Importance of the Exopolysaccharide Matrix in Antimicrobial Tolerance of Pseudomonas Aeruginosa Aggregates

is an opportunistic pathogen that can infect the lungs of cystic fibrosis (CF) patients and persist in the form of antibiotic-tolerant aggregates in the mucus. It has recently been suggested that such aggregates are formed due to restricted bacterial motility independent of the production of extracellular matrix components, and that they do not rely on an extracellular matrix for antimicrobial tolerance. However, we show here that biofilm matrix overexpression, as displayed by various clinical isolates, significantly protects aggregates against antimicrobial treatment. Alginate-overproducing mutant bacteria growing in aggregates showed highly increased antibiotic tolerance compared to wild-type bacteria in aggregates. Deletion of in the mutant strain abrogated alginate production and reversed the antibiotic tolerance displayed by the aggregates to a level similar to that observed for aggregates formed by the wild type. The Δ and Δ mutant strains both overproduce Pel and Psl exopolysaccharide, and when these bacteria grew in aggregates, they showed highly increased antibiotic tolerance compared to wild-type bacteria growing in aggregates. However, the Δ and Δ mutant strains, deficient in Pel/Psl production due to additional Δ Δ deletions, formed aggregates that displayed antibiotic tolerance levels close to those of wild-type aggregates. These results suggest that biofilm matrix components, such as alginate, Pel, and Psl, do play a role in the tolerance toward antimicrobials when bacteria grow as aggregates.