实时 PCR 与环介导等温扩增法直接从支气管肺泡灌洗液样本中检测肠杆菌科的碳青霉烯酶和/或 ESBL 基因的比较研究。
A comparative study between real-time PCR and loop-mediated isothermal amplification to detect carbapenemase and/or ESBL genes in Enterobacteriaceae directly from bronchoalveolar lavage fluid samples.
机构信息
Department of Clinical Microbiology - CDB, Hospital Clínic, University of Barcelona, Barcelona, Spain.
Institute for Global Health (ISGlobal), Hospital Clínic - Universitat de Barcelona, Barcelona, Spain.
出版信息
J Antimicrob Chemother. 2020 Jun 1;75(6):1453-1457. doi: 10.1093/jac/dkaa031.
OBJECTIVES
To evaluate and compare the efficacy of real-time PCR (Xpert Carba-R) and loop-mediated isothermal amplification (Eazyplex® SuperBug CRE) for detecting carbapenemase carriage in Enterobacteriaceae directly from bronchoalveolar lavage (BAL).
METHODS
Negative BAL samples were spiked with 21 well-characterized carbapenemase-producing Enterobacteriaceae strains to a final concentration of 102-104 cfu/mL. Xpert Carba-R (Cepheid, Sunnyvale, CA, USA), which detects five targets (blaKPC, blaNDM, blaVIM, blaOXA-48 and blaIMP-1), and the Eazyplex® SuperBug CRE system (Amplex-Diagnostics GmbH, Germany), which detects seven genes (blaKPC, blaNDM, blaVIM, blaOXA-48, blaOXA-181, blaCTXM-1 and blaCTXM-9), were evaluated for the detection of these genes directly from BAL samples.
RESULTS
Xpert Carba-R showed 100% agreement with carbapenemase characterization by PCR and sequencing for all final bacteria concentrations. Eazyplex® SuperBug CRE showed 100%, 80% and 27% agreement with PCR and sequencing when testing 104, 103 and 102 cfu/mL, respectively. False negative results for Eazyplex® SuperBug CRE matched the highest cycle threshold values for Xpert Carba-R. Hands-on time for both assays was about 15 min, but Eazyplex® SuperBug CRE results were available within 30 min, whereas Xpert Carba-R took around 50 min.
CONCLUSIONS
We here describe the successful use of two commercial diagnostic tests, Xpert Carba-R and Eazyplex® SuperBug CRE, to detect bacterial carbapenem resistance genes directly in lower respiratory tract samples. Our results could be used as proof-of-concept data for validation of these tests for this indication.
目的
评估并比较实时 PCR（Xpert Carba-R）和环介导等温扩增（Eazyplex® SuperBug CRE）直接从支气管肺泡灌洗液（BAL）检测肠杆菌科中产碳青霉烯酶菌的疗效。
方法
将 21 株特征明确的产碳青霉烯酶肠杆菌科菌株以 102-104 cfu/mL 的终浓度混入阴性 BAL 样本中。Xpert Carba-R（Cepheid，加利福尼亚州森尼韦尔）检测 5 个靶标（blaKPC、blaNDM、blaVIM、blaOXA-48 和 blaIMP-1），Eazyplex® SuperBug CRE 系统（德国 Amplex-Diagnostics GmbH）检测 7 个基因（blaKPC、blaNDM、blaVIM、blaOXA-48、blaOXA-181、blaCTXM-1 和 blaCTXM-9），评估这些基因直接从 BAL 样本中的检测情况。
结果
Xpert Carba-R 与所有最终细菌浓度的 PCR 和测序鉴定产碳青霉烯酶结果完全一致。Eazyplex® SuperBug CRE 以 104、103 和 102 cfu/mL 的浓度进行测试时，与 PCR 和测序的符合率分别为 100%、80%和 27%。Eazyplex® SuperBug CRE 的假阴性结果与 Xpert Carba-R 的最高循环阈值相对应。两种检测方法的实际操作时间约为 15 分钟，但 Eazyplex® SuperBug CRE 结果可在 30 分钟内获得，而 Xpert Carba-R 则需要大约 50 分钟。
结论
我们在此描述了两种商业诊断测试 Xpert Carba-R 和 Eazyplex® SuperBug CRE 成功地直接在下呼吸道样本中检测细菌碳青霉烯酶耐药基因。我们的结果可以用作这些测试针对该适应证的验证的概念验证数据。
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