Firstly, the biofilm formation of A

Firstly, the biofilm formation of AbIC IΔblp1 and AbIC IIΔblp1 mutants and their blp1-complemented strains was assessed. Bacteria were grown on the plastic surface and the formed structures were analyzed by confocal laser scanning microscopy (CLSM) using SYTO9 and propidium iodide (PI) staining as described in Materials and Methods. Interestingly, 2 h post-seeding, the initial attachment of bacteria to the plastic surface, based on the increase of PI-stained (red stain) versus total amount of bacteria (green stain) was affected in AbIC IΔblp1 mutant, demonstrating up to 55% increase in PI-stained bacteria compared with the parental strain (Fig. 2a, upper panels). Contrary to the AbIC IΔblp1 mutant, the AbIC IIΔblp1 strain showed no alterations in the initial attachment compared to the parent, evident through the lack of PI-stained bacteria and through the formation of more uniform biofilm structure similar to parental strain (Fig. 2a, lower panels). To verify that the observed phenotype of AbIC IΔblp1 was indeed blp1-dependent, we introduced plasmids pblp1IC I and pblp1IC II carrying respective blp1 alleles into AbIC IΔblp1 bacteria and assessed their attachment to the plastic surface. Notably,both blp1 alleles efficiently restored the phenotype of AbIC IΔblp1 mutant manifested by the reduction in PIstained cells compared to the AbIC IΔblp1 mutant carrying control plasmid (Fig. 2b and Additional file 3: Figure S2).