Biofilm development during ennoblement of stainless steel in Baltic Sea water: A microscopic study

This paper describes the development of biofilm on stainless steel during the increase of open circuit potential (ennoblement, Ecorr) that follows the exposure of AISI 316 stainless steel to natural seawater. The ennoblement occurred in the laboratory model ecosystem (mesocosm) reproducibly and similarly to that observed under field conditions in the Baltic Sea, provided that natural seawater was used as the feed, and the hydraulic flow was sufficiently high (10–30 mm s−1). Features distinguishing the ennobling biofilm from the non-ennobling, were studied using scanning (SEM) and transmission electron microscopy (TEM) as well as confocal laser scanning microscopy (CLSM). The increase of open circuit potential started in the laboratory (23°C±1) after 8 days and was complete (ΔE of 300–500 mV) after 21 days. In the field (10–14°C), ennoblement progressed from 21 days to 42 days (δE of 400–500 mV). When more than ten initially adhered cells per mm2 had grown into mushroom-like structures of > 50 μm in diameter and > 10 μm in height, covering 1–5% of the steel surface, the ennoblement started. The potential continued to increase while the biofilm mushrooms grew in height to 100 μm, and reached a plateau at 300–400 mV vs. SCE (standard calomel electrode) when the coverage of the steel surface reached 10–20%. Compared to the non-ennobling biofilms, those on the ennobled stainless steels attached more firmly to steel, were impermeable to lipophilic (acridine orange) and hydrophilic (lectin conjugates) dyes and grew as individual biofilm mushrooms of an average size of 0.5…2×106 μm3 containing 0.3–1×106 of tiny (< 1 μm) bacterial cells dispersed in interstitial matter. Biofilms on nonennobled stainless steels in the same seawater were easily removable, fluffy and permeable. The option of explaining the ennoblement (increase of cathodic reaction) of the stainless steel by microbial oxidation of organic residues inside the impermeable biofilm mushroom at the expense of Fe3+ reduction is discussed.