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We investigated Staphylococcus aureus diversity, genetic factors, and humoral immune responses against antigens via genome analysis of S. aureus isolates from chronic rhinosinusitis (CRS) patients in a long-term follow-up. Of the 42 patients who provided S. aureus isolates and serum for a previous study, 34 could be included for follow-up after a decade. Clinical examinations were performed and bacterial samples were collected from the maxillary sinus and nares. S. aureus isolates were characterized by whole-genome sequencing, and specific anti-staphylococcal IgG in serum was determined using protein arrays. S. aureus was detected in the nares and/or maxillary sinus at both initial inclusion and follow-up in 15 of the 34 respondents (44%). Three of these (20%) had S. aureus isolates from the same genetic lineage as at inclusion. A low number of single-nucleotide polymorphisms (SNPs) were identified when comparing isolates from nares and maxillary sinus collected at the same time point. The overall change of antibody responses to staphylococcal antigens over time showed great variability, and no correlation was found between the presence of genes encoding antigens and the corresponding anti-staphylococcal IgG in serum; thus our findings did not support a role, in CRS, of the specific S. aureus antigens investigated., Funding Agencies:ALF founding Region Örebro County Research Committee of Örebro County Council OLL-929623OLL-878861

A PVL-positive, methicillin-susceptible Staphylococcus aureus was cultured from pus from cervical lymphadenitis of a patient of East-African origin. Microarray hybridisation assigned the isolate to clonal complex (CC) 80 but revealed unusual features, including the presence of the ORF-CM14 enterotoxin homologue and of an ACME-III element as well as the absence of etD and edinB. The isolate was subjected to both, Illumina and Nanopore sequencing allowing characterisation of deviating regions within the strain´s genome. Atypical features of this strain were attributable to the presence of two genomic regions that originated from other S. aureus lineages and that comprised, respectively, 3% and 1.4% of the genome. One deviating region extended from walJ to sirB. It comprised ORF-CM14 and the ACME-III element. A homologous but larger fragment was also found in an atypical S. aureus CC1/ST567 strain whose lineage might have served as donor of this genomic region. This region itself is a chimera comprising fragments from CC1 as well as fragments of unknown origin. The other deviating region comprised the region from htsB to ecfA2, i.e., another 3% of the genome. It was very similar to CC1 sequences. Either this suggests an incorporation of CC1 DNA into the study strain, or alternatively a recombination event affecting "canonical" CC80. Thus, the study strain bears witness of several recombination events affecting supposedly core genomic genes. Although the exact mechanism is not yet clear, such chimerism seems to be an additional pathway in the evolution of S. aureus. This could facilitate also a transmission of virulence and resistance factors and therefore offer an additional evolutionary advantage.

Background: Staphylococcus aureus (S. aureus) can behave both as a harmless commensal and as a pathogen. Its significance in the pathogenesis of chronic rhinosinusitis (CRS) is not yet fully understood. This study aimed to determine serum antibody re-sponses to specific staphylococcal antigens in patients with CRS and healthy controls, and to investigate the correlation between specific antibody response and severity of symptoms. Methodology: Serum samples from 39 patients with CRS and 56 healthy controls were analysed using a protein microarray to investigate the antibody response to S. aureus specific antigens, with a focus on immunoglobulin G (IgG) directed toward stap-hylococcal components accessible to the immune system. Holm-Bonferroni corrections were applied in all analyses. Information about growth of S. aureus in nares and maxillary sinus was taken from a previous study based on the same individuals. Clinical symptoms were assessed using a scoring system. Results: IgG antibody levels toward staphylococcal TSST-1 and LukF-PV were significantly higher in the CRS patient group com-pared to healthy controls, and levels of anti-TSST-1 antibodies were significantly higher in the CRS patient group with S. aureus in maxillary sinus than in controls. There were no correlations between the severity of symptoms and levels of serum anti-staphylo-coccal IgG antibody levels for LukF-PV and TSST-1. Conclusions: TSST-1 and LukF-PV could be interesting markers for future studies of the pathogenesis of CRS., This study was funded by the Research Committee of Örebro County Council, Sweden, and ALF funding for Region Örebro County, Sweden. The funding was used to finance time for research work and analysis.

Background: Staphylococcus aureus (S. aureus) can behave both as a harmless commensal and as a pathogen. Its significance in the pathogenesis of chronic rhinosinusitis (CRS) is not yet fully understood. This study aimed to determine serum antibody re-sponses to specific staphylococcal antigens in patients with CRS and healthy controls, and to investigate the correlation between specific antibody response and severity of symptoms. Methodology: Serum samples from 39 patients with CRS and 56 healthy controls were analysed using a protein microarray to investigate the antibody response to S. aureus specific antigens, with a focus on immunoglobulin G (IgG) directed toward stap-hylococcal components accessible to the immune system. Holm-Bonferroni corrections were applied in all analyses. Information about growth of S. aureus in nares and maxillary sinus was taken from a previous study based on the same individuals. Clinical symptoms were assessed using a scoring system. Results: IgG antibody levels toward staphylococcal TSST-1 and LukF-PV were significantly higher in the CRS patient group com-pared to healthy controls, and levels of anti-TSST-1 antibodies were significantly higher in the CRS patient group with S. aureus in maxillary sinus than in controls. There were no correlations between the severity of symptoms and levels of serum anti-staphylo-coccal IgG antibody levels for LukF-PV and TSST-1. Conclusions: TSST-1 and LukF-PV could be interesting markers for future studies of the pathogenesis of CRS., This study was funded by the Research Committee of Örebro County Council, Sweden, and ALF funding for Region Örebro County, Sweden. The funding was used to finance time for research work and analysis.

The presence of the methicillin resistance gene mecC in coagulase-negative Staphylococcus spp. (CoNS) is scarce. The aim of this study was to characterize mecC-positive CoNS isolated from various wild and domestic animals. The presence of the mecC gene was screened in 4299 samples from wild animals and domestic animals. Fifteen coagulase-negative staphylococci, that displayed a cefoxitin-resistant phenotype, were tested mecC-positive by PCR. Antimicrobial susceptibility testing was performed for all isolates. The 15 isolates were genotyped by sequencing of the entire class E mec gene complex (blaZ-mecC-mecRI-mecI), the ccrA and ccrB recombinase genes and other determinants within the type XI SCCmec element. DNA microarray analysis was performed and five selected isolates were additionally whole genome sequenced and analyzed. S. stepanovicii (n = 3), S. caprae (n = 1), S. warneri (n = 1), S. xylosus (n = 1) and S. sciuri (n = 9) were detected. All but the S. sciuri isolates were found to be susceptible to all non-beta lactams. The entire class E mec gene complex was detected in all isolates but ccrA and ccrB genes were not identified in S. stepanovicii and S. xylosus. The genes erm(B) and fexA (n = 4, each) were the most predominant non-beta lactam resistance genes detected in the S. sciuri isolates. Even though the presence of the mecC gene among CoNS is a rare observation, this study further expands our knowledge by showing that the mecC gene, including its allotypes, are present in more staphylococcal species from different animal species than has been previously described.

The evolution and global transmission of antimicrobial resistance have been well documented for Gram-negative bacteria and health care-associated epidemic pathogens, often emerging from regions with heavy antimicrobial use. However, the degree to which similar processes occur with Gram-positive bacteria in the community setting is less well understood. In this study, we traced the recent origins and global spread of a multidrug-resistant, community-associated Staphylococcus aureus lineage from the Indian subcontinent, the Bengal Bay clone (ST772). We generated whole-genome sequence data of 340 isolates from 14 countries, including the first isolates from Bangladesh and India, to reconstruct the evolutionary history and genomic epidemiology of the lineage. Our data show that the clone emerged on the Indian subcontinent in the early 1960s and disseminated rapidly in the 1990s. Short-term outbreaks in community and health care settings occurred following intercontinental transmission, typically associated with travel and family contacts on the subcontinent, but ongoing endemic transmission was uncommon. Acquisition of a multidrug resistance integrated plasmid was instrumental in the emergence of a single dominant and globally disseminated clade in the early 1990s. Phenotypic data on biofilm, growth, and toxicity point to antimicrobial resistance as the driving force in the evolution of ST772. The Bengal Bay clone therefore combines the multidrug resistance of traditional health care-associated clones with the epidemiological transmission of community-associated methicillin-resistant S. aureus (MRSA). Our study demonstrates the importance of whole-genome sequencing for tracking the evolution of emerging and resistant pathogens. It provides a critical framework for ongoing surveillance of the clone on the Indian subcontinent and elsewhere. The Bengal Bay clone (ST772) is a community-associated and multidrug-resistant Staphylococcus aureus lineage first isolated from B

The presence of the methicillin resistance gene mecC in coagulase-negative Staphylococcus spp. (CoNS) is scarce. The aim of this study was to characterize mecC-positive CoNS isolated from various wild and domestic animals. The presence of the mecC gene was screened in 4299 samples from wild animals and domestic animals. Fifteen coagulase-negative staphylococci, that displayed a cefoxitin-resistant phenotype, were tested mecC-positive by PCR. Antimicrobial susceptibility testing was performed for all isolates. The 15 isolates were genotyped by sequencing of the entire class E mec gene complex (blaZ-mecC-mecRI-mecI), the ccrA and ccrB recombinase genes and other determinants within the type XI SCCmec element. DNA microarray analysis was performed and five selected isolates were additionally whole genome sequenced and analyzed. S. stepanovicii (n = 3), S. caprae (n = 1), S. warneri (n = 1), S. xylosus (n = 1) and S. sciuri (n = 9) were detected. All but the S. sciuri isolates were found to be susceptible to all non-beta lactams. The entire class E mec gene complex was detected in all isolates but ccrA and ccrB genes were not identified in S. stepanovicii and S. xylosus. The genes erm(B) and fexA (n = 4, each) were the most predominant non-beta lactam resistance genes detected in the S. sciuri isolates. Even though the presence of the mecC gene among CoNS is a rare observation, this study further expands our knowledge by showing that the mecC gene, including its allotypes, are present in more staphylococcal species from different animal species than has been previously described.

Background: Staphylococcus aureus (S. aureus) can behave both as a harmless commensal and as a pathogen. Its significance in the pathogenesis of chronic rhinosinusitis (CRS) is not yet fully understood. This study aimed to determine serum antibody re-sponses to specific staphylococcal antigens in patients with CRS and healthy controls, and to investigate the correlation between specific antibody response and severity of symptoms. Methodology: Serum samples from 39 patients with CRS and 56 healthy controls were analysed using a protein microarray to investigate the antibody response to S. aureus specific antigens, with a focus on immunoglobulin G (IgG) directed toward stap-hylococcal components accessible to the immune system. Holm-Bonferroni corrections were applied in all analyses. Information about growth of S. aureus in nares and maxillary sinus was taken from a previous study based on the same individuals. Clinical symptoms were assessed using a scoring system. Results: IgG antibody levels toward staphylococcal TSST-1 and LukF-PV were significantly higher in the CRS patient group com-pared to healthy controls, and levels of anti-TSST-1 antibodies were significantly higher in the CRS patient group with S. aureus in maxillary sinus than in controls. There were no correlations between the severity of symptoms and levels of serum anti-staphylo-coccal IgG antibody levels for LukF-PV and TSST-1. Conclusions: TSST-1 and LukF-PV could be interesting markers for future studies of the pathogenesis of CRS., This study was funded by the Research Committee of Örebro County Council, Sweden, and ALF funding for Region Örebro County, Sweden. The funding was used to finance time for research work and analysis.

Abiola Senok1, Ali M Somily2, Rania Nassar1, Ghada Garaween3, Garwin Kim Sing3, Elke Müller4,5, Annett Reissig4,5, Darius Gawlik6, Ralf Ehricht4,5, Stefan Monecke4,5,7 1Department of Basic Medical Sciences, College of Medicine, Mohammed Bin Rashid University of Medicine and Health Sciences, Dubai, United Arab Emirates; 2Department of Pathology and Laboratory Medicine, College of Medicine, King Khalid University Hospital and King Saud University, Riyadh, Saudi Arabia; 3Department of Microbiology and Immunology, College of Medicine, Alfaisal University, Riyadh, Saudi Arabia; 4InfectoGnostics Research Campus Jena, Jena, Germany; 5Leibniz Institute of Photonic Technology (IPHT), Jena, Germany; 6PTC - Phage Technology Center GmbH, Bönen, Germany; 7Medical Faculty “Carl Gustav Carus”, Institute for Medical Microbiology and Hygiene, Technische Universität Dresden, Dresden, GermanyCorrespondence: Abiola SenokCollege of Medicine, Mohammed Bin Rashid University of Medicine and Health Sciences, P.O. Box 505055, Dubai, United Arab EmiratesTel +971 4 383 8717Email abiola.senok@mbru.ac.aePurpose: There is a need for continuous surveillance of methicillin-resistant Staphylococcus aureus (MRSA) to identify emergence of new strains. We hypothesize that MRSA strains are evolving with ongoing acquisition of SCCmec elements. This study was carried out to evaluate the evolution of MRSA at a tertiary care facility in Saudi Arabia.Methods: MRSA isolates associated with invasive clinical infection, which were identified in 2017 at the microbiology laboratory, King Khalid University Hospital (KKUH) in Riyadh, Saudi Arabia, were studied. The molecular characterization of isolates was carried out using StaphyType DNA microarray (Alere Technologies GmbH/Abbott, Jena, Germany).Results: The 125 MRSA isolates studied belonged to 18 clonal complexes (CC) which were distributed into 32 strain assignments. The predominant CC were CC5 (n=30), CC6 (n=17)

The evolution and global transmission of antimicrobial resistance have been well documented for Gram-negative bacteria and health care-associated epidemic pathogens, often emerging from regions with heavy antimicrobial use. However, the degree to which similar processes occur with Gram-positive bacteria in the community setting is less well understood. In this study, we traced the recent origins and global spread of a multidrug-resistant, community-associated Staphylococcus aureus lineage from the Indian subcontinent, the Bengal Bay clone (ST772). We generated whole-genome sequence data of 340 isolates from 14 countries, including the first isolates from Bangladesh and India, to reconstruct the evolutionary history and genomic epidemiology of the lineage. Our data show that the clone emerged on the Indian subcontinent in the early 1960s and disseminated rapidly in the 1990s. Short-term outbreaks in community and health care settings occurred following intercontinental transmission, typically associated with travel and family contacts on the subcontinent, but ongoing endemic transmission was uncommon. Acquisition of a multidrug resistance integrated plasmid was instrumental in the emergence of a single dominant and globally disseminated clade in the early 1990s. Phenotypic data on biofilm, growth, and toxicity point to antimicrobial resistance as the driving force in the evolution of ST772. The Bengal Bay clone therefore combines the multidrug resistance of traditional health care-associated clones with the epidemiological transmission of community-associated methicillin-resistant S. aureus (MRSA). Our study demonstrates the importance of whole-genome sequencing for tracking the evolution of emerging and resistant pathogens. It provides a critical framework for ongoing surveillance of the clone on the Indian subcontinent and elsewhere. The Bengal Bay clone (ST772) is a community-associated and multidrug-resistant Staphylococcus aureus lineage first isolated from B

The presence of the methicillin resistance gene mecC in coagulase-negative Staphylococcus spp. (CoNS) is scarce. The aim of this study was to characterize mecC-positive CoNS isolated from various wild and domestic animals. The presence of the mecC gene was screened in 4299 samples from wild animals and domestic animals. Fifteen coagulase-negative staphylococci, that displayed a cefoxitin-resistant phenotype, were tested mecC-positive by PCR. Antimicrobial susceptibility testing was performed for all isolates. The 15 isolates were genotyped by sequencing of the entire class E mec gene complex (blaZ-mecC-mecRI-mecI), the ccrA and ccrB recombinase genes and other determinants within the type XI SCCmec element. DNA microarray analysis was performed and five selected isolates were additionally whole genome sequenced and analyzed. S. stepanovicii (n = 3), S. caprae (n = 1), S. warneri (n = 1), S. xylosus (n = 1) and S. sciuri (n = 9) were detected. All but the S. sciuri isolates were found to be susceptible to all non-beta lactams. The entire class E mec gene complex was detected in all isolates but ccrA and ccrB genes were not identified in S. stepanovicii and S. xylosus. The genes erm(B) and fexA (n = 4, each) were the most predominant non-beta lactam resistance genes detected in the S. sciuri isolates. Even though the presence of the mecC gene among CoNS is a rare observation, this study further expands our knowledge by showing that the mecC gene, including its allotypes, are present in more staphylococcal species from different animal species than has been previously described.

The presence of the methicillin resistance gene mecC in coagulase-negative Staphylococcus spp. (CoNS) is scarce. The aim of this study was to characterize mecC-positive CoNS isolated from various wild and domestic animals. The presence of the mecC gene was screened in 4299 samples from wild animals and domestic animals. Fifteen coagulase-negative staphylococci, that displayed a cefoxitin-resistant phenotype, were tested mecC-positive by PCR. Antimicrobial susceptibility testing was performed for all isolates. The 15 isolates were genotyped by sequencing of the entire class E mec gene complex (blaZ-mecC-mecRI-mecI), the ccrA and ccrB recombinase genes and other determinants within the type XI SCCmec element. DNA microarray analysis was performed and five selected isolates were additionally whole genome sequenced and analyzed. S. stepanovicii (n = 3), S. caprae (n = 1), S. warneri (n = 1), S. xylosus (n = 1) and S. sciuri (n = 9) were detected. All but the S. sciuri isolates were found to be susceptible to all non-beta lactams. The entire class E mec gene complex was detected in all isolates but ccrA and ccrB genes were not identified in S. stepanovicii and S. xylosus. The genes erm(B) and fexA (n = 4, each) were the most predominant non-beta lactam resistance genes detected in the S. sciuri isolates. Even though the presence of the mecC gene among CoNS is a rare observation, this study further expands our knowledge by showing that the mecC gene, including its allotypes, are present in more staphylococcal species from different animal species than has been previously described.

Background: Staphylococcus aureus (S. aureus) can behave both as a harmless commensal and as a pathogen. Its significance in the pathogenesis of chronic rhinosinusitis (CRS) is not yet fully understood. This study aimed to determine serum antibody re-sponses to specific staphylococcal antigens in patients with CRS and healthy controls, and to investigate the correlation between specific antibody response and severity of symptoms. Methodology: Serum samples from 39 patients with CRS and 56 healthy controls were analysed using a protein microarray to investigate the antibody response to S. aureus specific antigens, with a focus on immunoglobulin G (IgG) directed toward stap-hylococcal components accessible to the immune system. Holm-Bonferroni corrections were applied in all analyses. Information about growth of S. aureus in nares and maxillary sinus was taken from a previous study based on the same individuals. Clinical symptoms were assessed using a scoring system. Results: IgG antibody levels toward staphylococcal TSST-1 and LukF-PV were significantly higher in the CRS patient group com-pared to healthy controls, and levels of anti-TSST-1 antibodies were significantly higher in the CRS patient group with S. aureus in maxillary sinus than in controls. There were no correlations between the severity of symptoms and levels of serum anti-staphylo-coccal IgG antibody levels for LukF-PV and TSST-1. Conclusions: TSST-1 and LukF-PV could be interesting markers for future studies of the pathogenesis of CRS., This study was funded by the Research Committee of Örebro County Council, Sweden, and ALF funding for Region Örebro County, Sweden. The funding was used to finance time for research work and analysis.

Background: Staphylococcus aureus (S. aureus) can behave both as a harmless commensal and as a pathogen. Its significance in the pathogenesis of chronic rhinosinusitis (CRS) is not yet fully understood. This study aimed to determine serum antibody re-sponses to specific staphylococcal antigens in patients with CRS and healthy controls, and to investigate the correlation between specific antibody response and severity of symptoms. Methodology: Serum samples from 39 patients with CRS and 56 healthy controls were analysed using a protein microarray to investigate the antibody response to S. aureus specific antigens, with a focus on immunoglobulin G (IgG) directed toward stap-hylococcal components accessible to the immune system. Holm-Bonferroni corrections were applied in all analyses. Information about growth of S. aureus in nares and maxillary sinus was taken from a previous study based on the same individuals. Clinical symptoms were assessed using a scoring system. Results: IgG antibody levels toward staphylococcal TSST-1 and LukF-PV were significantly higher in the CRS patient group com-pared to healthy controls, and levels of anti-TSST-1 antibodies were significantly higher in the CRS patient group with S. aureus in maxillary sinus than in controls. There were no correlations between the severity of symptoms and levels of serum anti-staphylo-coccal IgG antibody levels for LukF-PV and TSST-1. Conclusions: TSST-1 and LukF-PV could be interesting markers for future studies of the pathogenesis of CRS., This study was funded by the Research Committee of Örebro County Council, Sweden, and ALF funding for Region Örebro County, Sweden. The funding was used to finance time for research work and analysis.

Background: Staphylococcus aureus (S. aureus) can behave both as a harmless commensal and as a pathogen. Its significance in the pathogenesis of chronic rhinosinusitis (CRS) is not yet fully understood. This study aimed to determine serum antibody re-sponses to specific staphylococcal antigens in patients with CRS and healthy controls, and to investigate the correlation between specific antibody response and severity of symptoms. Methodology: Serum samples from 39 patients with CRS and 56 healthy controls were analysed using a protein microarray to investigate the antibody response to S. aureus specific antigens, with a focus on immunoglobulin G (IgG) directed toward stap-hylococcal components accessible to the immune system. Holm-Bonferroni corrections were applied in all analyses. Information about growth of S. aureus in nares and maxillary sinus was taken from a previous study based on the same individuals. Clinical symptoms were assessed using a scoring system. Results: IgG antibody levels toward staphylococcal TSST-1 and LukF-PV were significantly higher in the CRS patient group com-pared to healthy controls, and levels of anti-TSST-1 antibodies were significantly higher in the CRS patient group with S. aureus in maxillary sinus than in controls. There were no correlations between the severity of symptoms and levels of serum anti-staphylo-coccal IgG antibody levels for LukF-PV and TSST-1. Conclusions: TSST-1 and LukF-PV could be interesting markers for future studies of the pathogenesis of CRS., This study was funded by the Research Committee of Örebro County Council, Sweden, and ALF funding for Region Örebro County, Sweden. The funding was used to finance time for research work and analysis.

The evolution and global transmission of antimicrobial resistance have been well documented for Gram-negative bacteria and health care-associated epidemic pathogens, often emerging from regions with heavy antimicrobial use. However, the degree to which similar processes occur with Gram-positive bacteria in the community setting is less well understood. In this study, we traced the recent origins and global spread of a multidrug-resistant, community-associated Staphylococcus aureus lineage from the Indian subcontinent, the Bengal Bay clone (ST772). We generated whole-genome sequence data of 340 isolates from 14 countries, including the first isolates from Bangladesh and India, to reconstruct the evolutionary history and genomic epidemiology of the lineage. Our data show that the clone emerged on the Indian subcontinent in the early 1960s and disseminated rapidly in the 1990s. Short-term outbreaks in community and health care settings occurred following intercontinental transmission, typically associated with travel and family contacts on the subcontinent, but ongoing endemic transmission was uncommon. Acquisition of a multidrug resistance integrated plasmid was instrumental in the emergence of a single dominant and globally disseminated clade in the early 1990s. Phenotypic data on biofilm, growth, and toxicity point to antimicrobial resistance as the driving force in the evolution of ST772. The Bengal Bay clone therefore combines the multidrug resistance of traditional health care-associated clones with the epidemiological transmission of community-associated methicillin-resistant S. aureus (MRSA). Our study demonstrates the importance of whole-genome sequencing for tracking the evolution of emerging and resistant pathogens. It provides a critical framework for ongoing surveillance of the clone on the Indian subcontinent and elsewhere. The Bengal Bay clone (ST772) is a community-associated and multidrug-resistant Staphylococcus aureus lineage first isolated from B

Streptococcus (S.) agalactiae sind grampositive, in Ketten gelagerte Kokken, die auf Blutagar eine Hämolyse zeigen. Aufgrund ihrer Zugehörigkeit zur Lancefield-Gruppe B werden sie auch als Gruppe B Streptokokken (GBS) bezeichnet (Hof, 2005). GBS sind die Hauptursache von Sepsis, Meningitis und Pneumonie bei Neugeborenen (Schrag, et al., 2000). Die Arbeit beschäftigte sich mit der Genotypisierung von GBS. Darüber hinaus konnten auch Einblicke in die Phylogenese sowie die Populationsstruktur von GBS gewonnen werden. Ziel war es, einen DNA-Microarray zu entwickeln und zur Genotypisierung von GBS einzusetzen. Während der Evaluierung des DNA Microarray konnten stammspezifische Muster beobachtet werden, diese wurden durch bereits etablierte Typisierungmethoden (MLST) bekannten Genotypen zugeordnet. Die Ergebnisse wurden in einer Datenbank zusammengefasst. Mithilfe der Datenbank konnte die Software zur Auswertung entwickelt werden. (siehe http://alere-technologies.com/en/products/lab-solutions.html). Der DNA-Microarray trägt Sonden für GBS-spezifische Virulenzfaktoren und Oberflächenmarker. Für die auf dem Microarray basierende Typisierung wurden 11 über das ganze Genom verteilte Gene bzw. Gencluster (bac, alp, pil1 locus, pepS8, fBsB, capsule locus, hylB, abiG-I/-II plus Q8DZ34, pil2 locus, nss plus srr plus rogB2 und rgfC/A/D/B) ausgewählt. Ubiquitär vorkommende, konservierte Gene (z. B. cylD/cylE) eignen sich nicht als Marker für eine Typisierung, wurden aber als Kontrollen und zur Normierung eingeschlossen. Zur vollständigen Charakterisierung wurden außerdem Sonden für hochmobile plasmidgebundene Resistenzgene wie z. B. erm(A), erm(B), erm(C), tet(M), emrB/qacA aufgetragen (Aracil, et al., 2002; Betriu, et al., 2003; Uh, et al., 2001). Diese Gene sind nicht für GBS spezifisch. Sie eignen sich z. B. für eine Unterscheidung einzelner Isolate, nicht jedoch für die Unterteilung der GBS Population in verschiedene Stämme. Für die vorliegende Arbeit wurden insgesamt 448 klini, Streptococcus (S.) agalactiae are Gram-positive, chain-forming cocci, which show hemolysis on blood agar. They are also referred to group B streptococci (GBS) because of their affiliation to Lancefield-group B (Hof, 2005). GBS are the main cause of sepsis, meningitis and pneumonia in newborns (Schrag, et al., 2000). The work focused on genotyping of GBS. The aim was to develop a DNA microarray and to use it for epidemiological typing of GBS. During the evaluation of the microarray, strain-specific patterns could be observed and these patterns assigned to genotypes as defined by other typing methods (MLST). The results were summarized in a database that subsequently was developed into software for automated analysis of experiments (http://alere-technologies.com/en/products/lab-solutions.html). The DNA microarray carries probes for GBS specific virulence factors and surface markers. For the microarray-based typing, 11 genes or gene clusters were selected that are distributed across the entire genome (bac, alp, pil1 locus, pepS8, fBsB, capsule locus, hylB, abiG-I/-II plus Q8DZ34, pil2 locus, nss plus srr plus rogB2 and rgfC/A/D/B). Ubiquitous, conserved genes (e.g. cylD/cylE) were included to be used as species markers and controls. Furthermore, probes for highly mobile plasmid-borne resistance genes such as erm(A), erm(B), erm(C), tet(M), emrB/qacA were also included (Aracil, et al., 2002; Betriu, et al., 2003; Uh, et al., 2001). These genes are not specific to GBS, but they are found in some isolates. They can be used to distinguish individual, related isolates, rather than for a definition of distinct strains. A total of 448 isolates of GBS was selected and examined for the present work. Among them were isolates from severe diseases, such as sepsis and meningitis, isolates from local infections as well as isolates from asymptomatic/healthy carriers. For comparison, isolates from veterinary medicine (from cases of bovine mastitis) and human isolates from a geographic

Streptococcus (S.) agalactiae sind grampositive, in Ketten gelagerte Kokken, die auf Blutagar eine Hämolyse zeigen. Aufgrund ihrer Zugehörigkeit zur Lancefield-Gruppe B werden sie auch als Gruppe B Streptokokken (GBS) bezeichnet (Hof, 2005). GBS sind die Hauptursache von Sepsis, Meningitis und Pneumonie bei Neugeborenen (Schrag, et al., 2000). Die Arbeit beschäftigte sich mit der Genotypisierung von GBS. Darüber hinaus konnten auch Einblicke in die Phylogenese sowie die Populationsstruktur von GBS gewonnen werden. Ziel war es, einen DNA-Microarray zu entwickeln und zur Genotypisierung von GBS einzusetzen. Während der Evaluierung des DNA Microarray konnten stammspezifische Muster beobachtet werden, diese wurden durch bereits etablierte Typisierungmethoden (MLST) bekannten Genotypen zugeordnet. Die Ergebnisse wurden in einer Datenbank zusammengefasst. Mithilfe der Datenbank konnte die Software zur Auswertung entwickelt werden. (siehe http://alere-technologies.com/en/products/lab-solutions.html). Der DNA-Microarray trägt Sonden für GBS-spezifische Virulenzfaktoren und Oberflächenmarker. Für die auf dem Microarray basierende Typisierung wurden 11 über das ganze Genom verteilte Gene bzw. Gencluster (bac, alp, pil1 locus, pepS8, fBsB, capsule locus, hylB, abiG-I/-II plus Q8DZ34, pil2 locus, nss plus srr plus rogB2 und rgfC/A/D/B) ausgewählt. Ubiquitär vorkommende, konservierte Gene (z. B. cylD/cylE) eignen sich nicht als Marker für eine Typisierung, wurden aber als Kontrollen und zur Normierung eingeschlossen. Zur vollständigen Charakterisierung wurden außerdem Sonden für hochmobile plasmidgebundene Resistenzgene wie z. B. erm(A), erm(B), erm(C), tet(M), emrB/qacA aufgetragen (Aracil, et al., 2002; Betriu, et al., 2003; Uh, et al., 2001). Diese Gene sind nicht für GBS spezifisch. Sie eignen sich z. B. für eine Unterscheidung einzelner Isolate, nicht jedoch für die Unterteilung der GBS Population in verschiedene Stämme. Für die vorliegende Arbeit wurden insgesamt 448 klini, Streptococcus (S.) agalactiae are Gram-positive, chain-forming cocci, which show hemolysis on blood agar. They are also referred to group B streptococci (GBS) because of their affiliation to Lancefield-group B (Hof, 2005). GBS are the main cause of sepsis, meningitis and pneumonia in newborns (Schrag, et al., 2000). The work focused on genotyping of GBS. The aim was to develop a DNA microarray and to use it for epidemiological typing of GBS. During the evaluation of the microarray, strain-specific patterns could be observed and these patterns assigned to genotypes as defined by other typing methods (MLST). The results were summarized in a database that subsequently was developed into software for automated analysis of experiments (http://alere-technologies.com/en/products/lab-solutions.html). The DNA microarray carries probes for GBS specific virulence factors and surface markers. For the microarray-based typing, 11 genes or gene clusters were selected that are distributed across the entire genome (bac, alp, pil1 locus, pepS8, fBsB, capsule locus, hylB, abiG-I/-II plus Q8DZ34, pil2 locus, nss plus srr plus rogB2 and rgfC/A/D/B). Ubiquitous, conserved genes (e.g. cylD/cylE) were included to be used as species markers and controls. Furthermore, probes for highly mobile plasmid-borne resistance genes such as erm(A), erm(B), erm(C), tet(M), emrB/qacA were also included (Aracil, et al., 2002; Betriu, et al., 2003; Uh, et al., 2001). These genes are not specific to GBS, but they are found in some isolates. They can be used to distinguish individual, related isolates, rather than for a definition of distinct strains. A total of 448 isolates of GBS was selected and examined for the present work. Among them were isolates from severe diseases, such as sepsis and meningitis, isolates from local infections as well as isolates from asymptomatic/healthy carriers. For comparison, isolates from veterinary medicine (from cases of bovine mastitis) and human isolates from a geographic

Streptococcus (S.) agalactiae sind grampositive, in Ketten gelagerte Kokken, die auf Blutagar eine Hämolyse zeigen. Aufgrund ihrer Zugehörigkeit zur Lancefield-Gruppe B werden sie auch als Gruppe B Streptokokken (GBS) bezeichnet (Hof, 2005). GBS sind die Hauptursache von Sepsis, Meningitis und Pneumonie bei Neugeborenen (Schrag, et al., 2000). Die Arbeit beschäftigte sich mit der Genotypisierung von GBS. Darüber hinaus konnten auch Einblicke in die Phylogenese sowie die Populationsstruktur von GBS gewonnen werden. Ziel war es, einen DNA-Microarray zu entwickeln und zur Genotypisierung von GBS einzusetzen. Während der Evaluierung des DNA Microarray konnten stammspezifische Muster beobachtet werden, diese wurden durch bereits etablierte Typisierungmethoden (MLST) bekannten Genotypen zugeordnet. Die Ergebnisse wurden in einer Datenbank zusammengefasst. Mithilfe der Datenbank konnte die Software zur Auswertung entwickelt werden. (siehe http://alere-technologies.com/en/products/lab-solutions.html). Der DNA-Microarray trägt Sonden für GBS-spezifische Virulenzfaktoren und Oberflächenmarker. Für die auf dem Microarray basierende Typisierung wurden 11 über das ganze Genom verteilte Gene bzw. Gencluster (bac, alp, pil1 locus, pepS8, fBsB, capsule locus, hylB, abiG-I/-II plus Q8DZ34, pil2 locus, nss plus srr plus rogB2 und rgfC/A/D/B) ausgewählt. Ubiquitär vorkommende, konservierte Gene (z. B. cylD/cylE) eignen sich nicht als Marker für eine Typisierung, wurden aber als Kontrollen und zur Normierung eingeschlossen. Zur vollständigen Charakterisierung wurden außerdem Sonden für hochmobile plasmidgebundene Resistenzgene wie z. B. erm(A), erm(B), erm(C), tet(M), emrB/qacA aufgetragen (Aracil, et al., 2002; Betriu, et al., 2003; Uh, et al., 2001). Diese Gene sind nicht für GBS spezifisch. Sie eignen sich z. B. für eine Unterscheidung einzelner Isolate, nicht jedoch für die Unterteilung der GBS Population in verschiedene Stämme. Für die vorliegende Arbeit wurden insgesamt 448 klini, Streptococcus (S.) agalactiae are Gram-positive, chain-forming cocci, which show hemolysis on blood agar. They are also referred to group B streptococci (GBS) because of their affiliation to Lancefield-group B (Hof, 2005). GBS are the main cause of sepsis, meningitis and pneumonia in newborns (Schrag, et al., 2000). The work focused on genotyping of GBS. The aim was to develop a DNA microarray and to use it for epidemiological typing of GBS. During the evaluation of the microarray, strain-specific patterns could be observed and these patterns assigned to genotypes as defined by other typing methods (MLST). The results were summarized in a database that subsequently was developed into software for automated analysis of experiments (http://alere-technologies.com/en/products/lab-solutions.html). The DNA microarray carries probes for GBS specific virulence factors and surface markers. For the microarray-based typing, 11 genes or gene clusters were selected that are distributed across the entire genome (bac, alp, pil1 locus, pepS8, fBsB, capsule locus, hylB, abiG-I/-II plus Q8DZ34, pil2 locus, nss plus srr plus rogB2 and rgfC/A/D/B). Ubiquitous, conserved genes (e.g. cylD/cylE) were included to be used as species markers and controls. Furthermore, probes for highly mobile plasmid-borne resistance genes such as erm(A), erm(B), erm(C), tet(M), emrB/qacA were also included (Aracil, et al., 2002; Betriu, et al., 2003; Uh, et al., 2001). These genes are not specific to GBS, but they are found in some isolates. They can be used to distinguish individual, related isolates, rather than for a definition of distinct strains. A total of 448 isolates of GBS was selected and examined for the present work. Among them were isolates from severe diseases, such as sepsis and meningitis, isolates from local infections as well as isolates from asymptomatic/healthy carriers. For comparison, isolates from veterinary medicine (from cases of bovine mastitis) and human isolates from a geographic

Abiola C Senok,1 Ali M Somily,2 Peter Slickers,3,4 Muhabat A Raji,5 Ghada Garaween,5 Atef Shibl,5 Stefan Monecke,3,4,6 Ralf Ehricht3,4 1Department of Basic Science, College of Medicine, Mohammed Bin Rashid University of Medicine and Health Sciences, Dubai, United Arab Emirates; 2Department of Pathology and Laboratory Medicine, College of Medicine, King Khalid University Hospital and King Saud University, Riyadh, Saudi Arabia; 3Alere Technologies GmbH, Jena, Germany, 4InfectoGnostics Research Campus, Jena, Germany; 5Department of Microbiology and Immunology, College of Medicine, Alfaisal University, Riyadh, Saudi Arabia; 6Institute for Medical Microbiology and Hygiene (IMMH), Technische Universität Dresden, Dresden, Germany Purpose: Methicillin resistant Staphylococcus aureus CC15 strains (CC15-MRSA) have only been sporadically described in literature. This study was carried out to describe the genetic make-up for this rare MRSA strain.Methods: Four CC15-MRSA isolates collected in Riyadh, Saudi Arabia, between 2013 and 2014 were studied. Two isolates were from clinical infection and 2 from retail meat products. Whole genome sequencing was carried out using Illumina HiSeq2500 genome analyzer.Results: All the CC15-MRSA isolates had the multilocus sequence typing profile ST1535, 13–13-1–1-81-11-13, which is a single locus variant of ST15. Of the 6 contigs related to the SCC element, one comprised a recombinase gene ccrAA, ccrC-PM1, fusC and a helicase, another one included mvaS, dru, mecA and 1 had yobV and Q4LAG7. The SCC element had 5 transposase genes, namely 3 identical paralogs of tnpIS431 and 2 identical paralogs of tnpIS256. Two identical copies of a tnpIS256-based insertion element flank the aacA-aphD gene. Two copies of this insertion element were present with 1 located in the SCC element and another inserted into the sasC gene. A short 3 kb region, which lacks any bacteriophage structural genes and site-specific DNA integrase, was

Streptococcus (S.) agalactiae sind grampositive, in Ketten gelagerte Kokken, die auf Blutagar eine Hämolyse zeigen. Aufgrund ihrer Zugehörigkeit zur Lancefield-Gruppe B werden sie auch als Gruppe B Streptokokken (GBS) bezeichnet (Hof, 2005). GBS sind die Hauptursache von Sepsis, Meningitis und Pneumonie bei Neugeborenen (Schrag, et al., 2000). Die Arbeit beschäftigte sich mit der Genotypisierung von GBS. Darüber hinaus konnten auch Einblicke in die Phylogenese sowie die Populationsstruktur von GBS gewonnen werden. Ziel war es, einen DNA-Microarray zu entwickeln und zur Genotypisierung von GBS einzusetzen. Während der Evaluierung des DNA Microarray konnten stammspezifische Muster beobachtet werden, diese wurden durch bereits etablierte Typisierungmethoden (MLST) bekannten Genotypen zugeordnet. Die Ergebnisse wurden in einer Datenbank zusammengefasst. Mithilfe der Datenbank konnte die Software zur Auswertung entwickelt werden. (siehe http://alere-technologies.com/en/products/lab-solutions.html). Der DNA-Microarray trägt Sonden für GBS-spezifische Virulenzfaktoren und Oberflächenmarker. Für die auf dem Microarray basierende Typisierung wurden 11 über das ganze Genom verteilte Gene bzw. Gencluster (bac, alp, pil1 locus, pepS8, fBsB, capsule locus, hylB, abiG-I/-II plus Q8DZ34, pil2 locus, nss plus srr plus rogB2 und rgfC/A/D/B) ausgewählt. Ubiquitär vorkommende, konservierte Gene (z. B. cylD/cylE) eignen sich nicht als Marker für eine Typisierung, wurden aber als Kontrollen und zur Normierung eingeschlossen. Zur vollständigen Charakterisierung wurden außerdem Sonden für hochmobile plasmidgebundene Resistenzgene wie z. B. erm(A), erm(B), erm(C), tet(M), emrB/qacA aufgetragen (Aracil, et al., 2002; Betriu, et al., 2003; Uh, et al., 2001). Diese Gene sind nicht für GBS spezifisch. Sie eignen sich z. B. für eine Unterscheidung einzelner Isolate, nicht jedoch für die Unterteilung der GBS Population in verschiedene Stämme. Für die vorliegende Arbeit wurden insgesamt 448 klini, Streptococcus (S.) agalactiae are Gram-positive, chain-forming cocci, which show hemolysis on blood agar. They are also referred to group B streptococci (GBS) because of their affiliation to Lancefield-group B (Hof, 2005). GBS are the main cause of sepsis, meningitis and pneumonia in newborns (Schrag, et al., 2000). The work focused on genotyping of GBS. The aim was to develop a DNA microarray and to use it for epidemiological typing of GBS. During the evaluation of the microarray, strain-specific patterns could be observed and these patterns assigned to genotypes as defined by other typing methods (MLST). The results were summarized in a database that subsequently was developed into software for automated analysis of experiments (http://alere-technologies.com/en/products/lab-solutions.html). The DNA microarray carries probes for GBS specific virulence factors and surface markers. For the microarray-based typing, 11 genes or gene clusters were selected that are distributed across the entire genome (bac, alp, pil1 locus, pepS8, fBsB, capsule locus, hylB, abiG-I/-II plus Q8DZ34, pil2 locus, nss plus srr plus rogB2 and rgfC/A/D/B). Ubiquitous, conserved genes (e.g. cylD/cylE) were included to be used as species markers and controls. Furthermore, probes for highly mobile plasmid-borne resistance genes such as erm(A), erm(B), erm(C), tet(M), emrB/qacA were also included (Aracil, et al., 2002; Betriu, et al., 2003; Uh, et al., 2001). These genes are not specific to GBS, but they are found in some isolates. They can be used to distinguish individual, related isolates, rather than for a definition of distinct strains. A total of 448 isolates of GBS was selected and examined for the present work. Among them were isolates from severe diseases, such as sepsis and meningitis, isolates from local infections as well as isolates from asymptomatic/healthy carriers. For comparison, isolates from veterinary medicine (from cases of bovine mastitis) and human isolates from a geographic

The biofilm-associated protein (Bap) of Staphylococcus aureus is a high molecular weight cell-wallanchored protein involved in biofilm formation, first described in bovine mastitis strains from Spain. So far, studies regarding Bap were mainly based on the Spanish strain V329 and its mutants, but no information on the genetic variability of bap-positive Staph. aureus strains is yet available in the literature. The present study investigated the molecular characteristics of 8 bap-positive Staph. aureus strains from subclinical bovine mastitis, isolated in 5 herds; somatic cell counts (SCC) of milk samples were also registered. Strains were characterised using MLST, SPA typing and microarray and the results were compared with V329. All isolates from this study and V329 were assigned to ST126, t605, but some molecular differences were observed. Only herd A and B strains harboured the genes for β-lactams resistance; the leukocidin D/E gene, a type I site-specific deoxyribonuclease subunit, 3rd locus gene and serin-protease A and B were carried by all strains, but not by V329, while serin-protease E was absent in V329 and in another isolate. Four isolates and V329 harboured the fibronectin-binding protein B gene. SCC showed the highest value in the milk sample affected by the only strain carrying all the virulence factors considered. Potential large variability of virulence was evidenced among V329 and all bap-positive Staph. aureus strains considered: the carriage of fnb could enhance the accumulation of biofilm, but the lack of lukD/E and splA, B or E might decrease the invasiveness of strain.

The biofilm-associated protein (Bap) of Staphylococcus aureus is a high molecular weight cell-wallanchored protein involved in biofilm formation, first described in bovine mastitis strains from Spain. So far, studies regarding Bap were mainly based on the Spanish strain V329 and its mutants, but no information on the genetic variability of bap-positive Staph. aureus strains is yet available in the literature. The present study investigated the molecular characteristics of 8 bap-positive Staph. aureus strains from subclinical bovine mastitis, isolated in 5 herds; somatic cell counts (SCC) of milk samples were also registered. Strains were characterised using MLST, SPA typing and microarray and the results were compared with V329. All isolates from this study and V329 were assigned to ST126, t605, but some molecular differences were observed. Only herd A and B strains harboured the genes for β-lactams resistance; the leukocidin D/E gene, a type I site-specific deoxyribonuclease subunit, 3rd locus gene and serin-protease A and B were carried by all strains, but not by V329, while serin-protease E was absent in V329 and in another isolate. Four isolates and V329 harboured the fibronectin-binding protein B gene. SCC showed the highest value in the milk sample affected by the only strain carrying all the virulence factors considered. Potential large variability of virulence was evidenced among V329 and all bap-positive Staph. aureus strains considered: the carriage of fnb could enhance the accumulation of biofilm, but the lack of lukD/E and splA, B or E might decrease the invasiveness of strain.

The anterior nares have been regarded as the major carriage site of Staphylococcus aureus. From here, the organism can spread to other parts of the body where it might act as harmless commensal or cause mild to severe infections. Nasal sinuses are normally sterile, but in patients with chronic rhinosinusitis (CRS), the finding of S. aureus in maxillary sinus cultures is common. Isolates were obtained from the nares and maxillary sinus of patients with CRS and the nares of healthy controls. A significantly higher frequency of S. aureus was found in nares samples from patients (24/42) compared to controls (16/57) (p = 0.004). There is no consensus regarding whether S. aureus is a relevant pathogen in CRS. A DNA microarray was used to investigate the prevalence of S. aureus virulence genes with focus on staphylococcal enterotoxins, toxic shock syndrome toxin-1, agr types, and cell wall-associated proteins. The genotyping of S. aureus isolates revealed only small and non-significant differences in gene prevalence between isolates collected from patients with CRS and those collected from healthy nasal carriers. This study provides an increased knowledge of the genetic pattern of virulence genes among S. aureus collected in CRS., Funding Agency:Research Committee of Orebro County Council

Staphylococcus aureus is one of the major pathogens that causes bacteremia; therefore, it is important to understand the long-term molecular epidemiology of S. aureus bacteremia infections. In particular, little is known about the population structure of methicillin-sensitive S. aureus (MSSA) compared to that of methicillin-resistant S. aureus. We investigated potential changes in the MSSA molecular epidemiology in Örebro County, Sweden, from 1980 through 2010. 400 MSSA bacteremia isolates, the first 100 isolated each decade from 1980 through 2010, were retrospectively identified and analyzed regarding assignment to clonal complexes (CCs), presence of virulence genes and antibiotic resistant determinants with DNA microarray-based genotyping. 24 different CCs were identified. Most isolates (80%) belonged to 6 predominant lineages. Of those, the number of isolates assigned to CC5 and CC15 increased, and those assigned to CC8, CC25, and CC30 decreased. The most prevalent clone, CC45, did not show a significant change in prevalence during the study period. A change in prevalence was observed for some of the virulence genes, mainly attributed with their association to certain CCs. With the exception of the common blaZ gene (encoding penicillinase), antibiotic resistance genes were only sporadically detected. In conclusion, the MSSA population structure was genetically diverse. We observed decadal changes in assignments to five predominant clones, and corresponding changes in the prevalence of some virulence genes linked to CC affiliation. In light of the restrictive antibiotics prescriptions and extensive infection control procedures in Sweden, antibiotic resistance genes were rarely detected and their prevalence unaffected during the study period., Funding Agency:Research Committee of the County Council of Örebro University Hospital

Staphylococcus aureus encodes a remarkable number of virulence factors which may contribute to its pathogenicity and ability to cause invasive disease. The main objective of this study was to evaluate the association between S. aureus invasiveness and bacterial genotype, in terms of the presence of virulence genes and affiliation to clonal complexes. Also, the significance of different virulence genes, mainly adhesins, for the development of infective endocarditis was investigated. DNA microarray technology was used to analyze 134 S. aureus isolates, all methicillin-susceptible, derived from three groups of clinically well-characterized patients: nasal carriers (n=46), bacteremia (n=55), and bacteremia with infective endocarditis (n=33). Invasive isolates were dominant in four of the major clonal complexes: 5, 8, 15, and 25. Of the 170 virulence genes examined, those encoding accessory gene regulator group II (agr II), capsule polysaccharide serotype 5 (cap5), and adhesins such as S. aureus surface protein G (sasG) and fibronectin-binding protein B (fnbB) were found to be associated with invasive disease. The same was shown for the leukocidin genes lukD/lukE, as well as the genes encoding serine protease A and B (splA/splB), staphylococcal complement inhibitor (scn) and the staphylococcal exotoxin-like protein (setC or selX). In addition, there was a trend of higher prevalence of certain genes or gene clusters (sasG, agr II, cap5) among isolates causing infective endocarditis compared to other invasive isolates. In most cases, the presence of virulence genes was linked to clonal complex affiliation. In conclusion, certain S. aureus clonal lineages harboring specific sets of virulence genes seem to be more successful in causing invasive disease., Funding Agency: Research Comitte of the County Council of Orebro University Hospital

Panton-Valentine leukocidin (PVL) is a virulence factor of Staphylococcus aureus, which is associated with skin and soft-tissue infections and necrotizing pneumonia. To develop a rapid phenotypic assay, recombinant PVL F component was used to generate monoclonal antibodies by phage display. These antibodies were spotted on protein microarrays and screened using different lukF-PV preparations and detection antibodies. This led to the identification of the optimal antibody combination that was then used to establish a lateral flow assay. This test was used to detect PVL in S. aureus cultures. The detection limit of the assay with purified native and recombinant antigens was determined to be around 1 ng/ml. Overnight cultures from various solid and liquid media proved suitable for PVL detection. Six hundred strains and clinical isolates from patients from America, Europe, Australia, Africa, and the Middle East were tested. Isolates were genotyped in parallel by DNA microarray hybridization for confirmation of PVL status and assignment to clonal complexes. The sensitivity, specificity, and positive and negative predictive values of the assay in this trial were 99.7, 98.3, 98.4, and 99.7%, respectively. A total of 302 clinical isolates and reference strains were PVL positive and were assigned to 21 different clonal complexes. In summary, the lateral flow test allows rapid and economical detection of PVL in a routine bacteriology laboratory. As the test utilizes cultures from standard media and does not require sophisticated equipment, it can be easily integrated into a laboratory's workflow and might contribute to timely therapy of PVL-associated infections.

In Australia the PVL - positive ST93-IV [2B], colloquially known as ‘‘Queensland CA-MRSA’’ has become the dominant CA-MRSA clone. First described in the early 2000s, ST93-IV [2B] is associated with skin and severe invasive infections including necrotizing pneumonia. A singleton by multilocus sequence typing (MLST) eBURST analysis ST93 is distinct from other S aureus clones. To determine if the increased prevalence of ST93-IV [2B] is due to the widespread transmission of a single strain of ST93-IV [2B] the genetic relatedness of 58 S. aureus ST93 isolated throughout Australia over an extended period were studied in detail using a variety of molecular methods including pulsed-field gel electrophoresis, spa typing, MLST, microarray DNA, SCCmec typing and dru typing. Identification of the phage harbouring the lukS-PV/lukF-PV Panton Valentine leucocidin genes, detection of allelic variations in lukS-PV/lukF-PV, and quantification of LukF-PV expression was also performed. Although ST93-IV [2B] is known to have an apparent enhanced clinical virulence, the isolates harboured few known virulence determinants. All PVL-positive isolates carried the PVL-encoding phage WSa2USA and the lukS-PV/lukF-PV genes had the same R variant SNP profile. The isolates produced similar expression levels of LukF-PV. Although multiple rearrangements of the spa sequence have occurred, the core genome in ST93 is very stable.The emergence of ST93-MRSA is due to independent acquisitions of different dru-defined type IV and type V SCCmec elements in several spa-defined ST93-MSSA backgrounds. Rearrangement of the spa sequence in ST93-MRSA has subsequently occurred in some of these strains. Although multiple ST93-MRSA strains were characterised, little genetic diversity was identified for most isolates, with PVLpositive ST93-IVa [2B]-t202-dt10 predominant across Australia. Whether ST93-IVa [2B] t202-dt10 arose from one PVL-positive ST93-MSSA-t202, or by independent acquisitions of SCCmec-IVa [2B]-dt10

Staphylococcus aureus kann bei etwa 25% der gesunden Normalbevölkerung nachgewiesen werden, ohne Symptome zu verursachen. Dieser Keim ist jedoch auch einer der wichtigsten Erreger bei Osteomyelitis und Infektionen von orthopädischen Implantaten wie z. B. von künstlichen Knie- oder Hüftgelenken. Diese Infektionen führen meist zu aufwendigen und risikobehafteten operativen Eingriffen sowie zu einer langfristigen Antibiotikagabe. In der vorliegenden Arbeit sollten S. aureus-Isolate charakterisiert werden, die aus Osteomyelitisherden oder infizierten orthopädischen Implantaten gewonnen wurden. Ziel war es, die Isolate daraufhin zu untersuchen, ob bestimmte Stämme dominieren und ob das Vorhandensein bestimmter Virulenzfaktoren mit einem besonderen Risiko für solche Infektionen korreliert. Für diese Untersuchungen wurden DNA-Arrays eingesetzt, welche es ermöglichen, alle relevanten Virulenzfaktoren in einem Experiment nachzuweisen, einen „genetischen Fingerabdruck“ zu erheben und die Isolate so Verwandtschaftsgruppen (klonalen Komplexen, CC) zuzuordnen. Insgesamt wurden 119 klinische Isolate charakterisiert. Sie gehörten zu 20 verschiedenen klonalen Komplexen. CC8 (19,3%), CC45 (17,7%) und CC30 (12,6%) dominierten. MRSA waren selten nachweisbar. Die sieben MRSA-Isolate gehörten zu den lokal dominierenden Stämmen (Rhein-Hessen, Süddeutscher, Barnimer und Berliner Epidemiestamm sowie Europäischer caMRSA-Klon). Die Populationsstruktur der klinischen Isolate und die Häufigkeiten der untersuchten Virulenz- und Adhäsionsfaktoren entsprachen weitestgehend Isolaten von asymptomatischen Trägern, die in einer früheren Studie bestimmt wurden (Molecular epidemiology of Staphylococcus aureus in asymptomatic carriers; Monecke, Lüdicke, Slickers, Ehricht; Eur J Clin Microbiol Infect Dis. 2009). Es konnte kein molekularer Marker identifiziert werden, der allein für eine Risikostratifizierung eingesetzt werden kann. Das Gen für Staphylokinase (sak) war jedoch bei den klinischen Isolaten (, Staphylococcus aureus is asymptomatically carried by approximately 25% of a normal population. It is also one of the most important causes of osteomyelitis and infections of orthopedic implants such as total hip or knee replacements. Such infections usually lead to complicated and risky surgical procedures as well as to long-term antibiotic treatment. In the present work, S. aureus isolates from osteomyelitis or implant infections were to be characterised. The aim of the study was to prove whether certain strains were overrepresented among patient isolates, and whether the presence of certain virulence factors might correlate with these infections. DNA arrays where used which facilitate to screen for all relevant virulence factors within a single experiment and which allow typing by obtaining a genetic fingerprint of the examined isolate. By this method, it was also possible to assign isolates to phylogenetic clusters, so-called clonal complexes. 119 clinical isolates were characterised in this way. They belonged to 20 different clonal complexes (CC). CC8 (19.3%), CC45 (17.7%) and CC30 (12.6%) dominated. MRSA were rarely detected. The seven MRSA isolates belonged to locally predominant epidemic strains (ST5-MRSA-II, ST228-MRSA-I, ST22-MRSA-IV, ST45-MRSA-IV and ST80-MRSA-IV). The population structure of the clinical isolates and the relative abundances of the examined virulence and adhesion factors corresponded largely to isolates from asymptomatic carriers, which has been examined in an earlier study (Molecular epidemiology of Staphylococcus aureus in asymptomatic carriers; Monecke, Lüdicke, Slickers, Ehricht; Eur J Clin Microbiol Infect Dis. 2009). No molecular maker was detected which could alone be used for risk assessment. The gene for staphylokinase (sak) was clearly more common among clinical isolates (90.8%) than in isolates from asymptomatic carriers (71.6%). Some other genes were also found to be more common in patient isolates, but were very rare so that a

Staphylococcus aureus kann bei etwa 25% der gesunden Normalbevölkerung nachgewiesen werden, ohne Symptome zu verursachen. Dieser Keim ist jedoch auch einer der wichtigsten Erreger bei Osteomyelitis und Infektionen von orthopädischen Implantaten wie z. B. von künstlichen Knie- oder Hüftgelenken. Diese Infektionen führen meist zu aufwendigen und risikobehafteten operativen Eingriffen sowie zu einer langfristigen Antibiotikagabe. In der vorliegenden Arbeit sollten S. aureus-Isolate charakterisiert werden, die aus Osteomyelitisherden oder infizierten orthopädischen Implantaten gewonnen wurden. Ziel war es, die Isolate daraufhin zu untersuchen, ob bestimmte Stämme dominieren und ob das Vorhandensein bestimmter Virulenzfaktoren mit einem besonderen Risiko für solche Infektionen korreliert. Für diese Untersuchungen wurden DNA-Arrays eingesetzt, welche es ermöglichen, alle relevanten Virulenzfaktoren in einem Experiment nachzuweisen, einen „genetischen Fingerabdruck“ zu erheben und die Isolate so Verwandtschaftsgruppen (klonalen Komplexen, CC) zuzuordnen. Insgesamt wurden 119 klinische Isolate charakterisiert. Sie gehörten zu 20 verschiedenen klonalen Komplexen. CC8 (19,3%), CC45 (17,7%) und CC30 (12,6%) dominierten. MRSA waren selten nachweisbar. Die sieben MRSA-Isolate gehörten zu den lokal dominierenden Stämmen (Rhein-Hessen, Süddeutscher, Barnimer und Berliner Epidemiestamm sowie Europäischer caMRSA-Klon). Die Populationsstruktur der klinischen Isolate und die Häufigkeiten der untersuchten Virulenz- und Adhäsionsfaktoren entsprachen weitestgehend Isolaten von asymptomatischen Trägern, die in einer früheren Studie bestimmt wurden (Molecular epidemiology of Staphylococcus aureus in asymptomatic carriers; Monecke, Lüdicke, Slickers, Ehricht; Eur J Clin Microbiol Infect Dis. 2009). Es konnte kein molekularer Marker identifiziert werden, der allein für eine Risikostratifizierung eingesetzt werden kann. Das Gen für Staphylokinase (sak) war jedoch bei den klinischen Isolaten (, Staphylococcus aureus is asymptomatically carried by approximately 25% of a normal population. It is also one of the most important causes of osteomyelitis and infections of orthopedic implants such as total hip or knee replacements. Such infections usually lead to complicated and risky surgical procedures as well as to long-term antibiotic treatment. In the present work, S. aureus isolates from osteomyelitis or implant infections were to be characterised. The aim of the study was to prove whether certain strains were overrepresented among patient isolates, and whether the presence of certain virulence factors might correlate with these infections. DNA arrays where used which facilitate to screen for all relevant virulence factors within a single experiment and which allow typing by obtaining a genetic fingerprint of the examined isolate. By this method, it was also possible to assign isolates to phylogenetic clusters, so-called clonal complexes. 119 clinical isolates were characterised in this way. They belonged to 20 different clonal complexes (CC). CC8 (19.3%), CC45 (17.7%) and CC30 (12.6%) dominated. MRSA were rarely detected. The seven MRSA isolates belonged to locally predominant epidemic strains (ST5-MRSA-II, ST228-MRSA-I, ST22-MRSA-IV, ST45-MRSA-IV and ST80-MRSA-IV). The population structure of the clinical isolates and the relative abundances of the examined virulence and adhesion factors corresponded largely to isolates from asymptomatic carriers, which has been examined in an earlier study (Molecular epidemiology of Staphylococcus aureus in asymptomatic carriers; Monecke, Lüdicke, Slickers, Ehricht; Eur J Clin Microbiol Infect Dis. 2009). No molecular maker was detected which could alone be used for risk assessment. The gene for staphylokinase (sak) was clearly more common among clinical isolates (90.8%) than in isolates from asymptomatic carriers (71.6%). Some other genes were also found to be more common in patient isolates, but were very rare so that a

Staphylococcus aureus kann bei etwa 25% der gesunden Normalbevölkerung nachgewiesen werden, ohne Symptome zu verursachen. Dieser Keim ist jedoch auch einer der wichtigsten Erreger bei Osteomyelitis und Infektionen von orthopädischen Implantaten wie z. B. von künstlichen Knie- oder Hüftgelenken. Diese Infektionen führen meist zu aufwendigen und risikobehafteten operativen Eingriffen sowie zu einer langfristigen Antibiotikagabe. In der vorliegenden Arbeit sollten S. aureus-Isolate charakterisiert werden, die aus Osteomyelitisherden oder infizierten orthopädischen Implantaten gewonnen wurden. Ziel war es, die Isolate daraufhin zu untersuchen, ob bestimmte Stämme dominieren und ob das Vorhandensein bestimmter Virulenzfaktoren mit einem besonderen Risiko für solche Infektionen korreliert. Für diese Untersuchungen wurden DNA-Arrays eingesetzt, welche es ermöglichen, alle relevanten Virulenzfaktoren in einem Experiment nachzuweisen, einen „genetischen Fingerabdruck“ zu erheben und die Isolate so Verwandtschaftsgruppen (klonalen Komplexen, CC) zuzuordnen. Insgesamt wurden 119 klinische Isolate charakterisiert. Sie gehörten zu 20 verschiedenen klonalen Komplexen. CC8 (19,3%), CC45 (17,7%) und CC30 (12,6%) dominierten. MRSA waren selten nachweisbar. Die sieben MRSA-Isolate gehörten zu den lokal dominierenden Stämmen (Rhein-Hessen, Süddeutscher, Barnimer und Berliner Epidemiestamm sowie Europäischer caMRSA-Klon). Die Populationsstruktur der klinischen Isolate und die Häufigkeiten der untersuchten Virulenz- und Adhäsionsfaktoren entsprachen weitestgehend Isolaten von asymptomatischen Trägern, die in einer früheren Studie bestimmt wurden (Molecular epidemiology of Staphylococcus aureus in asymptomatic carriers; Monecke, Lüdicke, Slickers, Ehricht; Eur J Clin Microbiol Infect Dis. 2009). Es konnte kein molekularer Marker identifiziert werden, der allein für eine Risikostratifizierung eingesetzt werden kann. Das Gen für Staphylokinase (sak) war jedoch bei den klinischen Isolaten (, Staphylococcus aureus is asymptomatically carried by approximately 25% of a normal population. It is also one of the most important causes of osteomyelitis and infections of orthopedic implants such as total hip or knee replacements. Such infections usually lead to complicated and risky surgical procedures as well as to long-term antibiotic treatment. In the present work, S. aureus isolates from osteomyelitis or implant infections were to be characterised. The aim of the study was to prove whether certain strains were overrepresented among patient isolates, and whether the presence of certain virulence factors might correlate with these infections. DNA arrays where used which facilitate to screen for all relevant virulence factors within a single experiment and which allow typing by obtaining a genetic fingerprint of the examined isolate. By this method, it was also possible to assign isolates to phylogenetic clusters, so-called clonal complexes. 119 clinical isolates were characterised in this way. They belonged to 20 different clonal complexes (CC). CC8 (19.3%), CC45 (17.7%) and CC30 (12.6%) dominated. MRSA were rarely detected. The seven MRSA isolates belonged to locally predominant epidemic strains (ST5-MRSA-II, ST228-MRSA-I, ST22-MRSA-IV, ST45-MRSA-IV and ST80-MRSA-IV). The population structure of the clinical isolates and the relative abundances of the examined virulence and adhesion factors corresponded largely to isolates from asymptomatic carriers, which has been examined in an earlier study (Molecular epidemiology of Staphylococcus aureus in asymptomatic carriers; Monecke, Lüdicke, Slickers, Ehricht; Eur J Clin Microbiol Infect Dis. 2009). No molecular maker was detected which could alone be used for risk assessment. The gene for staphylokinase (sak) was clearly more common among clinical isolates (90.8%) than in isolates from asymptomatic carriers (71.6%). Some other genes were also found to be more common in patient isolates, but were very rare so that a

Staphylococcus aureus kann bei etwa 25% der gesunden Normalbevölkerung nachgewiesen werden, ohne Symptome zu verursachen. Dieser Keim ist jedoch auch einer der wichtigsten Erreger bei Osteomyelitis und Infektionen von orthopädischen Implantaten wie z. B. von künstlichen Knie- oder Hüftgelenken. Diese Infektionen führen meist zu aufwendigen und risikobehafteten operativen Eingriffen sowie zu einer langfristigen Antibiotikagabe. In der vorliegenden Arbeit sollten S. aureus-Isolate charakterisiert werden, die aus Osteomyelitisherden oder infizierten orthopädischen Implantaten gewonnen wurden. Ziel war es, die Isolate daraufhin zu untersuchen, ob bestimmte Stämme dominieren und ob das Vorhandensein bestimmter Virulenzfaktoren mit einem besonderen Risiko für solche Infektionen korreliert. Für diese Untersuchungen wurden DNA-Arrays eingesetzt, welche es ermöglichen, alle relevanten Virulenzfaktoren in einem Experiment nachzuweisen, einen „genetischen Fingerabdruck“ zu erheben und die Isolate so Verwandtschaftsgruppen (klonalen Komplexen, CC) zuzuordnen. Insgesamt wurden 119 klinische Isolate charakterisiert. Sie gehörten zu 20 verschiedenen klonalen Komplexen. CC8 (19,3%), CC45 (17,7%) und CC30 (12,6%) dominierten. MRSA waren selten nachweisbar. Die sieben MRSA-Isolate gehörten zu den lokal dominierenden Stämmen (Rhein-Hessen, Süddeutscher, Barnimer und Berliner Epidemiestamm sowie Europäischer caMRSA-Klon). Die Populationsstruktur der klinischen Isolate und die Häufigkeiten der untersuchten Virulenz- und Adhäsionsfaktoren entsprachen weitestgehend Isolaten von asymptomatischen Trägern, die in einer früheren Studie bestimmt wurden (Molecular epidemiology of Staphylococcus aureus in asymptomatic carriers; Monecke, Lüdicke, Slickers, Ehricht; Eur J Clin Microbiol Infect Dis. 2009). Es konnte kein molekularer Marker identifiziert werden, der allein für eine Risikostratifizierung eingesetzt werden kann. Das Gen für Staphylokinase (sak) war jedoch bei den klinischen Isolaten (, Staphylococcus aureus is asymptomatically carried by approximately 25% of a normal population. It is also one of the most important causes of osteomyelitis and infections of orthopedic implants such as total hip or knee replacements. Such infections usually lead to complicated and risky surgical procedures as well as to long-term antibiotic treatment. In the present work, S. aureus isolates from osteomyelitis or implant infections were to be characterised. The aim of the study was to prove whether certain strains were overrepresented among patient isolates, and whether the presence of certain virulence factors might correlate with these infections. DNA arrays where used which facilitate to screen for all relevant virulence factors within a single experiment and which allow typing by obtaining a genetic fingerprint of the examined isolate. By this method, it was also possible to assign isolates to phylogenetic clusters, so-called clonal complexes. 119 clinical isolates were characterised in this way. They belonged to 20 different clonal complexes (CC). CC8 (19.3%), CC45 (17.7%) and CC30 (12.6%) dominated. MRSA were rarely detected. The seven MRSA isolates belonged to locally predominant epidemic strains (ST5-MRSA-II, ST228-MRSA-I, ST22-MRSA-IV, ST45-MRSA-IV and ST80-MRSA-IV). The population structure of the clinical isolates and the relative abundances of the examined virulence and adhesion factors corresponded largely to isolates from asymptomatic carriers, which has been examined in an earlier study (Molecular epidemiology of Staphylococcus aureus in asymptomatic carriers; Monecke, Lüdicke, Slickers, Ehricht; Eur J Clin Microbiol Infect Dis. 2009). No molecular maker was detected which could alone be used for risk assessment. The gene for staphylokinase (sak) was clearly more common among clinical isolates (90.8%) than in isolates from asymptomatic carriers (71.6%). Some other genes were also found to be more common in patient isolates, but were very rare so that a