron microscopy and virus isolation
1.8.2 Serological essay
1.8.3 Biotechnology
1.9 Astrovirus immunization and control
1.10 Bovine astrovirus（BAstV）
1.11 Brief history of water buffalo
1.12 Aims and importance of the research
1.12.1 Importance
1.12.2 Aims
Chapter Ⅱ.MOLECULAR EPIDEMIOLOGY OF BOVINE ASTROVIRUS IN CATTLEAND WATER BUFFALO IN GUANGXI
2.0 Experimental design and technical route
2.1 Materials
2.1.1 Samples
2.1.2 Samples preparation
2.1.3 Principal Reagents
2.1.4 Reagents and culture media preparation
2.1.5 Principal Instruments and Equipments
2.2 Methods
2.2.1 Primers design
2.2.2 Total RNA extraction
2.2.3 1st strand cDNA synthesis （RT）
2.2.4 PCR detection for Astrovirus and gene amplification
2.2.5 Agarose Gel Electrophoresis （AGE）
2.2.6 Gene cloning
2.2.7 Gene sequencing
2.2.8 Results analysis and computation
2.3 Results analysis
2.3.1 RT-PCR detection results
2.3.2 Partial ORF2 amplification results
2.3.3 Trial study on co-infection of Astrovirus and other known viral enteropathogens
2.3.4 Phylogenetic analysis based on partial ORF2 of the Guangxi BAstVs
2.4 Discussion
Chapter Ⅲ. COMPLETE GENOME CLONING AND SEQUENCE ANALYSIS OF THEGUANGXI BOVINE ASTROVIRUSES （BAstV-GX/CHN/2014）
3.0 Experimental design and technical route
3.1 Introduction
3.2 Materials
3.2.1 Samples
3.2.2 Principal Reagents, instruments and equipments
3.3 Methods
3.3.1 Primers design
3.3.2 Total RNA extraction
st strand cDNA bank synthesis"> 3.3.3 1st strand cDNA bank synthesis
st strand cDNA bank synthesis'> 3.3.4 3'-end amplification 1st strand cDNA bank synthesis
st strand cDNA bank synthesis'> 3.3.5 5'-end amplification 1st strand cDNA bank synthesis
3.3.6 The complete genome PCR amplification
3.3.7 Gene cloning and sequencing
3.3.8 Computation and analysis
3.4 Results analysis
3.4.1 ORF1a analysis
3.4.2 ORF1b analysis
3.4.3 ORF2 analysis
3.4.4 Analysis of the ORF1b/ORF2 junction and the overlap region
3.4.5 Complete genome sequences analysis
3.5 Discussion
Chapter Ⅳ. GENERAL CONCLUSION
REFERENCES
ANNEXES
ACKOWLEDMENTS
【参考文献】：
期刊论文
[1]牛病毒性腹泻病毒RT-PCR检测方法的建立及应用[J]. 张淑琴,郭利,张亭亭,吴永旺,武华. 中国畜牧兽医. 2011(06)
本文编号：3043027
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