2种养殖模式下蛋鸭大肠埃希菌毒力基因和耐药特征分析

doi:10.3785/j.issn.1008-9209.2019.03.281

浙江大学学报（农业与生命科学版）

2020, Vol. 46

Issue (2): 254-262 DOI: 10.3785/j.issn.1008-9209.2019.03.281

动物科学与动物医学

2种养殖模式下蛋鸭大肠埃希菌毒力基因和耐药特征分析

赵燕¹(

),金俊杰¹,任敏敏²,侯凤香¹,刘素贞¹,薛成俊³,肖英平²(

)

^1.温州市农业科学研究院动物科学研究所，浙江温州 325000
^2.浙江省农业科学院农产品质量标准研究所，杭州 310021
^3.苍南县畜牧兽医局，浙江苍南 325800

Virulence genes and drug resistance characteristics of Escherichia coli in laying duck under the two different breeding modes

Yan ZHAO¹(

),Junjie JIN¹,Minmin REN²,Fengxiang HOU¹,Suzhen LIU¹,Chengjun XUE³,Yingping XIAO²(

)

^1.Institute of Animal Sciences, Wenzhou Academy of Agricultural Sciences, Wenzhou 325000, Zhejiang, China
^2.Institute of Quality and Standard for Agricultural Products, Zhejiang Academy of Agricultural Sciences, Hangzhou 310021, China
^3.Cangnan County Animal Husbandry and Veterinary Bureau, Cangnan 325800, Zhejiang, China

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摘要：

为比较分析在网床水面圈养与网床全旱养2种不同养殖模式下蛋鸭肠道中大肠埃希菌的毒力基因状况和耐药特征，分别在温州地区选择3家网床水面圈养和3家网床全旱养的蛋鸭场，每个蛋鸭场各选取10只蛋鸭，用棉拭子在直肠中采集微生物，用于大肠埃希菌分离，采用VITEK 2 COMPACT全自动细菌鉴定及药敏分析系统对分离株进行鉴定和开展耐药表型分析，并通过聚合酶链式反应（polymerase chain reaction, PCR）扩增毒力基因、耐药基因和Ⅰ型整合酶基因。结果表明：蛋鸭中大肠埃希菌的分离率为100%；网床全旱养蛋鸭肠道大肠埃希菌的毒力基因eae、ipa H、invE、aggR和pic检出率高于网床水面圈养蛋鸭。对于耐药表型，网床全旱养蛋鸭肠道大肠埃希菌耐药数≥3的菌株比例为23.33%，而网床水面圈养中耐药数≥3的菌株比例为10.00%。同样地，耐药基因和Ⅰ型整合酶基因的检出率也为网床全旱养高于网床水面圈养。10株Ⅰ型整合子阳性大肠埃希菌中4株携带基因盒插入片段，其中：1株来源于网床水面圈养蛋鸭，其大肠埃希菌Ⅰ型整合子基因盒插入的为dfrA12-aadA2；3株来源于网床全旱养蛋鸭，均为dfrA1-aadA1。网床全旱养蛋鸭肠道大肠埃希菌的毒力基因携带率和耐药性表型均高于网床水面圈养蛋鸭，说明养殖模式变化对水禽肠道微生物耐药性可产生一定的影响。

关键词： 蛋鸭; 大肠埃希菌; 耐药表型; 耐药基因; 毒力基因; Ⅰ型整合子

Abstract:

This study mainly compared the virulence genes and drug resistance characteristics of Escherichia coli in laying ducks under the two different breeding modes: net bed with pool and net bed dry-system. Six farms including three net beds with pool and three net bed dry-systems were selected in Wenzhou, Zhejiang Province, and ten laying ducks were collected from each farm. The cotton swabs were used to collect microorganisms in the rectum forthe isolation of E. coli. The VITEK 2 COMPACT analysis system was used to identify the isolates and analyze the drug resistance profile. The virulence genes, drug-resistant genes and class Ⅰ integron genes were amplified by polymerase chain reaction. The results showed that the isolation rate of E. coli in laying ducks was 100%. The detection rates of virulence genes eae, ipa H, invE, aggR and pic of E. coli in the net bed dry-system were higher than those in the net bed with pool. For the drug-resistant phenotype, the proportion of strains with ≥3 resistance in E. coli in the net bed dry-system was 23.33%, while the proportion of strains with ≥3 in the net bed with pool was 10.00%. Similarly, the detection rates of the drug-resistant genes and the class Ⅰ integron genes in the net bed dry-system were also higher than those in the net bed with pool. Four of the 10 strains of class Ⅰ integron-positive E. coli carried the gene cassette insert, one of which derived from the E. coli class Ⅰ integron gene cassette of laying duck in the net bed with pool was dfrA12-aadA2, and the other three strains derived from the E. coli class Ⅰ integron gene cassette of laying duck in the net bed dry-system were dfrA1-aadA1. The above results indicate that the virulence gene carrying rate and drug resistance of E. coli in the intestinal tract of laying duck in the net bed dry-system are higher than those in the net bed with pool. It is suggested tha the changes of breeding mode may have some effect on the drug resistance characteristics of bacteria in waterfowls.

Key words: laying duck Escherichia coli drug-resistant phenotype drug-resistant gene virulence gene class Ⅰ integron

收稿日期: 2019-03-28 出版日期: 2020-05-22

CLC:

S 834

基金资助: 浙江省公益技术研究农业项目(2016C32078);浙江省农业科学院农产品质量安全国家重点实验室培育项目(2010DS700124-ZZ1905);浙江省温州市科技计划项目(X20170035);浙江省温州市农业丰收计划项目(FSJH2018011)

通讯作者: 肖英平 E-mail: yanzhao_wk@163.com;ypxiaozju@126.com

作者简介: 赵燕（https://orcid.org/0000-0002-4750-3129），Tel：+86-577-88413634，E-mail：yanzhao_wk@163.com

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引用本文:

赵燕,金俊杰,任敏敏,侯凤香,刘素贞,薛成俊,肖英平. 2种养殖模式下蛋鸭大肠埃希菌毒力基因和耐药特征分析[J]. 浙江大学学报（农业与生命科学版）, 2020, 46(2): 254-262.

Yan ZHAO,Junjie JIN,Minmin REN,Fengxiang HOU,Suzhen LIU,Chengjun XUE,Yingping XIAO. Virulence genes and drug resistance characteristics of Escherichia coli in laying duck under the two different breeding modes. Journal of Zhejiang University (Agriculture and Life Sciences), 2020, 46(2): 254-262.

链接本文:

http://www.zjujournals.com/agr/CN/10.3785/j.issn.1008-9209.2019.03.281 或 http://www.zjujournals.com/agr/CN/Y2020/V46/I2/254

表1 毒力基因引物信息

表2 耐药基因引物信息

表3 β-内酰胺酶基因引物信息

图1 eae毒力基因的PCR产物电泳结果M：DL5000标志物；NC：阴性对照；1～2：分离自网床水面圈养蛋鸭中的大肠埃希菌；3～4：分离自网床全旱养蛋鸭中的大肠埃希菌；PC：阳性对照。

图2 大肠埃希菌的毒力基因检出率

图3 网床水面圈养和网床全旱养蛋鸭中大肠埃希菌耐药菌株分布情况

图4 网床水面圈养和网床全旱养蛋鸭中大肠埃希菌耐药基因检出率

图5 CTX-M耐药基因的PCR产物电泳结果M：DL5000标志物；NC：阴性对照；1～2：分离自网床水面圈养蛋鸭中的菌株；3～4：分离自网床全旱养蛋鸭中的菌株；PC：阳性对照。

图6 ESBLs阳性大肠埃希菌β-内酰胺酶基因检出率

图7 大肠埃希菌的Ⅰ型整合子检出率

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