Listeria monocytogenes is well known for its durable physiological characteristics, which allow the organism to grow at low temperature and pH and high osmolarity. Growth under high osmolarity depends on the accumulation of compatible solutes, among which glycine betaine and carnitine are the preferred solutes for this organism. Three different transport systems, Gbu, BetL, and OpuC, have been identified in L. monocytogenes which serve to scavenge the preferred compatible solutes. The general stress response regulator sigma(B) has been shown to play an important role in osmotic adaptation in L. monocytogenes, presumably by directing transcription from one or more of the solute transport genes. In the studies presented here, we have used primer extension analyses to identify the promoter elements responsible for transcription of the opuC, gbuA, and betL genes. All three genes are osmotically inducible to some degree. betL is transcribed from a sigma(B)-independent promoter, while gbuA is transcribed from dual promoters, one of which is sigma(B) dependent. opuC is transcribed exclusively from a sigma(B)-dependent promoter. The betL promoter is similar in sequence to the sigma(B)-independent gbuAP1 promoter. Kinetic analysis of transcript accumulation after osmotic upshift demonstrated that sigma(B)-dependent transcripts from gbuAP2 and sigB accumulate for an extended period after upshift, suggesting that sigma(B) activity may provide a mechanism for sustained high-level expression during osmotic challenge. In contrast to osmotic upshift, expression from the sigma(B)-dependent opuC and gbuAP2 promoters after temperature upshift and ethanol stress was minimal, suggesting that additional mechanisms may also participate in regulating transcription from these sigma(B)-dependent promoters.