METHODS article
Front. Microbiol.
Sec. Microbiotechnology
Volume 16 - 2025 | doi: 10.3389/fmicb.2025.1554660
This article is part of the Research TopicNavigating Challenges and Innovations in Antimicrobial Resistance, Environmental Microbiology, and Industrial SolutionsView all 8 articles
Application and evaluation of a rapid detection method based on twodimensional PCR technology for Hypervirulent Klebsiella pneumoniae
Provisionally accepted
- Department of Laboratory Medicine, Jinshan Hospital, Fudan University, Shanghai, China
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Objectives: Hypervirulent K. pneumoniae (hvKp) is an emerging pathogen that is more virulent than classical K. pneumoniae (cKp). This study aimed to develop an economical, high-throughput, and accurate two-dimensional polymerase chain reaction (2D-PCR) assay for the rapid detection of hvKp.Recombinant plasmids containing the iucA, peg-344, rmpA2, and rmpA virulence genes were constructed and used for assessing the sensitivity and specificity of the 2D-PCR. Clinical samples (n=105) were collected and evaluated the performance of the 2D-PCR to comparison with conventional PCR methods.The minimum detection limit of the 2D-PCR assay for iucA, peg-344, rmpA2, and rmpA were 10³, 10², 10³, and 10³ copies/µl, respectively. Additionally, the concordance rates between the 2D-PCR and conventional PCR for detecting iucA, peg-344, rmpA2, and rmpA were all over 95%. The analysis revealed a sensitivity of 100.0% and a specificity of 96.2% when compared to conventional PCR.A 2D-PCR-based multiplex method for virulence genes of hvKp was successfully developed, demonstrating its outstanding features of high specificity, high sensitivity, and high throughput capability. This method could be used for the rapid diagnosis of infectious diseases caused by hvKp in clinical settings.
Keywords: 2D-PCR, hypervirulent Klebsiella pneumoniae, Virulence gene, diagnosis, biomarkers
Received: 02 Jan 2025; Accepted: 26 May 2025.
Copyright: © 2025 Zhu, Wang, Jiang and Zhao. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) or licensor are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.
* Correspondence: Yan Zhao, Department of Laboratory Medicine, Jinshan Hospital, Fudan University, Shanghai, China
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