Stress Relaxation Analysis Facilitates a Quantitative Approach towards Antimicrobial Penetration into Biofilms

Abstract

Biofilm-related infections can develop everywhere in the human body and are rarely cleared by the host immune system. Moreover, biofilms are often tolerant to antimicrobials, due to a combination of inherent properties of bacteria in their adhering, biofilm mode of growth and poor physical penetration of antimicrobials through biofilms. Current understanding of biofilm recalcitrance toward antimicrobial penetration is based on qualitative descriptions of biofilms. Here we hypothesize that stress relaxation of biofilms will relate with antimicrobial penetration. Stress relaxation analysis of single-species oral biofilms grown in vitro identified a fast, intermediate and slow response to an induced deformation, corresponding with outflow of water and extracellular polymeric substances, and bacterial re-arrangement, respectively. Penetration of chlorhexidine into these biofilms increased with increasing relative importance of the slow and decreasing importance of the fast relaxation element. Involvement of slow relaxation elements suggests that biofilm structures allowing extensive bacterial re-arrangement after deformation are more open, allowing better antimicrobial penetration. Involvement of fast relaxation elements suggests that water dilutes the antimicrobial upon penetration to an ineffective concentration in deeper layers of the biofilm. Next, we collected biofilms formed in intra-oral collection devices bonded to the buccal surfaces of the maxillary first molars of human volunteers. Ex situ chlorhexidine penetration into two weeks old in vivo formed biofilms followed a similar dependence on the importance of the fast and slow relaxation elements as observed for in vitro formed biofilms. This study demonstrates that biofilm properties can be derived that quantitatively explain antimicrobial penetration into a biofilm.

Static biofilm removal around ultrasonic tips in vitro

“…The extent to which specific different hydrodynamic factors contribute is still dependent on the specifics of the instruments. On the other hand, He and co-workers found that powered brushing in non-contact mode changed the viscoelastic properties of the oral biofilm resulting in increased penetration of antimicrobial compounds into the biofilm [24,25]. Therefore, even if biofilms are not totally removed by sonic brushing, they may be more susceptible to antimicrobials thereafter.…”

Biomechanical Analysis of Infectious Biofilms

“…), with the motion of one controlled to impose a predefined strain or stress schedule. Biofilms can be grown ex situ and transplanted to the rheometer, either intact on plates [41,66] or by a destructive process [55,104], or the rheometer modified to permit growth in situ [69,70]. Macrorheometry can extract bulk properties [68,82] but is not well suited to studying heterogeneities.…”

“…The free surfaces of biofilms in their native state can be compressed globally [41,71], or locally such as performed by atomic force microscopy [14,54], and force-displacement relations and relaxation curves measured. Heterogeneity can be probed by multi-site sampling [48].…”

“…Fitting stress relaxation in compressed biofilms to a generalised Maxwell model has been used to infer the relevance of extracellular DNA to the viscoelasticity of single species S. mutans, P. aeruginosa, S. aureus and S. epidermidis systems, where principal component analysis was employed to determine the number of modes [71]. Biomass rearrangement and flow modes were inferred from similar analysis of two oral species (S. oralis and A. naeslundii) and intact dental plaque [41]. Despite the intuitive appeal of these findings, it should be noted that spring-dashpot models are idealised and admit only a small number of relaxation times, in contrast to P. aeruginosa matrix extracts which have been shown to relax over a broad distribution of time scales [104].…”

Dental Plak Oluşumundan Sorumlu Mikroorganizmalar ve Bunların Tüketilen Gıdalarla İlişkisi

“…Yaptığı incelemelerde çok sayıda küçük canlının insan diş yüzeyinde yaşadığını fark eden Leeuwenhoek, dişlerini sirke ile yıkadıktan sonra da örnek alarak inceleme yapmış ve dişindeki tortu dışında yaşayan hayvancıkların öldüğünü ancak tortu içerisinde bulunanlara sirkenin etki etmediğini görmüştür [1].…”