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Objectives The development of daptomycin resistance in Staphylococcus aureus is associated with clinical treatment failures. The mechanism(s) of such resistance have not been clearly defined. Methods We studied an isogenic daptomycin-susceptible (DAPS) and daptomycin-resistant (DAPR) S. aureus strain pair (616; 701) from a patient with relapsing endocarditis during daptomycin treatment, using comparative transcriptomic and proteomic techniques. Results Minor differences in the genome content were found between strains by DNA hybridization. Transcriptomic analyses identified a number of genes differentially expressed in important functional categories: cell division; metabolism of bacterial envelopes; and global regulation. Of note, the DAPR isolate exhibited reduced expression of the major cell wall autolysis gene coincident with the up-regulation of genes involved in cell wall teichoic acid production. Using quantitative (q)RT-PCR on the gene cadre putatively involved in cationic peptide resistance, we formulated a putative regulatory network compatible with microarray data sets, mainly implicating bacterial envelopes. Of interest, qRT-PCR of this same gene cadre from two distinct isogenic DAPS/DAPR clinical strain pairs revealed evidence of other strain-dependent networks operative in the DAPR phenotype. Comparative proteomics of 616 versus 701 revealed a differential abundance of proteins in various functional categories, including cell wall-associated targets and biofilm formation proteins. Phenotypically, strains 616 and 701 showed major differences in their ability to develop bacterial biofilms in the presence of the antibacterial lipid, oleic acid. Conclusions Compatible with previous in vitro observations, in vivo-acquired DAPR in S. aureus is a complex, multistep phenomenon involving: (i) strain-dependent phenotypes; (ii) transcriptome adaptation; and (iii) modification of the lipid and protein contents of cellular envelopes

Background: Ecs is an ATP-binding cassette (ABC) transporter present in aerobic and facultative anaerobic Gram-positive Firmicutes. Inactivation of Bacillus subtilis Ecs causes pleiotropic changes in the bacterial phenotype including inhibition of intramembrane proteolysis. The molecule(s) transported by Ecs is (are) still unknown.Methodology/Principal Findings: In this study we mutated the ecsAB operon in two Staphylococcus aureus strains, Newman and LS-1. Phenotypic and functional characterization of these Ecs deficient mutants revealed a defect in growth, increased autolysis and lysostaphin sensitivity, altered composition of cell wall proteins including the precursor form of staphylokinase and an altered bacterial surface texture. DNA microarray analysis indicated that the Ecs deficiency changed expression of the virulence factor regulator protein Rot accompanied by differential expression of membrane transport proteins, particularly ABC transporters and phosphate-specific transport systems, protein A, adhesins and capsular polysaccharide biosynthesis proteins. Virulence of the ecs mutants was studied in a mouse model of hematogenous S. aureus infection. Mice inoculated with the ecs mutant strains developed markedly milder infections than those inoculated with the wild-type strains and had consequently lower mortality, less weight loss, milder arthritis and decreased persistence of staphylococci in the kidneys. The ecs mutants had higher susceptibility to ribosomal antibiotics and plant alkaloids chelerythrine and sanguinarine.Conclusions/Significance: Our results show that Ecs is essential for staphylococcal virulence and antimicrobial resistance probably since the transport function of Ecs is essential for the normal structure and function of the cell wall. Thus targeting Ecs may be a new approach in combating staphylococcal infection.

BACKGROUND: Staphylococcus aureus secretes numerous exotoxins which may exhibit superantigenic properties. Whereas the virulence of several of them is well documented, their exact biological effects are not fully understood. Exotoxins may influence the immune and inflammatory state of various organs, including the sinonasal mucosa: their possible involvement in chronic rhinosinusitis has been suggested and is one of the main trends in current research. The aim of this study was to investigate whether the presence of any of the 22 currently known staphylococcal exotoxin genes could be correlated with chronic rhinosinusitis. METHODOLOGY/PRINCIPAL FINDINGS: We conducted a prospective, multi-centred European study, analysing 93 Staphylococcus aureus positive swabs taken from the middle meatus of patients suffering from chronic rhinosinusitis, with or without nasal polyposis, and controls. Strains were systematically tested for the presence of the 22 currently known exotoxin genes and genotyped according to their agr groups. No direct correlation was observed between chronic rhinosinusitis, with or without nasal polyposis, and either agr groups or the presence of the most studied exotoxins genes (egc, sea, seb, pvl, exfoliatins or tsst-1). However, genes for enterotoxins P and Q were frequently observed in nasal polyposis for the first time, but absent in the control group. The number of exotoxin genes detected was not statistically different among the 3 patient groups. CONCLUSIONS/SIGNIFICANCE: Unlike many previous studies have been suggesting, we did not find any evident correlation between staphylococcal exotoxin genes and the presence or severity of chronic rhinosinusitis with or without nasal polyposis.

Methicillin-resistant Staphylococcus aureus (MRSA) is an emerging community pathogen. Community-acquired MRSA (CA-MRSA) has been associated with virulent strains producing Panton-Valentine leukocidin (PVL) and a variety of other exotoxins. In Geneva, PVL-producing CA-MRSA was first reported in 2002 and a surveillance system based on voluntary reporting was set up. Each MRSA-positive culture result with an antibiotic resistance profile different from the endemic strain prevailing in the Geneva healthcare setting diagnosed in a patient without a history of hospital admission in the previous 12 months was notified to the local health department. A questionnaire was completed by the attending physician with demographic, clinical and exposure information. From January 2002 until December 2004, data on 58 cases were reported, including 26 cases grouped in 13 distinct transmission clusters. Most were family related and for two of them, colonisation persisted over a 12 month period despite treatment. Thirty three patients (57%) were male. Median age was 32 years, 22% being younger than 10 years. Forty one cases (71%) were infected and 17 (29%) colonised. Symptomatic skin lesions such as furunculosis, impetigo or abscess were present in 40 (97%) of the 41 infected cases. Most cases had no underlying disease. Thirty eight cases (65%) had travelled abroad. Forty (69%) of 58 isolates carried the PVL toxin. CA-MRSA infections in Geneva appear to be an emerging problem in the canton. Surveillance should continue and should possibly be extended to other parts of the country to better describe transmission patterns and the spread of this pathogen. Prevention and control of CA-MRSA infections represent a challenge for the future, requiring contact tracing, education and treatment of infected and colonised contacts.

Introduction Le microbiote intestinal participe au developpent de l'obesite, du diabete de type 2 et de l'inflammation de faible grade. Nos travaux precedents ont associe la composition du microbiote intestinal avec certains stigmates de l'obesite et du diabete de type 2 (par exemple : la masse grasse, la tolerance au glucose, l'inflammation). Nous avons montre que modifier le microbiote intestinal (par ex : augmenter Bifidobacterium spp.) d'animaux obeses avec des prebiotiques ameliore l'obesite et ses desordres. Cependant, a ce jour, il n'existe aucune analyse complete et precise du microbiome dans ce contexte. Quelles sont les bacteries veritablement impliquees ? Materiels et methodes 20 souris obeses (ob/ob) traitees 5 semaines avec ou sans prebiotiques (n=10/groupe). Le microbiome est analyse par 4 techniques: 1e pyro-sequencage, 2 e Mouse-Intestinal-CHIP-array, 3 e DGGE et 4 e PCR-quantitative ; toutes ses techniques sont basees sur l'analyse de l'ARN 16S-ribosomal (16SrRNA). De plus, nous avons etudies l'homeostasie du glucose, l'inflammation, le stress oxydatif et la barriere intestinale. Resultats Les analyses multivariees ("heat-map profile", "dendrogram", et analyse en composantes principales) revelent deux clusters distincts dependant des traitements nutritionnels. Le sequencage et le microarray (soit plus de 72 000 sequences bacteriennes) mettent en evidence des changements drastiques du microbiote par les prebiotiques, avec 102 sequences 16S-rRNA significativement modifiees dont 16 taxa bacteriens sont changes plus de 10 fois (8 diminues et 8 augmentes). En outre les prebiotiques diminuent la permeabilite intestinale, l'inflammation, le stress oxydatif et l'intolerance au glucose, ces phenomenes sont associes a une augmentation de la differentiation des cellules souches intestinales en cellules enteroendocrines de type-L. Enfin, nous montrons de nettes correlations entre certains parametres metaboliques et des bacteries jusqu'alors jamais identifiees dans ce contexte. Conclusion En etablissant le profil complet du microbiome de souris obeses traitees ou non avec des prebiotiques, nous avons identifies de nouvelles cibles bacteriennes qui faconnent le metabolisme de l'hote.

Background Identification of unexpected taxa in 16S rRNA surveys of low-density microbiota, diluted mock communities and cultures demonstrated that a variable fraction of sequence reads originated from exogenous DNA. The sources of these contaminants are reagents used in DNA extraction, PCR, and next-generation sequencing library preparation, and human (skin, oral and respiratory) microbiota from the investigators. Results For in silico removal of reagent contaminants, a pipeline was used which combines the relative abundance of operational taxonomic units (OTUs) in V3–4 16S rRNA gene amplicon datasets with bacterial DNA quantification based on qPCR targeting of the V3 segment of the 16S rRNA gene. Serially diluted cultures of Escherichia coli and Staphylococcus aureus were used for 16S rDNA profiling, and DNA from each of these species was used as a qPCR standard. OTUs assigned to Escherichia or Staphylococcus were virtually unaffected by the decontamination procedure, whereas OTUs from Pseudomonas, which is a major reagent contaminant, were completely or nearly completely removed. The decontamination procedure also attenuated the trend of increase in OTU richness in serially diluted cultures. Conclusions Removal of contaminant sequences derived from reagents based on use of qPCR data may improve taxonomic representation in samples with low DNA concentration. Using the described pipeline, OTUs derived from cross-contamination of negative extraction controls were not recognized as contaminants and not removed from the sample dataset. Electronic supplementary material The online version of this article (doi:10.1186/s12866-016-0689-4) contains supplementary material, which is available to authorized users.