The antibacterial activity of thiamphenicol was compared to that of chloramphenicol against 313 strains of gram-negative bacilli isolated from various clinical specimens. These two antibiotics were equally active against the 106 isolates of Haemophilus (MIC = 0.1–1.56 μg/ml) and against 40 strains of Bacteroides fragilis (almost all strains being inhibited by 12.5 μg/ml of the two drugs). In contrast, when compared with chloramphenicol, 2–16 times as much of thiamphenicol was required to inhibit Enterobacteriaceae, making prediction of the susceptibility of these strains to thiamphenicol on the basis of chloramphenicol testing alone likely to be hazardous. Disc diffusion test using 30-μg discs and 12 mm as cut-off point was a reliable technique to determine susceptibility of bacteria either to chloramphenicol or thiamphenicol. When thiamphenicol discs of greater potency (50 μg) were employed, many strains exhibited wide zones of inhibition although most of them were resistant by the agar dilution method (MIC > 12.5 μg/mL). This practice is not advisable for testing organisms isolated outside of the urinary tract.