Characterization of Unexpressed Extended-Spectrum Beta-Lactamase Genes in Antibiotic-Sensitive Klebsiella pneumoniae Isolates
Characterization of Unexpressed Extended-Spectrum Beta-Lactamase Genes in Antibiotic-Sensitive Klebsiella pneumoniae Isolates
复制标题
抗生素敏感肺炎克雷伯菌分离株中未表达的广谱 β-内酰胺酶基因的表征
DOI：
10.1089/mdr.2017.0018
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发表时间：
2017-11-01
影响因子：
2.6
通讯作者：
Gao, Zhancheng
中科院分区：
文献类型：
--
作者：
Zhang, Zhao;Zhai, Yao;Gao, Zhancheng
目的：本研究旨在探讨临床非产超广谱内酰胺酶（ESBL）肺炎克雷伯菌分离株中是否存在ESBL基因。方法：对202株临床分离的非产ESBL克雷伯菌进行检测。pneumoniae的分离株。通过PCR筛选13种β-内酰胺酶基因（bla（SHV）、（CTX-M）、（TEM）、（OXA-2）、（OXA-10）、（VEB）、（PER）、（SFO）、（GES）、（CSP）、（TLA）、（BEL）和（IBC）），并通过PCR产物测序确认它们的身份。双纸片协同试验证实9株产ESBL表型。同时对产ESBL基因型（+）/表型（-）的分离株的ESBL基因上游序列进行PCR和测序。将ESBL基因及其上游序列克隆到大肠杆菌DH 5中进行功能鉴定。结果：202株大肠埃希菌中18株（8.9%）携带ESBL基因。18株大肠埃希菌均为单一ESBL基因，其中bla（SHV）占33.3%（6/18），bla（CTX-M）占66.7%（12/18）。在携带ESBL基因的菌株中，9株被证实为ESBL表型（-）。ESBL基因型（+）/表型（-）菌株中，bla（SHV-27、38、41、42）占66.7%（6/9），bla（CTX-M-3、15、24）占33.3%（3/9）。上游基因序列，包括这些未表达的ESBL基因的启动子，是完整的，没有任何突变或间隔区和有效的8株。在一个携带未表达bla（CTX-M-15）基因的分离株中未发现上游区的ISEcp 1元件。结论：临床上产非ESBL的克雷伯菌pneumoniae菌株可携带启动子完整的ESBL基因，但不携带相关表型。特异性沉默机制可能在调节ESBL基因表达中发挥重要作用。这类菌株有可能将其ESBL基因转移到具有有效启动子的其他细菌中，从而产生ESBL表型。
Objective: The current investigation explores whether extended-spectrum -lactamase (ESBL) genes exist in clinical non-ESBL-producing Klebsiella pneumoniae isolates. Methods: A total of 202 clinical isolates with non-ESBL-producing K. pneumoniae were collected from southern and middle of China. Thirteen -lactamase genes (bla(SHV), (CTX-M), (TEM), (OXA-2), (OXA-10), (VEB), (PER), (SFO), (GES), (CSP), (TLA), (BEL), (and IBC)) were screened by PCR and their identity confirmed by sequencing of PCR products. The ESBL-producing phenotype of the isolates that carried ESBL genes was tested and confirmed in 9 of the 18 isolates by a double-disc synergy test. The sequences upstream of ESBL genes of isolates with ESBL-producing genotype (+)/phenotype (-) were also subjected to PCR and sequencing. The ESBL genes and their upstream regions were cloned into Escherichia coli DH5 for functional evaluation. Results: A total of 8.9% (18/202) isolates carried ESBL genes. All of them harbored only one ESBL gene, including 33.3% (6/18) bla(SHV) and 66.7% (12/18) bla(CTX-M). Among the isolates carrying ESBL genes, nine isolates were confirmed as ESBL phenotype (-). The ESBL genotype (+)/phenotype (-) isolates had bla(SHV-27,38,41,42) (66.7%, 6/9) and bla(CTX-M-3,15,24) (33.3%, 3/9). The upstream gene sequences, including promoters of these unexpressed ESBL genes, were intact without any mutations or spacers and effective among eight strains. The ISEcp1 element in the upstream region was not found in one isolate carrying an unexpressed bla(CTX-M-15) gene. Conclusions: Clinical non-ESBL-producing K. pneumoniae isolates could carry ESBL genes with intact promoter, but without the correlated phenotype. Specific silencing mechanisms may play an important role in regulating ESBL gene expression. This kind of isolates has the potential to transfer their ESBL genes to other bacteria with effective promoters, resulting in ESBL phenotype.