Raman Spectroscopy-Based Measurements of Single-Cell Phenotypic Diversity in Microbial Populations

ABSTRACT Microbial cells experience physiological changes due to environmental change, such as pH and temperature, the release of bactericidal agents, or nutrient limitation. This has been shown to affect community assembly and physiological processes (e.g., stress tolerance, virulence, or cellular...

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Autores principales:	Cristina García-Timermans, Ruben Props, Boris Zacchetti, Myrsini Sakarika, Frank Delvigne, Nico Boon
Formato:	article
Lenguaje:	EN
Publicado:	American Society for Microbiology 2020
Materias:	Raman spectroscopy microbial population stress phenotypic diversity single-cell analysis Hill numbers Microbiology QR1-502
Acceso en línea:	https://doaj.org/article/0530660be79b4bd58c2c707bfd59a29a
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spelling	oai:doaj.org-article:0530660be79b4bd58c2c707bfd59a29a2021-11-15T15:30:58ZRaman Spectroscopy-Based Measurements of Single-Cell Phenotypic Diversity in Microbial Populations10.1128/mSphere.00806-202379-5042https://doaj.org/article/0530660be79b4bd58c2c707bfd59a29a2020-10-01T00:00:00Zhttps://journals.asm.org/doi/10.1128/mSphere.00806-20https://doaj.org/toc/2379-5042ABSTRACT Microbial cells experience physiological changes due to environmental change, such as pH and temperature, the release of bactericidal agents, or nutrient limitation. This has been shown to affect community assembly and physiological processes (e.g., stress tolerance, virulence, or cellular metabolic activity). Metabolic stress is typically quantified by measuring community phenotypic properties such as biomass growth, reactive oxygen species, or cell permeability. However, bulk community measurements do not take into account single-cell phenotypic diversity, which is important for a better understanding and the subsequent management of microbial populations. Raman spectroscopy is a nondestructive alternative that provides detailed information on the biochemical makeup of each individual cell. Here, we introduce a method for describing single-cell phenotypic diversity using the Hill diversity framework of Raman spectra. Using the biomolecular profile of individual cells, we obtained a metric to compare cellular states and used it to study stress-induced changes. First, in two Escherichia coli populations either treated with ethanol or nontreated and then in two Saccharomyces cerevisiae subpopulations with either high or low expression of a stress reporter. In both cases, we were able to quantify single-cell phenotypic diversity and to discriminate metabolically stressed cells using a clustering algorithm. We also described how the lipid, protein, and nucleic acid compositions changed after the exposure to the stressor using information from the Raman spectra. Our results show that Raman spectroscopy delivers the necessary resolution to quantify phenotypic diversity within individual cells and that this information can be used to study stress-driven metabolic diversity in microbial populations. IMPORTANCE Microbial cells that live in the same community can exist in different physiological and morphological states that change as a function of spatiotemporal variations in environmental conditions. This phenomenon is commonly known as phenotypic heterogeneity and/or diversity. Measuring this plethora of cellular expressions is needed to better understand and manage microbial processes. However, most tools to study phenotypic diversity only average the behavior of the sampled community. In this work, we present a way to quantify the phenotypic diversity of microbial samples by inferring the (bio)molecular profile of its constituent cells using Raman spectroscopy. We demonstrate how this tool can be used to quantify the phenotypic diversity that arises after the exposure of microbes to stress. Raman spectroscopy holds potential for the detection of stressed cells in bioproduction.Cristina García-TimermansRuben PropsBoris ZacchettiMyrsini SakarikaFrank DelvigneNico BoonAmerican Society for MicrobiologyarticleRaman spectroscopymicrobial populationstressphenotypic diversitysingle-cell analysisHill numbersMicrobiologyQR1-502ENmSphere, Vol 5, Iss 5 (2020)
institution	DOAJ
collection	DOAJ
language	EN
topic	Raman spectroscopy microbial population stress phenotypic diversity single-cell analysis Hill numbers Microbiology QR1-502
spellingShingle	Raman spectroscopy microbial population stress phenotypic diversity single-cell analysis Hill numbers Microbiology QR1-502 Cristina García-Timermans Ruben Props Boris Zacchetti Myrsini Sakarika Frank Delvigne Nico Boon Raman Spectroscopy-Based Measurements of Single-Cell Phenotypic Diversity in Microbial Populations
description	ABSTRACT Microbial cells experience physiological changes due to environmental change, such as pH and temperature, the release of bactericidal agents, or nutrient limitation. This has been shown to affect community assembly and physiological processes (e.g., stress tolerance, virulence, or cellular metabolic activity). Metabolic stress is typically quantified by measuring community phenotypic properties such as biomass growth, reactive oxygen species, or cell permeability. However, bulk community measurements do not take into account single-cell phenotypic diversity, which is important for a better understanding and the subsequent management of microbial populations. Raman spectroscopy is a nondestructive alternative that provides detailed information on the biochemical makeup of each individual cell. Here, we introduce a method for describing single-cell phenotypic diversity using the Hill diversity framework of Raman spectra. Using the biomolecular profile of individual cells, we obtained a metric to compare cellular states and used it to study stress-induced changes. First, in two Escherichia coli populations either treated with ethanol or nontreated and then in two Saccharomyces cerevisiae subpopulations with either high or low expression of a stress reporter. In both cases, we were able to quantify single-cell phenotypic diversity and to discriminate metabolically stressed cells using a clustering algorithm. We also described how the lipid, protein, and nucleic acid compositions changed after the exposure to the stressor using information from the Raman spectra. Our results show that Raman spectroscopy delivers the necessary resolution to quantify phenotypic diversity within individual cells and that this information can be used to study stress-driven metabolic diversity in microbial populations. IMPORTANCE Microbial cells that live in the same community can exist in different physiological and morphological states that change as a function of spatiotemporal variations in environmental conditions. This phenomenon is commonly known as phenotypic heterogeneity and/or diversity. Measuring this plethora of cellular expressions is needed to better understand and manage microbial processes. However, most tools to study phenotypic diversity only average the behavior of the sampled community. In this work, we present a way to quantify the phenotypic diversity of microbial samples by inferring the (bio)molecular profile of its constituent cells using Raman spectroscopy. We demonstrate how this tool can be used to quantify the phenotypic diversity that arises after the exposure of microbes to stress. Raman spectroscopy holds potential for the detection of stressed cells in bioproduction.
format	article
author	Cristina García-Timermans Ruben Props Boris Zacchetti Myrsini Sakarika Frank Delvigne Nico Boon
author_facet	Cristina García-Timermans Ruben Props Boris Zacchetti Myrsini Sakarika Frank Delvigne Nico Boon
author_sort	Cristina García-Timermans
title	Raman Spectroscopy-Based Measurements of Single-Cell Phenotypic Diversity in Microbial Populations
title_short	Raman Spectroscopy-Based Measurements of Single-Cell Phenotypic Diversity in Microbial Populations
title_full	Raman Spectroscopy-Based Measurements of Single-Cell Phenotypic Diversity in Microbial Populations
title_fullStr	Raman Spectroscopy-Based Measurements of Single-Cell Phenotypic Diversity in Microbial Populations
title_full_unstemmed	Raman Spectroscopy-Based Measurements of Single-Cell Phenotypic Diversity in Microbial Populations
title_sort	raman spectroscopy-based measurements of single-cell phenotypic diversity in microbial populations
publisher	American Society for Microbiology
publishDate	2020
url	https://doaj.org/article/0530660be79b4bd58c2c707bfd59a29a
work_keys_str_mv	AT cristinagarciatimermans ramanspectroscopybasedmeasurementsofsinglecellphenotypicdiversityinmicrobialpopulations AT rubenprops ramanspectroscopybasedmeasurementsofsinglecellphenotypicdiversityinmicrobialpopulations AT boriszacchetti ramanspectroscopybasedmeasurementsofsinglecellphenotypicdiversityinmicrobialpopulations AT myrsinisakarika ramanspectroscopybasedmeasurementsofsinglecellphenotypicdiversityinmicrobialpopulations AT frankdelvigne ramanspectroscopybasedmeasurementsofsinglecellphenotypicdiversityinmicrobialpopulations AT nicoboon ramanspectroscopybasedmeasurementsofsinglecellphenotypicdiversityinmicrobialpopulations
_version_	1718427867264581632

Raman Spectroscopy-Based Measurements of Single-Cell Phenotypic Diversity in Microbial Populations

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