Selection of Fecal Enterococci Exhibiting tcrB-Mediated Copper Resistance in Pigs Fed Diets Supplemented with Copper
Selection of Fecal Enterococci Exhibiting tcrB-Mediated Copper Resistance in Pigs Fed Diets Supplemented with Copper
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DOI：
10.1128/aem.00364-11
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发表时间：
2011-08-01
影响因子：
4.4
通讯作者：
Nagaraja, T. G.
中科院分区：
文献类型：
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作者：
Amachawadi, R. G.;Shelton, N. W.;Nagaraja, T. G.
铜（以硫酸铜的形式）越来越多地被用作断奶仔猪促生长的饲料抗生素替代品。在屎肠球菌和粪肠球菌中已报道了由质粒携带的可转移铜抗性（tcrB）基因赋予的获得性铜抗性。进行了一项纵向田间研究以确定补铜与仔猪中tcrB阳性肠球菌流行率之间的关系。该研究以断奶仔猪为对象，将其饲养在10个栏中，每个栏6头仔猪，分别饲喂添加正常水平（16.5 ppm；对照）或高水平（125 ppm）铜的日粮。在第0、14、28和42天从每个栏的3头仔猪中随机采集粪便样本，接种在M -肠球菌琼脂上，从每个样本中获得3株肠球菌分离株。在饲喂高铜日粮的仔猪中，tcrB阳性肠球菌的总体流行率为21.1%（38/180），而在对照组中为2.8%（5/180）。在43株tcrB阳性分离株中，35株为屎肠球菌，8株为粪肠球菌。tcrB阴性和tcrB阳性肠球菌对铜的平均最低抑菌浓度（MIC）分别为6.2 mM和22.2 mM。用S1核酸酶对屎肠球菌或粪肠球菌的基因组DNA进行限制性内切酶消化，产生了一条约194 - kbp大小的条带，tcrB和erm（B）基因探针均可与之杂交。接合试验表明tcrB和erm（B）基因在屎肠球菌和粪肠球菌菌株之间可共同转移。与饲喂正常铜水平日粮的仔猪相比，饲喂高铜日粮的仔猪中tcrB阳性肠球菌的流行率更高，这表明在猪日粮中添加铜会筛选出抗性菌株。
Copper, as copper sulfate, is increasingly used as an alternative to in-feed antibiotics for growth promotion in weaned piglets. Acquired copper resistance, conferred by a plasmid-borne, transferable copper resistance (tcrB) gene, has been reported in Enterococcus faecium and E. faecalis. A longitudinal field study was undertaken to determine the relationship between copper supplementation and the prevalence of tcrB-positive enterococci in piglets. The study was done with weaned piglets, housed in 10 pens with 6 piglets per pen, fed diets supplemented with a normal (16.5 ppm; control) or an elevated (125 ppm) level of copper. Fecal samples were randomly collected from three piglets per pen on days 0, 14, 28, and 42 and plated on M-Enterococcus agar, and three enterococcal isolates were obtained from each sample. The overall prevalence of tcrB-positive enterococci was 21.1% (38/180) in piglets fed elevated copper and 2.8% (5/180) in the control. Among the 43 tcrB-positive isolates, 35 were E. faecium and 8 were E. faecalis. The mean MICs of copper for tcrB-negative and tcrB-positive enterococci were 6.2 and 22.2 mM, respectively. The restriction digestion of the genomic DNA of E. faecium or E. faecalis with S1 nuclease yielded a band of similar to 194-kbp size to which both tcrB and the erm(B) gene probes hybridized. A conjugation assay demonstrated cotransfer of tcrB and erm(B) genes between E. faecium and E. faecalis strains. The higher prevalence of tcrB-positive enterococci in piglets fed elevated copper compared to that in piglets fed normal copper suggests that supplementation of copper in swine diets selected for resistance.