万州零售淡水鱼中副溶血性弧菌的污染状况与特征研究

Study on pollution status and characteristics of Vibrio parahaemolyticus inretail freshwater fish in Wanzhou

为了解万州零售淡水鱼中副溶血性弧菌(Vibrio parahaemolyticus,Vp)污染状况、血清型、耐药性及亲缘关系,于2022年—2023年采集万州零售淡水鱼共141份,根据GB 4789.7—2013《食品安全国家标准食品微生物学检验副溶血性弧菌检验》分离Vp和定量,PCR法特异性扩增tlh、tdh、trh、toxR、pirA、T3SS1、T3SS2和T3SS2基因,用多重PCR检测血清型,纸片扩散法检测耐药性以及肠杆菌科基因间重复序列(enterobacterial repetitive intergenic consensus, ERIC)-PCR分析亲缘关系。结果显示,141份样品中总污染率为38.3%,阳性样品定量范围为1.5~1 100 MPN/g,花鲢污染率最高(66.7%)。Vp受季节和温度的影响,夏季污染率最高(57.1%)。54株Vp全部携带tlh、toxR和T3SS1基因,1株同时携带tdh和T3SS2α基因,未检测到trh、pirA和T3SS2β基因。血清分布较分散,分为9个O群,O1为优势群(33.3%)。54株Vp对苯唑西林、青霉素和氨苄西林耐药率分别为:100%、98.1%和90.7%,对头孢他啶、头孢曲松、庆大霉素、四环素、多西环素和米诺环素100%敏感。ERIC-PCR分析表明,存在与临床分离株有遗传相似的致病菌株。该研究结果可为淡水鱼养殖、消费和临床治疗Vp导致的食源性疾病提供理论依据。

To investigate the prevalence,serotype,antibiotic resistance,and genetic relationship of Vibrio parahaemolyticus(Vp)in the retail freshwater fish in Wanzhou,141 samples of retail freshwater fish were collected from^(2)022 to 2023.The Vp was isolated and quantified according to the method of GB 4789.7—2013,and then PCR was used to amplify the thermolabile hemolysin gene(tlh),heat-resistant direct hemolysin gene(tdh),heat-resistant-related hemolysin gene(trh),transmembrane transcriptional regulator gene(toxR),Photorhabdus insect-related toxin gene(pirA),typeШsecretion system 1 gene(T3SS1),and typeШsecretion system^(2) genes(T3SS2αand T3SS2β)that were associated with acute hepatopancreas necrosis.Multiplex PCR was used to detect serotypes.The Kirby-Bauer and enterobacterial repeat intergenic consensus sequence PCR(ERIC-PCR)were used to detect antimicrobial susceptibility and genetic relationship,respectively.Results showed that 54 samples were Vp positive out of 141 samples(38.3%),When the quantitative range of positive samples was in the range of 1.5 MPN/g to 1100 MPN/g,the pollution of Aristichys nobilis was the highest(66.7%).Temperature significantly affected the abundance of Vp,and the seasonal differences in sample pollution were significant,with the highest pollution rate of 57.1%in summer.All 54 Vp isolates carried tlh,toxR,and T3SS1 genes,one isolate carried both tdh and T3SS2αgenes,and trh,pirA,and T3SS2βgenes were not found.The serotypes distribution was more dispersed and divided into nine O groups,with an O1 dominant cluster(33.3%).The resistance rates of 54 Vp strains to benzoxacillin,penicillin,and ampicillin were 100%,98.1%,and 90.7%,respectively.54 Vp strains were 100%susceptible to ceftazidime,ceftriaxone,gentamicin,tetracycline,doxycycline,and minocycline.ERIC-PCR showed that the isolate carrying pathogenic genes was genetically similar to clinical isolates.The results of this study might provide a theoretical basis for freshwater fish farming,consumption and clinical treatment of foodborne diseases caused by Vp.