Reversible antibiotic tolerance induced in <i>Staphylococcus aureus</i> by concurrent drug exposure

ABSTRACT   Resistance of Staphylococcus aureus to beta-lactam antibiotics has led to increasing use of the glycopeptide antibiotic vancomycin as a life-saving treatment for major S. aureus infections. Coinfection by an unrelated bacterial species may necessitate concurrent treatment with a second antibiotic that targets the coinfecting pathogen. While investigating factors that affect bacterial antibiotic sensitivity, we discovered that susceptibility of S. aureus to vancomycin is reduced by concurrent exposure to colistin, a cationic peptide antimicrobial employed to treat infections by Gram-negative pathogens. We show that colistin-induced vancomycin tolerance persists only as long as the inducer is present and is accompanied by gene expression changes similar to those resulting from mutations that produce stably inherited reduction of vancomycin sensitivity (vancomycin-intermediate S. aureus [VISA] strains). As colistin-induced vancomycin tolerance is reversible, it may not be detected by routine sensitivity testing and may be responsible for treatment failure at vancomycin doses expected to be clinically effective based on such routine testing. IMPORTANCE   Commonly, antibiotic resistance is associated with permanent genetic changes, such as point mutations or acquisition of resistance genes. We show that phenotypic resistance can arise where changes in gene expression result in tolerance to an antibiotic without any accompanying genetic changes. Specifically, methicillin-resistant Staphylococcus aureus (MRSA) behaves like vancomycin-intermediate S. aureus (VISA) upon exposure to colistin, which is currently used against infections by Gram-negative bacteria. Vancomycin is a last-resort drug for treatment of serious S. aureus infections, and VISA is associated with poor clinical prognosis. Phenotypic and reversible resistance will not be revealed by standard susceptibility testing and may underlie treatment failure

Human antimicrobial peptide, LL-37, induces non-inheritable reduced susceptibility to vancomycin in <i>Staphylococcus aureus</i>

Abstract Antimicrobial peptides (AMPs) are central components of the innate immune system providing protection against pathogens. Yet, serum and tissue concentrations vary between individuals and with disease conditions. We demonstrate that the human AMP LL-37 lowers the susceptibility to vancomycin in the community-associated methicillin-resistant S. aureus (CA-MRSA) strain FPR3757 (USA300). Vancomycin is used to treat serious MRSA infections, but treatment failures occur despite MRSA strains being tested susceptible according to standard susceptibility methods. Exposure to physiologically relevant concentrations of LL-37 increased the minimum inhibitory concentration (MIC) of S. aureus towards vancomycin by 75%, and resulted in shortened lag-phase and increased colony formation at sub-inhibitory concentrations of vancomycin. Computer simulations using a mathematical antibiotic treatment model indicated that a small increase in MIC might decrease the efficacy of vancomycin in clearing a S. aureus infection. This prediction was supported in a Galleria mellonella infection model, where exposure of S. aureus to LL-37 abolished the antimicrobial effect of vancomycin. Thus, physiological relevant concentrations of LL-37 reduce susceptibility to vancomycin, indicating that tissue and host specific variations in LL-37 concentrations may influence vancomycin susceptibility in vivo

Norlichexanthone Reduces Virulence Gene Expression and Biofilm Formation in Staphylococcus aureus

Staphylococcus aureus is a serious human pathogen and antibiotic resistant, community-associated strains, such as the methicillin resistant S. aureus (MRSA) strain USA300, continue to spread. To avoid resistance, anti-virulence therapy has been proposed where toxicity is targeted rather than viability. Previously we have shown that norlichexanthone, a small non-reduced tricyclic polyketide produced by fungi and lichens, reduces expression of hla encoding α-hemolysin as well as the regulatory RNAIII of the agr quorum sensing system in S. aureus 8325-4. The aim of the present study was to further characterise the mode of action of norlichexanthone and its effect on biofilm formation. We find that norlichexanthone reduces expression of both hla and RNAIII also in strain USA300. Structurally, norlichexanthone resembles ω-hydroxyemodin that recently was shown to bind the agr two component response regulator, AgrA, which controls expression of RNAIII and the phenol soluble modulins responsible for human neutrophil killing. We show that norlichexanthone reduces S. aureus toxicity towards human neutrophils and interferes directly with AgrA binding to its DNA target. In contrast to ω-hydroxyemodin however, norlichexanthone reduces staphylococcal biofilm formation. Transcriptomic analysis revealed that genes regulated by the SaeRS two-component system are repressed by norlichexanthone when compared to untreated cells, an effect that was mitigated in strain Newman carrying a partially constitutive SaeRS system. Our data show that norlichexanthone treatment reduces expression of key virulence factors in CA-MRSA strain USA300 via AgrA binding and represses biofilm formation

Deliberation after consensus

This editorial introduction presents an overview of the themes explored in the symposium on Deliberation after Consensus. For all the talk of its obsolescence and irrelevance, the concept of consensus still remains centrally contested through generations of deliberative democracy scholarship. In face of criticism for being neither empirically feasible nor normatively desirable, some deliberative theorists have moved away from consensus-oriented teleology and argued in favor of other legitimate outcomes of deliberations. Other theorists have resisted this move, claiming that the aim of deliberation implies that consensus should remain as a regulative ideal for deliberative outcomes. Engaging with these debates about the role of consensus in theories of deliberative democracy, this symposium brings together a selection of innovative, original research articles that raise novel questions about the role consensus could and should play in democratic deliberation and in a deliberative democracy. This introduction offers an overview of the debate over consensus drawing on the notion of successive generations of deliberative democracy research. Our aim is to demonstrate that the view of consensus has changed during generations of deliberative scholarships, but also that some scholars still defend the normative importance of the meaning of consensus once developed by the first generation. Consequently, there are tendencies of both change and continuity in the debate over consensus in deliberative theory. We conclude this introduction by providing a brief synopsis of each paper

Deliberation after Consensus: Introduction to the Symposium

This editorial introduction presents an overview of the themes explored in the symposium on Deliberation after Consensus. For all the talk of its obsolescence and irrelevance, the concept of consensus still remains centrally contested through generations of deliberative democracy scholarship. In face of criticism for being neither empirically feasible nor normatively desirable, some deliberative theorists have moved away from consensus-oriented teleology and argued in favor of other legitimate outcomes of deliberations. Other theorists have resisted this move, claiming that the aim of deliberation implies that consensus should remain as a regulative ideal for deliberative outcomes. Engaging with these debates about the role of consensus in theories of deliberative democracy, this symposium brings together a selection of innovative, original research articles that raise novel questions about the role consensus could and should play in democratic deliberation and in a deliberative democracy. This introduction offers an overview of the debate over consensus drawing on the notion of successive generations of deliberative democracy research. Our aim is to demonstrate that the view of consensus has changed during generations of deliberative scholarships, but also that some scholars still defend the normative importance of the meaning of consensus once developed by the first generation. Consequently, there are tendencies of both change and continuity in the debate over consensus in deliberative theory. We conclude this introduction by providing a brief synopsis of each paper

The agr quorum sensing system in Staphylococcus aureus cells mediates death of sub-population

Abstract Objective In the human pathogen, Staphylococcus aureus, the agr quorum sensing system controls expression of a multitude of virulence factors and yet, agr negative cells frequently arise both in the laboratory and in some infections. The aim of this study was to examine the possible reasons behind this phenomenon. Results We examined viability of wild type and agr mutant cell cultures using a live-dead stain and observed that in stationary phase, 3% of the wild type population became non-viable whereas for agr mutant cells non-viable cells were barely detectable. The effect appears to be mediated by RNAIII, the effector molecule of agr, as ectopic overexpression of RNAIII resulted in 60% of the population becoming non-viable. This effect was not due to toxicity from delta toxin that is encoded by the hld gene located within RNAIII as hld overexpression did not cause cell death. Importantly, lysed S. aureus cells promoted bacterial growth. Our data suggest that RNAIII mediated cell death of agr positive but not agr negative cells provides a selective advantage to the agr negative cell population and may contribute to the common appearance of agr negative cells in S. aureus populations

MOESM1 of The agr quorum sensing system in Staphylococcus aureus cells mediates death of sub-population

Additional file 1: Figure S1. Hemolysis negative cells arise in S. aureus Newman. Five parallel cultures of WT were passaged in TSB for 24 days. Every second day suitable dilutions were plated out on TSB agar with 5 % calf’s blood and each colony was scored for hemolysis by comparing to WT freshly inoculated from the freeze stock. Zones of hemolysis smaller than ~0,5 mm were scored as hemolysis negative. Figure S2. RNAIII overexpression with pTXΔRNAIII. RT-qPCR was used to measure expression of RNAIII in S. aureus Newman carrying vector (pTXΔ) or RNAIII overproducing plasmid (pTXΔRNAIII) after 6 hours of growth in TSB. Data represent three biological replicates and are shown as mean ratios normalized to a run calibrator of genomic DNA. Error bars represent the standard deviation. Figure S3. Lysed bacteria supports growth. WT cells were grown in diluted 0.1xTSB supplemented with increasing amounts of bacterial lysate, and growth was measured in a Bioscreen at OD600. The experiment was performed with biological triplicates for each condition and the data represent the mean OD600 and standard deviation. Table S1. Strains and plasmids used in this study. Table S2. Oligonucleotides used in this study

Psychometric properties and concurrent validity of the Transgender Congruence Scale (TCS) in the Swedish setting

The Transgender Congruence Scale (TCS) is a non-binary tool used in Sweden for gender dysphoria (GD) assessment; however, its Swedish version has not been validated. To investigate the psychometric properties of the TCS, its capacity to distinguish individuals with GD and its concurrent validity compared to other scales. Patients with GD (n=135) and controls (n=443) filled in a questionnaire containing sociodemographic questions, the TCS, the Utrecht Gender Dysphoria Scale (UGDS), and the Gender Identity/Gender Dysphoria Questionnaire for Adolescents and Adults (GIDYQ-AA). TCS had good discriminatory validity and internal consistency. Patients with GD, stratified by birth-assigned sex, had lower TCS scores compared to controls. Confirmatory factor analysis (CFA) supported the two-factor model of the TCS. Multiple-group CFA suggested measurement invariance between birth-assigned sexes and configural invariance between patients with GD and controls. Area under the ROC curve for birth-assigned males was 0.991 and for females 0.994. A TCS mean value of three provided sensitivity 94.3% and 95.1% as well as specificity 98.6% and 98% for aM and aF, respectively. The TCS was significantly correlated to UGDS and GIDYQ-AA. The TCS may be a valuable tool in the clinical assessment of individuals with GD

Norlichexanthone interferes with AgrA binding to DNA.

<p>Samples including AgrAc, DNA probe, and/or compound were loaded in TBE buffer containing 10 mM dithiothreitol. Assays including the P2-P3 49 bp probe were analyzed in 4.5% native polyacrylamide gels. Lanes: 1. Size marker; 2. P2-P3 probe alone; 3. Probe and norlichexanthone (Nor) in the absence of AgrAc protein; 4. 100 uM I-d (potential hit compound and positive binding control); 5–10: norlichexanthone (Nor) increased by two-fold concentrations from 0–100 μg/ml.</p