Confocal spectral microscopy, a non-destructive approach to follow contamination and biofilm formation of mCherry Staphylococcus aureus on solid surfaces

Muhammad Tanveer Munir,Mark Irle,Chantal Thorin,Ismael Oumarou-Mahamane,Matthieu Eveillard,Laurence Dubreil,Florence Aviat,Julien Pichon,Hélène Pailhoriès,Mohammed Gharbia,Christophe Belloncle,Nattar Maneewan,Michel Federighi,Patrice Le Pape,Jessica Baude,Didier Lepelletier

doi:10.1038/s41598-021-94939-2

Abstract

Methods to test the safety of wood material for hygienically sensitive places are indirect, destructive and limited to incomplete microbial recovery via swabbing, brushing and elution-based techniques. Therefore, we chose mCherry Staphylococcus aureus as a model bacterium for solid and porous surface contamination. Confocal spectral laser microscope (CSLM) was employed to characterize and use the autofluorescence of Sessile oak (Quercus petraea), Douglas fir (Pseudotsuga menziesii) and poplar (Populus euramericana alba L.) wood discs cut into transversal (RT) and tangential (LT) planes. The red fluorescent area occupied by bacteria was differentiated from that of wood, which represented the bacterial quantification, survival and bio-distribution on surfaces from one hour to one week after inoculation. More bacteria were present near the surface on LT face wood as compared to RT and they persisted throughout the study period. Furthermore, this innovative methodology identified that S. aureus formed a dense biofilm on melamine but not on oak wood in similar inoculation and growth conditions. Conclusively, the endogenous fluorescence of materials and the model bacterium permitted direct quantification of surface contamination by using CSLM and it is a promising tool for hygienic safety evaluation.

Highlights

Methods to test the safety of wood material for hygienically sensitive places are indirect, destructive and limited to incomplete microbial recovery via swabbing, brushing and elution-based techniques
This study aimed to develop a methodology to visualize firstly the spatial distribution of the S. aureus bacterium on different wood species to be able to analyse the bacterial contamination on wooden surfaces over time and secondly to apply this methodology to investigate biofilm formation potential of bacterium on wood as compared to melamine
Measurements were done on wood disc by using 405 nm excitation and fluorescence collection between 405 and 690 nm

Summary

Introduction

Methods to test the safety of wood material for hygienically sensitive places are indirect, destructive and limited to incomplete microbial recovery via swabbing, brushing and elution-based techniques. Wood is an organic and porous material that is abundantly used in hygienically important places, such as in the architecture or indoor construction of the interiors of healthcare and residential buildings, and in the food industry as contact surfaces for food preparation, packaging and storage. Regarding the hygienically important microbes such as Staphylococcus aureus, which is one of the most common bacteria responsible for hospital-acquired infections (HAI)[3] and foodborne o utbreaks[4] This bacterium survives on inanimate surfaces from days to months depending upon environmental c onditions[5,6,7,8]. Studying the microbial distribution inside the porous wood material will provide clues whether the microbes can attach and stay hidden inside the surfaces or are killed due to the antimicrobial activity of wood

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