Development and Validation of Two Diagnostic Real-Time PCR (TaqMan) Assays for the Detection of <i>Bordetella avium</i> from Clinical Samples and Comparison to the Currently Available Real-Time TaqMan PCR Assay

<i>Bordetella avium</i> (BA) is one of many pathogens that cause respiratory diseases in turkeys. However, other bacterial species can easily overgrow it during isolation attempts. This makes confirming the diagnosis of BA as the causative agent of turkey coryza more difficult. Currently...

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Autores principales:	Amro Hashish, Avanti Sinha, Amr Mekky, Yuko Sato, Nubia R. Macedo, Mohamed El-Gazzar
Formato:	article
Lenguaje:	EN
Publicado:	MDPI AG 2021
Materias:	<i>Bordetella avium</i> (BA) bordetellosis TaqMan real-time PCR (qPCR) bacterial detection clinical samples analytical validation Biology (General) QH301-705.5
Acceso en línea:	https://doaj.org/article/fb56210901c9481ba0b618382247585e
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spelling	oai:doaj.org-article:fb56210901c9481ba0b618382247585e2021-11-25T18:24:25ZDevelopment and Validation of Two Diagnostic Real-Time PCR (TaqMan) Assays for the Detection of <i>Bordetella avium</i> from Clinical Samples and Comparison to the Currently Available Real-Time TaqMan PCR Assay10.3390/microorganisms91122322076-2607https://doaj.org/article/fb56210901c9481ba0b618382247585e2021-10-01T00:00:00Zhttps://www.mdpi.com/2076-2607/9/11/2232https://doaj.org/toc/2076-2607<i>Bordetella avium</i> (BA) is one of many pathogens that cause respiratory diseases in turkeys. However, other bacterial species can easily overgrow it during isolation attempts. This makes confirming the diagnosis of BA as the causative agent of turkey coryza more difficult. Currently, there are two PCR assays for the molecular detection of BA. One is conventional gel-based PCR and the other is TaqMan real-time PCR (qPCR) assay. However, multiple pitfalls were detected in both assays regarding their specificity, sensitivity, and efficiency, which limits their utility as diagnostic tools. In this study, we developed and validated two TaqMan qPCR assays and compared their performance to the currently available TaqMan qPCR. The two assays were able to correctly identify all BA isolates and showed negative results against a wide range of different microorganisms. The two assays were found to have high efficiency with a detection limit of approximately 1 × 10<sup>3</sup> plasmid DNA Copies/mL with high repeatability and reproducibility. In comparison to the currently available TaqMan qPCR assay, the newly developed assays showed significantly higher PCR efficiencies due to superior primers and probes design. The new assays can serve as a reliable tool for the sensitive, specific, and efficient diagnosis of BA.Amro HashishAvanti SinhaAmr MekkyYuko SatoNubia R. MacedoMohamed El-GazzarMDPI AGarticle<i>Bordetella avium</i> (BA)bordetellosisTaqMan real-time PCR (qPCR)bacterial detectionclinical samplesanalytical validationBiology (General)QH301-705.5ENMicroorganisms, Vol 9, Iss 2232, p 2232 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	<i>Bordetella avium</i> (BA) bordetellosis TaqMan real-time PCR (qPCR) bacterial detection clinical samples analytical validation Biology (General) QH301-705.5
spellingShingle	<i>Bordetella avium</i> (BA) bordetellosis TaqMan real-time PCR (qPCR) bacterial detection clinical samples analytical validation Biology (General) QH301-705.5 Amro Hashish Avanti Sinha Amr Mekky Yuko Sato Nubia R. Macedo Mohamed El-Gazzar Development and Validation of Two Diagnostic Real-Time PCR (TaqMan) Assays for the Detection of <i>Bordetella avium</i> from Clinical Samples and Comparison to the Currently Available Real-Time TaqMan PCR Assay
description	<i>Bordetella avium</i> (BA) is one of many pathogens that cause respiratory diseases in turkeys. However, other bacterial species can easily overgrow it during isolation attempts. This makes confirming the diagnosis of BA as the causative agent of turkey coryza more difficult. Currently, there are two PCR assays for the molecular detection of BA. One is conventional gel-based PCR and the other is TaqMan real-time PCR (qPCR) assay. However, multiple pitfalls were detected in both assays regarding their specificity, sensitivity, and efficiency, which limits their utility as diagnostic tools. In this study, we developed and validated two TaqMan qPCR assays and compared their performance to the currently available TaqMan qPCR. The two assays were able to correctly identify all BA isolates and showed negative results against a wide range of different microorganisms. The two assays were found to have high efficiency with a detection limit of approximately 1 × 10<sup>3</sup> plasmid DNA Copies/mL with high repeatability and reproducibility. In comparison to the currently available TaqMan qPCR assay, the newly developed assays showed significantly higher PCR efficiencies due to superior primers and probes design. The new assays can serve as a reliable tool for the sensitive, specific, and efficient diagnosis of BA.
format	article
author	Amro Hashish Avanti Sinha Amr Mekky Yuko Sato Nubia R. Macedo Mohamed El-Gazzar
author_facet	Amro Hashish Avanti Sinha Amr Mekky Yuko Sato Nubia R. Macedo Mohamed El-Gazzar
author_sort	Amro Hashish
title	Development and Validation of Two Diagnostic Real-Time PCR (TaqMan) Assays for the Detection of <i>Bordetella avium</i> from Clinical Samples and Comparison to the Currently Available Real-Time TaqMan PCR Assay
title_short	Development and Validation of Two Diagnostic Real-Time PCR (TaqMan) Assays for the Detection of <i>Bordetella avium</i> from Clinical Samples and Comparison to the Currently Available Real-Time TaqMan PCR Assay
title_full	Development and Validation of Two Diagnostic Real-Time PCR (TaqMan) Assays for the Detection of <i>Bordetella avium</i> from Clinical Samples and Comparison to the Currently Available Real-Time TaqMan PCR Assay
title_fullStr	Development and Validation of Two Diagnostic Real-Time PCR (TaqMan) Assays for the Detection of <i>Bordetella avium</i> from Clinical Samples and Comparison to the Currently Available Real-Time TaqMan PCR Assay
title_full_unstemmed	Development and Validation of Two Diagnostic Real-Time PCR (TaqMan) Assays for the Detection of <i>Bordetella avium</i> from Clinical Samples and Comparison to the Currently Available Real-Time TaqMan PCR Assay
title_sort	development and validation of two diagnostic real-time pcr (taqman) assays for the detection of <i>bordetella avium</i> from clinical samples and comparison to the currently available real-time taqman pcr assay
publisher	MDPI AG
publishDate	2021
url	https://doaj.org/article/fb56210901c9481ba0b618382247585e
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Development and Validation of Two Diagnostic Real-Time PCR (TaqMan) Assays for the Detection of <i>Bordetella avium</i> from Clinical Samples and Comparison to the Currently Available Real-Time TaqMan PCR Assay

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