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Using Fluorescence Intensity of Enhanced Green Fluorescent Protein to Quantify Pseudomonas aeruginosa

A variety of direct and indirect methods have been used to quantify planktonic and biofilm bacterial cells. Direct counting methods to determine the total number of cells include plate counts, microscopic cell counts, Coulter cell counting, flow cytometry, and fluorescence microscopy. However, indir...

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Autores principales: Wilson, Erin, Okuom, Macduff, Kyes, Nathan, Mayfield, Dylan, Wilson, Christina, Sabatka, Derek, Sandoval, Jasmin, Foote, Jared R., Kangas, Michael J., Holmes, Andrea E., Sutlief, Arin L.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: 2018
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6860918/
https://www.ncbi.nlm.nih.gov/pubmed/31741893
http://dx.doi.org/10.3390/chemosensors6020021
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author Wilson, Erin
Okuom, Macduff
Kyes, Nathan
Mayfield, Dylan
Wilson, Christina
Sabatka, Derek
Sandoval, Jasmin
Foote, Jared R.
Kangas, Michael J.
Holmes, Andrea E.
Sutlief, Arin L.
author_facet Wilson, Erin
Okuom, Macduff
Kyes, Nathan
Mayfield, Dylan
Wilson, Christina
Sabatka, Derek
Sandoval, Jasmin
Foote, Jared R.
Kangas, Michael J.
Holmes, Andrea E.
Sutlief, Arin L.
author_sort Wilson, Erin
collection PubMed
description A variety of direct and indirect methods have been used to quantify planktonic and biofilm bacterial cells. Direct counting methods to determine the total number of cells include plate counts, microscopic cell counts, Coulter cell counting, flow cytometry, and fluorescence microscopy. However, indirect methods are often used to supplement direct cell counting, as they are often more convenient, less time-consuming, and require less material, while providing a number that can be related to the direct cell count. Herein, an indirect method is presented that uses fluorescence emission intensity as a proxy marker for studying bacterial accumulation. A clinical strain of Pseudomonas aeruginosa was genetically modified to express a green fluorescent protein (PA14/EGFP). The fluorescence intensity of EGFP in live cells was used as an indirect measure of live cell density, and was compared with the traditional cell counting methods of optical density (OD(600)) and plate counting (colony-forming units (CFUs)). While both OD(600) and CFUs are well-established methods, the use of fluorescence spectroscopy to quantify bacteria is less common. This study demonstrates that EGFP intensity is a convenient reporter for bacterial quantification. In addition, we demonstrate the potential for fluorescence spectroscopy to be used to measure the quantity of PA14/EGFP biofilms, which have important human health implications due to their antimicrobial resistance. Therefore, fluorescence spectroscopy could serve as an alternative or complementary quick assay to quantify bacteria in planktonic cultures and biofilms.
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spelling pubmed-68609182019-11-18 Using Fluorescence Intensity of Enhanced Green Fluorescent Protein to Quantify Pseudomonas aeruginosa Wilson, Erin Okuom, Macduff Kyes, Nathan Mayfield, Dylan Wilson, Christina Sabatka, Derek Sandoval, Jasmin Foote, Jared R. Kangas, Michael J. Holmes, Andrea E. Sutlief, Arin L. Chemosensors (Basel) Article A variety of direct and indirect methods have been used to quantify planktonic and biofilm bacterial cells. Direct counting methods to determine the total number of cells include plate counts, microscopic cell counts, Coulter cell counting, flow cytometry, and fluorescence microscopy. However, indirect methods are often used to supplement direct cell counting, as they are often more convenient, less time-consuming, and require less material, while providing a number that can be related to the direct cell count. Herein, an indirect method is presented that uses fluorescence emission intensity as a proxy marker for studying bacterial accumulation. A clinical strain of Pseudomonas aeruginosa was genetically modified to express a green fluorescent protein (PA14/EGFP). The fluorescence intensity of EGFP in live cells was used as an indirect measure of live cell density, and was compared with the traditional cell counting methods of optical density (OD(600)) and plate counting (colony-forming units (CFUs)). While both OD(600) and CFUs are well-established methods, the use of fluorescence spectroscopy to quantify bacteria is less common. This study demonstrates that EGFP intensity is a convenient reporter for bacterial quantification. In addition, we demonstrate the potential for fluorescence spectroscopy to be used to measure the quantity of PA14/EGFP biofilms, which have important human health implications due to their antimicrobial resistance. Therefore, fluorescence spectroscopy could serve as an alternative or complementary quick assay to quantify bacteria in planktonic cultures and biofilms. 2018-05-03 2018-06 /pmc/articles/PMC6860918/ /pubmed/31741893 http://dx.doi.org/10.3390/chemosensors6020021 Text en http://creativecommons.org/licenses/by/4.0/ 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
Wilson, Erin
Okuom, Macduff
Kyes, Nathan
Mayfield, Dylan
Wilson, Christina
Sabatka, Derek
Sandoval, Jasmin
Foote, Jared R.
Kangas, Michael J.
Holmes, Andrea E.
Sutlief, Arin L.
Using Fluorescence Intensity of Enhanced Green Fluorescent Protein to Quantify Pseudomonas aeruginosa
title Using Fluorescence Intensity of Enhanced Green Fluorescent Protein to Quantify Pseudomonas aeruginosa
title_full Using Fluorescence Intensity of Enhanced Green Fluorescent Protein to Quantify Pseudomonas aeruginosa
title_fullStr Using Fluorescence Intensity of Enhanced Green Fluorescent Protein to Quantify Pseudomonas aeruginosa
title_full_unstemmed Using Fluorescence Intensity of Enhanced Green Fluorescent Protein to Quantify Pseudomonas aeruginosa
title_short Using Fluorescence Intensity of Enhanced Green Fluorescent Protein to Quantify Pseudomonas aeruginosa
title_sort using fluorescence intensity of enhanced green fluorescent protein to quantify pseudomonas aeruginosa
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6860918/
https://www.ncbi.nlm.nih.gov/pubmed/31741893
http://dx.doi.org/10.3390/chemosensors6020021
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