High throughput chemical file screening with an enzymatic assay to detect inhibitors of the ErmC methyltransferase enzyme from macrolide-lincosamide-streptogramin B (MLS_B) resistant pathogenic bacteria identified low molecular weight compounds that had IC_50s (50% inhibitory concentration) in the nMolar to μMlar range. These same inhibitors were assessed in vitro for their capacity to inhibit the liver enzyme, cathechol-O-methyltransferase and the prokaryotic enzyme, EcoRI methylase. Selective inhibitors of the ErmC methyltransferase were tested in tertiary assays to determine their minimal inhibitory concentrations (MICs), as single agents and in combination with the macrolide, azithromycin, against strains of pathogenic bacteria expressing MLS_B-resistance. Compounds that were active in vitro, alone or in combination with azithromycin, against strains of macrolide-resistant pathogens were tested in a mouse model of infection using an MLS_B-resistant strain of Staphylococcus aureus or a macrolide-susceptible strain of Streptococcus pyogenes.