Objective: This study was undertaken to establish whether methylation of inorganic Hg occurs in dental-unit wastewater, and if so, to determine which bacteria might be responsible. Methods: Grab samples were obtained from holding tanks at two dental treatment facilities on two separate occasions six months apart and from a stand-alone dental-unit on four consecutive days. Samples were prepared for Me-Hg analysis by acidic K/Br extraction and analyzed using aqueous phase ethylation, purge/trap, isothermal GC separation, and Cold Vapor Atomic Fluorescence detection. Original bacterial communities, as well as sulfate reducing enrichment cultures from 3 samples were characterized by profiling a conserved region of the 16S rRNA gene using denaturing gradient gel electrophoresis (DGGE). Results: Me-Hg from the first samples at clinic #1 (100-chairs) averaged 99.40 mg/liter (n=2) with total Hg averaging 32,700 mg/liter. Six months later mean Me-Hg concentrations from this clinic averaged 30.46 mg/liter (n=4, SD=1.55). Samples taken from clinic #2 (35-chairs) showed an average methyl-Hg concentration of 0.32 mg/liter (n=3, SD=0.138) with total Hg averaging 2567 mg/liter (n=3, SD=205). Six months later methyl-Hg levels averaged 3.69 mg/liter (n=4, SD=0.13). Chairside samples taken from a dental-unit at the end of 4-consecutive days showed non-detectable levels (n=4, MDL=6.54 mg/liter) of Me-Hg with total Hg ranging from to 3,760 to 167,000 mg/liter. DGGE identified 7 genera of bacteria, some known to reside in the oral cavity. Conclusions: Results show methylation doesn’t occur directly at the dental-unit, but can take place in holding tanks downstream by bacteria present in the oral flora. The considerable variation in Me-Hg concentrations between and within-clinics on the sampling occasions may be due to the age of the particular sample within the holding tank (which was not controlled in this study). This study was supported by a grant from the USEPA (IAG# DW17947929-01-0).