鸡源巴氏杆菌GZ-TZ2017株的分离及其16S rRNA序列分析
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摘要
为了探明贵州某肉鸡养殖场突发疾病病因,试验通过解剖、细菌分离、染色镜检、药敏试验、生化试验、16S rRNA的PCR扩增及序列分析等方法对病原进行分析。结果表明:从病鸡内脏中分离得到1株革兰氏阴性、两极浓染的球杆菌,将其命名为GZ-TZ2017;该菌株的培养特性、菌体形态、生化试验特征均与已报道的巴氏杆菌相似;GZ-TZ2017菌株对米诺环素、氧氟沙星和诺氟沙星等抗生素较为敏感,对头孢拉啶、苯唑西林、卡那霉素、青霉素等抗生素耐药;16S rRNA PCR扩增得到大小为1 400 bp左右的目的片段,分离菌GZ-TZ2017与各地分离的多杀性巴氏杆菌的同源性极高,均在99.0%以上;GZ-TZ2017与多杀性巴氏杆菌属细菌属于一个分支,和重庆分离株PM-LK(GenBank登录号为KX579746)、安徽滁州分离株CZ-6(GenBank登录号为KY673151)在同一小分支上,属于禽巴氏杆菌。说明该肉鸡场鸡群感染了禽巴氏杆菌。
In order to find out the causes of sudden illness in a broiler farm in Guizhou, the pathogen was analyzed by pathological examination, bacterial isolation, microscopic examination, drug sensitivity test, biochemical test and PCR amplication and sequence analysis of 16 S rRNA.The results showed that a Gram negative and two polar dense Bacillus from the viscera of the sick chicken was named GZ-TZ2017.The culture characteristics, the morphology of the bacteria and the physiological and biochemical characteristics of the strain were similar to the reported Pasteurella. GZ-TZ2017 strains were sensitive to minocycline, ofloxacin and norfloxacin, and resistant to cefradine, oxacillin, kanamycin and penicillin.16 S rRNA PCR amplified the target fragment about 1 400 bp.The isolated bacteria were very homologous to the isolated Pasteurella multocida, and all homology were above 99.0%. GZ-TZ2017 belongs to one branch of the genus Bacteria with Pasteurella multocida, and the Chongqing isolator PM-LK(GenBank No. KX579746) and the Anhui Chuzhou isolator CZ-6(GenBank No. KY673151) on the same small branch, belongs to poultry Bacillus.It shows that the chicken colony in the chicken farm is infected with poultry Bacillus.
In order to find out the causes of sudden illness in a broiler farm in Guizhou, the pathogen was analyzed by pathological examination, bacterial isolation, microscopic examination, drug sensitivity test, biochemical test and PCR amplication and sequence analysis of 16 S rRNA.The results showed that a Gram negative and two polar dense Bacillus from the viscera of the sick chicken was named GZ-TZ2017.The culture characteristics, the morphology of the bacteria and the physiological and biochemical characteristics of the strain were similar to the reported Pasteurella. GZ-TZ2017 strains were sensitive to minocycline, ofloxacin and norfloxacin, and resistant to cefradine, oxacillin, kanamycin and penicillin.16 S rRNA PCR amplified the target fragment about 1 400 bp.The isolated bacteria were very homologous to the isolated Pasteurella multocida, and all homology were above 99.0%. GZ-TZ2017 belongs to one branch of the genus Bacteria with Pasteurella multocida, and the Chongqing isolator PM-LK(GenBank No. KX579746) and the Anhui Chuzhou isolator CZ-6(GenBank No. KY673151) on the same small branch, belongs to poultry Bacillus.It shows that the chicken colony in the chicken farm is infected with poultry Bacillus.
引文
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[14] 管宇,沈志强,刘吉山,等.应用16S rRNA基因测序法鉴定禽多杀性巴氏杆菌的研究[J].中国预防兽医学报,2003,25(5):349-352.
[2] 王红勋.禽多杀性巴氏杆菌PlpB基因克隆、表达及表达蛋白生物学特性研究[D].武汉:华中农业大学,2007.
[3] 姜昆.禽巴氏杆菌病的防治措施[J].当代畜禽养殖业,2017(5):25.
[4] 于成接,丁优玲,华亚峰,等.禽霍乱巴氏杆菌的分离鉴定与致病性研究[J].中国兽医杂志,2013,49(2):21-24.
[5] BRENDA A,MENGFEI H.Pasteurella multocida:from zoonosis to cellular microbiology[J].Clin Microbiol Rev, 2013,26(3):631-655.
[6] RHOADES K R,RIMLER R B.Capsular groups of Pasteurella multocida isolated from avian hosts[J].Avian Dis,1987,31(4):895-898.
[7] 胡桂学.兽医微生物学实验教程(动物医学专业)[M].北京:中国农业大学出版社, 2006.
[8] SOPHIE K, MICHAELA L, FELIX L,et al.Pasteurella multocida involved in respiratory disease of wild chimpanzees[J].PLoS One,2011, 6(9): e24236.
[9] 杨敬.鸭源多杀性巴氏杆菌的分离鉴定和生物学特性[D].雅安:四川农业大学,2015.
[10] MBUTHIA P G,NJAQI L W,NYAQA P N,et al.Pasteurella multocida in scavenging family chickens and ducks:carrier status,age susceptibility and transmission between species[J].Avian Pathol,2008,37(1):51-57.
[11] 罗青平,杨峻,温国元,等.禽巴氏杆菌病疫苗研究进展[J].湖北畜牧兽医,2012(12):14-16.
[12] 王建兵,王凤叶.禽巴氏杆菌病病原的分离与鉴定[J].畜牧兽医杂志,2015,34(1):90-91.
[13] 杨霞,陈陆,王川庆.16S rRNA基因序列分析技术在细菌分类中应用的研究进展[J].西北农林科技大学学报(自然科学版),2008,36(2):55-60.
[14] 管宇,沈志强,刘吉山,等.应用16S rRNA基因测序法鉴定禽多杀性巴氏杆菌的研究[J].中国预防兽医学报,2003,25(5):349-352.