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Development of a rapid phage-based method for the detection of viable Mycobacterium avium subsp. paratuberculosis in blood within 48 h()
The aim of this study was to develop a methodology to rapidly detect viable Mycobacterium avium subsp. paratuberculosis (MAP) in clinical blood samples. MAP cells spiked into commercially available blood were recovered using optimised peptide-mediated magnetic separation (PMMS) and detected using a...
Autores principales: | , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Elsevier Biomedical
2013
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3783900/ https://www.ncbi.nlm.nih.gov/pubmed/23811207 http://dx.doi.org/10.1016/j.mimet.2013.06.015 |
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author | Swift, Benjamin M.C. Denton, Emily J. Mahendran, Sophie A. Huxley, Jonathan N. Rees, Catherine E.D. |
author_facet | Swift, Benjamin M.C. Denton, Emily J. Mahendran, Sophie A. Huxley, Jonathan N. Rees, Catherine E.D. |
author_sort | Swift, Benjamin M.C. |
collection | PubMed |
description | The aim of this study was to develop a methodology to rapidly detect viable Mycobacterium avium subsp. paratuberculosis (MAP) in clinical blood samples. MAP cells spiked into commercially available blood were recovered using optimised peptide-mediated magnetic separation (PMMS) and detected using a phage-based method, and the identity of the cells detected confirmed using nested-PCR amplification of MAP signature sequences (IS900). The limit of detection was determined to be 10 MAP cells per ml of blood and was used to detect MAP present in clinical bovine blood samples. Using the PMMS-phage method there was no difference when detecting MAP from whole blood or from isolated buffy coat. MAP was detected in animals that were milk-ELISA positive (15 animals) by PMMS-phage and no MAP was detected in blood samples from an accredited Johne's disease free herd (5 animals). In a set of samples from one herd (10 animals) that came from animals with variable milk ELISA status, the PMMS-phage results agreed with the positive milk-ELISA results in all but one case. These results show that the PMMS-phage method can detect MAP present in naturally infected blood. Total assay time is 48 h and, unlike PCR-based detection tests, only viable cells are detected. A rapid method for detecting MAP in blood could further the understanding of disseminated infection in animals with Johne's disease. |
format | Online Article Text |
id | pubmed-3783900 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2013 |
publisher | Elsevier Biomedical |
record_format | MEDLINE/PubMed |
spelling | pubmed-37839002013-09-30 Development of a rapid phage-based method for the detection of viable Mycobacterium avium subsp. paratuberculosis in blood within 48 h() Swift, Benjamin M.C. Denton, Emily J. Mahendran, Sophie A. Huxley, Jonathan N. Rees, Catherine E.D. J Microbiol Methods Article The aim of this study was to develop a methodology to rapidly detect viable Mycobacterium avium subsp. paratuberculosis (MAP) in clinical blood samples. MAP cells spiked into commercially available blood were recovered using optimised peptide-mediated magnetic separation (PMMS) and detected using a phage-based method, and the identity of the cells detected confirmed using nested-PCR amplification of MAP signature sequences (IS900). The limit of detection was determined to be 10 MAP cells per ml of blood and was used to detect MAP present in clinical bovine blood samples. Using the PMMS-phage method there was no difference when detecting MAP from whole blood or from isolated buffy coat. MAP was detected in animals that were milk-ELISA positive (15 animals) by PMMS-phage and no MAP was detected in blood samples from an accredited Johne's disease free herd (5 animals). In a set of samples from one herd (10 animals) that came from animals with variable milk ELISA status, the PMMS-phage results agreed with the positive milk-ELISA results in all but one case. These results show that the PMMS-phage method can detect MAP present in naturally infected blood. Total assay time is 48 h and, unlike PCR-based detection tests, only viable cells are detected. A rapid method for detecting MAP in blood could further the understanding of disseminated infection in animals with Johne's disease. Elsevier Biomedical 2013-09 /pmc/articles/PMC3783900/ /pubmed/23811207 http://dx.doi.org/10.1016/j.mimet.2013.06.015 Text en © 2013 The Authors https://creativecommons.org/licenses/by/3.0/ Open Access under CC BY 3.0 (https://creativecommons.org/licenses/by/3.0/) license |
spellingShingle | Article Swift, Benjamin M.C. Denton, Emily J. Mahendran, Sophie A. Huxley, Jonathan N. Rees, Catherine E.D. Development of a rapid phage-based method for the detection of viable Mycobacterium avium subsp. paratuberculosis in blood within 48 h() |
title | Development of a rapid phage-based method for the detection of viable Mycobacterium avium subsp. paratuberculosis in blood within 48 h() |
title_full | Development of a rapid phage-based method for the detection of viable Mycobacterium avium subsp. paratuberculosis in blood within 48 h() |
title_fullStr | Development of a rapid phage-based method for the detection of viable Mycobacterium avium subsp. paratuberculosis in blood within 48 h() |
title_full_unstemmed | Development of a rapid phage-based method for the detection of viable Mycobacterium avium subsp. paratuberculosis in blood within 48 h() |
title_short | Development of a rapid phage-based method for the detection of viable Mycobacterium avium subsp. paratuberculosis in blood within 48 h() |
title_sort | development of a rapid phage-based method for the detection of viable mycobacterium avium subsp. paratuberculosis in blood within 48 h() |
topic | Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3783900/ https://www.ncbi.nlm.nih.gov/pubmed/23811207 http://dx.doi.org/10.1016/j.mimet.2013.06.015 |
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