Sulfate-reducing bacteria-dominated biofilms that precipitate ZnS in a subsurface circumneutral-pH mine drainage system

The microbial diversity of ZnS-forming biofilms in 8degreesC, circumneutral-pH groundwater in tunnels within the abandoned Piquette Zn, Pb mine (Tennyson, Wisconsin, USA) has been investigated by molecular methods, fluorescence in situ hybridization (FISH), and cultivation techniques. These biofilms are growing on old mine timbers that generate locally anaerobic zones within the mine drainage system. Sulfate-reducing bacteria (SRB) exclusively of the family Desulfobacteriaceae comprise a significant fraction of the active microbiota. Desulfosporosinus strains were isolated, but could not be detected by molecular methods. Other important microbial clusters belonged to the beta-, gamma-, and epsilon-Proteobacteria, the Cytophaga/Flexibacter/Bacteroides-group (CFB), Plancto-mycetales, Spirochaetales, Clostridia, and green nonsulfur bacteria. Our investigations indicated a growth dependence of SRB on fermentative, cellulolytic, and organic acid-producing Clostridia. A few clones related to sulfuroxidizing bacteria were detected, suggesting a sulfur cycle related to redox gradients within the biofilm. Sulfur oxidation prevents sulfide accumulation that would lead to precipitation of other sulfide phases. FISH analyses indicated that Desulfobacteriaceae populations were not early colonizers in freshly grown and ZnS-poor biofilms, whereas them were abundant in older, naturally established, and ZnS-rich biofilms. Gram-negative SRB have been detected in situ over a period of 6 months, supporting the important role of these organisms in selective ZnS precipitation in Tennyson mine. Results demonstrate the complex nature of biofilms responsible for In situ bioremediation of toxic metals in a subsurface mine drainage system.