Biofilm disruption by an air bubble reveals heterogeneous age-dependent detachment patterns dictated by initial extracellular matrix distribution

Abstract

Bacteria often adhere to surfaces, where they form communities known as biofilms. Recently, it has been shown that biofilm formation initiates with the microscopically heterogeneous deposition of a skeleton of extracellular polymeric substances (EPS) by individual cells crawling on the surface, followed by growth of the biofilm into a surface-covering continuum. Here we report microfluidic experiments with Pseudomonas aeruginosa biofilms showing that their “hidden” heterogeneity can affect the later dynamics of their disruption. Using controlled air bubbles as a model for mechanical insult, we demonstrate that biofilm disruption is strongly dependent on biofilm age, and that disruption to early-stage biofilms can take the shape of a semi-regular pattern of ~15 µm diameter holes from which bacteria have been removed. We explain hole formation in terms of the rupture and retreat of the thin liquid layer created by the long bubble, which scrapes bacteria off the surface and rearranges their distribution. We find that the resulting pattern correlates with the spatial distribution of EPS: holes form where there is less EPS, whereas regions with more EPS act as strongholds against the scraping liquid front. These results show that heterogeneity in the microscale EPS skeleton of biofilms has profound consequences for later dynamics, including disruption. Because few attached cells suffice to regrow a biofilm, these results point to the importance of considering microscale heterogeneity when designing and assessing the effectiveness of biofilm removal strategies by mechanical forces.

Bubble-mediated transport of benthic microorganisms into the water column: Identification of methanotrophs and implication of seepage intensity on transport efficiency

“…Microbial transport efficiency may be affected by the specific cell surface activity of microorganisms, which results, for example, from inter-species differences in outer membrane composition and extracellular components 58,59 but also depends on the growth conditions (e.g., nutrient levels) 60 . Also, the cell position in a biofilm can influence their anchorage on particles and their likelihood of detachment by external forces 61 . In addition, a recent study on the aerosolization of bacteria and viruses at the ocean-atmosphere interface suggested that cell morphology is a critical parameter that influences bubble-induced transport between these two environments 62 .…”

Chemical efficacy of several NaOCl concentrations on biofilms of different architecture: new insights on NaOCl working mechanisms

“…Arguably, the low reactivity of the 2% NaOCl with the poor EPS content of the upper layers of the four‐day CDFF biofilms causes initially superficial changes leading to the formation of small bubbles. As the bubbles leave the biofilm out they cause evaporation of the thin layer of ‘free’ water, while displacing and packing bacteria to adjacent regions (Jang et al ). This is supported by the significant decrease of the influence free water component (E 1 ) and increase of the influence of the bacterial component (E 4 ) noted for the 2% NaOCl‐treated CDFF biofilms.…”

Ultrasound‐mediated therapies for the treatment of biofilms in chronic wounds: a review of present knowledge

“…The fluorescent signal emitted increases proportionally with the quantity of PI that has intercalated with DNA; therefore, it can be utilized as a direct means of assessing how MBs facilitate uptake of macromolecules (Stiefel et al ., ). It could be therefore hypothesized that US‐activated MBs facilitate the uptake of antibiotics by biofilms, principally through increasing membrane permeability of bacterial cells and via heterogeneous alterations to the biofilm architecture, which can include the development of pores in the EPS matrix (Dong et al ., ; Jang et al ., ; Hu et al ., ). Fluid shear stress has also been shown to significantly affect biofilm morphology; at shear stresses under 1 Pa, biofilms are shown to grow in a laboratory‐typical mushroom‐like shape with interstitial channels and voids (Salta et al ., ).…”

“…The stress imparted on a cell membrane can equally cause membrane perforation. of antibiotics by biofilms, principally through increasing membrane permeability of bacterial cells and via heterogeneous alterations to the biofilm architecture, which can include the development of pores in the EPS matrix Jang et al, 2017;Hu et al, 2018).…”