We analyzed a representative sample of methicillin-resistant Staphylococcus aureus (MRSA) from 11 European countries (referred to as the HARMONY collection) using three molecular typing methods used within the HARMONY group to examine their usefulness for large, multicenter MRSA surveillance networks that use these different laboratory methodologies. MRSA isolates were collected based on their prevalence in each center and their genetic diversity, assessed by pulsed-field gel electrophoresis (PFGE). PFGE groupings (<3 bands difference between patterns) were compared to those made by sequencing of the variable repeats in the protein A gene spa and clonal designations based on multilocus sequence typing (MLST), combined with PCR analysis of the staphylococcal chromosome cassette containing the mec genes involved in methicillin resistance (SCCmec). A high level of discrimination was achieved using each of the three methodologies, with discriminatory indices between 89.5% and 91.9% with overlapping 95% confidence intervals. There was also a high level of concordance of groupings made using each method. MLST/SCCmec typing distinguished 10 groups containing at least two isolates, and these correspond to the majority of nosocomial MRSA clones described in the literature. PFGE and spa typing resolved 34 and 31 subtypes, respectively, within these 10 MRSA clones, with each subtype differing only slightly from the most common pattern using each method. The HARMONY group has found that the methods used in this study differ in their availability and affordability to European centers involved in MRSA surveillance. Here, we demonstrate that the integration of such technologies is achievable, although common protocols (such as we have developed for PFGE) may also be important, as is the use of centralized Internet sites to facilitate data analysis. PFGE and spa-typing data from analysis of MRSA isolates from the many centers that have access to the relevant equipment can be compared to reference patterns/sequences, and clonal designations can be made. In the majority of cases, these will correspond to those made by the (more expensive) method of choice-MLST/SCCmec typing-and these alternative methods can therefore be used as frontline typing systems for multicenter surveillance of MRSA.Methicillin-resistant Staphylococcus aureus (MRSA) is among the most common nosocomial pathogens globally and is generally acknowledged as the most significant due to the burden of disease it causes and to the evolution and global spread of multidrug-resistant clones. MRSA isolation rates in the United States, parts of Europe, and Asia have been increasing for more than 4 decades (36), and recent figures show that in some areas Ͼ50% of S. aureus bacteremias are caused by MRSA (4, 5, 6). Emerging intermediate, and more recently high-level (vanA-encoded), vancomycin resistance (8,22) and increasing numbers of multidrug-resistant MRSA emphasize the importance of effective antimicrobial prescribing and infection control measures that can be informed...

The aim of the present study was to gain an insight into the role of virulence determinants and biofilm production in bacteraemia of urinary tract origin. For this purpose 105 Escherichia coli isolates from patients with bacteraemia of urinary tract origin, isolated at the Institute of Microbiology and Immunology, University of Ljubljana, Slovenia, were investigated. A total of 88 strains (84 %) were isolated from immunocompromised patients and 17 (16 %) from nonimmunocompromised patients. The prevalence of virulence factor (VF)-encoding genes and associations with phylogenetic background, antibiotic resistance, biofilm production and patient status were analysed by PCR and bioassay. Biofilm was produced by 55 (53 %) of the strains. No combination of VFs was highly associated with biofilm production. Of the tested VF-encoding genes, usp, papC and the adhesin-encoding sfa/foc were significantly more prevalent among strains from non-immunocompromised patients. Our results indicate that the uropathogenic specific protein (USP) may be, as judged by predominance and associations of the usp gene, an important VF contributing significantly to bacteraemia of urinary tract origin.

Mur ligases participate in the intracellular path of bacterial peptidoglycan biosynthesis and constitute attractive, although so far underexploited, targets for antibacterial drug discovery. A series of hydroxy-substituted 5-benzylidenethiazolidin-4-ones were synthesized and tested as inhibitors of Mur ligases. The most potent compound 5 a was active against MurD-F with IC(50) values between 2 and 6 microm, making it a promising multitarget inhibitor of Mur ligases. Antibacterial activity against different strains, inhibitory activity against protein kinases, mutagenicity and genotoxicity of 5 a were also investigated, and kinetic and NMR studies were conducted.

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MurD and MurE ligases, consecutive enzymes participating in the intracellular steps of bacterial peptidoglycan biosynthesis, are important targets for antibacterial drug discovery. We have designed, synthesized, and evaluated the first d-glutamic acid-containing dual inhibitor of MurD and MurE ligases from Escherichia coli and Staphylococcus aureus (IC50 values between 6.4 and 180 μM) possessing antibacterial activity against Gram-positive S. aureus and its methicillin-resistant strain (MRSA) with minimal inhibitory concentration (MIC) values of 8 μg/mL. The inhibitor was also found to be noncytotoxic for human HepG2 cells at concentrations below 200 μM.