OprF impactsPseudomonas aeruginosabiofilm matrix eDNA levels in a nutrient-dependent manner

Abstract

The biofilm matrix is composed of exopolysaccharides, eDNA, membrane vesicles, and proteins. While proteomic analyses have identified numerous matrix proteins, their functions in the biofilm remain understudied compared to the other biofilm components. In thePseudomonas aeruginosabiofilm, several studies have identified OprF as an abundant matrix protein and, more specifically, as a component of biofilm membrane vesicles. OprF is a major outer membrane porin ofP. aeruginosacells. However, current data describing the effects of OprF in theP. aeruginosabiofilm is limited. Here we identify a nutrient-dependent effect of OprF in static biofilms, whereby ΔoprFcells form significantly less biofilm than wild type when grown in media containing glucose or low sodium chloride concentrations. Interestingly, this biofilm defect occurs during late static biofilm formation and is not dependent on the production of PQS, which is responsible for outer membrane vesicle production. Furthermore, while biofilms lacking OprF contain approximately 60% less total biomass than those of wild type, the number of cells in these two biofilms is equivalent. We demonstrate thatP. aeruginosaΔoprFbiofilms with reduced biofilm biomass contain less eDNA than wild-type biofilms. These results suggest that the nutrient-dependent effect of OprF is involved in the maintenance of matureP. aeruginosabiofilms by retaining eDNA in the matrix.IMPORTANCEMany pathogens form biofilms, which are bacterial communities encased in an extracellular matrix that protects them against antibacterial treatments. The roles of several matrix components of the opportunistic pathogenPseudomonas aeruginosahave been characterized. However, the effects ofP. aeruginosamatrix proteins remain understudied and are untapped potential targets for antibiofilm treatments. Here we describe a conditional effect of the abundant matrix protein OprF on late-stageP. aeruginosabiofilms. A ΔoprFstrain formed significantly less biofilm in low sodium chloride or with glucose. Interestingly, the defective ΔoprFbiofilms did not exhibit fewer resident cells but contained significantly less extracellular DNA (eDNA) than wild type. These results suggest that OprF is involved in matrix eDNA retention in mature biofilms.

OprF functions as a latch to direct Outer Membrane Vesicle release inPseudomonas aeruginosa

“…In a similar vein, OMVs have long been believed to interact with eDNA in the biofilm matrix (119)(120)(121). Though no direct interaction with OprF has been proposed, loss of the protein was shown to affect overall eDNA production under some conditions (122). Thus, the role of OprF in OMV-matrix interaction is an area ripe for deeper investigation, and a new understanding that OprF in OMVs adopts a different conformation than OprF in cells provides a new wrinkle to be further explored.…”