Methodological Evaluation of Carbapenemase Detection by Different Methods.

Polish journal of microbiology Pub Date : 2024-09-13 eCollection Date: 2024-09-01 DOI:10.33073/pjm-2024-034
Nana Gao, Jing Zhou, Ge Li, Runde Liu, Guoping Lu, Jilu Shen
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Abstract

The global proliferation of carbapenemase-producing bacteria (CPB) has garnered significant attention worldwide. Early diagnosis of CPB and accurate identification of carbapenemases are crucial for preventing the spread of CPB and ensuring targeted antibiotic therapy. Therefore, efficient and accurate identification of carbapenemases is paramount in clinically treating diseases associated with CPB. In this study, 58 CPB strains were collected and detected using the DNA endonuclease-targeted CRISPR trans reporter (DETECTR) method, a rapid detection platform based on CRISPR-Cas12a gene editing and isothermal amplification. Additionally, four conventional methods (the APB/EDTA method, PCR, NG-test Carba 5, and GeneXpert Carba-R) were employed and compared against whole genome sequencing (WGS) results, considered the gold standard, to evaluate their efficacy in detecting carbapenemases. Detection by the APB/EDTA method revealed that 29 strains were positive for Class A serine endopeptidases, while 29 strains were positive for Class B metalloenzymes. The classification of these zymotypes was consistent with the sequencing result. All target carbapenemases for KPC were identified with 100% sensitivity using NG-test Carba 5, PCR, DETECTR, and GeneXpert Carba-R. In the case of NDM, both Xpert Carba-R and DETECTR showed a sensitivity of 100%. In contrast, NG-test Carba 5 and PCR had a slightly lower sensitivity of 96.7%, each missing one target carbapenemase. n this study, the APB/EDTA method is capable of identifying the zymotype classification but not the specific resistant genes, while Xpert Carba-R and DETECTR are able to detect all target carbapenemases.

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不同方法检测碳青霉烯酶的方法学评估
产碳青霉烯酶细菌(CPB)在全球的扩散引起了全世界的高度关注。CPB 的早期诊断和碳青霉烯酶的准确鉴定对于防止 CPB 的传播和确保有针对性的抗生素治疗至关重要。因此,高效、准确地鉴定碳青霉烯酶对临床治疗 CPB 相关疾病至关重要。本研究收集了 58 株 CPB 菌株,并使用 DNA 核酸内切酶靶向 CRISPR 反式报告器(DETECTR)方法进行检测,该方法是一种基于 CRISPR-Cas12a 基因编辑和等温扩增的快速检测平台。此外,还采用了四种传统方法(APB/EDTA 法、PCR、NG-test Carba 5 和 GeneXpert Carba-R),并将其与被视为金标准的全基因组测序(WGS)结果进行比较,以评估它们在检测碳青霉烯酶方面的功效。通过 APB/EDTA 法检测发现,29 株菌株的 A 类丝氨酸内肽酶呈阳性,29 株菌株的 B 类金属酶呈阳性。这些酶型的分类与测序结果一致。使用 NG-test Carba 5、PCR、DETECTR 和 GeneXpert Carba-R 对 KPC 的所有目标碳青霉烯酶进行鉴定,灵敏度均为 100%。对于 NDM,Xpert Carba-R 和 DETECTR 的灵敏度均为 100%。相比之下,NG-test Carba 5 和 PCR 的灵敏度稍低,仅为 96.7%,各自漏检了一种目标碳青霉烯酶。在这项研究中,APB/EDTA 方法能够识别酶切型分类,但不能识别特定的耐药基因,而 Xpert Carba-R 和 DETECTR 能够检测所有目标碳青霉烯酶。
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