In recent years, society interest in health promoting food has considerably increased the demand for functional foods. Gamma-amino-butyric acid (GABA), a non-protein amino acid, is an inhibitory neurotransmitter in the brain with many physiological functions such as antihypertensive, and tranquilizer effects. Consequently, it has a strong potential as bioactive ingredient and its production by microbial fermentation has emerged as a way to increase GABA content in food. Some microorganisms are indeed able to biosynthesize GABA by decarboxylation of glutamate through the GAD acid resistance system, composed of a glutamate decarboxylase (encoded by gadB gene) and a GABA/glutamate antiport (encoded by gadC gene). Streptococcus thermophilus (ST) is a lactic acid bacterium with a long history of use in traditional and industrial manufacture of fermented dairy products. GABA production by some ST strains was reported, though at different levels [1,2]. Previous work on ST genomics revealed a high polymorphism of the gadB upstream sequence, suggesting a role of the gadB promoter in GABA production diversity between strains [3]. The objective of this work was to screen ST strains for GABA production and to investigate the role of the gadB promoter region in the ability of ST to produce GABA. First, 26 ST strains were screened for their GAD activity using fast colorimetric assay in resting cell conditions. Screening results revealed a strong strain-dependency: five strains were classified as strong GABA producers, four as moderate producers and 17 did not produce GABA or at low level. The screening results were confirmed by GABA quantification (HPLC). To better understand the diversity observed between strains, gadB promoter region gene was sequenced for all strains. Strains were clustered into four groups according to the polymorphism of the gadB promoter. Interestingly, strong GABA producer strains seemed to share the same promoter sequence, hinting that sequence variation could contribute to differential expression level of gadB. To further investigate this hypothesis, gadB promoter was characterized and its activity in strains from different clusters was monitored by transcriptional fusion. This work provides better insights of the GABA producing ability in ST and could contribute to better select high GABA producers.