qnrD, a Novel Gene Conferring Transferable Quinolone Resistance in Salmonella enterica Serovar Kentucky and Bovismorbificans Strains of Human Origin
qnrD, a Novel Gene Conferring Transferable Quinolone Resistance in Salmonella enterica Serovar Kentucky and Bovismorbificans Strains of Human Origin
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DOI：
10.1128/aac.00997-08
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发表时间：
2009-02-01
影响因子：
4.9
通讯作者：
Aarestrup, F. M.
中科院分区：
文献类型：
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作者：
Cavaco, L. M.;Hasman, H.;Aarestrup, F. M.
在先前的一项研究中，中国河南省的4株人源沙门氏菌分离株对环丙沙星的敏感性降低（最低抑菌浓度为0.125 - 0.25微克/毫升），但对萘啶酸敏感（最低抑菌浓度为4 - 8微克/毫升）。所有分离株已知的qnr基因（A、B和S）、aac(6')Ib - cr以及gyrA和parC的突变均为阴性。从所有4株分离株中提取质粒DNA，并通过电穿孔法转化到大肠杆菌TG1和DH10B细胞中，在含0.06微克/毫升环丙沙星的脑心浸液琼脂平板上筛选转化子。对环丙沙星的抗性可通过电穿孔转移，并且在所有转化子中都发现了一个相似的4270bp的质粒。通过序列分析，发现该质粒携带一个开放阅读框，其与其他qnr基因具有相似性，并编码一个含214个氨基酸的五肽重复蛋白。这个基因被命名为qnrD，它与qnrA1有48%的相似性，与qnrB1有61%的相似性，与qnrS1有41%的相似性。进一步将qnrD编码区亚克隆到载体pBR322的组成型表达的tetA基因中，结果表明该基因使环丙沙星的最低抑菌浓度增加了32倍（从0.002微克/毫升的最低抑菌浓度增加到0.06微克/毫升）。作为对比，qnrA1和qnrS1也被亚克隆到pBR322中并转化到DH10B细胞中，分别赋予0.125微克/毫升和0.5微克/毫升的最低抑菌浓度。对所有已知的qnr序列进行了系统发育分析，结果表明qnrD与qnrB变体关系更密切，但形成了一个独立的簇。据我们所知，这是对qnrD基因的首次描述。
In a previous study, four Salmonella isolates from humans in the Henan province of China showed reduced susceptibility to ciprofloxacin (MIC, 0.125 to 0.25 mu g/ml) but were susceptible to nalidixic acid ( MIC, 4 to 8 mu g/ml). All isolates were negative for known qnr genes ( A, B, and S), aac(6')Ib-cr, and mutations in gyrA and parC. Plasmid DNA was extracted from all four isolates and transformed into Escherichia coli TG1 and DH10B cells by electroporation, and transformants were selected on 0.06 mu g/ml ciprofloxacin containing brain heart infusion agar plates. Resistance to ciprofloxacin could be transferred by electroporation, and a similar 4,270-bp plasmid was found in all transformants. By sequence analysis, the plasmid was found to carry an open reading frame that had similarities to other qnr genes and that encoded a 214-amino-acid pentapeptide repeat protein. This gene, designated qnrD, showed 48% similarity to qnrA1, 61% similarity to qnrB1, and 41% similarity to qnrS1. Further subcloning of the qnrD coding region into the constitutively expressed tetA gene of vector pBR322 showed that the gene conferred an increase in the MIC of ciprofloxacin by a factor of 32 ( from an MIC of 0.002 to an MIC of 0.06 mu g/ml). For comparison, qnrA1 and qnrS1 were also subcloned into pBR322 and transformed into DH10B cells, conferring MICs of 0.125 and 0.5 mu g/ml, respectively. A phylogenetic analysis of all known qnr sequences was performed and showed that qnrD was more closely related to the qnrB variants but formed an independent cluster. To our knowledge, this is the first description of this qnrD gene.