Cargando…

Quantitative Imaging of Bacteriophage Amplification for Rapid Detection of Bacteria in Model Foods

Rapid detection of bacteria in water and food samples is a critical need. The current molecular methods like real-time PCR can provide rapid detection after initial enrichment. However, these methods require significant preparation steps, specialized facilities to reduce contamination, and relativel...

Descripción completa

Detalles Bibliográficos
Autores principales: Wisuthiphaet, Nicharee, Yang, Xu, Young, Glenn M., Nitin, Nitin
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Frontiers Media S.A. 2022
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8931685/
https://www.ncbi.nlm.nih.gov/pubmed/35308341
http://dx.doi.org/10.3389/fmicb.2022.853048
_version_ 1784671312893968384
author Wisuthiphaet, Nicharee
Yang, Xu
Young, Glenn M.
Nitin, Nitin
author_facet Wisuthiphaet, Nicharee
Yang, Xu
Young, Glenn M.
Nitin, Nitin
author_sort Wisuthiphaet, Nicharee
collection PubMed
description Rapid detection of bacteria in water and food samples is a critical need. The current molecular methods like real-time PCR can provide rapid detection after initial enrichment. However, these methods require significant preparation steps, specialized facilities to reduce contamination, and relatively expensive reagents. This study evaluates a novel approach for detecting bacteria based on imaging of bacteriophage amplification upon infection of the target host bacteria to mitigate some of these constraints and improve the specificity of discriminating live vs. dead bacteria. Thus, this research leverages the natural ability of lytic bacteriophages to rapidly amplify their genetic material and generate progeny phages upon infecting the host bacterium. This study uses a nucleic acid staining dye, a conventional fluorescence microscope, and quantitative image analysis for imaging the amplification of bacteriophages. The sensitivity and assay time for imaging-based quantification of phage amplification for detecting Escherichia coli were compared with RT-PCR and the standard plaque-forming assay for detection phage amplification in model systems, including coconut water and spinach wash water. The results demonstrate that the imaging approach matches both the sensitivity and speed for detecting E. coli using the RT-PCR method without requiring isolation of nucleic acids, expensive reagents, and specialized facilities. The quantitative imaging results demonstrate the detection of 10 CFU/ml of E. coli in coconut water and simulated spinach wash water with a chemical oxygen demand (COD) of 3,000 ppm within 8 h, including initial enrichment of the bacteria. In summary, the results of this study illustrate a novel phage amplification-based approach for detecting target bacteria in complex food and water samples using simple sample preparation methods and low-cost reagents.
format Online
Article
Text
id pubmed-8931685
institution National Center for Biotechnology Information
language English
publishDate 2022
publisher Frontiers Media S.A.
record_format MEDLINE/PubMed
spelling pubmed-89316852022-03-19 Quantitative Imaging of Bacteriophage Amplification for Rapid Detection of Bacteria in Model Foods Wisuthiphaet, Nicharee Yang, Xu Young, Glenn M. Nitin, Nitin Front Microbiol Microbiology Rapid detection of bacteria in water and food samples is a critical need. The current molecular methods like real-time PCR can provide rapid detection after initial enrichment. However, these methods require significant preparation steps, specialized facilities to reduce contamination, and relatively expensive reagents. This study evaluates a novel approach for detecting bacteria based on imaging of bacteriophage amplification upon infection of the target host bacteria to mitigate some of these constraints and improve the specificity of discriminating live vs. dead bacteria. Thus, this research leverages the natural ability of lytic bacteriophages to rapidly amplify their genetic material and generate progeny phages upon infecting the host bacterium. This study uses a nucleic acid staining dye, a conventional fluorescence microscope, and quantitative image analysis for imaging the amplification of bacteriophages. The sensitivity and assay time for imaging-based quantification of phage amplification for detecting Escherichia coli were compared with RT-PCR and the standard plaque-forming assay for detection phage amplification in model systems, including coconut water and spinach wash water. The results demonstrate that the imaging approach matches both the sensitivity and speed for detecting E. coli using the RT-PCR method without requiring isolation of nucleic acids, expensive reagents, and specialized facilities. The quantitative imaging results demonstrate the detection of 10 CFU/ml of E. coli in coconut water and simulated spinach wash water with a chemical oxygen demand (COD) of 3,000 ppm within 8 h, including initial enrichment of the bacteria. In summary, the results of this study illustrate a novel phage amplification-based approach for detecting target bacteria in complex food and water samples using simple sample preparation methods and low-cost reagents. Frontiers Media S.A. 2022-03-04 /pmc/articles/PMC8931685/ /pubmed/35308341 http://dx.doi.org/10.3389/fmicb.2022.853048 Text en Copyright © 2022 Wisuthiphaet, Yang, Young and Nitin. https://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 Microbiology
Wisuthiphaet, Nicharee
Yang, Xu
Young, Glenn M.
Nitin, Nitin
Quantitative Imaging of Bacteriophage Amplification for Rapid Detection of Bacteria in Model Foods
title Quantitative Imaging of Bacteriophage Amplification for Rapid Detection of Bacteria in Model Foods
title_full Quantitative Imaging of Bacteriophage Amplification for Rapid Detection of Bacteria in Model Foods
title_fullStr Quantitative Imaging of Bacteriophage Amplification for Rapid Detection of Bacteria in Model Foods
title_full_unstemmed Quantitative Imaging of Bacteriophage Amplification for Rapid Detection of Bacteria in Model Foods
title_short Quantitative Imaging of Bacteriophage Amplification for Rapid Detection of Bacteria in Model Foods
title_sort quantitative imaging of bacteriophage amplification for rapid detection of bacteria in model foods
topic Microbiology
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8931685/
https://www.ncbi.nlm.nih.gov/pubmed/35308341
http://dx.doi.org/10.3389/fmicb.2022.853048
work_keys_str_mv AT wisuthiphaetnicharee quantitativeimagingofbacteriophageamplificationforrapiddetectionofbacteriainmodelfoods
AT yangxu quantitativeimagingofbacteriophageamplificationforrapiddetectionofbacteriainmodelfoods
AT youngglennm quantitativeimagingofbacteriophageamplificationforrapiddetectionofbacteriainmodelfoods
AT nitinnitin quantitativeimagingofbacteriophageamplificationforrapiddetectionofbacteriainmodelfoods