河南地区猪源性大肠杆菌耐药性及耐药基因型分析
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摘要
【目的】掌握河南地区猪源性大肠杆菌的耐药情况及不同的耐药基因类型,为临床防治用药提供依据。【方法】以从河南地区规模化猪场中分离鉴定的23株大肠杆菌为研究对象,选用21种抗生素药物对大肠杆菌耐药性进行分析,用PCR方法对耐药质粒DNA进行扩增,并对氨基糖苷类(aac(3)-Ⅳ、aph(3′)-Ⅱ)、四环素类(tetD、tetB)、β-内酰胺类(TEM、SHV)、大环内酯类(ermB、ermC)等4类耐药基因型进行检测。【结果】23株致病性大肠杆菌对抗生素类药物均有不同程度的耐药性,其中对青霉素、红霉素、四环素、卡那霉素、多西环素、氟哌酸、氨苄西林等7种抗生素的耐药性较强,耐药率分别为100.0%,91.3%,86.9%,78.2%,78.2%,73.9%和73.9%,且均在70%以上。23株大肠杆菌检出的耐药基因包括氨基糖苷类aac(3)-Ⅳ和aph(3′)-Ⅱ、四环素类tetD和tetB及β-内酰胺类SHV、大环内酯类ermC等6个基因型,其与GenBank数据库中相关序列的同源性均在97%以上,未检出TEM和ermB。【结论】河南地区大肠杆菌耐药基因类型多而复杂,耐药性严重并存在多重耐药性。
【Objective】This study investigated the drug-resistance of pig-derived Escherichia coli in Henan and types of drug resistance genes to provide basis for its clinical prevention and treatment.【Method】A total of 23 strains of E.coli isolated from Henan pig farms were selected and their drug resistances were analyzed using 21 antibiotics.The resistant plasmid DNA was amplified by PCR and 4 types of resistant genotypes including aminoglycosides(aac(3)-Ⅳ and aph(3′)-Ⅱ),tetracycline(tetDand tetB),beta lactam(TEMand SHV),and macrolide(ermBand ermC)were detected.【Result】The 23 pathogenic E.coli showed different resistances to the antibiotics.The strains had high resistance to seven antibiotics including penicillin,erythromycin,tetracycline,kanamycin,doxycycline,norfloxacin and ampicillin with resistance rates of 100%,91.3%,86.9%,78.2%,78.2%,73.9% and 73.9%,respectively.Resistance genes detected in the 23 strains of E.coli included aminoglycosides aac(3)-Ⅳ and aph(3′)-Ⅱ,tetracycline tetD and tetB,β-lactamSHV,and macrolide ermC,and they had above 97% homology with related sequences in the GenBank database.TEM and ermB were not detected.【Conclusion】The types of E.coli resistance genes in Henan had large number and were complex,and the drug resistance was severe and mixed.
【Objective】This study investigated the drug-resistance of pig-derived Escherichia coli in Henan and types of drug resistance genes to provide basis for its clinical prevention and treatment.【Method】A total of 23 strains of E.coli isolated from Henan pig farms were selected and their drug resistances were analyzed using 21 antibiotics.The resistant plasmid DNA was amplified by PCR and 4 types of resistant genotypes including aminoglycosides(aac(3)-Ⅳ and aph(3′)-Ⅱ),tetracycline(tetDand tetB),beta lactam(TEMand SHV),and macrolide(ermBand ermC)were detected.【Result】The 23 pathogenic E.coli showed different resistances to the antibiotics.The strains had high resistance to seven antibiotics including penicillin,erythromycin,tetracycline,kanamycin,doxycycline,norfloxacin and ampicillin with resistance rates of 100%,91.3%,86.9%,78.2%,78.2%,73.9% and 73.9%,respectively.Resistance genes detected in the 23 strains of E.coli included aminoglycosides aac(3)-Ⅳ and aph(3′)-Ⅱ,tetracycline tetD and tetB,β-lactamSHV,and macrolide ermC,and they had above 97% homology with related sequences in the GenBank database.TEM and ermB were not detected.【Conclusion】The types of E.coli resistance genes in Henan had large number and were complex,and the drug resistance was severe and mixed.
引文
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[12]Guillaume G,Verbrugge D,Chasseur-Libotte M,et al.PCR typing of tetracycline resistance determinants(Tet A-E)in Salmonella enterica serotype Hadar and in the microbial community of activated sludges from hospital and urban wastewater treatment facilities in Belgium[J].FEMS Microbiol Ecol,2000,32(1):77-85.
[13]董杰.细菌获得性抗生素耐药基因研究进展[J].中国预防医学杂志,2015(1):71-74.Dong J.Research progress on antibiotic resistant gene of bacterial acquired antibiotics[J].Chinese Journal of Preventive Medicine,2015(1):71-74.
[14]戴秀美,常维山.2012年我国部分地区猪源大肠杆菌耐药性分析[J].畜禽业,2013(1):50-52.Dai X M,Chang W S.Analysis of resistance of Escherichia coli in pig sources in some areas of China in 2012[J].Livestock and Poultry Industry,2013(1):50-52.
[15]朱力军.动物大肠杆菌耐药性的变化趋势[J].中国兽医杂志,2001,35(2):16-18.Zhu L J.The trend of resistance to Escherichia coli in animals[J].Chinese Journal of Veterinary Medicine,2001,35(2):16-18.
[16]Yan J J,Ko W C,Tsai S H,et al.Dissemination of CTX-M-3and CMY-2beta-lactamaases among clinical isolat es of Escherichia coli in southern Taiwan[J].J Clin Microbiol,2000,38:4320-4325.
[17]Kozak G K,Boerlin P,Janecko N,et al.Antimicrobial resistance in Escherichia coli isolates from swine and wild small mammals in the proximity of swine farms and in natural environments in Ontario,Canada[J].Appl Environ Microbiol,2009,75(3):559-566.
[18]Holzel C S,Harms K S,Bauer J,et al.Diversity of antimicrobial resistance genes and class-1-integrons in phylogenetically related porcine and human Escherichia coli[J].Vet Microbiol,2012,160(3/4):403-412.
[19]汤景元,王红宁,张鹏举,等.95个猪场大肠杆菌耐药表型及氨基糖苷类药物耐药基因型调查[J].畜牧兽医学报,2008,39(4):472-477.Tang J Y,Wang H N,Zhang P J,et al.Investigation of drug resistance phenotypes of Escherichia coli and aminoglycoside resistant genotypes in 95pig farms[J].Chinese Journal of Animaland Veterinary Sciences,2008,39(4):472-477.
[20]邱军,张子威,樊瑞峰,等.呼伦贝尔地区猪源大肠杆菌血清型鉴定与耐药基因检测[J].东北农业大学学报,2015,46(7):50-56.Qiu J,Zhang Z W,Fan R F,et al.Identification of Escherichia coli in swine origin and detection of drug resistance gene in Hulunbeier Region[J].Journal of Northeast Agricultural U-niversity,2015,46(7):50-56.
[2]Ho T Y,Lo H Y,Li C C,et al.In vitro and in vivo bioluminescent imaging to evaluate anti Escherichia coli activity of Galla chinensis[J].Bio Medicine,2013,3(4):160-166.
[3]张晓丹,高丽丽,胡家卿,等.猪源携带质粒介导喹诺酮类耐药基因大肠杆菌气源性传播的研究[J].中国畜牧兽医,2015,42(7):1842-1850.Zhang X D,Gao L L,Hu J Q,et al.Study on plasmid-mediated aerosol delivery of fluoroquinolone-resistant Escherichia coli in swine origin[J].China Animal Husbandry and Veterinary Medicine,2015,42(7):1842-1850.
[4]张彬,韩志霞,马金玉,等.河南省部分地区鸡大肠杆菌的分离鉴定及耐药性研究[J].河南农业科学,2015,44(2):127-131.Zhang B,Han Z X,Ma J Y,et al.Isolation and identification of chicken Escherichia coli and drug resistance in some areas of Henan Province[J].Journal of Henan Agricultural Sciences,2015,44(2):127-131.
[5]李海华,郭蔚冰,杨海明,等.内蒙古中东部地区不同来源大肠杆菌耐药性调查[J].中国畜牧兽医,2017,44(4):1182-1188.Li H H,Guo W B,Yang H M,et al.Antibiotic resistance of different origins of E.coliin central and eastern Inner Mongolia[J].China Animal Husbandry and Veterinary Medicine,2017,44(4):1182-1188.
[6]雷连成,江文正,韩文瑜,等.致病性大肠杆菌的耐药性监测[J].中国兽医杂志,2001(1):12-13.Lei L C,Jiang W Z,Han W Y,et al.Drug resistance monitoring of pathogenic Escherichia coli[J].Chinese Journal of Veterinary Medicine,2001(1):12-13.
[7]叶满玉,王红宁,田国宝,等.猪鸡致病性大肠杆菌沙门氏菌对β-内酰胺类药耐药表型和基因检测[J].中国兽医杂志,2010,46(3):15-18.Ye M Y,Wang H N,Tian G B,et al.Detection of beta lactam resistant phenotype and gene of Salmonella enterotoxigenic Escherichia coli in pigs and chickens[J].Chinese Journal of Veterinary Medicine,2010,46(3):15-18.
[8]王利勤,王晶钰,董睿,等.鸡源致病性大肠埃希菌中氨基糖苷类抗生素耐药基因的检测[J].动物医学进展,2012,33(7):49-53.Wang L Q,Wang J Y,Dong R,et al.Detection of aminoglycoside resistant genes in pathogenic Escherichia coli of chicken[J].Progressin Veterinary Medicine,2012,33(7):49-53.
[9]孙慧,雷战,邹俊峰,等.山东地区禽源致病性大肠杆菌氨基糖苷类药物耐药性及耐药基因的检测[J].中国兽医学报,2011,31(9):1279-1282.Sun H,Lei Z,Zou J F,et al.Detection of drug resistance and drug resistance genes of avian pathogenic Escherichia coli aminoglycosides in Shandong province[J].Chinese Journal of Veterinary Science,2011,31(9):1279-1282.
[10]黄明钱,孔祥峰,肖莉春,等.中药水提物对猪源耐四环素大肠杆菌的抑制活性[J].中国农业科学,2013,46(11):2370-2376.Huang M Q,Kong X F,Xiao L C,et al.Inhibitory activity of Chinese medicine water extract on tetracycline resistant Escherichia coli in pig source[J].Scientia Agricultura Sinica,2013,46(11):2370-2376.
[11]田国宝,王红宁,张安云,等.规模化猪场大肠杆菌β-内酰胺类药物耐药性检测及其超广谱β-内酰胺酶调查[J].中国预防兽医学报,2011,33(10):776-780.Tian G B,Wang H N,Zhang A Y,et al.Detection of Escherichia coliβ-lactams in large-scale pig farms and investigation of their extended-spectrumβ-lactamases[J].Chinese Journal of Preventive Veterinary Medici,2011,33(10):776-780.
[12]Guillaume G,Verbrugge D,Chasseur-Libotte M,et al.PCR typing of tetracycline resistance determinants(Tet A-E)in Salmonella enterica serotype Hadar and in the microbial community of activated sludges from hospital and urban wastewater treatment facilities in Belgium[J].FEMS Microbiol Ecol,2000,32(1):77-85.
[13]董杰.细菌获得性抗生素耐药基因研究进展[J].中国预防医学杂志,2015(1):71-74.Dong J.Research progress on antibiotic resistant gene of bacterial acquired antibiotics[J].Chinese Journal of Preventive Medicine,2015(1):71-74.
[14]戴秀美,常维山.2012年我国部分地区猪源大肠杆菌耐药性分析[J].畜禽业,2013(1):50-52.Dai X M,Chang W S.Analysis of resistance of Escherichia coli in pig sources in some areas of China in 2012[J].Livestock and Poultry Industry,2013(1):50-52.
[15]朱力军.动物大肠杆菌耐药性的变化趋势[J].中国兽医杂志,2001,35(2):16-18.Zhu L J.The trend of resistance to Escherichia coli in animals[J].Chinese Journal of Veterinary Medicine,2001,35(2):16-18.
[16]Yan J J,Ko W C,Tsai S H,et al.Dissemination of CTX-M-3and CMY-2beta-lactamaases among clinical isolat es of Escherichia coli in southern Taiwan[J].J Clin Microbiol,2000,38:4320-4325.
[17]Kozak G K,Boerlin P,Janecko N,et al.Antimicrobial resistance in Escherichia coli isolates from swine and wild small mammals in the proximity of swine farms and in natural environments in Ontario,Canada[J].Appl Environ Microbiol,2009,75(3):559-566.
[18]Holzel C S,Harms K S,Bauer J,et al.Diversity of antimicrobial resistance genes and class-1-integrons in phylogenetically related porcine and human Escherichia coli[J].Vet Microbiol,2012,160(3/4):403-412.
[19]汤景元,王红宁,张鹏举,等.95个猪场大肠杆菌耐药表型及氨基糖苷类药物耐药基因型调查[J].畜牧兽医学报,2008,39(4):472-477.Tang J Y,Wang H N,Zhang P J,et al.Investigation of drug resistance phenotypes of Escherichia coli and aminoglycoside resistant genotypes in 95pig farms[J].Chinese Journal of Animaland Veterinary Sciences,2008,39(4):472-477.
[20]邱军,张子威,樊瑞峰,等.呼伦贝尔地区猪源大肠杆菌血清型鉴定与耐药基因检测[J].东北农业大学学报,2015,46(7):50-56.Qiu J,Zhang Z W,Fan R F,et al.Identification of Escherichia coli in swine origin and detection of drug resistance gene in Hulunbeier Region[J].Journal of Northeast Agricultural U-niversity,2015,46(7):50-56.