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Mucins trigger dispersal of Pseudomonas aeruginosa biofilms.

ABSTRACT: Mucus is a biological gel that lines all wet epithelia in the body, including the mouth, lungs, and digestive tract, and has evolved to protect the body from pathogenic infection. However, microbial pathogenesis is often studied in mucus-free environments that lack the geometric constraints and microbial interactions in physiological three-dimensional mucus gels. We developed fluid-flow and static test systems based on purified mucin polymers, the major gel-forming constituents of the mucus barrier, to understand how the mucus barrier influences bacterial virulence, particularly the integrity of Pseudomonas aeruginosa biofilms, which can become resistant to immune clearance and antimicrobial agents. We found that mucins separate the cells in P. aeruginosa biofilms and disperse them into suspension. Other viscous polymer solutions did not match the biofilm disruption caused by mucins, suggesting that mucin-specific properties mediate the phenomenon. Cellular dispersion depended on functional flagella, indicating a role for swimming motility. Taken together, our observations support a model in which host mucins are key players in the regulation of microbial virulence. These mucins should be considered in studies of mucosal pathogenesis and during the development of novel strategies to treat biofilms.

SUBMITTER: Co JY

PROVIDER: S-EPMC6180003 | biostudies-literature | 2018

REPOSITORIES: biostudies-literature

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Mucins trigger dispersal of <i>Pseudomonas aeruginosa</i> biofilms.

Co Julia Y JY Cárcamo-Oyarce Gerardo G Billings Nicole N Wheeler Kelsey M KM Grindy Scott C SC Holten-Andersen Niels N Ribbeck Katharina K

NPJ biofilms and microbiomes 20181010

Mucus is a biological gel that lines all wet epithelia in the body, including the mouth, lungs, and digestive tract, and has evolved to protect the body from pathogenic infection. However, microbial pathogenesis is often studied in mucus-free environments that lack the geometric constraints and microbial interactions in physiological three-dimensional mucus gels. We developed fluid-flow and static test systems based on purified mucin polymers, the major gel-forming constituents of the mucus barr ...[more]

PMID: 30323945

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Project description:BackgroundBiofilms disperse in response to specific environmental cues, such as reduced oxygen concentration, changes in nutrient concentration and exposure to nitric oxide. Interestingly, biofilms do not completely disperse under these conditions, which is generally attributed to physiological heterogeneity of the biofilm. However, our results suggest that genetic heterogeneity also plays an important role in the non-dispersing population of P. aeruginosa in biofilms after nutrient starvation.ResultsIn this study, 12.2% of the biofilm failed to disperse after 4 d of continuous starvation-induced dispersal. Cells were recovered from the dispersal phase as well as the remaining biofilm. For 96 h starved biofilms, rugose small colony variants (RSCV) were found to be present in the biofilm, but were not observed in the dispersal effluent. In contrast, wild type and small colony variants (SCV) were found in high numbers in the dispersal phase. Genome sequencing of these variants showed that most had single nucleotide mutations in genes associated with biofilm formation, e.g. in wspF, pilT, fha1 and aguR. Complementation of those mutations restored starvation-induced dispersal from the biofilms. Because c-di-GMP is linked to biofilm formation and dispersal, we introduced a c-di-GMP reporter into the wild-type P. aeruginosa and monitored green fluorescent protein (GFP) expression before and after starvation-induced dispersal. Post dispersal, the microcolonies were smaller and significantly brighter in GFP intensity, suggesting the relative concentration of c-di-GMP per cell within the microcolonies was also increased. Furthermore, only the RSCV showed increased c-di-GMP, while wild type and SCV were no different from the parental strain.ConclusionsThis suggests that while starvation can induce dispersal from the biofilm, it also results in strong selection for mutants that overproduce c-di-GMP and that fail to disperse in response to the dispersal cue, starvation.

Project description:BackgroundMicrobial biofilms, as a hallmark of cystic fibrosis (CF) lung disease and other chronic infections, remain a desirable target for antimicrobial therapy. These biopolymer-based viscoelastic structures protect pathogenic organisms from immune responses and antibiotics. Consequently, treatments directed at disrupting biofilms represent a promising strategy for combating biofilm-associated infections. In CF patients, the viscoelasticity of biofilms is determined mainly by their polymicrobial nature and species-specific traits, such as Pseudomonas aeruginosa filamentous (Pf) bacteriophages. Therefore, we examined the impact of microbicidal ceragenins (CSAs) supported by mucolytic agents-DNase I and poly-aspartic acid (pASP), on the viability and viscoelasticity of mono- and bispecies biofilms formed by Pf-positive and Pf-negative P. aeruginosa strains co-cultured with Staphylococcus aureus or Candida albicans.MethodsThe in vitro antimicrobial activity of ceragenins against P. aeruginosa in mono- and dual-species cultures was assessed by determining minimum inhibitory concentration (MIC) and minimum bactericidal/fungicidal concentration (MBC/MFC). Inhibition of P. aeruginosa mono- and dual-species biofilms formation by ceragenins alone and in combination with DNase I or poly-aspartic acid (pASP) was estimated by the crystal violet assay. Additionally, the viability of the biofilms was measured by colony-forming unit (CFU) counting. Finally, the biofilms' viscoelastic properties characterized by shear storage (G') and loss moduli (G"), were analyzed with a rotational rheometer.ResultsOur results demonstrated that ceragenin CSA-13 inhibits biofilm formation and increases its fluidity regardless of the Pf-profile and species composition; however, the Pf-positive biofilms are characterized by elevated viscosity and elasticity parameters.ConclusionDue to its microbicidal and viscoelasticity-modifying properties, CSA-13 displays therapeutic potential in biofilm-associated infections, especially when combined with mucolytic agents.

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Mucins trigger dispersal of Pseudomonas aeruginosa biofilms.

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Mucins trigger dispersal of <i>Pseudomonas aeruginosa</i> biofilms.

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