SummarySerratia marcescens SS-1 produces at least four N -acylhomoserine lactones (AHLs) which were identified using high-resolution mass spectrometry and chemical synthesis, as N-(3-oxohexanoyl) homoserine lactone (3-oxo-C6-HSL), N -hexanoyl-(C6-HSL), N -heptanoyl (C7-HSL) and N -octanoyl-(C8-HSL) homoserine lactone. These AHLs are synthesized via the LuxI homologue SpnI, and regulate via the LuxR homologue SpnR, the production of the red pigment, prodigiosin, the nuclease, NucA, and a biosurfactant which facilitates surface translocation. spnR overexpression and spnR gene deletion show that SpnR, in contrast to most LuxR homologues, acts as a negative regulator. spnI overexpression, the provision of exogenous AHLs and spnI gene deletion suggest that SpnR is de-repressed by 3-oxo-C6-HSL. In addition, long chain AHLs antagonize the biosurfactantmediated surface translocation of S. marcescens SS-1. Upstream of spnI there is a gene which we have termed spnT . spnI and spnT form an operon and although database searches failed to reveal any spnT homologues, overexpression of this novel gene negatively affected both sliding motility and prodigiosin production.