Figure 2

Assessing biofilm formation of heterologously expressed bslA variants in B. subtilis. (A) Amino acid alignment of BslA variants found in other Bacillus species. Abbreviations used are as follows: B_sub, Bacillus subtilis; B_pum, Bacillus pumilis; B_amy, Bacillus amyloliquefaciens; B_lic, Bacillus licheniformis. Underlined and bolded amino acids signify the signal sequence, black represents 100% sequence identity. Blue amino acids represent the hydrophobic cap regions, where dark blue are conserved amino acids and light blue are conservative substitutions. The purple highlighted amino acids are glycine which are conserved across all species and always follow the amino acids comprising the caps. Aspartic acid is shown in red and serine residues within caps 1 and 2 are shown in green. Note that YweA is differentiated from Bs_BslA and the BslA orthologues by the fact it lacks both the N-terminal region following the signalling sequence and the C-terminal domain. The * symbols indicate the cap regions containing serine residues. Biofilms and pellicles were formed from the wild type strain (NCIB3610), a strain possessing a B. subtilis bslA gene deletion mutant (bslA ⁻), strains that heterologously express the bslA gene of each species studied (B. subtilis, B_sub_bslA; B. amyloliquefaciens, B_sub_amy; B. licheniformis, B_lich_bslA; B. pumilus, B_amy_bslA) complemented into the bslA ⁻ mutant strain, and a strain expressing the yweA gene complemented into the bslA ⁻ mutant. Biofilm phenotypes were characterized by assessing (B) complex colony morphology; (C) pellicle formation; (D,E) colony surface hydrophobicity (see Table S3 for details on strains).