A novel metabolomic approach used for the comparison of Staphylococcus aureus planktonic cells and biofilm samples

Stipetic, L. H., Dalby, M. J. , Davies, R. L. , Morton, F. r., Ramage, G. and Burgess, K. (2016) A novel metabolomic approach used for the comparison of Staphylococcus aureus planktonic cells and biofilm samples. Metabolomics, 12, 75. (doi:10.1007/s11306-016-1002-0) (PMID:27013931) (PMCID:PMC4783440)

Stipetic, L. H., Dalby, M. J. , Davies, R. L. , Morton, F. r., Ramage, G. and Burgess, K. (2016) A novel metabolomic approach used for the comparison of Staphylococcus aureus planktonic cells and biofilm samples. Metabolomics, 12, 75. (doi:10.1007/s11306-016-1002-0) (PMID:27013931) (PMCID:PMC4783440)

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Abstract

Introduction: Bacterial cell characteristics change significantly during differentiation between planktonic and biofilm states. While established methods exist to detect and identify transcriptional and proteomic changes, metabolic fluctuations that distinguish these developmental stages have been less amenable to investigation. Objectives: The objectives of the study were to develop a robust reproducible sample preparation methodology for high throughput biofilm analysis and to determine differences between Staphylococcus aureus in planktonic and biofilm states. Methods: The method uses bead beating in a chloroform/methanol/water extraction solvent to both disrupt cells and quench metabolism. Verification of the method was performed using liquid-chromatography-mass spectrometry. Raw mass-spectrometry data was analysed using an in-house bioinformatics pipe-line incorporating XCMS, MzMatch and in-house R-scripts, with identifications matched to internal standards and metabolite data-base entries. Results: We have demonstrated a novel mechanical bead beating method that has been optimised for the extraction of the metabolome from cells of a clinical Staphylococcus aureus strain existing in a planktonic or biofilm state. This high-throughput method is fast and reproducible, allowing for direct comparison between different bacterial growth states. Significant changes in arginine biosynthesis were identified between the two cell populations. Conclusions: The method described herein represents a valuable tool in studying microbial biochemistry at a molecular level. While the methodology is generally applicable to the lysis and extraction of metabolites from Gram positive bacteria, it is particularly applicable to biofilms. Bacteria that exist as a biofilm are shown to be highly distinct metabolically from their ‘free living’ counterparts, thus highlighting the need to study microbes in different growth states. Metabolomics can successfully distinguish between a planktonic and biofilm growth state. Importantly, this study design, incorporating metabolomics, could be optimised for studying the effects of antimicrobials and drug modes of action, potentially providing explanations and mechanisms of antibiotic resistance and to help devise new antimicrobials.

Item Type:Articles
Status:Published
Refereed:Yes
Glasgow Author(s) Enlighten ID:Burgess, Dr Karl and Ramage, Professor Gordon and Morton, Mr Fraser and Davies, Dr Robert and Dalby, Professor Matthew
Authors: Stipetic, L. H., Dalby, M. J., Davies, R. L., Morton, F. r., Ramage, G., and Burgess, K.
College/School:College of Medical Veterinary and Life Sciences > Institute of Infection Immunity and Inflammation
College of Medical Veterinary and Life Sciences > Institute of Molecular Cell and Systems Biology
Journal Name:Metabolomics
Publisher:Springer
ISSN:1573-3882
ISSN (Online):1573-3890
Published Online:08 March 2016
Copyright Holders:Copyright © 2016 The Authors
First Published:First published in Metabolomics 12:75
Publisher Policy:Reproduced under a Creative Commons licence

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Project CodeAward NoProject NamePrincipal InvestigatorFunder's NameFunder RefLead Dept
594271Integrated Health - Polyomics and Systems Biomedicine (ISSF Bid)Anna DominiczakWellcome Trust (WELLCOME)097821/Z/11/ZRI CARDIOVASCULAR & MEDICAL SCIENCES
623593Institutional Strategic Support Fund (ISSF)Anna DominiczakWellcome Trust (WELLCOME)105614/Z/14/ZRI CARDIOVASCULAR & MEDICAL SCIENCES
470561DTC in cell and proteomic technologies (continuation)Jonathan CooperEngineering & Physical Sciences Research Council (EPSRC)EP/F500424/1ENG - BIOMEDICAL ENGINEERING