鸡源山夫登堡沙门菌分离鉴定及耐药性和毒力基因检测

doi:10.16431/j.cnki.1671-7236.2024.04.044

摘要/Abstract

摘要： 【目的】确定广东某肉鸡养殖场疑似沙门菌感染病例的病原及其药物敏感性,为制定有效的防控和净化措施及合理的用药方案提供科学依据。【方法】本研究对广东省某规模化鸡场若干份病鸡的肝脏和肠道样品进行沙门菌的分离培养、革兰染色、生化试验、invA特异性基因PCR鉴定、16S rRNA鉴定、血清学分型,采用K-B纸片法检测菌株对20种抗菌药物的敏感性,并通过PCR方法检测菌株的耐药基因和毒力基因。【结果】细菌分离培养可见无色透明、呈细小的露珠样菌落或光滑圆整、透明湿润、中心为黑色的菌落,疑似菌落革兰染色呈粉红色、散在分布、两端稍圆的短杆菌,无荚膜,无芽孢;分离菌生化试验呈产气、底层产酸、产硫化氢呈黑色且斜面产碱变红现象;对疑似菌落DNA进行PCR扩增,invA特异性基因扩增出大小约为285 bp的条带,16S rRNA基因扩增出大小约为1 500 bp的条带,本研究共分离鉴定出8株山夫登堡沙门菌。药物敏感性试验结果显示,8株菌对头孢他啶、庆大霉素、阿米卡星、卡那霉素、米诺环素、强力霉素、多黏菌素B均无耐药性,但对青霉素、头孢氨苄、万古霉素、红霉素、林可霉素、氨苄西林、哌拉西林、头孢唑林、头孢呋辛钠、头孢曲松、头孢哌酮、链霉素、四环素的耐药率较高,介于87.5%~100%,且分离株均为多重耐药菌;有7株菌均检出qnrS和stcM耐药基因,有1株未检出任何耐药基因;毒力基因检测结果显示,毒力基因invA、hilA、sipA、sipC、ssrA、stnP1的检出率均为100%,sopB和ssaR基因检出率分别为50.0%和62.5%。【结论】本研究分离的鸡源山夫登堡沙门菌均具有多重耐药性,对头孢他啶、阿米卡星等7种药物敏感,invA、hilA、sipA、sipC、ssrA、stnP1 6种毒力基因的携带率均为100%。本研究结果补充了鸡源沙门菌病的流行病学资料,为临床沙门菌的防控和净化提供了理论参考。

关键词: 山夫登堡沙门菌; 分离鉴定; 耐药性; 毒力基因

Abstract: 【Objective】 This study was aimed to determine the pathogen and drug sensitivity of a suspected Salmonella infection in a broiler farm in Guangdong province,and provide scientific basis for formulating effective prevention and control and purification measures and rational antibiotic using program.【Method】 In this study,Salmonella isolation and culture,Gram staining,biochemical test,invA-specific gene PCR identification,16S rRNA identification and serotyping were performed on liver and intestinal samples of several sick chickens from a large-scale chicken farm in Guangdong province.K-B drug sensitivity method was used to detect the sensitivity of the strains to 20 kinds of antibiotics.The drug resistance genes and virulence genes were detected by PCR.【Result】 In isolation and culture,it could be seen that colorless transparent,small dewdrop-like colonies or smooth,round,transparent and moist colonies with black center could be found.The suspected colonies were pink in Gram staining,scattered in distribution and slightly round at both ends,no capsule,no spore. The biochemical test showed gas production,acid production at the bottom,hydrogen sulfide production was black,and alkali production on the inclined surface turned red.PCR was performed to amplify the suspected colonies DNA,and the invA specific gene amplified a band of about 285 bp,and the 16S rRNA amplified a band of about 1 500 bp.In this study,a total of 8 strains of Salmonella Senftenberg were isolated and identified.The results of drug susceptibility test showed that 8 strains had no resistance to ceftazidime,gentamicin,amikacin,kanamycin,minocycline,doxycycline and polymyxins B.However,the resistance rates to penicillin,cephalexin,vancomycin,erythromycin,lincomycin,ampicillin,piperacillin,cefazolin,cefuroxime sodium,ceftriaxone,cefoperazone,streptomycin and tetracycline were high,ranging from 87.5% to 100%,and the isolates were all multi-drug resistant bacteria, qnrS and stcM resistance genes were detected in 7 strains,and no resistance genes were detected in 1 strain.The test results of virulence genes showed that detection rates of invA,hilA, sipA,sipC,ssrA and stnP1 genes were 100%,and sopB and ssaR genes were 50.0% and 62.5%,respectively.【Conclusion】 All isolates of chicken-derived Salmonella Senftenberg had multi-drug resistance and were sensitive to 7 species drugs,including ceftazidme and amicarcin,but the carrying rates of six virulence genes invA,hilA,sipA,sipC,ssrA and stnP1 were 100%.The results of this study supplemented the epidemiological data of Salmonella of chicken origin and provided theoretical reference for the prevention and control and purification of clinical Salmonella.

Key words: Salmonella Senftenberg; isolation and identification; drug resistance; virulence gene

中图分类号:

S852.61

刘春红, 吕志航, 张玉倩, 廉春杨, 郑煦灿, 严常燕, 张雪莲. 鸡源山夫登堡沙门菌分离鉴定及耐药性和毒力基因检测[J]. 中国畜牧兽医, 2024, 51(4): 1773-1783.

LIU Chunhong, LYU Zhihang, ZHANG Yuqian, LIAN Chunyang, ZHENG Xucan, YAN Changyan, ZHANG Xuelian. Isolation and Identification of Chicken-derived Salmonella Senftenberg and Detection of Drug Resistance and Virulence Genes[J]. China Animal Husbandry and Veterinary Medicine, 2024, 51(4): 1773-1783.

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