驴乳房炎大肠杆菌的分离鉴定及生物学特性研究

doi:10.16431/j.cnki.1671-7236.2018.11.028

摘要/Abstract

摘要：

为了查明导致新疆某规模化奶驴场乳房炎的主要病原菌及其生物学特性，试验采用常规微生物学方法对病原菌进行分离，采用生化鉴定管和16S rRNA的PCR扩增方法对病原菌进行鉴定，采用K-B药敏纸片法测定其耐药性，D_{600 nm}值绘制细菌生长曲线，最后分析同源性并构建系统进化树。结果显示，分离株为革兰氏阴性短杆菌，能在麦康凯培养基上长出粉红色菌落，在伊红－美兰培养基上长出有金属光泽的菌落；生化鉴定分离株的葡磷胨水、赖氨酸、鸟氨酸、棉子糖、山梨醇、侧金盏花醇、木胶糖试验均呈阳性，而苯丙氨酸和硫化氢试验均为阴性；PCR扩增后测序得到205 bp的1号菌株和203 bp的2号菌株，其特性符合大肠杆菌的特性；药敏结果表明，分离株对链霉素和氯霉素高度敏感，对青霉素、阿莫西林和庆大霉素耐药；生长曲线表明，分离株在2~16 h时为高度繁殖期，16~32 h处于稳定期，32 h以后开始进入衰退期；1号菌株与2号菌株同源性高达99%，1号菌株与大肠杆菌11J和RCB800的同源性最高（100%）；2号菌株与大肠杆菌675SK2和DTU-1的同源性最高（100%）；由系统进化树分析可知，1号菌株与大肠杆菌11J和RCB800处于同一个小分支，2号菌株与大肠杆菌675SK2和DTU-1处于同一个小分支。因此，导致本次驴乳房炎的病原菌为大肠杆菌，本研究结果为进一步系统研究导致驴乳房炎大肠杆菌的致病机制提供参考。

关键词: 驴乳房炎; 大肠杆菌; 分离鉴定; 生物学特性

Abstract:

In order to investigate the main pathogenic bacteria of mastitis and biological characteristics from a large-scale donkey farm in Xinjiang,routine microbiological methods were used to isolate pathogens,biochemical identification and 16S rRNA PCR amplification methods were used to identify pathogens,K-B susceptibility paper method was used to determine their drug resistance,D_{600 nm} value was measured,bacterial growth curves were drawn,the homology was analyzed,and the phylogenetic tree was constructed.The results showed that the isolates were short gram-negative bacilli,could grow pink and metallic luster colonies on MacConkey culture medium and Eosin-Meilan medium,respectively.The biochemical identification of isolates of phosphohydrazine,lysine,ornithine,raffinose,sorbitol,side ginsenoside and xyloglucan tested positively,while the phenylalanine and hydrogen sulfide tests were negative,and after the PCR amplification,the 205 bp No.1 strain and the 203 bp No.2 strain were obtained,their characteristics accorded with the characteristics of Escherichia coli(E.coli).Antibiotic sensitive test showed that the isolates were highly sensitive to streptomycin and chloramphenicol and resistant to penicillin,amoxicillin and gentamicin.The growth curve showed that the isolates were highly prolificated at 2 to 16 h,stabilized at 16 to 32 h,and entered the recession after 32 h.The homology of No.1 and No.2 strains was as high as 99%.The homology with E.coli 11J and RCB800 was the highest (100%);The No.2 strain had the highest homology with E.coli 675SK2 and DTU-1 (100%).From the phylogenetic tree,No.1 strain was in the same small branch with E.coli 11J and RCB800,and No.2 strain was in the same small branch with E.coli 675SK2 and DTU-1.Therefore,the pathogen of the donkey mastitis was E.coli,and its biological characteristics laid a solid theoretical basis for the further systematic study of the pathogenic mechanism of donkey mastitis E.coli.

Key words: donkey mastitis; Escherichia coli; isdation and identification; biological characteristics

中图分类号:

S858.22

高静雯, 曾宗波, 张莉, 齐亚银. 驴乳房炎大肠杆菌的分离鉴定及生物学特性研究[J]. 《中国畜牧兽医》, 2018, 45(11): 3211-3220.

GAO Jingwen, ZENG Zongbo, ZHANG Li, QI Yayin. Isolation, Identification and Biological Characteristic of Donkey Mastitis Escherichia coli[J]. , 2018, 45(11): 3211-3220.

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