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GeXP analyzer-based multiplex reverse-transcription PCR assay for the simultaneous detection and differentiation of eleven duck viruses

BACKGROUND: Duck viral pathogens primarily include the avian influenza virus (AIV) subtypes H5, H7, and H9; duck hepatitis virus (DHV); duck tembusu virus (DTMUV); egg drop syndrome virus (EDSV); duck enteritis virus (DEV); Newcastle disease virus (NDV); duck circovirus (DuCV); muscovy duck reovirus...

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Autores principales: Zhang, Yan-fang, Xie, Zhi-xun, Xie, Li-ji, Deng, Xian-wen, Xie, Zhi-qin, Luo, Si-si, Huang, Li, Huang, Jiao-ling, Zeng, Ting-ting
Formato: Online Artículo Texto
Lenguaje:English
Publicado: BioMed Central 2015
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4628294/
https://www.ncbi.nlm.nih.gov/pubmed/26518004
http://dx.doi.org/10.1186/s12866-015-0590-6
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author Zhang, Yan-fang
Xie, Zhi-xun
Xie, Li-ji
Deng, Xian-wen
Xie, Zhi-qin
Luo, Si-si
Huang, Li
Huang, Jiao-ling
Zeng, Ting-ting
author_facet Zhang, Yan-fang
Xie, Zhi-xun
Xie, Li-ji
Deng, Xian-wen
Xie, Zhi-qin
Luo, Si-si
Huang, Li
Huang, Jiao-ling
Zeng, Ting-ting
author_sort Zhang, Yan-fang
collection PubMed
description BACKGROUND: Duck viral pathogens primarily include the avian influenza virus (AIV) subtypes H5, H7, and H9; duck hepatitis virus (DHV); duck tembusu virus (DTMUV); egg drop syndrome virus (EDSV); duck enteritis virus (DEV); Newcastle disease virus (NDV); duck circovirus (DuCV); muscovy duck reovirus (MDRV); and muscovy duck parvovirus (MDPV). These pathogens cause great economic losses to China’s duck breeding industry. RESULT: A rapid, specific, sensitive and high-throughput GeXP-based multiplex PCR assay consisting of chimeric primer-based PCR amplification with fluorescent labeling and capillary electrophoresis separation was developed and optimized to simultaneously detect these eleven viral pathogens. Single and mixed pathogen cDNA/DNA templates were used to evaluate the specificity of the GeXP-multiplex assay. Corresponding specific DNA products were amplified from each pathogen. Other pathogens, including duck Escherichia coli, duck Salmonella, duck Staphylococcus aureus, Pasteurella multocida, infectious bronchitis virus, and Mycoplasma gallisepticum, did not result in amplification products. The detection limit of GeXP was 10(3)copies when all twelve pre-mixed plasmids containing the target genes of eleven types of duck viruses were present. To further evaluate the reliability of GeXP, 150 clinical field samples were evaluated. Comparison with the results of conventional PCR methods for the field samples, the GeXP-multiplex PCR method was more sensitive and accurate. CONCLUSION: This GeXP-based multiplex PCR method can be utilized for the rapid differential diagnosis of clinical samples as an effective tool to prevent and control duck viruses with similar clinical symptoms.
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spelling pubmed-46282942015-11-01 GeXP analyzer-based multiplex reverse-transcription PCR assay for the simultaneous detection and differentiation of eleven duck viruses Zhang, Yan-fang Xie, Zhi-xun Xie, Li-ji Deng, Xian-wen Xie, Zhi-qin Luo, Si-si Huang, Li Huang, Jiao-ling Zeng, Ting-ting BMC Microbiol Methodology Article BACKGROUND: Duck viral pathogens primarily include the avian influenza virus (AIV) subtypes H5, H7, and H9; duck hepatitis virus (DHV); duck tembusu virus (DTMUV); egg drop syndrome virus (EDSV); duck enteritis virus (DEV); Newcastle disease virus (NDV); duck circovirus (DuCV); muscovy duck reovirus (MDRV); and muscovy duck parvovirus (MDPV). These pathogens cause great economic losses to China’s duck breeding industry. RESULT: A rapid, specific, sensitive and high-throughput GeXP-based multiplex PCR assay consisting of chimeric primer-based PCR amplification with fluorescent labeling and capillary electrophoresis separation was developed and optimized to simultaneously detect these eleven viral pathogens. Single and mixed pathogen cDNA/DNA templates were used to evaluate the specificity of the GeXP-multiplex assay. Corresponding specific DNA products were amplified from each pathogen. Other pathogens, including duck Escherichia coli, duck Salmonella, duck Staphylococcus aureus, Pasteurella multocida, infectious bronchitis virus, and Mycoplasma gallisepticum, did not result in amplification products. The detection limit of GeXP was 10(3)copies when all twelve pre-mixed plasmids containing the target genes of eleven types of duck viruses were present. To further evaluate the reliability of GeXP, 150 clinical field samples were evaluated. Comparison with the results of conventional PCR methods for the field samples, the GeXP-multiplex PCR method was more sensitive and accurate. CONCLUSION: This GeXP-based multiplex PCR method can be utilized for the rapid differential diagnosis of clinical samples as an effective tool to prevent and control duck viruses with similar clinical symptoms. BioMed Central 2015-10-30 /pmc/articles/PMC4628294/ /pubmed/26518004 http://dx.doi.org/10.1186/s12866-015-0590-6 Text en © Zhang et al. 2015 Open AccessThis article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.
spellingShingle Methodology Article
Zhang, Yan-fang
Xie, Zhi-xun
Xie, Li-ji
Deng, Xian-wen
Xie, Zhi-qin
Luo, Si-si
Huang, Li
Huang, Jiao-ling
Zeng, Ting-ting
GeXP analyzer-based multiplex reverse-transcription PCR assay for the simultaneous detection and differentiation of eleven duck viruses
title GeXP analyzer-based multiplex reverse-transcription PCR assay for the simultaneous detection and differentiation of eleven duck viruses
title_full GeXP analyzer-based multiplex reverse-transcription PCR assay for the simultaneous detection and differentiation of eleven duck viruses
title_fullStr GeXP analyzer-based multiplex reverse-transcription PCR assay for the simultaneous detection and differentiation of eleven duck viruses
title_full_unstemmed GeXP analyzer-based multiplex reverse-transcription PCR assay for the simultaneous detection and differentiation of eleven duck viruses
title_short GeXP analyzer-based multiplex reverse-transcription PCR assay for the simultaneous detection and differentiation of eleven duck viruses
title_sort gexp analyzer-based multiplex reverse-transcription pcr assay for the simultaneous detection and differentiation of eleven duck viruses
topic Methodology Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4628294/
https://www.ncbi.nlm.nih.gov/pubmed/26518004
http://dx.doi.org/10.1186/s12866-015-0590-6
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