Application of Molecular Techniques to Define Mechanisms of Microbially Influenced Corrosion of Stainless Steel and Copper in Marine Systems.

Bacteria play a major role in the processes of biofouling and biocorrosion as members of biofilms and regulators of nutrient cycling in these systems. In these capacities, bacterial communities change temporally and spatially due to consortial interactions. Some bacterial species may facilitate or inhibit development of metabolic capacity of other bacterial species in the biofilm. To fully understand the processes of consortial interactions, therefore, the identity and activity of component species in a consortium must be resolved at the relevant temporal and spatial scales. The objectives of our research were to resolve the temporal and spatial patterns of N2-fixing and sulfate reducing bacteria within a bacterial consortium. Specifically: (1) Develop in situ hybridization techniques (in situ PCR) with fluorescent probes to detect the temporal abundance and spatial distribution of N2-fixing, and sulfate-reducing bacteria in biofilms; (2) Map the spatial distribution of the bacteria using congruently in situ hybridization techniques and confocal microscopy. Several approaches were directed toward Objective I. First, a biofilm was simulated by smearing fixed bacterial cells onto a glass microscope slide. The first experiment used immuno in situ techniques, whereby the anti- nitrogenase antibody was applied to a slide smeared with: (a) V natriegens cells that were harvested when fixing N2, (b) V natriegens cells that were not fixing N2, and (c) the sulfate-reducing bacterium (SRB), D. vulgaris. A strong fluorescent signal was present in the N2-fixing cells, but not in the non-N2-fixing cells or the SRB. The same assemblage of cells also was probed with EUB, a probe specific for a 18 bp conserved region of the SSU in all eubacteria. The 18 bp/rhodamine-coupled probe produced a strong signal in both V natriegens and D. vulgaris.