Atomic force microscopy analysis of Pel polysaccharide- and type IV pili-mediated adhesion of PA14 to an abiotic surface.

Type IV pili (TFP) contribute to the ability of microbes such as to engage with and move across surfaces. We reported previously that TFP generate retractive forces of ∼30 pN and provided indirect evidence that TFP-mediated surface attachment was enhanced in the presence of the Pel polysaccharide. Here, we use different mutants defective in flagellar, Pel production or TFP production - alone or in combination - to decipher the relative contribution of these biofilm-promoting factors for adhesion. By means of atomic force microscopy (AFM), we show that mutating the flagellum (Δ mutant) results in an increase in Pel polysaccharide production, but this increase in Pel does not result in an increase in surface adhesive properties compared to those previously described for the WT strain. By blocking Pel production in the Δ mutant (ΔΔ), we directly show that TFP play a major role in the adhesion of the bacteria to hydrophobic AFM tips, but that the adhesion force is only slightly impaired by the absence of Pel. Inversely, performing single-cell force spectroscopy measurements with the mutant lacking TFP (ΔΔ) reveals that the Pel can modulate the attachment of the bacteria to a hydrophobic substrate in a time-dependent manner. Finally, little adhesion was detected for the ΔΔΔ triple mutant, suggesting that both TFP and Pel polysaccharide make a substantial contribution to bacteria-substratum interaction events. Altogether, our data allow us to decipher the relative contribution of Pel and TFP in the early attachment by .