Physicochemical characterization, automated ribosomal intergenic spacer analysis (ARISA) community profiling, and 16S rRNA gene sequencing approaches were used to study bacterial communities inhabiting submerged Lake Huron sinkholes inundated with hypoxic, sulfate-rich groundwater. Photosynthetic cyanobacterial mats on the sediment surface were dominated by Phormidium autumnale, while deeper, organically rich sediments contained diverse and active bacterial communities.Groundwater intrusion is becoming recognized as an important source of nutrients, contaminants, and trace elements in aquatic ecosystems (7). Recent reports regarding marine habitats suggest that groundwater influences nitrogen inputs (29) and may have a significant impact on nutrient dynamics over seasonal (27) and longer (25) time scales. To date, investigations of groundwater effects have focused primarily on marine habitats. To better understand the impact of groundwater intrusion into freshwater habitats, we have been studying submerged sinkholes in the Laurentian Great Lakes.Sinkholes typically develop in areas of terrestrial karst when underground caverns collapse (36). We recently discovered submerged sinkholes that occur beneath the surface of Lake Huron in water up to a depth of 93 m (9, 41). These unique habitats are formed by groundwater dissolution of Paleozoic limestone and marine evaporite sediments in the Michigan Basin (16). Some sinkholes actively release cold, dense groundwater through underwater vents onto the lake floor. The venting groundwater has a lower pH (ϳ7.1), higher specific conductivity (ϳ2.3 mS ⅐ cm Ϫ1 , due to high levels of dissolved sulfate [Ͼ1,000 mg ⅐ liter Ϫ1 ], carbonate, and chloride ions), and lower concentrations of dissolved oxygen (Ͻ0.4 mg ⅐ liter Ϫ1 ) and nitrate than Lake Huron water (2,3,41,42). The intrusion of cold, hypoxic, sulfate-rich groundwater greatly alters the local lake habitat and has a significant impact on the sediment microbial community. Sediments at nearby control sites are sandy, but submerged sinkhole sediments (with carbon accounting for 5 to 35% of the sediment dry weight) are rich in organic matter originating from phytoplankton in the overlying water column