3 years ago

Extracellular Polymeric Matrix Production and Relaxation under Fluid Shear and Mechanical Pressure in Staphylococcus aureus Biofilms.

Betsy van de Belt-Gritter, Jiapeng Hou, Deepak H Veeregowda, Henk J Busscher, Henny C van der Mei
The viscoelasticity of a biofilm's EPS (extracellular-polymeric-substance) matrix conveys protection against mechanical challenges, but adaptive responses of biofilm inhabitants to produce EPS are not well known. Here, we compare the response of a biofilm of an EPS producing (ATCC 12600) and non-EPS producing (5298) Staphylococcus aureus strain to fluid shear and mechanical challenge. Confocal-Laser-Scanning-Microscopy confirmed absence of calcofluorwhite-stainable EPS in biofilms of S. aureus 5298. ATR-FTIR spectroscopy combined with tribometry indicated that the polysaccharide production per bacterium in the initial adhering layer was higher during growth at high shear than at low shear and this increased EPS production extended to entire biofilms, as indicated by tribometrically measured coefficients of friction (CoF). CoFs of biofilms grown under high fluid shear were higher than when grown under low shear, likely due to wash-off of polysaccharides. Measurement of a biofilm's CoF implies application of mechanical pressure that yielded an immediate increase in polysaccharide band area of S. aureus ATCC 12600 biofilms due to their compression that decreased after relieving pressure to the level observed prior to mechanical pressure. For biofilms grown under high shear, this coincided with a higher %whiteness in Optical-Coherence-Tomography-images indicative of water outflow, returning back into the biofilm during stress relaxation. Biofilms grown under low shear however, were stimulated during tribometry to produce EPS, also after stress relieve. Knowledge of factors that govern EPS production and water flow in biofilms will allow better control of biofilms under mechanical challenge and understanding of the barrier properties of biofilms toward antimicrobial penetration.IMPORTANCE Adaptive responses of biofilm inhabitants in nature to environmental challenges such as fluid shear and mechanical pressure, often involve EPS production with the aim of protecting biofilm inhabitants. EPS can assist biofilm bacteria to remain attached or impede antimicrobial penetration. The tribochemist is a recently introduced instrument, allowing to study initially adhering bacteria to a Germanium crystal using ATR-FTIR spectroscopy, while simultaneously allowing measurement of the coefficient of friction of a biofilm, serving as an indicator of the EPS content of a biofilm. EPS production can be stimulated by both fluid shear during growth and mechanical pressure, while increased EPS production can continue after pressure relaxation of the biofilm. Since EPS is pivotal in the protection of biofilm inhabitants against mechanical and chemical challenges, knowledge the factors that make biofilm inhabitants decide to produce EPS as provided in this study, are important for the development of biofilm control measures.

Publisher URL: http://doi.org/10.1128/AEM.01516-17

DOI: 10.1128/AEM.01516-17

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