番鸭源鸭疫里默氏杆菌的分离鉴定及其耐药性分析

为了解番鸭源鸭疫里默氏杆菌的耐药性，本研究对疑似鸭疫里默氏杆菌感染的发病番鸭进行细菌分离培养、革兰氏染色镜检、病鸭病原检测、生化试验、16S rRNA序列分析、PCR鉴定、药敏试验和耐药基因检测。细菌分离结果显示，分离菌在鲜血琼脂培养基上长出表面光滑、边缘整齐、有光泽、半透明的奶油状针尖大小菌落；革兰氏染色镜检呈革兰氏阴性短小杆菌，命名为GZQN201907。GZQN201907生化试验中尿素反应阳性，葡萄糖、麦芽糖、乳糖等生化反应呈阴性。其16S rRNA系统进化树与鸭疫里默氏杆菌处于同一分支；并且分离菌鸭疫里默氏杆菌OmpA基因PCR鉴定结果为阳性。其对头孢呋辛、红霉素、头孢他啶等18种抗菌药耐药，对羧苄西林和环丙沙星中度敏感，对新霉素和复方新诺明敏感。而耐药基因能检测出β-内酰胺类耐药基因VIM、TEM，四环素类耐药基因tetB，大环内酯类耐药基因ermB和ermF。药敏试验与耐药基因检测结果说明，GZQN201907对β-内酰胺类、四环素类、大环内酯类3类药物的耐药表型和耐药基因检测结果一致。动物回归试验中接种GZQN201907的雏鸭在72 h内全部死亡，而对照组雏鸭未出现任何症状，说明GZQN201907对雏鸭有致病力。试验成功分离到1株番鸭源鸭疫里默氏杆菌，为鸭疫里默氏杆菌病的防治奠定基础。

Isolation,Identification and Drug Resistance Analysis of Riemerella anatipestifer from Muscovy Duck

In order to understand the drug resistance of Riemerella anatipestifer from Muscovy duck,this study carried out bacterial isolation and culture,Gram staining microscopy,pathogen detection,biochemical test,16S rRNA sequence analysis,PCR identification,drug sensitivity test and drug resistance gene detection for Muscovy duck suspected of Riemerella anatipestifer infection.The results of bacterial isolation showed that on the blood agar medium,the isolated bacteria grew creamy needle tip size colonies with smooth surface,neat edge,luster and translucency.The Gram-negative bacillus brevis was detected by Gram-negative staining microscopy,and it was named GZQN201907.In the biochemical test of GZQN201907,urea reaction was positive,but glucose,maltose,lactose and other biochemical reactions were negative.The 16S rRNA phylogenetic tree was in the same branch as Riemerella anatipestifer.And the OmpA gene of PCR identification results of the isolated strain were positive.The drug sensitivity test was sensitive to 18 antibiotics including cefuroxime,erythromycin and ceftazidime,moderately sensitive to carboxypicillin and ciprofloxacin,and sensitive to neomycin and cotrimoxazole.And the resistance genes could detect the β-lactam resistance genes VIM and TEM,tetracycline resistance genes tetB,macrolide resistance genes ermB and ermF.The results of drug sensitivity test and drug resistance gene detection indicated that GZQN201907 showed the same resistance phenotype and gene detection results for β-lactam,tetracycline and macrolide.In the animal regression test,all the ducklings inoculated with GZQN201907 died within 72 h,while the control group showed no symptoms,indicating that GZQN201907 was virulent to the ducklings.One strain of Riemerella anatipestifer from Muscovy duck was successfully isolated,which laid a foundation for the prevention and treatment of Riemerella anatipestifer from muscovich.