鸡源大肠杆菌生物被膜形成与耐药性、毒力基因的关联性分析  

Association analysis between biofilm formation capacity of Escherichia coli derived from chickens and drug resistance and virulence genes

作　　者：陶梦珂 李苗苗 石晴晴 赵恒 张鲁星 胡功政[1] 潘玉善[1] 刘建华[1] TAO Mengke;LI Miaomiao;SHI Qingqing;ZHAO Heng;ZHANG Luxing;HU Gongzheng;PAN Yushan;LIU Jianhua(College of Veterinary Medicine,Henan Agricultural University,Zhengzhou 450046,China)

机构地区：[1]河南农业大学动物医学院,河南郑州450046

出　　处：《畜牧与兽医》2024年第6期86-93,共8页Animal Husbandry & Veterinary Medicine

基　　金：国家自然科学基金面上项目(32072914);河南省重点研发项目(221111111300)。

摘　　要：旨在了解鸡源大肠杆菌临床分离株生物被膜形成能力与耐药性、毒力基因之间的关系。通过刚果红试验、结晶紫染色法测定生物被膜形成能力;采用微量肉汤稀释法检测分离株对阿莫西林、氟苯尼考、庆大霉素等8种抗菌药的耐药性;PCR检测常见耐药、毒力基因携带情况;采用实时荧光定量PCR检测毒力基因在生物被膜阳性菌中的表达量。结果表明,在51株分离株中,生物被膜阳性菌株比例为19.61%,阳性菌株中,OD_(600)最大值为2.233±0.702,最小值为0.374±0.099,其他介于1.455~1.604之间;耐药性检测显示,生物被膜阳性菌株对头孢喹肟、氟苯尼考、庆大霉素、阿米卡星、黏菌素的耐药率均高于阴性菌株,其中对阿米卡星和黏菌素差异显著(P<0.05),但对多西环素耐药率低于阴性菌株,差异显著(P<0.05),二者对阿莫西林+克拉维酸、恩诺沙星耐药率无差异;OD_(600)值最大的E13对8种测定药物全部耐药,6株OD_(600)值介于1.455~1.604的菌株对5种及以上抗菌药产生耐药性,OD_(600)值为0.374的菌株E39为敏感菌株。PCR结果显示耐药基因bla_(CTX-M-9)、bla_(CTX-M-U)、bla_(OXA-1)、mcr-1、rmtB、floR在生物被膜阳性菌株中的检出率高于阴性菌株,菌株E13同时携带9种耐药基因;共检出16种毒力基因,其中fimH、astA、aggR、irp1、iucD、fyuA、ybtA、ompA在生物被膜阳性菌株中的检出率高于阴性菌株,且阳性菌株毒力基因型复杂;以OD_(600)值为0.374的E39菌株为对照,OD_(600)值较大的生物被膜阳性菌株中,fimH、ompA、ompA、iucD、hlyF毒力基因表达呈现上调趋势。综上,与生物被膜阴性菌株相比,大肠杆菌生物被膜阳性菌株耐药更为严重,携带毒力基因型更加复杂多样,且毒力基因表达呈现上调趋势。This study was to identify the correlation between biofilm-forming capacity,and drug resistance and virulence genes of the clini⁃cal isolates of Escherichia coli from chickens.The biofilm formation ability was assessed by crystal violet staining,while the broth microdilu⁃tion method was utilized to determine the susceptibility of the bacterial isolates to eight antimicrobial agents,including amoxicillin,florfenicol and gentamicin,etc.PCR tests were performed to detect the presence of common drug resistance and virulence genes of the isolates,while real-time PCR was utilized to quantify the expression of virulence genes in biofilm-positive bacteria.The results revealed that among the 51 isolates,10 strains were positive for biofilm formation,with a positivity rate of 19.61%.Among the positive strains,the highest OD_(600) value recorded was 2.233±0.702,while the lowest value was 0.374±0.099,and all the other values ranged between 1.455 and 1.604.The re⁃sults of the resistance test showed that the biofilm-positive strains demonstrated significantly higher levels of resistance to cefquinome,flor⁃fenicol,gentamicin,amikacin and colistin(P<0.05),but lower resistance to doxycycline,as compared with the negative strains(P<0.05).The resistance rates of amoxicillin+clavulanic acid and enrofloxacin did not exhibit significant differences.E13 displayed the highest OD_(600) value and presented resistance to all the eight measured drugs.Additionally,six of the strains exhibiting OD_(600) values ranging from 1.455 to 1.604 were found to possess resistance to five or more antimicrobial agents.Strain E39,with an OD_(600) value of 0.374,was deter⁃mined susceptible.The PCR results showed a higher prevalence of the resistance genes bla_(CTX-M-9),bla_(CTX-M-U),bla_(OXA-1),mcr-1,rmtB,and floR in the biofilm-positive strains,compared with the negative ones.Additionally,E13 was found to harbor 9 resistance genes.A total of 16 virulence genes were identified,with fimH,astA,aggR,irp1,iucD,fyuA,ybtA and ompA exhibting higher pr

关 键 词：大肠杆菌 生物被膜 毒力基因 耐药基因 耐药性 

分 类 号：S855[农业科学—临床兽医学]