Quorum sensing by means of N-acyl-L-homoserine lactones (AHLs) is widespread in Gram-negative bacteria, where diverse AHLs influence a wide variety of functions, even in a single genus such as Serratia. Here we report the identification and characterization of the quorum sensing system of Serratia plymuthica strain RVH1. This strain isolated from a raw vegetable processing line produces at least three AHLs which were identified as N-butanoyl- (C4-HSL), N-hexanoyl- (C6-HSL) and N-(3-oxo-hexanoyl)-homoserine lactone (3-oxo-C6-HSL). The identified LuxI homolog SplI synthesizes 3-oxo-C6-HSL, and influences the production of C4-HSL and C6-HSL, as splI gene inactivation resulted in loss of 3- oxo-C6-HSL production and smaller amounts of C4-HSL and C6-HSL produced. SplI-dependent quorum sensing controls 2,3-butanediol fermentation (previously reported) and the production of an extracellular chitinase, nuclease, protease and antibacterial compound. The identity of the latter is not yet elucidated, but appears to be different from the known antibacterial compounds produced by Serratia strains. SplR, the homolog of the LuxR regulator, appears to act as a repressor of synthesis of extracellular enzymes and antibacterial compound and to autorepress its own expression, probably by binding to a 21 bp lux box sequence.