Specificity in the settlement -- modifying response of bacterial biofilms towards zoospores of the marine alga Enteromorpha

Previous studies have shown that the rate of settlement of zoospores of the green alga Enteromorpha is stimulated by mixed microbial biofilms and that the number of zoospores settling is positively correlated with the number of bacteria in the biofilm. In the present study the specificity of this relationship has been investigated. Ninety-nine strains of marine bacteria were isolated from natural biofilms on rocks and the surface of Enteromorpha plants. Isolates were screened by denaturing gradient gel electrophoresis (DGGE) to eliminate replicates and 16S rDNA sequencing identified a total of 37 unique strains. Phylogenetic analysis revealed that the isolated bacterial strains belonged to three groups gamma-Proteobacteria (28 strains), Cytophaga-Flavobacteria-Bacteroid (CFB) group (six strains) and alpha-Proteobacteria (one strain). Two strains were unassigned, showing < 93% sequence similarity with the CFB group. The main genera of gamma-Proteobacteria were Pseudoalteromonas (14 strains), Vibrio (five strains), Shewanella (five strains), Halomonas (three strains) and Pseudomonas (one strain). Spore settlement experiments were conducted on single-species biofilms, developed for different times on glass slides. The effect of correcting spore settlement values for biofilm density was evaluated. Results showed that the effect of bacterial strains on spore settlement was strain- but not taxon-specific and activity varied with the age of the biofilm. However, most of the strains belonging to genera Vibrio and Shewanella showed stimulation. Pseudoalteromonas strains showed a range of effects including settlement-inhibiting, paralysing and lysing activities. Spatial analysis of bacterial density in the presence and absence of spores revealed a range of different types of association between spores and bacteria. Overall, the spatial association between spores and bacteria appears to be independent of the overall quantitative influence of bacterial cells on spore settlement.