Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, State Key Laboratory for Diagnosis and Treatment of Infectious Diseases, The First Affiliated Hospital, College of Medicine, Zhejiang University, Hangzhou, China.
Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, State Key Laboratory for Diagnosis and Treatment of Infectious Diseases, The First Affiliated Hospital, College of Medicine, Zhejiang University, Hangzhou, China; School of Laboratory Medicine and Life Science, Wenzhou Medical University, Wenzhou, China.
Environ Pollut. 2018 Apr;235:931-937. doi: 10.1016/j.envpol.2017.12.084. Epub 2018 Feb 21.
The wide spread of carbapenemase-producing Enterobacteriaceae (CPE) in the environment is an emerging environmental issue with potentially-serious public health implications. However, carbapenemase-producing Citrobacter from environment has rarely been investigated. Here we report the isolation and comparative genomics of carbapenemase-producing Citrobacter isolates from river sediment in China. Potential CPE was isolated by selective MacConkey agar plates containing 2 mg/L meropenem. The presence of carbapenemase genes was detected by PCR and sequencing. The clonal relatedness of Klebsiella pneumoniae carbapenemase (KPC-2)-producing Citrobacter isolates was assessed by pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing. Plasmid analysis of KPC-2-producing Citrobacter isolates was performed by S1-PFGE, Southern blotting, and whole genome sequencing. A total of four KPC-2-producing Citrobacter and three Aeromonas isolates were recovered from 54 sediment cultures of Shifeng River. Notably, all KPC-producing isolates were isolated from sampling sites near a waste water treatment plant. Antimicrobial susceptibility testing showed that three of the four sequenced isolates (C1710, C191, and C196) resistant to multiple antibiotics. Genotyping and pan-genome analyses revealed that the C191 and C196 C. freundii isolates exhibited a high level of genetic similarity. Plasmid analysis confirmed that the bla gene is located on either IncF or IncN3 plasmids in all isolates. The bla gene of C1710, C181 and C191 was successfully transferred with E. coli EC600 as the recipient strain. In silico analysis further suggested that pKPC-191 is a novel IncF plasmid, with 99% identity to two previously described IncFII plasmids at 71% coverage. We report here the presence of diverse conjugative bla plasmids from environmental Citrobacter isolates, which poses the possible dissemination of antimicrobial resistance into clinical isolates. To our knowledge, this is the first study to culture and characterize KPC-2-producing Citrobacter isolates from river sediments in China.
碳青霉烯酶产生肠杆菌科（CPE）在环境中的广泛传播是一个新出现的环境问题，具有潜在的严重公共卫生影响。然而，环境中产碳青霉烯酶的柠檬酸杆菌却很少被研究。在这里，我们报告了从中国河流沉积物中分离和比较产碳青霉烯酶柠檬酸杆菌的情况。通过含有 2mg/L 美罗培南的选择性 MacConkey 琼脂平板分离出潜在的 CPE。通过 PCR 和测序检测碳青霉烯酶基因的存在。通过脉冲场凝胶电泳（PFGE）和多位点序列分型评估肺炎克雷伯菌碳青霉烯酶（KPC-2）产生的柠檬酸杆菌分离株的克隆相关性。通过 S1-PFGE、Southern 印迹和全基因组测序对 KPC-2 产生的柠檬酸杆菌分离株进行质粒分析。从石门河的 54 个沉积物培养物中总共回收了 4 株产 KPC-2 的柠檬酸杆菌和 3 株气单胞菌。值得注意的是，所有产 KPC 的分离株均来自污水处理厂附近的采样点。药敏试验表明，4 株测序分离株中的 3 株（C1710、C191 和 C196）对多种抗生素耐药。基因分型和泛基因组分析表明，C191 和 C196 弗氏柠檬酸杆菌分离株具有高度的遗传相似性。质粒分析证实，所有分离株中的 bla 基因均位于 IncF 或 IncN3 质粒上。bla 基因的 C1710、C181 和 C191 成功地转移到 E. coli EC600 作为受体菌。计算机分析进一步表明，pKPC-191 是一种新型 IncF 质粒，在 71%的覆盖率下与之前描述的两种 IncFII 质粒具有 99%的同一性。我们在这里报告了从环境中分离的产碳青霉烯酶柠檬酸杆菌中存在多种可转移的 bla 质粒，这可能会导致抗生素耐药性传播到临床分离株。据我们所知，这是首次在中国河流沉积物中培养和鉴定产 KPC-2 的柠檬酸杆菌分离株。
