9株Ⅰ群禽腺病毒的分离鉴定及hexon基因的遗传变异分析
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摘要
为了解山东省肉鸡场禽腺病毒的感染流行特点,为其科学防治提供依据,通过临床症状观察、病毒分离鉴定、鸡胚致病性试验、血凝试验、PCR检测及克隆测序等方法进行病原检测分析。结果表明,发病肉鸡主要表现为精神沉郁、采食减少等临床症状。鸡胚致病性试验可见死亡胚体呈全身出血、体型小、肝肿大现象。血凝试验显示,分离株不具凝集鸡、鸭、大鼠红细胞的特性。对hexon基因进行克隆测序分析显示,LY9株与Ⅰ群禽腺病毒血清8型亲缘性近,与参考株(58株)的核苷酸同源性为98.8%,氨基酸同源性为98.6%。其余8株分离株间的核苷酸同源性为91.3%~99.8%,氨基酸同源性为97.8%~100%;与Ⅰ群禽腺病毒血清4型参考株(HB1510株)属同一进化分支,核苷酸同源性为93.1%~100%,氨基酸同源性为98.9%~100%。结论:从山东省4家肉鸡场分离到的9株病毒均为Ⅰ群禽腺病毒。
The aim of the experiment was to provide basis for scientific prevention and control of the disease and to understand infection prevalence of FAdv in broiler farms in Shandong province. The main pathogen identification methods included clinical symptoms observation,virus isolation and identification,chicken embryo pathogenicity test,blood clotting test,PCR detection and cloning sequencing analysis. The results showed that,the main symptoms in chicken were depressed spirit,appetite reduction,swelling degeneration liver and other clinical symptoms. The chicken embryo pathogenicity test showed that the embryos were systemic bleeding,smaller and hepatomegaly. They hadn't agglutination properties to red cells of chicken,duck and rat. The results of hexon gene amplification and cloning sequencing analysis showed that the relationship between LY9 isolate and FAdv groupⅰserum 8 reference strain( 58strain) was close,nucleotide homology was 98. 8%,and amino acids homology was 98. 6%. Among the other 8 isolates,nucleotide homology was 91. 3% ~ 99. 8%,and amino acids homology was 97. 8% ~ 100%. These isolates and FAdv groupⅰserum 4 reference strain( HB1510 strain) belonged to the same evolutionary branch. Their nucleotide homology was 93. 1% ~ 100%,and amino acids homology was 98. 9% ~ 100%. The conclusion is that 9 isolates from 4 broiler farms in Shandong province are FAdv groupⅰisolates.
The aim of the experiment was to provide basis for scientific prevention and control of the disease and to understand infection prevalence of FAdv in broiler farms in Shandong province. The main pathogen identification methods included clinical symptoms observation,virus isolation and identification,chicken embryo pathogenicity test,blood clotting test,PCR detection and cloning sequencing analysis. The results showed that,the main symptoms in chicken were depressed spirit,appetite reduction,swelling degeneration liver and other clinical symptoms. The chicken embryo pathogenicity test showed that the embryos were systemic bleeding,smaller and hepatomegaly. They hadn't agglutination properties to red cells of chicken,duck and rat. The results of hexon gene amplification and cloning sequencing analysis showed that the relationship between LY9 isolate and FAdv groupⅰserum 8 reference strain( 58strain) was close,nucleotide homology was 98. 8%,and amino acids homology was 98. 6%. Among the other 8 isolates,nucleotide homology was 91. 3% ~ 99. 8%,and amino acids homology was 97. 8% ~ 100%. These isolates and FAdv groupⅰserum 4 reference strain( HB1510 strain) belonged to the same evolutionary branch. Their nucleotide homology was 93. 1% ~ 100%,and amino acids homology was 98. 9% ~ 100%. The conclusion is that 9 isolates from 4 broiler farms in Shandong province are FAdv groupⅰisolates.
引文
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[8]国纪垒,刁有祥,薛聪,等.Ⅰ群禽腺病毒山东株的分离鉴定及hexon基因的克隆与分析[J].中国兽医学报,2012,32(12):1773-1777.
[9]王小辉.Ⅰ群禽腺病毒的分离鉴定及其生物学特性研究[D].扬州:扬州大学,2008.
[10]刘东,刘红祥,于静,等.Ⅰ亚群腺病毒在我国鸡群的流行病学调查[J].中国家禽,2015,37(15):70-73.
[11]Niczyporuk J S,Wozniakowski G,Samorek-Salamonowicz E.Application of cross-priming amplification(CPA)for detection of fowl adenovirus(FAdv)strains[J].Archives of Virology,2015(4):157-163.
[12]Niczyporuk,Jowital.Phylogenetic and geographic analysis of fowl adenovirus field strains isolated from poultry in Poland[J].Archives of Virology,2016(1):859-862.
[2]Niczyporuk,Jowital.Phylogenetic and geographic analysis of fowl adenovirus field strains isolated from poultry in Poland[J].Archives of Virology,2016(1):33-42.
[3]Dinesh Mittal,Naresh Jindal,Ashok Kumar Tiwari,et al.Characterization of fowl adenoviruses associated with hydropericardium syndrome and inclusion body hepatitis in broiler chickens[J].Virus Disease,2014(1):114-119.
[4]宗玉霞,王李辉,张和平.一种新型鸡腺病毒病的流行特点及防治措施[J].当代畜牧,2016,06:52.
[5]Ayse Günes,Ana Marek,Beatrice Grafl,et al.Real-time PCR assay for universal detection and quantitation of all five species of fowl adenoviruses(FAd V-A to FAd V-E)[J].Journal of Virological Methods,2012(No.2):147-153.
[6]Helena Grgil,Peter J Krell,Eva Nagy.Comparison of fiber gene sequences of inclusion body hepatitis(IBH)and non-IBH strains of serotype 8 and 11 fowl adenoviruses[J].Virus Genes,2014(No.1):74-80.
[7]郝东敏.山东地区Ⅰ群禽腺病毒的分离鉴定及多价油乳剂灭活疫苗的研制[D].泰安:山东农业大学,2014.
[8]国纪垒,刁有祥,薛聪,等.Ⅰ群禽腺病毒山东株的分离鉴定及hexon基因的克隆与分析[J].中国兽医学报,2012,32(12):1773-1777.
[9]王小辉.Ⅰ群禽腺病毒的分离鉴定及其生物学特性研究[D].扬州:扬州大学,2008.
[10]刘东,刘红祥,于静,等.Ⅰ亚群腺病毒在我国鸡群的流行病学调查[J].中国家禽,2015,37(15):70-73.
[11]Niczyporuk J S,Wozniakowski G,Samorek-Salamonowicz E.Application of cross-priming amplification(CPA)for detection of fowl adenovirus(FAdv)strains[J].Archives of Virology,2015(4):157-163.
[12]Niczyporuk,Jowital.Phylogenetic and geographic analysis of fowl adenovirus field strains isolated from poultry in Poland[J].Archives of Virology,2016(1):859-862.