Micron Scale Spatial Measurement of the O<sub>2</sub> Gradient Surrounding a Bacterial Biofilm in Real Time

ABSTRACT Bacteria alter their local chemical environment through both consumption and the production of a variety of molecules, ultimately shaping the local ecology. Molecular oxygen (O2) is a key metabolite that affects the physiology and behavior of virtually all bacteria, and its consumption ofte...

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Autores principales:	Alexander D. Klementiev, Zhaoyu Jin, Marvin Whiteley
Formato:	article
Lenguaje:	EN
Publicado:	American Society for Microbiology 2020
Materias:	biofilm oxygen antibiotics electrochemistry Pseudomonas aeruginosa antibiotic resistance Microbiology QR1-502
Acceso en línea:	https://doaj.org/article/810c897ac7164acf9063faa34151c27d
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spelling	oai:doaj.org-article:810c897ac7164acf9063faa34151c27d2021-11-15T16:19:09ZMicron Scale Spatial Measurement of the O<sub>2</sub> Gradient Surrounding a Bacterial Biofilm in Real Time10.1128/mBio.02536-202150-7511https://doaj.org/article/810c897ac7164acf9063faa34151c27d2020-10-01T00:00:00Zhttps://journals.asm.org/doi/10.1128/mBio.02536-20https://doaj.org/toc/2150-7511ABSTRACT Bacteria alter their local chemical environment through both consumption and the production of a variety of molecules, ultimately shaping the local ecology. Molecular oxygen (O2) is a key metabolite that affects the physiology and behavior of virtually all bacteria, and its consumption often results in O2 gradients within sessile bacterial communities (biofilms). O2 plays a critical role in several bacterial phenotypes, including antibiotic tolerance; however, our understanding of O2 levels within and surrounding biofilms has been hampered by the difficulties in measuring O2 levels in real-time for extended durations and at the micron scale. Here, we developed electrochemical methodology based on scanning electrochemical microscopy to quantify the O2 gradients present above a Pseudomonas aeruginosa biofilm. These results reveal that a biofilm produces a hypoxic zone that extends hundreds of microns from the biofilm surface within minutes and that the biofilm consumes O2 at a maximum rate. Treating the biofilm with levels of the antibiotic ciprofloxacin that kill 99% of the bacteria did not affect the O2 gradient, indicating that the biofilm is highly resilient to antimicrobial treatment in regard to O2 consumption. IMPORTANCE O2 is a fundamental environmental metabolite that affects all life on earth. While toxic to many microbes and obligately required by others, those that have appropriate physiological responses survive and can even benefit from various levels of O2, particularly in biofilm communities. Although most studies have focused on measuring O2 within biofilms, little is known about O2 gradients surrounding biofilms. Here, we developed electrochemical methodology based on scanning electrochemical microscopy to measure the O2 gradients surrounding biofilms in real time on the micron scale. Our results reveal that P. aeruginosa biofilms produce a hypoxic zone that can extend hundreds of microns from the biofilm surface and that this gradient remains even after the addition of antibiotic concentrations that eradicated 99% of viable cells. Our results provide a high resolution of the O2 gradients produced by P. aeruginosa biofilms and reveal sustained O2 consumption in the presence of antibiotics.Alexander D. KlementievZhaoyu JinMarvin WhiteleyAmerican Society for MicrobiologyarticlebiofilmoxygenantibioticselectrochemistryPseudomonas aeruginosaantibiotic resistanceMicrobiologyQR1-502ENmBio, Vol 11, Iss 5 (2020)
institution	DOAJ
collection	DOAJ
language	EN
topic	biofilm oxygen antibiotics electrochemistry Pseudomonas aeruginosa antibiotic resistance Microbiology QR1-502
spellingShingle	biofilm oxygen antibiotics electrochemistry Pseudomonas aeruginosa antibiotic resistance Microbiology QR1-502 Alexander D. Klementiev Zhaoyu Jin Marvin Whiteley Micron Scale Spatial Measurement of the O<sub>2</sub> Gradient Surrounding a Bacterial Biofilm in Real Time
description	ABSTRACT Bacteria alter their local chemical environment through both consumption and the production of a variety of molecules, ultimately shaping the local ecology. Molecular oxygen (O2) is a key metabolite that affects the physiology and behavior of virtually all bacteria, and its consumption often results in O2 gradients within sessile bacterial communities (biofilms). O2 plays a critical role in several bacterial phenotypes, including antibiotic tolerance; however, our understanding of O2 levels within and surrounding biofilms has been hampered by the difficulties in measuring O2 levels in real-time for extended durations and at the micron scale. Here, we developed electrochemical methodology based on scanning electrochemical microscopy to quantify the O2 gradients present above a Pseudomonas aeruginosa biofilm. These results reveal that a biofilm produces a hypoxic zone that extends hundreds of microns from the biofilm surface within minutes and that the biofilm consumes O2 at a maximum rate. Treating the biofilm with levels of the antibiotic ciprofloxacin that kill 99% of the bacteria did not affect the O2 gradient, indicating that the biofilm is highly resilient to antimicrobial treatment in regard to O2 consumption. IMPORTANCE O2 is a fundamental environmental metabolite that affects all life on earth. While toxic to many microbes and obligately required by others, those that have appropriate physiological responses survive and can even benefit from various levels of O2, particularly in biofilm communities. Although most studies have focused on measuring O2 within biofilms, little is known about O2 gradients surrounding biofilms. Here, we developed electrochemical methodology based on scanning electrochemical microscopy to measure the O2 gradients surrounding biofilms in real time on the micron scale. Our results reveal that P. aeruginosa biofilms produce a hypoxic zone that can extend hundreds of microns from the biofilm surface and that this gradient remains even after the addition of antibiotic concentrations that eradicated 99% of viable cells. Our results provide a high resolution of the O2 gradients produced by P. aeruginosa biofilms and reveal sustained O2 consumption in the presence of antibiotics.
format	article
author	Alexander D. Klementiev Zhaoyu Jin Marvin Whiteley
author_facet	Alexander D. Klementiev Zhaoyu Jin Marvin Whiteley
author_sort	Alexander D. Klementiev
title	Micron Scale Spatial Measurement of the O<sub>2</sub> Gradient Surrounding a Bacterial Biofilm in Real Time
title_short	Micron Scale Spatial Measurement of the O<sub>2</sub> Gradient Surrounding a Bacterial Biofilm in Real Time
title_full	Micron Scale Spatial Measurement of the O<sub>2</sub> Gradient Surrounding a Bacterial Biofilm in Real Time
title_fullStr	Micron Scale Spatial Measurement of the O<sub>2</sub> Gradient Surrounding a Bacterial Biofilm in Real Time
title_full_unstemmed	Micron Scale Spatial Measurement of the O<sub>2</sub> Gradient Surrounding a Bacterial Biofilm in Real Time
title_sort	micron scale spatial measurement of the o<sub>2</sub> gradient surrounding a bacterial biofilm in real time
publisher	American Society for Microbiology
publishDate	2020
url	https://doaj.org/article/810c897ac7164acf9063faa34151c27d
work_keys_str_mv	AT alexanderdklementiev micronscalespatialmeasurementoftheosub2subgradientsurroundingabacterialbiofilminrealtime AT zhaoyujin micronscalespatialmeasurementoftheosub2subgradientsurroundingabacterialbiofilminrealtime AT marvinwhiteley micronscalespatialmeasurementoftheosub2subgradientsurroundingabacterialbiofilminrealtime
_version_	1718426910634016768

Micron Scale Spatial Measurement of the O<sub>2</sub> Gradient Surrounding a Bacterial Biofilm in Real Time

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