Colonization and degradation of polyurethane coatings by Pseudomonas protegens biofilms is promoted by PueA and PueB hydrolases

The ability of bacteria and fungi to degrade polymeric substrates is often assessed in liquid-based assays conducted by exposing polymers to planktonic cells, or cell-free supernatants or enzymes generated from them. These assessments miss the opportunity to specifically examine the role of biofilm formation and physiology in degradative processes. In a previous study, we examined the ability of cell-free supernatants from wildtype and hydrolase-deficient Pseudomonas protegens Pf-5 strains to degrade a commercial polyester polyurethane (PU). In this study, we developed the methodology to conduct spatial-temporal analysis of both biofilm colonization and degradation of polyester PU coatings using the same strains. Wild-type Pf-5 grew as confluent biofilms that produced tendrils containing viable cells; in contrast, Pf-5ΔpueAB, a mutant lacking PueA and PueB hydrolases, colonized the PU only by growing as a confluent biofilm but lacked tendril expansion. Degradation of PU by the wild-type hydrolases may alter the substrata, facilitating colonization by the biofilm. Chemical analysis by in situ Fourier transform infrared (FTIR) and Raman spectroscopies revealed the polyester block of the PU was preferentially degraded to varying degrees by the wild-type Pf-5 and mutant biofilms, and showed degradation due to PueB that was undetectable in liquid-based assays. Raman spectroscopy analysis of biofilms of Pf-5ΔpueAB grown on PU revealed a detectable, but the lowest, level of PU degradation relative to other strains. Degradation of PU by this mutant had previously been undetectable and this study revealed the existence of biofilm-associated hydrolytic mechanisms other than those driven by PueA and PueB. Taken together, our data suggest that both PueA and PueB hydrolases play a role in the colonization and degradation of a model polyester PU by P. protegens biofilms. This study demonstrates the importance of using biofilm-based assays to identify mechanisms of microbiologically-influenced degradation. •PueA and PueB hydrolase activities mediate ester-bond cleavage during PU degradation by biofilm bacteria of P. protegens.•Air-surface biofilms permit non-invasive analysis of PU degradation.•Secreted PueA and PueB hydrolases facilitate motility of bacteria over solid PU surfaces.•Degradation of PU by Pseudomonas hydrolases creates tendril structures and allows bacterial expansion of colonization.•Additional hydrolases produced by biofilm bacteria also play a role in l