*2.6. Biofilm Formation and Chitin Surface Attachment of P. rubra S4059 Was Not Affected by Deletion of GH19 Chitinase Gene*

Many *Pseudoalteromonas* species are good biofilm formers that are able to colonize crustaceans in marine environments [23], and since chitin colonization has been linked to biofilm formation in other bacteria [24], the biofilm formation of S4059 wild type and the mutant were compared in chitin containing media. The surface formed biofilm of the mutant and wild type was detected at the same level in all media (Figure S3).

The growth and attachment of the wild type and the mutant on natural chitin (shrimp shells) was assessed, and both strains grew from an initial density of 10<sup>4</sup> CFU/mL to a final cell density of 10<sup>7</sup> CFU/mL in the liquid surrounding the shells (Figure 3). Surface-attached bacteria were removed from shrimp shells by sonication resulting in an increase in cell density to 108 CFU/mL with similar levels reached by wild type and the mutant (Figure 3).

**Figure 3.** Growth and attachment of *Pseudoalteromonas rubra* S4059 wild type (red) and mutant Δ*GH19* (black) on Vannamei shrimp shells. Long dash lines: samples from liquid surrounding shrimp shells before sonication. Solid lines: samples from liquid surrounding shrimp shells after sonication. Short dash lines: samples from 3% Sigma sea salt without shrimp shells as a control. Data were analyzed on three biological replicates, and the error bars represent standard deviation.
