No abstract

Seawater-compensated fuel ballast systems maintain ship stability as fuel is spent but can introduce microorganisms that form biofilms, biodegrade fuel components, or enhance corrosion; all of which increase operating and maintenance costs. The aim of this study was to isolate planktonic bacteria from ballast tank fluids and taxonomically classify those that formed biofilm in culture. Twenty-two isolates were identified as belonging to seven genera based on 16S rRNA gene sequencing. Of the seven genera represented Alteromonas, Pseudoalteromonas, and Brevundimonas strains produced quantifiable biofilm in crystal violet assays. To test the hypothesis that the level of bulk nutrients would influence the extent of biofilm formation, isolates were grown in a conventional marine medium and marine medium supplemented with tryptone and yeast extract to represent standard and nutrient-replete media, respectively. While 80% of the Pseudoalteromonas strains produced 7 to 11-fold more biofilm in conventional medium vs. the supplemented medium, 64% of Alteromonas strains produced up to 50-fold less in the same medium. These results suggest that bulk nutrients influence the extent of biofilm formation in a taxa-specific fashion in these marine organisms. The sole Dasania marina isolate failed to display considerable biofilm growth in either media but was the only isolate to produce quorum sensing molecule(s), N-acyl homoserine lactone(s), in assays using an Agrobacterium tumefaciens reporter strain. Whether N-acyl homoserine lactones produced by D. marina could modulate biofilm formation in the other organisms isolated in this study would require further investigation.

We report the initial characterization of the αribazole (α-R) kinase enzyme of Geobacillus kaustophilus (GkCblS), which converts α-R to α-R-phosphate (α-RP) during the synthesis of cobamides. We implemented a continuous spectrophotometric assay to obtain kinetic parameters for several potential substrates and to study the specificity of the enzyme for α-N-linked ribosides. The apparent K m values for α-R and ATP were 358 and 297 μM, respectively. We also report methods for synthesizing and quantifying non-commercially available α-ribosides and β-ribazole (β-R). Purified GkCblS activated α-R and other α-ribosides, including α-adenosine (α-Ado). GkCblS did not phosphorylate β-N-linked glycosides like β-adenosine or β-R. Expression of G. kaustophilus cblS + in a Salmonella enterica subsp. enterica sv Typhimurium LT2 (S. enterica) strain lacking the nicotinate mononucleotide:5,6-dimethylbenzimidazole phosphoribosyl transferase (CobT) enzyme resulted in the activation of various benzimidazole α-ribosides, and the synthesis of benzimidazolyl cobamides to levels that supported robust growth. Notably, α-Ado did not support growth under similar conditions, in spite of the fact that GkCblS phosphorylated α-Ado in vitro. When α-Ado was provided at a very high concentration, growth was observed. This result suggested that in S. enterica α-Ado transport may be inefficient. We conclude that GkCblS has specificity for α-N-glycosidic bonds, but not for the base in α-ribosides.