Relationship between bacterial flora and H 2 S levels in breath air was investigated in this study. Oral malodor levels in 10 subjects (age: 21–56 years) were assessed with gas chromatography, Breathtron® and organoleptic score. Based on these assessments, subjects were divided into 2 groups; oral malodorous and nonodorous groups. Tongue coating were sampled and spread onto Fastidious Anaerobe Agar plates containing 0.05% cysteine, 0.12% glutathione and 0.02% lead acetate, and were then incubated anaerobically at 37 °C for 2 weeks. Bacteria forming black or gray colonies were designated as H 2 S-producing phenotypes, and were identified by 16S rRNA gene sequencing. The numbers of total bacteria (p<0.005) and H 2 S-producing bacteria (p<0.05) in the oral malodorous group were significantly larger than those in the nonodorous group. Veillonella, Actinomyces and Prevotella species were predominant in the H 2 S-producing bacteria both in the oral malodorous and nonodorous groups. These results suggest that an increase in the number of H 2 S-producing bacteria in the tongue biofilm is responsible for oral malodor.