On Feasibility of Fluorescence-based Bacteria Presence Quantification: P.Aeruginosa

Alexander Caschera, Gennadi Saiko

2021

Abstract

Introduction: Wound healing typically occurs in the presence of bacteria at levels ranging from contamination to colonization to infection. The role of bacteria in wound healing depends on multiple factors, including bacterial concentration, species present, and host response. Thus, the determination of bacterial load is of great importance. However, existing clinical bacteria load assessment methods (biopsy or swabbing combined with culture methods) are slow, labor- and time-consuming. Pseudomonas aeruginosa is a known pathogen implicated in numerous healthcare-associated infections and can express fluorescent metabolites during proliferation. In particular, the siderophore pyoverdine produces a fluorescent emission between 450-520 nm when excited at 400nm and can be measured quantitatively via fluorescence spectroscopy. The current project aims to investigate the possibility of quantifying bacterial presence using fluorescence measurements. Methods: Cultures of P.aeruginosa (PA01) were grown at various temperatures (ambient temperature, 30, 37-43C̊), inoculum starting condition (5*107 -5*108 CFUmL-1), and initial nutrient’s concentration (0.6, 1.5, 3.0 gL-1) in Tryptic Soy Broth media. Media optical density (OD, as a proxy of bacterial concentration) and fluorescence (ex. 400nm, em. 420-520nm) were measured hourly for 10 hours. Results: Cultures remained metabolically active in the whole temperature range, producing pyoverdine fluorescence (emission max at 455nm). We have correlated optical density with a fluorescent signal to establish a dependence between fluorescence and growth stage. Noticeable pyoverdine accumulation started approximately 3 hours after the beginning of the log growth phase and experienced saturation at the beginning of the stationary phase. Three distinct regimes (a sigmoid curve) were observed: linear dependence of fluorescence on OD for low concentrations, more rapid nonlinear dependence, and saturation when approaching the stationary phase. Conclusions: The sigmoid dependence of bacterial fluorescence on their concentration persisted through variations in temperature and inoculum starting condition; thus, it may have the potential for determining culture growth phase progression. These results, combined with classical knowledge on disease progression, could also lead to an advanced infection diagnosis before current pathogenesis observation techniques.

Paper Citation

in Harvard Style

Caschera A. and Saiko G. (2021). On Feasibility of Fluorescence-based Bacteria Presence Quantification: P.Aeruginosa. In Proceedings of the 14th International Joint Conference on Biomedical Engineering Systems and Technologies (BIOSTEC 2021) - Volume 2: BIOIMAGING; ISBN 978-989-758-490-9, SciTePress, pages 193-200. DOI: 10.5220/0010344000002865

in Bibtex Style

@conference{bioimaging21,
author={Alexander Caschera and Gennadi Saiko},
title={On Feasibility of Fluorescence-based Bacteria Presence Quantification: P.Aeruginosa},
booktitle={Proceedings of the 14th International Joint Conference on Biomedical Engineering Systems and Technologies (BIOSTEC 2021) - Volume 2: BIOIMAGING},
year={2021},
pages={193-200},
publisher={SciTePress},
organization={INSTICC},
doi={10.5220/0010344000002865},
isbn={978-989-758-490-9},
}

in EndNote Style

TY - CONF

JO - Proceedings of the 14th International Joint Conference on Biomedical Engineering Systems and Technologies (BIOSTEC 2021) - Volume 2: BIOIMAGING
TI - On Feasibility of Fluorescence-based Bacteria Presence Quantification: P.Aeruginosa
SN - 978-989-758-490-9
AU - Caschera A.
AU - Saiko G.
PY - 2021
SP - 193
EP - 200
DO - 10.5220/0010344000002865
PB - SciTePress