-Acyl homoserine lactone (AHL) quorum sensing (QS) controls expression of over 200 genes in There are two AHL regulatory systems: the LasR-LasI circuit and the RhlR-RhlI system. We mapped transcription termination sites affected by AHL QS in , and in doing so we identified AHL-regulated small RNAs (sRNAs). Of interest, we noted that one particular sRNA was located within the locus. We found that , which encodes the enzyme that produces the AHL -butanoyl-homoserine lactone (C4-HSL), is controlled by a 5' untranslated region (UTR)-derived sRNA we name RhlS. We also identified an antisense RNA encoded opposite the beginning of the open reading frame, which we name asRhlS. RhlS accumulates as wild-type cells enter stationary phase and is required for the production of normal levels of C4-HSL through activation of translation. RhlS also directly posttranscriptionally regulates at least one other unlinked gene, The asRhlS appears to be expressed at maximal levels during logarithmic growth, and we suggest RhlS may act antagonistically to the asRhlS to regulate translation. The -encoded sRNAs represent a novel aspect of RNA-mediated tuning of QS. The opportunistic human pathogen possesses multiple quorum sensing systems that regulate and coordinate production of virulence factors and adaptation to different environments. Despite extensive research, the regulatory elements that play a role in this complex network are still not fully understood. By using several RNA sequencing techniques, we were able to identify a small regulatory RNA we named RhlS. RhlS increases translation of RhlI, a key enzyme in the quorum sensing pathway, and represses the mRNA encoding one of the siderophore pyoverdine receptors. Our results highlight a new regulatory layer of quorum sensing and contribute to the growing understanding of the role regulatory RNAs play in bacterial physiology.