Differential Lipopolysaccharide Core Capping Leads to Quantitative and Correlated Modifications of Mechanical and Structural Properties in Pseudomonas aeruginosa Biofilms

Abstract

Bacterial biofilms are responsible for the majority of all microbial infections and have profound impact on industrial and geochemical processes. While many studies documented phenotypic differentiation and gene regulation of biofilms, the importance of their structural and mechanical properties is poorly understood. Here we investigate how changes in lipopolysaccharide (LPS) core capping in Pseudomonas aeruginosa affect biofilm structure through modification of adhesive, cohesive, and viscoelastic properties at an early stage of biofilm development. Microbead force spectroscopy and atomic force microscopy were used to characterize P. aeruginosa biofilm interactions with either glass substrata or bacterial lawns. Using isogenic migA, wapR, and rmlC mutants with defined LPS characteristics, we observed significant changes in cell mechanical properties among these strains compared to wild-type strain PAO1. Specifically, truncation of core oligosaccharides enhanced both adhesive and cohesive forces by up to 10-fold, whereas changes in instantaneous elasticity were correlated with the presence of O antigen. Using confocal laser scanning microscopy to quantify biofilm structural changes with respect to differences in LPS core capping, we observed that textural parameters varied with adhesion or the inverse of cohesion, while areal and volumetric parameters were linked to adhesion, cohesion, or the balance between them. In conclusion, this report demonstrated for the first time that changes in LPS expression resulted in quantifiable cellular mechanical changes that were correlated with structural changes in bacterial biofilms. Thus, the interplay between architectural and functional properties may be an important contributor to bacterial community survival.Biofilms are sessile microbial communities growing on a surface or at an interface, often enmeshed in polymeric substances. Being the predominant mode of microbial growth in nature, bacterial biofilms are particularly problematic in the context of human health, accounting for up to 80% of all bacterial infections. In industrial processes, bacterial biofilms cause corrosion and biofouling, resulting in considerable loss of productivity. In the natural environment, biofilms play a role in modulating worldwide geochemical cycles. Given the impact of biofilms in these diverse areas, the need for developing effective strategies to control them is of paramount importance. Since bacterial cell surface structures are convenient targets for control agents, their roles in influencing biofilm function and architecture warrant in-depth investigations. To date, most studies of biofilms have focused on genetic regulation, phenotypic differentiation and their contribution to antibiotic resistance. In contrast, the mechanical and structural properties that link the genotypes to phenotypes of bacterial biofilms are not well understood and rarely studied in a quantitative and correlated manner.Pseudomonas aeruginosa is a gram-negative opportunistic pathogen implicated in serious infe...

APorphyromonas gingivalisMutant Defective in a Putative Glycosyltransferase Exhibits Defective Biosynthesis of the Polysaccharide Portions of Lipopolysaccharide, Decreased Gingipain Activities, Strong Autoaggregation, and Increased Biofilm Formation

“…A B-band-deficient mutant forms luxurious biofilms compared to the wild-type strain (48). Very recently, P. aeruginosa mutants with LPS core variants have been reported to show enhanced cell adhesion and cohesion and altered viscoelasticity in early biofilms (29). In the P. gingivalis gtfB mutant, polysaccharide portions of A-LPS and O-LPS were deficient and bacterial cells were found to adhere tightly to one an- other by TEM analysis.…”

“…Pseudomonas aeruginosa is one of the major causative agents of mortality and morbidity in hospitalized patients due to a multiplicity of virulence factors associated with both chronic and acute infections. LPS of P. aeruginosa consists of lipid A, core oligosaccharide, and two distinct O side chains: the shorter A-band homopolymer is the so-called "common polysaccharide antigen" among the bacterium and consists of repeating D-rhamnose subunits, while the longer B-band heteropolymer is composed of repeating tri-to pentasaccharide subunits that vary among serotypes of P. aeruginosa (29). A B-band-deficient mutant forms luxurious biofilms compared to the wild-type strain (48).…”

Adaptive Divergence in Experimental Populations ofPseudomonas fluorescens. V. Insight into the Niche Specialist Fuzzy Spreader Compels Revision of the ModelPseudomonasRadiation

“…The role of LPS as a major component of WS biofilm strength and integrity, as a consequence of interactions between LPS and the cellulose matrix of the biofilm, was revealed in an earlier study (Spiers and Rainey 2005) and is well supported (Lau et al 2009;Nielsen et al 2011;Nilsson et al 2011). Alteration of the bacterial cell surface, with consequent changes to cell hydrophobicity, adhesive properties, and motility, likely perturbs the cell-cell interactions that maintain biofilm structure.…”

“…Alteration of the bacterial cell surface, with consequent changes to cell hydrophobicity, adhesive properties, and motility, likely perturbs the cell-cell interactions that maintain biofilm structure. The relationship between LPS defects and the adhesive, cohesive, and viscoelastic properties of biofilms and how these properties correlate with biofilm structure has been extensively characterized in P. aeruginosa PAO1 (Lau et al 2009). Essentially, more dramatic LPS defects (such as full truncation of O antigen) result in stronger adhesion of cells to glass and other abiotic surfaces and stronger cell-cell interactions (evident by flocculation).…”

Polymyxin B Resistance and Biofilm Formation in Vibrio cholerae Are Controlled by the Response Regulator CarR

“…Significance was determined by one-way ANOVA followed by Dunnett's multiple-comparison test, comparing the deletion mutants to the C6706 wild-type strain. NS, not significant; *, P Յ 0.05; ***, P Յ 0.001. brane of Gram-negative bacteria, have been shown to affect biofilm formation in P. aeruginosa and E. coli (40,41). The impact of modifications to LPS on V. cholerae biofilm formation and biofilm physiology has not been previously studied.…”