The accessory sigma factor B controls a general stress response that is thought to be important for Staphylococcus aureus survival and may contribute to virulence. The strain of choice for genetic studies, 8325-4, carries a small deletion in rsbU, which encodes a positive regulator of B activity. Consequently, to enable the role of B in virulence to be addressed, we constructed an rsbU ؉ derivative, SH1000, using a method that does not leave behind an antibiotic resistance marker. The phenotypic properties of SH1000 (8325-4 rsbU ؉ ) were characterized and compared to those of 8325-4, the rsbU mutant, parent strain. A recognition site for B was located in the promoter region of katA, the gene encoding the sole catalase of S. aureus, by primer extension analysis. However, catalase expression and activity were similar in SH1000 (8325-4 rsbU ؉ ), suggesting that this promoter may have a minor role in catalase expression under normal conditions. Restoration of B activity in SH1000 (8325-4 rsbU ؉ ) resulted in a marked decrease in the levels of the exoproteins SspA and Hla, and this is likely to be mediated by reduced expression of agr in this strain. By using Western blotting and a sarA-lacZ reporter assay, the levels of SarA were found to be similar in strains 8325-4 and SH1000 (8325-4 rsbU ؉ ) and sigB mutant derivatives of these strains. This finding contrasts with previous reports that suggested that SarA expression levels are altered when they are measured transcriptionally. Inactivation of sarA in each of these strains resulted in an expected decrease in agr expression; however, the relative level of agr in SH1000 (8325-4 rsbU ؉ ) remained less than the relative levels in 8325-4 and the sigB mutant derivatives. We suggest that SarA is not likely to be the effector in the overall B -mediated effect on agr expression.

N‐(3‐hydroxy‐7‐cis‐tetradecenoyl)‐l‐homoserine lactone (3OH,C14:1‐HSL) is a quorum‐sensing signalling molecule produced by Rhizobium leguminosarum. It is unusual in that it inhibits the growth of several strains of R. leguminosarum and was previously known as ‘small bacteriocin’. The cinRI locus responsible for the production of 3OH,C14:1‐HSL has been characterized; it is predicted to be on the chromosome, based on DNA hybridization. The cinR and cinI genes are in different transcriptional units, separated by a predicted transcription terminator. CinR regulates cinI expression to a very high level in a cell‐density dependent manner, and cinI expression is positively autoregulated by 3OH,C14:1‐HSL, the only identified N‐acyl homoserine lactone (AHL) produced by CinI. No other AHLs were identified that strongly induced cinI expression. Mutation of cinI or cinR abolishes the production of 3OH,C14:1‐HSL and also reduces the production of several other AHLs. This is thought to result from the expression of three other AHL production loci being affected by the absence of 3OH,C14:1‐HSL. AHLs produced by these other loci include N‐hexanoyl‐ and N‐octanoyl‐l‐homoserine lactones and, unexpectedly, N‐heptanoyl‐l‐homoserine lactone (C7‐HSL). The expression of the rhiI gene on the symbiotic plasmid is greatly reduced in a cinI mutant, and the major regulatory effect appears to be mediated at least in part as a result of an effect on expression of RhiR, the regulator of rhiI. Thus, cinR and cinI appear to be at the top of a regulatory cascade or network that influences several AHL‐regulated quorum‐sensing loci. The expression of cinI–lacZ fusions is significantly reduced (but not abolished) when the symbiosis plasmid pRL1JI is present, resulting in a reduction in the level of 3OH,C14:1‐HSL produced. Mutation of cinI had little effect on growth or nodulation. However, plasmid transfer was affected, and the results obtained indicate that 3OH,C14:1‐HSL produced by either the donor or the recipient in mating experiments can stimulate transfer of pRL1JI.

• We provide the first critical quantitative review of this rapidly developing area of research to serve as a basis for subsequent research and overviews.• We summarize data on population prevalence and functional effects of CYP2C19*17.• We argue on the basis of current evidence that potentially significant clinical effects are unlikely except for drugs with very narrow therapeutic windows. Of studied substrates, only clopidogrel may fall into this category. Even then, only homozygotes of the variant allele are likely to be at significantly increased risk.• The assignment of CYP2C19*17 homozygotes as EM, rather than UM, is adequate as the metabolic ratios of all probe drugs studied so far overlap completely the range of values seen in wild-type homozygotes.• The implications of CYP2C19*17 on the clinical effects of tamoxifen require further study. AIMSCytochrome P450 2C19 metabolizes many important drugs. In 2006, a variant allele (CYP2C19*17) associated with increased activity was discovered, but its likely clinical significance is controversial. Investigators disagree about the phenotype to be assigned to the two CYP2C19*17 genotypes. The aim of this study was to provide a critical summary, helpful to prescribers. METHODSWe searched MEDLINE for papers on the allele from 2006 and then undertook historical searches through the reference lists of papers retrieved. The relevant information was critically assessed and summarized. RESULTSCYP2C19*17 was associated with increased enzymic activity. Substrates studied were omeprazole, pantoprazole, escitalopram, sertraline, voriconazole, tamoxifen and clopidogrel. Most studies used pharmacokinetic variables as outcome measure. For clopidogrel, activated by CYP2C19, pharmacodynamic consequences focused on platelet aggregation. While for most pharmacokinetic parameters of the substrates studied the average value was altered, the range of values showed mostly complete overlap for CYP2C19*1/*17 heterozygotes and wild-type homozygotes. Even for CYP2C19*17 homozygotes, the absolute effect was modest compared with the effect of previously identified loss-of-function alleles. In Helicobacter pylori eradication CYP2C19*2 carriage was associated with an altered eradication rate (odds ratio 4.20, 95% confidence interval 1.23, 16.44) relative to the wild-type, but CYP2C19*17 homozygosity was not. Prevalence of the variant allele was typically <5% in Asians and about four times higher in White and African populations. CONCLUSIONSAssignment of CYP2C19*17 homozygotes as extensive metabolizers rather than ultrarapid metabolizers is adequate. CYP2C19*17 genotyping is unlikely to have clinical utility except for drugs with very narrow therapeutic indices.

The growth of some strains of Rhizobium leguminosarum bv. viciae is inhibited by N-(3-hydroxy-7-cis tetradecenoyl)-L-homoserine lactone (3OH-C 14:1 -HSL), which was previously known as the small bacteriocin before its characterization as an N-acyl homoserine lactone (AHL). Tn5-induced mutants of R. leguminosarum bv. viciae resistant to 3OH-C 14:1 -HSL were isolated, and mutations in two genes were identified. These genes, bisR and triR, which both encode LuxR-type regulators required for plasmid transfer, were found downstream of an operon containing trb genes involved in the transfer of the symbiotic plasmid pRL1JI. The first gene in this operon is traI, which encodes an AHL synthase, and the trbBCDEJKLFGHI genes were found between traI and bisR. Mutations in bisR, triR, traI, or trbL blocked plasmid transfer. Using gene fusions, it was demonstrated that bisR regulates triR in response to the presence of 3OH-C 14:1 -HSL. In turn, triR is then required for the induction of the traI-trb operon required for plasmid transfer. bisR also represses expression of cinI, which is chromosomally located and determines the level of production of 3OH-C 14:1 -HSL. The cloned bisR and triR genes conferred 3OH-C 14:1 -HSL sensitivity to strains of R. leguminosarum bv. viciae normally resistant to this AHL. Furthermore, bisR and triR made Agrobacterium tumefaciens sensitive to R. leguminosarum bv. viciae strains producing 3OH-C 14:1 -HSL. Analysis of patterns of growth inhibition using mutant strains and synthetic AHLs revealed that maximal growth inhibition required, in addition to 3OH-C 14:1 -HSL, the presence of other AHLs such as N-octanoyl-L-homoserine lactone and/or N-(3-oxo-octanoyl)-L-homoserine lactone. In an attempt to identify the causes of growth inhibition, a strain of R. leguminosarum bv. viciae carrying cloned bisR and triR was treated with an AHL extract containing 3OH-C 14:1 -HSL. N-terminal sequencing of induced proteins revealed one with significant similarity to the protein translation factor Ef-Ts.

The gram-positive human pathogen Staphylococcus aureus is often isolated with media containing potassium tellurite, to which it has a higher level of resistance than Escherichia coli. The S. aureus cysM gene was isolated in a screen for genes that would increase the level of tellurite resistance of E. coli DH5␣. The protein encoded by S. aureus cysM is sequentially and functionally homologous to the O-acetylserine (thiol)-lyase B family of cysteine synthase proteins. An S. aureus cysM knockout mutant grows poorly in cysteine-limiting conditions, and analysis of the thiol content in cell extracts showed that the cysM mutant produced significantly less cysteine than wild-type S. aureus SH1000. S. aureus SH1000 cannot use sulfate, sulfite, or sulfonates as the source of sulfur in cysteine biosynthesis, which is explained by the absence of genes required for the uptake and reduction of these compounds in the S. aureus genome. S. aureus SH1000, however, can utilize thiosulfate, sulfide, or glutathione as the sole source of sulfur. Mutation of cysM caused increased sensitivity of S. aureus to tellurite, hydrogen peroxide, acid, and diamide and also significantly reduced the ability of S. aureus to recover from starvation in amino acid-or phosphate-limiting conditions, indicating a role for cysteine in the S. aureus stress response and survival mechanisms.