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Ultrasensitive and Fast Diagnostics of Viable Listeria Cells by CBD Magnetic Separation Combined with A511::luxAB Detection

The genus Listeria includes foodborne pathogens that cause life-threatening infections in those at risk, and sensitive and specific methods for detection of these bacteria are needed. Based on their unrivaled host specificity and ability to discriminate viable cells, bacteriophages represent an idea...

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Autores principales: Kretzer, Jan W., Schmelcher, Mathias, Loessner, Martin J.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2018
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6266503/
https://www.ncbi.nlm.nih.gov/pubmed/30428537
http://dx.doi.org/10.3390/v10110626
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author Kretzer, Jan W.
Schmelcher, Mathias
Loessner, Martin J.
author_facet Kretzer, Jan W.
Schmelcher, Mathias
Loessner, Martin J.
author_sort Kretzer, Jan W.
collection PubMed
description The genus Listeria includes foodborne pathogens that cause life-threatening infections in those at risk, and sensitive and specific methods for detection of these bacteria are needed. Based on their unrivaled host specificity and ability to discriminate viable cells, bacteriophages represent an ideal toolbox for the development of such methods. Here, the authors describe an ultrasensitive diagnostic protocol for Listeria by combining two phage-based strategies: (1) specific capture and concentration of target cells by magnetic separation, harnessing cell wall-binding domains from Listeria phage endolysins (CBD-MS); and (2) highly sensitive detection using an adaptation of the A511::luxAB bioluminescent reporter phage assay in a microwell plate format. The combined assay enabled direct detection of approximately 100 bacteria per ml of pure culture with genus-level specificity in less than 6 h. For contaminated foods, the procedure included a 16 h selective enrichment step, followed by CBD-MS separation and A511::luxAB detection. It was able to consistently detect extremely low numbers (0.1 to 1.0 cfu/g) of viable Listeria cells, in a total assay time of less than 22 h. These results demonstrate the superiority of this phage-based assay to standard culture-based diagnostic protocols for the detection of viable bacteria, with respect to both sensitivity and speed.
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spelling pubmed-62665032018-12-07 Ultrasensitive and Fast Diagnostics of Viable Listeria Cells by CBD Magnetic Separation Combined with A511::luxAB Detection Kretzer, Jan W. Schmelcher, Mathias Loessner, Martin J. Viruses Article The genus Listeria includes foodborne pathogens that cause life-threatening infections in those at risk, and sensitive and specific methods for detection of these bacteria are needed. Based on their unrivaled host specificity and ability to discriminate viable cells, bacteriophages represent an ideal toolbox for the development of such methods. Here, the authors describe an ultrasensitive diagnostic protocol for Listeria by combining two phage-based strategies: (1) specific capture and concentration of target cells by magnetic separation, harnessing cell wall-binding domains from Listeria phage endolysins (CBD-MS); and (2) highly sensitive detection using an adaptation of the A511::luxAB bioluminescent reporter phage assay in a microwell plate format. The combined assay enabled direct detection of approximately 100 bacteria per ml of pure culture with genus-level specificity in less than 6 h. For contaminated foods, the procedure included a 16 h selective enrichment step, followed by CBD-MS separation and A511::luxAB detection. It was able to consistently detect extremely low numbers (0.1 to 1.0 cfu/g) of viable Listeria cells, in a total assay time of less than 22 h. These results demonstrate the superiority of this phage-based assay to standard culture-based diagnostic protocols for the detection of viable bacteria, with respect to both sensitivity and speed. MDPI 2018-11-13 /pmc/articles/PMC6266503/ /pubmed/30428537 http://dx.doi.org/10.3390/v10110626 Text en © 2018 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/).
spellingShingle Article
Kretzer, Jan W.
Schmelcher, Mathias
Loessner, Martin J.
Ultrasensitive and Fast Diagnostics of Viable Listeria Cells by CBD Magnetic Separation Combined with A511::luxAB Detection
title Ultrasensitive and Fast Diagnostics of Viable Listeria Cells by CBD Magnetic Separation Combined with A511::luxAB Detection
title_full Ultrasensitive and Fast Diagnostics of Viable Listeria Cells by CBD Magnetic Separation Combined with A511::luxAB Detection
title_fullStr Ultrasensitive and Fast Diagnostics of Viable Listeria Cells by CBD Magnetic Separation Combined with A511::luxAB Detection
title_full_unstemmed Ultrasensitive and Fast Diagnostics of Viable Listeria Cells by CBD Magnetic Separation Combined with A511::luxAB Detection
title_short Ultrasensitive and Fast Diagnostics of Viable Listeria Cells by CBD Magnetic Separation Combined with A511::luxAB Detection
title_sort ultrasensitive and fast diagnostics of viable listeria cells by cbd magnetic separation combined with a511::luxab detection
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6266503/
https://www.ncbi.nlm.nih.gov/pubmed/30428537
http://dx.doi.org/10.3390/v10110626
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