Antimicrobial targets localize to the extracellular vesicle-associated proteome of Pseudomonas aeruginosa grown in a biofilm

Abstract

Microbial biofilms are particularly resistant to antimicrobial therapies. These surface-attached communities are protected against host defenses and pharmacotherapy by a self-produced matrix that surrounds and fortifies them. Recent proteomic evidence also suggests that some bacteria, including the opportunistic pathogen Pseudomonas aeruginosa, undergo modifications within a biofilm that make them uniquely resistant compared to their planktonic (free-living) counterparts. This study examines 50 proteins in the resistance subproteome of both surface-associated and free-living P. aeruginosa PAO1 over three time points. Proteins were grouped into categories based on their roles in antimicrobial: (i) binding, (ii) efflux, (iii) resistance, and (iv) susceptibility. In addition, the extracellular outer membrane vesicle-associated proteome is examined and compared between the two growth modes. We show that in whole cells between 12–24% of the proteins are present at significantly different abundance levels over time, with some proteins being unique to a specific growth mode; however, the total abundance levels in the four categories remain consistent. In contrast, marked differences are seen in the protein content of the outer membrane vesicles, which contain a greater number of drug-binding proteins in vesicles purified from late-stage biofilms. These results show how the method of analysis can impact the interpretation of proteomic data (i.e., individual proteins vs. systems), and highlight the advantage of using protein-based methods to identify potential antimicrobial resistance mechanisms in extracellular sample components. Furthermore, this information has the potential to inform the development of specific antipseudomonal therapies that quench possible drug-sequestering vesicle proteins. This strategy could serve as a novel approach for combating the high-level of antimicrobial resistance in P. aeruginosa biofilms.

Vesicles-mediated resistance to antibiotics in bacteria

“…A recent proteomic analysis of OMVs obtained from planktonic growth and biofilm in Pseudomonas aeruginosa revealed that drug-binding proteins (notably efflux proteins) were more concentrated in the biofilm OMVs. It is another example of possible involvement of the vesicles in antibiotic resistance (Park et al, 2014 ).…”

Bacteriological Study of Pseudomonas Aeruginosa Isolated from Different Infections and Study Antimicrobial Activities of Plant Extract Solanum Nigrum Against It

“…Pseudomonas represents the phenomenon of resistance to antibiotics, as it is difficult to address the injuries caused by the drug resistance in this type of bacteria, as it possesses natural resistance, including changing the permeability of the outer wall, and the possession of several groups of stream systems, Pseudomonas aeruginosa also produces many enzymes responsible for the decomposition of many antimicrobials, especially Metallo ß-Lactamases (MßLs), Extended Spectrum Beta-Lactamases (ESßLs), and Ampler Molecular Class (AMPC) [4,5].…”

The Contribution of Membrane Vesicles to Bacterial Pathogenicity in Cystic Fibrosis Infections and Healthcare Associated Pneumonia

“…The MV-related proteins identified were OM enzymes and proteins involved in the transport of small molecules, the uptake of iron and antibiotic resistance (Couto et al, 2015). In addition, vesicles purified from late-stage P. aeruginosa biofilms are enriched for drug-binding proteins, which makes the bacterial species inside these biofilms even better protected against antibiotics (Park et al, 2014). It has been suggested further that MVs secreted by P. aeruginosa are under the control of the quorum sensing system and supply the forming biofilm with extracellular DNA (eDNA) and LPS (Nakamura et al, 2008).…”