鸭源鸡杆菌ompW基因缺失菌株的构建及其药物敏感性分析  

Construction of ompW Gene Deletion Strain of Gallibacterium anatis and Its Susceotibility to Antibacterial Agents

作　　者：彭志锋[1] 崔丹丹[1] 杨霞[1] 敬文宪 王忠田[2] 王坤芃 王川庆[1] 

机构地区：[1]河南农业大学禽病研究所,郑州450002 [2]中国兽医药品监察所,北京100086

出　　处：《畜牧兽医学报》2017年第7期1306-1313,共8页ACTA VETERINARIA ET ZOOTECHNICA SINICA

基　　金：国家自然科学基金(31302090);河南省自然科学基金(162300410153);河南省高校科技创新团队与支持计划资助项目(14IRTSHN015)

摘　　要：探索OmpW在鸭源鸡杆菌耐药性形成中的作用。运用M-IV培养基诱导鸭源鸡杆菌自然感受态细胞,用含有氯霉素抗性(Cmpr)筛选标记和同源臂的线性DNA打靶片段替换目的基因;以PCR、SDS-PAGE及Western blot鉴定阳性转化子ΔompW。通过微量稀释法测定12种抗菌药物对鸭源鸡杆菌亲本菌株RU和突变株ΔompW的最小抑菌浓度(MIC)。结果显示:鸭源鸡杆菌阳性转化子缺失了ompW,成功构建了ompW缺失突变菌株ΔompW;相对于RU,土霉素和四环素对ΔompW的MIC值分别降低到1/8(16/128,P<0.01)和1/32(4/128,P<0.01),其他药物的MIC值无明显变化。本研究首次利用自然转化法构建了鸭源鸡杆菌ompW缺失菌株ΔompW;OmpW在鸭源鸡杆菌抗四环素类药物中发挥重要作用。The study was conducted to explore the role of ompW gene in the drug resistance of Gallibacterium anatis（G.anatis）.The ompW gene knock-out mutant of G.anatis was constructed by natural transformation.The competent cells of G.anatis were induced following transfer of cells to M-IV medium from colonies on a blood agar plate.Then,transformation was performed by incubating the M-IV-compentent cells together with linear DNA fragment consisting of homologous arms and cmpr gene.The transformants were identified by using PCR,SDS-PAGE and Western blot analysis.The MIC-determinations of 12 kinds of antibacterial agents were performed for both wild type and mutantΔompW G.anatis using Mueller Hinton-II.The ompW gene deletion strainΔompW was successfully constructed.Comparing with wild type G.anatis,the MICs of terramycin and tetracycline forΔompW were decreased to 1/8-fold（16/128）and1/32-fold（4/128）,respectively.The MICs of other drugs in this study showed no obvious change.In conclusion,ompW deletion strain of G.anatis was constructed by natural transformation for the first time in this study.OmpW plays an important role in the formation of tetracyclines resistance phenotype of G.anatis strains.

关 键 词：鸭源鸡杆菌 OmpW 突变菌株 抗菌药物 MIC 

分 类 号：S855.12[农业科学—临床兽医学]