The production of most toxins and other exoproteins in Staphylococcus aureus is controlled globally by a complex polycistronic regulatory locus, agr. Secretory proteins are up‐regulated by agr whereas surface proteins are down‐regulated. agr contains two divergent promoters, one of which directs the synthesis of a 514 nucleotide (nt) transcript, RNAIII. In this report, we show that the cloned RNAIII determinant restores both positive and negative regulatory functions of agr to an agr‐null strain and that the RNA itself, rather than any protein, is the effector molecule. RNAIII acts primarily on the initiation of transcription and, secondarily in some cases, at the level of translation. In these cases, translation and transcription are regulated independently. RNAIII probably regulates translation directly by interacting with target gene transcripts and transcription indirectly by means of intermediary protein factors.

No abstract

Ethambutol (EMB) is a central component of drug regimens used worldwide for the treatment of tuberculosis. To gain insight into the molecular genetic basis of EMB resistance, approximately 2 Mb of five chromosomal regions with 12 genes in 75 epidemiologically unassociated EMB-resistant and 33 EMB-susceptibleMycobacterium tuberculosis strains isolated from human patients were sequenced. Seventy-six percent of EMBresistant organisms had an amino acid replacement or other molecular change not found in EMB-susceptible strains. Thirty-eight (51%) EMB-resistant isolates had a resistance-associated mutation in only 1 of the 12 genes sequenced. Nineteen EMB-resistant isolates had resistance-associated nucleotide changes that conferred amino acid replacements or upstream potential regulatory region mutations in two or more genes. Most isolates (68%) with resistance-associated mutations in a single gene had nucleotide changes in embB, a gene encoding an arabinosyltransferase involved in cell wall biosynthesis. The majority of these mutations resulted in amino acid replacements at position 306 or 406 of EmbB. Resistance-associated mutations were also identified in several genes recently shown to be upregulated in response to exposure of M. tuberculosis to EMB in vitro, including genes in the iniA operon. Approximately one-fourth of the organisms studied lacked mutations inferred to participate in EMB resistance, a result indicating that one or more genes that mediate resistance to this drug remain to be discovered. Taken together, the results indicate that there are multiple molecular pathways to the EMB resistance phenotype. Ethambutol [EMB; (S, S)-2,2Ј-(ethylenediimino)di-1-buta-nol] is used worldwide as one of the primary antituberculosis agents. The mechanism of action and the molecular genetic basis of resistance to EMB are not fully understood. Only the dextro isomer of EMB is biologically active, an observation consistent with the idea that the drug binds to a specific cellular target (7, 37). Several studies have implicated membrane-associated arabinosyltransferases as targets for EMB (1,5,20,22). These enzymes are well conserved in mycobacteria and are involved in the biosynthesis of arabinan, a component of arabinogalactan present in cell walls (6,8,17,33,34,39). Inhibition of arabinan synthesis leads to accumulation of mycolic acids and eventually to cell death.Three contiguous genes encoding arabinosyltransferases and designated embC, embA, and embB have been identified in Mycobacterium tuberculosis (35). The proteins encoded by these genes are about 65% identical to each other. Previous studies based on limited sequencing of the 10-kb region containing the embCAB genes have identified mutations that result in replacement of amino acid residues and are found only in EMB-resistant organisms cultured from humans. The most commonly affected amino acid was Met306 of EmbB. For example, Sreevatsan et al. (31) identified five distinct mutant codons that resulted in replacement of wild-type Met306 with Ile, Leu, or Va...

No abstract

The implementation of multiple molecular techniques in conjunction with surveillance data enabled us to identify a previously undetected outbreak in a defined geographical setting. The outbreak isolates comprise members of a distinct branch of the W family phylogenetic lineage. The use of molecular strain typing provides a proactive approach that may be used to initiate, and not just augment, traditional surveillance outbreak investigations.