鸭源弯曲菌的分离鉴定及其耐药性、毒力基因分析

Isolation,antimicrobial resistance profile and virulence gene analysis of Campylobacter spp.originated from duck

目的了解鸭肉生产链中弯曲菌的污染情况及其耐药性和毒力基因分布情况。方法根据GB 4789.9-2014,从肉鸭屠宰链中分离疑似弯曲菌,采用三重PCR方法,对疑似菌株进行准确鉴定;采用微量肉汤稀释法,对弯曲菌分离菌株进行8种抗菌药物敏感性试验,参考NARMS标准判定药敏试验结果;利用PCR检测与弯曲菌致病相关的4个毒力基因。结果根据形态特征、生化指标及PCR鉴定结果,从489份样品中,鉴定出空肠弯曲菌100株、结肠弯曲菌79株,其他弯曲菌8株,弯曲菌的总体分离率为38.24%,肉鸭屠宰前、脱毛环节、掏膛环节、屠宰后样品弯曲菌分离率分别为76.33%、5.62%、24.00%、0%;药敏试验结果表明弯曲菌对TET(95.72%)、CLI(90.91%)的耐药率较高,对AZI(63.64%)耐药率居中,对CIP(31.02%)、GEN(34.76%)、NAL(37.43%)、ERY(41.18%)、CHL(41.18%)的耐药率相对较低,所分离菌株多重耐药现象较为普遍,多重耐药率达72.19%。弯曲菌分离株对黏附相关基因cadF、鞭毛基因flaA、侵袭蛋白基因iamA、毒素调节基因cdtB的携带率分别为100%、80.75%、71.12%、94.65%。结论肉鸭屠宰过程中存在不同程度的弯曲菌污染,且其耐药情况较为严重,毒力相关基因广泛存在于弯曲菌中,应该加强卫生管理和抗菌药物使用监督。

To investigate the contamination conditions of Campylobacter spp. in duck production chain and its antimicrobial resistance, virulence gene distribution, samples were collected at the duck slaughterhouse according to GB 4789.9-2014. Trip- lex PCR assay was applied to identify the species of Campylobacter and the recovered Carnpylobacter strains were tested for the antimicrobial susceptibility against 8 kinds of antimicrobial agents using a broth microdilution method, the susceptibility results were determined according to the NARMS criteria （2011）. Subsequently, 4 virulence genes were detected by PCR method. The result showed that 187 Campylobacter isolates were obtained from 489 samples, including 160 C. jejuni, 130 C. coli and10 unidentified Campylobacter isolates. The total isolation rate of Campylobacter was 38. 24%. The prevalence of Campylobacter before slaughtering, at depilation stage, evis- ceration stage and duck products was 76. 33%, 5. 62%, 24.00%, and 0%, respectively. The Campylobacter isolates were most frequently resistant to tetracycline （95.72 %）, fol- lowed by resistance to clindamycin（90. 91% ）, the resistance rate of azithromycin （63.64%） was in the middle, the resist- ance rates of eiprofloxacin（31.02%）, gentamicin（34. 76%）, nalidixic acid （ 3 7. 43 %）, erythromycin （ 41. 18 % ）, chloram-phenicol （41.18%） were relatively low. The multi-drug resistance was common among Campylobacter isolates with a rate of 72.19%. The prevalence of adhesion-associated gene cadF, flagellin gene flaA, invasion associated protein gene iarnA, toxin regulation gene cdtB was 100%, 80. 75%, 71.12% and 94.65%, respectively. The results indicated that the Campylobacter contamination occurred in the slaughtering procedures of duck, and the antimicrobial resistance of Campylobacter isolates was relatively serious. In addition, the virulence-associated genes were detected widely among Campylobacter isolates. Therefore, the supervision of antimicrobial agents using at rearing stage should be strengthened, along with health management in duck production chain.