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A Novel Molecular Method for Simultaneous Identification of Vibrio parahaemolyticus 57 K-Serogroups Using Probe Melting Curve Analysis

The serotyping of Vibrio parahaemolyticus, which is crucial to the surveillance and detection of outbreaks of vibriosis infection, has been widely used in many countries. In this study, we developed a molecular assay, named multiplex ligation reaction based on probe melting curve analysis (MLMA), fo...

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Autores principales: Lu, Linying, Li, Minxu, Li, Yinghui, Jiang, Min, Jiang, Yixiang, Shi, Xiaolu, Zuo, Le, Wang, Lei, Bian, Shengzhe, Qiu, Yaqun, Cai, Rui, Liao, Yiqun, Li, Qingge, Li, Liqiang, Hu, Qinghua
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Frontiers Media S.A. 2021
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7953158/
https://www.ncbi.nlm.nih.gov/pubmed/33718262
http://dx.doi.org/10.3389/fcimb.2021.594808
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author Lu, Linying
Li, Minxu
Li, Yinghui
Jiang, Min
Jiang, Yixiang
Shi, Xiaolu
Zuo, Le
Wang, Lei
Bian, Shengzhe
Qiu, Yaqun
Cai, Rui
Liao, Yiqun
Li, Qingge
Li, Liqiang
Hu, Qinghua
author_facet Lu, Linying
Li, Minxu
Li, Yinghui
Jiang, Min
Jiang, Yixiang
Shi, Xiaolu
Zuo, Le
Wang, Lei
Bian, Shengzhe
Qiu, Yaqun
Cai, Rui
Liao, Yiqun
Li, Qingge
Li, Liqiang
Hu, Qinghua
author_sort Lu, Linying
collection PubMed
description The serotyping of Vibrio parahaemolyticus, which is crucial to the surveillance and detection of outbreaks of vibriosis infection, has been widely used in many countries. In this study, we developed a molecular assay, named multiplex ligation reaction based on probe melting curve analysis (MLMA), for simultaneous identification of V. parahaemolyticus 57 K-serogroups. Based on the previous genomes of 418 strains including 39 K-serogroups and the 18 K-serogroups sequences from public databases, we obtained 57 K-serogroups specific gene sequences for designing primers and probes. The developed MLMA assay for identifying the V. parahaemolyticus 57 K-serogroups showed high reproducibility, with the intra- and inter-assay standard deviations and coefficients of variation of no more than 1°C and 1%, respectively. The limit of detection for all gene targets ranged from 0.1 to 1.0 ng/µl. We validated the MLMA assay with a double-blind test identifying 595 V. parahaemolyticus isolates using conventional serotyping methods for comparison. The results showed the kappa value between the MLMA assay and the traditional serological method was 0.936 and that there was a 96.97% consistency rate with conventional serotyping methods for all detected isolates. Additionally, five rare K-serogroups were identified using the MLMA assay, as well as 18 strains that could not be identified using the traditional serotyping method. Thus, the MLMA assay provides a rapid, robust, and promising tool for the molecular serotyping of V. parahaemolyticus K-serogroups and has the potential application to the detection of outbreaks and surveillance of V. parahaemolyticus infection.
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spelling pubmed-79531582021-03-13 A Novel Molecular Method for Simultaneous Identification of Vibrio parahaemolyticus 57 K-Serogroups Using Probe Melting Curve Analysis Lu, Linying Li, Minxu Li, Yinghui Jiang, Min Jiang, Yixiang Shi, Xiaolu Zuo, Le Wang, Lei Bian, Shengzhe Qiu, Yaqun Cai, Rui Liao, Yiqun Li, Qingge Li, Liqiang Hu, Qinghua Front Cell Infect Microbiol Cellular and Infection Microbiology The serotyping of Vibrio parahaemolyticus, which is crucial to the surveillance and detection of outbreaks of vibriosis infection, has been widely used in many countries. In this study, we developed a molecular assay, named multiplex ligation reaction based on probe melting curve analysis (MLMA), for simultaneous identification of V. parahaemolyticus 57 K-serogroups. Based on the previous genomes of 418 strains including 39 K-serogroups and the 18 K-serogroups sequences from public databases, we obtained 57 K-serogroups specific gene sequences for designing primers and probes. The developed MLMA assay for identifying the V. parahaemolyticus 57 K-serogroups showed high reproducibility, with the intra- and inter-assay standard deviations and coefficients of variation of no more than 1°C and 1%, respectively. The limit of detection for all gene targets ranged from 0.1 to 1.0 ng/µl. We validated the MLMA assay with a double-blind test identifying 595 V. parahaemolyticus isolates using conventional serotyping methods for comparison. The results showed the kappa value between the MLMA assay and the traditional serological method was 0.936 and that there was a 96.97% consistency rate with conventional serotyping methods for all detected isolates. Additionally, five rare K-serogroups were identified using the MLMA assay, as well as 18 strains that could not be identified using the traditional serotyping method. Thus, the MLMA assay provides a rapid, robust, and promising tool for the molecular serotyping of V. parahaemolyticus K-serogroups and has the potential application to the detection of outbreaks and surveillance of V. parahaemolyticus infection. Frontiers Media S.A. 2021-02-26 /pmc/articles/PMC7953158/ /pubmed/33718262 http://dx.doi.org/10.3389/fcimb.2021.594808 Text en Copyright © 2021 Lu, Li, Li, Jiang, Jiang, Shi, Zuo, Wang, Bian, Qiu, Cai, Liao, Li, Li and Hu http://creativecommons.org/licenses/by/4.0/ 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) and the copyright owner(s) 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.
spellingShingle Cellular and Infection Microbiology
Lu, Linying
Li, Minxu
Li, Yinghui
Jiang, Min
Jiang, Yixiang
Shi, Xiaolu
Zuo, Le
Wang, Lei
Bian, Shengzhe
Qiu, Yaqun
Cai, Rui
Liao, Yiqun
Li, Qingge
Li, Liqiang
Hu, Qinghua
A Novel Molecular Method for Simultaneous Identification of Vibrio parahaemolyticus 57 K-Serogroups Using Probe Melting Curve Analysis
title A Novel Molecular Method for Simultaneous Identification of Vibrio parahaemolyticus 57 K-Serogroups Using Probe Melting Curve Analysis
title_full A Novel Molecular Method for Simultaneous Identification of Vibrio parahaemolyticus 57 K-Serogroups Using Probe Melting Curve Analysis
title_fullStr A Novel Molecular Method for Simultaneous Identification of Vibrio parahaemolyticus 57 K-Serogroups Using Probe Melting Curve Analysis
title_full_unstemmed A Novel Molecular Method for Simultaneous Identification of Vibrio parahaemolyticus 57 K-Serogroups Using Probe Melting Curve Analysis
title_short A Novel Molecular Method for Simultaneous Identification of Vibrio parahaemolyticus 57 K-Serogroups Using Probe Melting Curve Analysis
title_sort novel molecular method for simultaneous identification of vibrio parahaemolyticus 57 k-serogroups using probe melting curve analysis
topic Cellular and Infection Microbiology
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7953158/
https://www.ncbi.nlm.nih.gov/pubmed/33718262
http://dx.doi.org/10.3389/fcimb.2021.594808
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