广东省某屠宰场猪源肠球菌的分离鉴定及耐药性和毒力基因分析

doi:10.16431/j.cnki.1671-7236.2023.11.038

摘要/Abstract

摘要： 【目的】了解猪源性肠球菌分离株的耐药情况及毒力因子携带情况，为肠球菌的耐药监测提供数据支持并为制定合理的治疗方案提供科学依据。【方法】本研究对广州市某大型屠宰场屠宰环节156份猪小肠样品进行肠球菌的分离培养、革兰染色、PCR鉴定，采用琼脂扩散法对菌株进行14种抗菌药物和2种消毒剂的最小抑菌浓度（MIC）测定，利用PCR技术检测耐药基因、毒力基因和消毒剂抗性基因的携带情况。【结果】本研究分离到84株肠球菌（分离率为53.85%），包括44株粪肠球菌和40株屎肠球菌；分离培养可见菌落边缘光滑整齐、圆形或卵圆形排列、菌落周围培养基呈现黑色的菌落；疑似菌落革兰染色后呈圆形的革兰阳性球菌，单个或成堆排列在一起。对疑似菌落DNA进行PCR扩增，其中粪肠球菌引物扩增出大小约941 bp的条带，16S rRNA引物扩增出大小约1 500 bp的条带。药敏试验结果显示，44株粪肠球菌对克林霉素、多西环素、头孢噻呋、庆大霉素、红霉素、苯扎氯铵和氟苯尼考的耐药率较高，分别为97.7%、90.9%、88.6%、88.6%、81.8%、81.8%和77.3%；40株屎肠球菌对克林霉素、头孢噻呋、多西环素、庆大霉素、氟苯尼考和红霉素的耐药率分别为90.0%、90.0%、85.0%、77.5%、72.5%和67.5%。毒力基因与消毒剂抗性基因检测结果显示，粪肠球菌efa、gelE、fsrC毒力基因的检出率较高，分别为95.5%、84.1%和84.1%；emeA、gsp65消毒剂抗性基因的检出率较高，分别为95.5%和90.9%。屎肠球菌仅检测到fsrB毒力基因，检出率为2.5%，无消毒剂抗性基因的检出。【结论】本研究分离的猪源肠球菌对大部分抗菌药物具有较高的耐药率，部分菌株还对利奈唑胺产生耐药，亟需加强对猪源肠球菌的流行病学监测。

关键词: 肠球菌; 耐药性; 毒力基因; 抗菌药物; 消毒剂抗性基因

Abstract: 【Objective】 The purpose of this experiment was to understand the antibiotic resistance present status and virulence factor carrier of swine origin Enterococcus, provide data support for the monitoring of Enterococcus antibiotic resistance status and provide scientific basis for formulating reasonable treatment plan.【Method】 In this study, 156 swine small intestine samples from a large slaughterhouse in Guangzhou were isolated, cultured, Gram stained, and identified by PCR.The minimum inhibitory concentration of 14 kinds of antibiotics and 2 kinds of disinfectants were determined by agar diffusion method, and the carrying of drug resistance genes, virulence genes, and antiseptic resistance genes were detected by PCR technology.【Result】 The results showed that 84 strains of Enterococcus were isolated in this research (the separation rate was 53.85%), including 44 E.faecalis and 40 E.faecium.Isolation and culture showed that the colony edges were smooth and neat, round or oval arranged, and the medium around the colony showed black colonies.Suspected colonies of Gram positive cocci that appear circular after Gram staining, arranged individually or in piles.The DNA of the suspected colony was amplified by PCR.The bands of about 941 bp were amplified by E.faecalis primer, and the bands of about 1 500 bp were amplified by 16S rRNA primer.Drug sensitivity test results showed that 44 strains of E.faecalis were highly resistant to clindamycin, doxycycline, ceftiofur, gentamicin, erythromycin, benzalkonium chloride and flufenicol, and antibiotic resistance rates were 97.7%, 90.9%, 88.6%, 88.6%, 81.8%, 81.8% and 77.3%, respectively.40 strains of E.faecium were resistant to clindamycin, ceftiofur, doxycycline, gentamicin, flufenicol and erythromycin, and the resistance rates were 90.0%, 90.0%, 85.0%, 77.5%, 72.5% and 67.5%, respectively.The frequency of efa, gelE and fsrC genes were found to be high in the E.faecalis virulence genes, and the detection rates were 95.5%, 84.1% and 84.1%, respectively.The detection antiseptic resistance gene rates of emeA and gsp65 genes were 95.5% and 90.9%, respectively.Only fsrB virulence gene was detected in E.faecium, the detection rate was 2.5%, while the isolates were negative for any antiseptic resistance genes.【Conclusion】 The isolated Enterococcus showed high resistant to most antibacterial drugs, and a part of Enterococcus were resistant to linezolid, so swine-derived Enterococcus epidemiology needed to be monitored.

Key words: Enterococcus; drug resistance; virulence genes; antibacterial drugs; antiseptic resistance gene

中图分类号:

S852.61^＋1

张永祥, 胡健欣, 陆毅兴, 胡悦莹, 卢嘉璇, 辛琳, 曾振灵, 曾东平. 广东省某屠宰场猪源肠球菌的分离鉴定及耐药性和毒力基因分析[J]. 中国畜牧兽医, 2023, 50(11): 4714-4723.

ZHANG Yongxiang, HU Jianxin, LU Yixing, HU Yueying, LU Jiaxuan, XIN Lin, ZENG Zhenling, ZENG Dongping. Isolation and Identification of Swine-derived Enterococcus and Analysis of Drug Resistance and Virulence Genes from a Slaughterhouse in Guangdong Province[J]. China Animal Husbandry and Veterinary Medicine, 2023, 50(11): 4714-4723.

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