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Functionality of the accessory gene regulator (agr) quorum sensing system is an important factor promoting either acute or chronic infections by the notorious opportunistic human and veterinary pathogen Staphylococcus aureus. Spontaneous alterations of the agr system are known to frequently occur in human healthcare-associated S. aureus lineages. However, data on agr integrity and function are sparse regarding other major clonal lineages. Here we report on the agr system functionality and activity level in mecC-carrying methicillin resistant S. aureus (MRSA) of various animal origins (n = 33) obtained in Europe as well as in closely related human isolates (n = 12). Whole genome analysis assigned all isolates to four clonal complexes (CC) with distinct agr types (CC599 agr I, CC49 agr II, CC130 agr III and CC1943 agr IV). Agr functionality was assessed by a combination of phenotypic assays and proteome analysis. In each CC, isolates with varying agr activity levels were detected, including the presence of completely non-functional variants. Genomic comparison of the agr I–IV encoding regions associated these phenotypic differences with variations in the agrA and agrC genes. The genomic changes were detected independently in divergent lineages, suggesting that agr variation might foster viability and adaptation of emerging MRSA lineages to distinct ecological niches.

X.D. was funded by a grant from the National Institute for Health Research (NIHR) Health Protection Research Unit in Genomics and Enabling Data (no. NIHR200892). M.A.H. was supported by grants from the Medical Research Council (nos. G1001787/1, MR/N002660/1 and MR/P007201/1) and the Economic and Social Research Council (no. ES/S000186/1). E.M.H. was supported by a UK Research and Innovation (UKRI) Fellowship (no. MR/S00291X/1). The discovery of antibiotics more than 80 years ago has led to considerable improvements in human and animal health. Although antibiotic resistance in environmental bacteria is ancient, resistance in human pathogens is thought to be a modern phenomenon that is driven by the clinical use of antibiotics1. Here we show that particular lineages of methicillin-resistant Staphylococcus aureus—a notorious human pathogen—appeared in European hedgehogs in the pre-antibiotic era. Subsequently, these lineages spread within the local hedgehog populations and between hedgehogs and secondary hosts, including livestock and humans. We also demonstrate that the hedgehog dermatophyte Trichophyton erinacei produces two β-lactam antibiotics that provide a natural selective environment in which methicillin-resistant S. aureus isolates have an advantage over susceptible isolates. Together, these results suggest that methicillin resistance emerged in the pre-antibiotic era as a co-evolutionary adaptation of S. aureus to the colonization of dermatophyte-infected hedgehogs. The evolution of clinically relevant antibiotic-resistance genes in wild animals and the connectivity of natural, agricultural and human ecosystems demonstrate that the use of a One Health approach is critical for our understanding and management of antibiotic resistance, which is one of the biggest threats to global health, food security and development. Publisher PDF

Das Handbuch Qualitatsmanagement der Berliner Jugendfreizeiteinrichtungen (QM-Handbuch) ist ein fur alle offentlich geforderten Einrichtungen der OKJA in Berlin verbindlich anzuwendendes Instrument der Selbstevaluation. Es beinhaltet Ziele, Handlungsorientierungen und Beschreibungen typischer Angebotsschwerpunkte der OKJA. Seine Inhalte entstanden in einem dialogischen Prozess zwischen padagogischen Fachkraften, den Tragern, den Jugendamtern und der fur Jugend zustandigen Senatsverwaltung (Landesjugendamt) und werden regelmasig aktualisiert. Das QM-Handbuch dokumentiert das fachliche Selbstverstandnis der OKJA und gilt fur die Jugendpolitik als Beleg seiner fachlichen Qualitat. Der Beitrag enthalt grundlegende Ausfuhrungen zum methodischen Ansatz und zum Entstehungshintergrund sowie Beispiele fur die Anwendung im Rahmen von Selbstevaluation.

Wie konnen Studierende der Sozialen Arbeit und der Sozialpadagogik fur die Herausbildung einer professionellen Wahrnehmung ihrer zukunftigen beruflichen Rolle durch Supervision unterstutzt werden? Die Ausbildung muss Raum fur die Reflexion der Beziehung zwischen Klienten und Helfer/innen und fur die Personlichkeitsbildung der Studierenden bieten. Hierfur werden methodische Settings fur die Ausbildung an Hochschulen diskutiert. In welchem Verhaltnis stehen Wissensvermittlung, Anleitung und Supervision? Der Beitrag pladiert fur Supervision als Bildungsangebot und benennt die Qualifikationen fur Supervisoren.

There are only few data on the persistence and transmission of methicillin resistant Staphylococcus aureus (MRSA) of the clonal complex (CC) 398 among veterinarians and their household members. We therefore investigated the long-term colonisation with MRSA CC398 among participants of veterinary conferences in Germany in 2008/2009 and their household members. Forty-five initially MRSA CC398 positive and 180 initially MRSA CC398 negative conference participants were included in a longitudinal study. These persons and their household members were tested for nasal colonisation in 2011, 2012 and 2014. Of 31 continuously tested and initially MRSA CC398 positive participants only 8 (26%) were colonized with MRSA CC398 at all 4 time points, 4 (13%) of them consistently with the same spa type. Among initially MRSA CC398 negative participants, 13 (7%) were tested MRSA CC398 positive at least once during the follow-up period. Data for household members at least at one time point were available for 185 households. Of these 21 (11%) households had one or more household member who tested positive for MRSA CC398 at least once. The odds of household members to be MRSA CC398 positive was 12 times higher (95% confidence interval 4–37) when the conference participant tested MRSA CC398 positive in 2008/2009. This association remained strong when household members working in veterinary medicine or livestock farms were excluded. In summary, these data suggest that colonisation with MRSA CC398 is partially transient and that household members of MRSA CC398 colonized persons are at an increased risk of colonisation with MRSA CC398.

Livestock-associated methicillin-resistant Staphylococcus aureus (LA-MRSA) is widely disseminated as a nasal colonizer of conventionally raised livestock and of humans subjected to occupational exposure. Reports on contamination of raw meat raise the question as to whether occupationally exposed food handlers are at particular risk of nasal colonization by LA-MRSA. Here, we report the results from a cross-sectional study on nasal S. aureus/MRSA colonization of butchers, meat sellers, and cooks in Germany. We sampled 286 butchers and meat sellers in 26 butcheries and 319 cooks handling meat in 16 professional canteen kitchens. Swabs were processed on both blood agar plates and MRSA-selective plates. MRSA were confirmed by PCR for mec genes and by broth microdilution. All isolates were subjected to molecular typing. PCR for markers useful to differentiate human-adapted and animal-adapted subpopulations was performed due to the presence of clonal complexes known to occur in both livestock and humans (CC5, CC7, CC8, CC9, and CC398). Only two participants (0.33%) were colonized by MRSA (Hospital-associated MRSA ST22). Nasal colonization by methicillin-susceptible S. aureus (MSSA) was detected in 16.6% of cooks and in 26.2% of butchers and meat sellers. Among 16 of the isolates attributed to CC7, three were negative for the immune evasion gene cluster, suggesting an animal origin. Isolates attributed to CC5, CC8, and CC398 were negative for markers typical of animal-adapted subpopulations. The occupational handling of raw meat and raw meat products was not associated with nasal colonization by LA-MRSA.

Population-based studies on Staphylococcus aureus nasal colonization are scarce. We examined the prevalence, resistance, and molecular diversity of S. aureus in the general population in Northeast Germany. Nasal swabs were obtained from 3,891 adults in the large-scale population-based Study of Health in Pomerania (SHIP-TREND). Isolates were characterized using spa genotyping, as well as antibiotic resistance and virulence gene profiling. We observed an S. aureus prevalence of 27.2%. Nasal S. aureus carriage was associated with male sex and inversely correlated with age. Methicillin-resistant S. aureus (MRSA) accounted for 0.95% of the colonizing S. aureus strains. MRSA carriage was associated with frequent visits to hospitals, nursing homes, or retirement homes within the previous 24 months. All MRSA strains were resistant to multiple antibiotics. Most MRSA isolates belonged to the pandemic European hospital-acquired MRSA sequence type 22 (HA-MRSA-ST22) lineage. We also detected one livestock-associated MRSA ST398 (LA-MRSA-ST398) isolate, as well as six livestock-associated methicillin-susceptible S. aureus (LA-MSSA) isolates (clonal complex 1 [CC1], CC97, and CC398). spa typing revealed a diverse but also highly clonal S. aureus population structure. We identified a total of 357 spa types, which were grouped into 30 CCs or sequence types. The major seven CCs (CC30, CC45, CC15, CC8, CC7, CC22, and CC25) included 75% of all isolates. Virulence gene patterns were strongly linked to the clonal background. In conclusion, MSSA and MRSA prevalences and the molecular diversity of S. aureus in Northeast Germany are consistent with those of other European countries. The detection of HA-MRSA and LA-MRSA within the general population indicates possible transmission from hospitals and livestock, respectively, and should be closely monitored.

We investigated the evolution and epidemiology of a novel livestock-associated methicillin-resistant Staphylococcus aureus strain, which colonizes and infects urban-dwelling Danes even without a Danish animal reservoir. Genetic evidence suggests both poultry and human adaptation, with poultry meat implicated as a probable source.

The proteogenomes and secretomes of dominant human and zoonotic S. aureus lineages CC8, CC22 and CC398 were compared revealing genomic and regulatory differences in the secretion of 869 proteins. In the core secretome, 101 secreted or cell surface anchored virulence factors contribute with 82.4% to total secretome abundance. CC398 isolates showed higher secretion of α- and ß-hemolysins and lower secretion of surface proteins resulting in strong hemolysis and decreased biofilm formation because of lower SigB activity compared to human-specific CC8 and CC22., Graphical Abstract Highlights Proteogenomics and secretome comparison of human and zoonotic Staphylococcus aureus lineages.869 secreted proteins identified in eight S. aureus isolates of CC8, CC22 and CC398.CC398 lower secretion of surface proteins and higher secretion of hemolysins and exoenzymes.Regulatory differences in the secretomes could be linked to lower SigB activity in CC398., The spread of methicillin-resistant Staphylococcus aureus (MRSA) in the community, hospitals and in livestock is mediated by highly diverse virulence factors that include secreted toxins, superantigens, enzymes and surface-associated adhesins allowing host adaptation and colonization. Here, we combined proteogenomics, secretome and phenotype analyses to compare the secreted virulence factors in selected S. aureus isolates of the dominant human- and livestock-associated genetic lineages CC8, CC22, and CC398. The proteogenomic comparison revealed 2181 core genes and 1306 accessory genes in 18 S. aureus isolates reflecting the high genome diversity. Using secretome analysis, we identified 869 secreted proteins with 538 commons in eight isolates of CC8, CC22, and CC398. These include 64 predicted extracellular and 37 cell surface proteins that account for 82.4% of total secretome abundance. Among the top 10 most abundantly secreted virulence factors are the major autolysins (Atl, IsaA, Sle1, SAUPAN006375000), lipases and lipoteichoic acid hydrolases (Lip, Geh, LtaS), cytolytic toxins (Hla, Hlb, PSMβ1) and proteases (SspB). The CC398 isolates showed lower secretion of cell wall proteins, but higher secretion of α- and β-hemolysins (Hla, Hlb) which correlated with an increased Agr activity and strong hemolysis. CC398 strains were further characterized by lower biofilm formation and staphyloxanthin levels because of decreased SigB activity. Overall, comparative secretome analyses revealed CC8- or CC22-specific enterotoxin and Spl protease secretion as well as Agr- and SigB-controlled differences in exotoxin and surface protein secretion between human-specific and zoonotic lineages of S. aureus.

The spread of methicillin-resistant Staphylococcus aureus (MRSA) in the community, hospitals and in livestock is mediated by highly diverse virulence factors that include secreted toxins, superantigens, enzymes and surface-associated adhesins allowing host adaptation and colonization. Here, we combined proteogenomics, secretome and phenotype analyses to compare the secreted virulence factors in selected S. aureus isolates of the dominant human- and livestock-associated genetic lineages CC8, CC22, and CC398. The proteogenomic comparison revealed 2181 core genes and 1306 accessory genes in 18 S. aureus isolates reflecting the high genome diversity. Using secretome analysis, we identified 869 secreted proteins with 538 commons in eight isolates of CC8, CC22, and CC398. These include 64 predicted extracellular and 37 cell surface proteins that account for 82.4% of total secretome abundance. Among the top 10 most abundantly secreted virulence factors are the major autolysins (Atl, IsaA, Sle1, SAUPAN006375000), lipases and lipoteichoic acid hydrolases (Lip, Geh, LtaS), cytolytic toxins (Hla, Hlb, PSMβ1) and proteases (SspB). The CC398 isolates showed lower secretion of cell wall proteins, but higher secretion of α- and β-hemolysins (Hla, Hlb) which correlated with an increased Agr activity and strong hemolysis. CC398 strains were further characterized by lower biofilm formation and staphyloxanthin levels because of decreased SigB activity. Overall, comparative secretome analyses revealed CC8- or CC22-specific enterotoxin and Spl protease secretion as well as Agr- and SigB-controlled differences in exotoxin and surface protein secretion between human-specific and zoonotic lineages of S. aureus.

During the past 25 years an increase in the prevalence of methicillin-resistant Staphylococcus aureus (HA-MRSA) was recorded worldwide. Additionally, MRSA infections may occur outside and independent of hospitals, caused by community associated MRSA (CA-MRSA). In Germany, we found that at least 10% of these sporadic infections are due to livestock-associated MRSA (LA-MRSA), which is initially associated with livestock. The majority of these MRSA cases are attributed to clonal complex CC398. LA-MRSA CC398 colonizes the animals asymptomatically in about half of conventional pig farms. For about 77%–86% of humans with occupational exposure to pigs, nasal carriage has been reported; it can be lost when exposure is interrupted. Among family members living at the same farms, only 4%–5% are colonized. Spread beyond this group of people is less frequent. The prevalence of LA-MRSA in livestock seems to be influenced by farm size, farming systems, usage of disinfectants, and in-feed zinc. LA-MRSA CC398 is able to cause the same kind of infections in humans as S. aureus and MRSA in general. It can be introduced to hospitals and cause nosocomial infections such as postoperative surgical site infections, ventilator associated pneumonia, septicemia, and infections after joint replacement. For this reason, screening for MRSA colonization at hospital admittance is recommended for farmers and veterinarians with livestock contacts. Intrahospital dissemination, typical for HA-MRSA in the absence of sufficient hygiene, has only rarely been observed for LA-MRSA to date. The proportion of LA-MRSA among all MRSA from nosocomial infections is about 3% across Germany. In geographical areas with a comparatively high density of conventional farms, LA-MRSA accounts for up to 10% of MRSA from septicemia and 15% of MRSA from wound infections. As known from comparative genome analysis, LA-MRSA has evolved from human-adapted methicillin-susceptible S. aureus, and the jump to livestock was obviously associated with several genetic changes. Reversion of the genetic changes and readaptation to humans bears a potential health risk and requires tight surveillance. Although most LA-MRSA (>80%) is resistant to several antibiotics, there are still sufficient treatment options.

Methicillin-resistant Staphylococcus aureus (MRSA) is a major cause of bacteraemia. We aimed to obtain a complete picture of severe MRSA infections by characterizing all MRSA isolates from bloodstream infections in the largest German federal state (North Rhine-Westphalia, 18 million inhabitants) using S. aureus protein A (spa) sequence-typing and antimicrobial susceptibility testing. MRSA isolates (n = 1952) were collected prospectively (2011–2013) and spa-typed. Among 181 different spa types, t003 (n = 746 isolates; 38.2%) and t032 (n = 594; 30.4%) were predominant. Analysis of the geographical occurrence of spa clonal complexes (spa-CCs) and spa types revealed divergent distribution between federal state districts for spa-CCs 003 (p

The immune evasion gene cluster (IEC) is typical for Staphylococcus aureus isolated from humans but is usually absent in S. aureus isolated from animals. Previous studies have shown that methicillin resistant S. aureus (MRSA) CC398 obviously lost the IEC when evolving as livestock-associated MRSA from a human-adapted, methicillin-susceptible ancestor. This study aimed to look for the presence of IEC in MRSA from pigs and horses as well as from the colonization of humans with occupational animal contact and from infections in humans. For comparison, methicillin susceptible S. aureus (MSSA) isolates from infections in humans were included. We did not detect the IEC among 94 isolates from the nasal colonization of pigs; however, the IEC was found in 6 of 61 isolates from nosocomial infections in horses. MRSA CC398 isolates from the nasal colonization of 138 pig farmers were negative for the IEC. It was detected, however, in 4 of 69 veterinarians treating horses. Among 99 epidemiologically unrelated MRSA isolates attributed to CC398 originating from infections in humans, 19 were positive for the IEC. Only three of these isolates which also contained luk-PV were attributed to the ancestral, human-adapted subpopulation of CC398 by means of PCR for detection of canonical SNPs. A considerable proportion of LA-MRSA CC398 attributed to the animal subpopulation and originating from infections in humans had acquired the IEC; this acquisition is, however, obviously not a prerequisite to the capacity of LA-MRSA CC398 to cause infections in this host. Among 15 MSSA CC398 isolates from infections in humans, 11 contained the IEC, and of these, two were attributed to the animal subpopulation. Six isolates containing both the IEC and luk-PV were attributed to the ancestral, human subpopulation. Re-acquisition of the IEC by LA-MRSA CC398 suggests readaptation to the human host. In epidemiological surveillance, discrimination from the ancestral human subpopulation is important.

To elucidate the evolutionary history of Staphylococcus aureus clonal complex (CC) 8, which encompasses several globally distributed epidemic lineages, including hospital-associated methicillin-resistant S. aureus (MRSA) and the highly prevalent community-associated MRSA clone USA300.We reconstructed the phylogeny of S. aureus CC8 by mutation discovery at 112 genetic housekeeping loci from each of 174 isolates, sampled on five continents between 1957 and 2008. The distribution of antimicrobial resistance traits and of diverse mobile genetic elements was investigated in relation to the isolates' phylogeny.Our analyses revealed the existence of nine phylogenetic clades within CC8. We identified at least eight independent events of methicillin resistance acquisition in CC8 and dated the origin of a methicillin-resistant progenitor of the notorious USA300 clone to the mid-1970s. Of the S. aureus isolates in our collection, 88% carried plasmidic rep gene sequences, with up to five different rep genes in individual isolates and a total of eight rep families. Mapping the plasmid content onto the isolates' phylogeny illustrated the stable carriage over decades of some plasmids and the more volatile nature of others. Strikingly, we observed trends of increasing antibiotic resistance during the evolution of several lineages, including USA300.We propose a model for the evolution of S. aureus CC8, involving a split into at least nine phylogenetic lineages and a subsequent series of acquisitions and losses of mobile genetic elements that carry diverse virulence and antimicrobial resistance traits. The evolution of MRSA USA300 towards resistance to additional antibiotic classes is of major concern.

Staphylococcus aureus sequence type (ST)398, which is commonly found as a colonization strain in pig farming, is emerging more frequently as the cause of human infections. In this study, we analysed ST398 of porcine and human origin at the genetic, protein and immunogenic levels. Although genetic analysis of the genes encoding the major virulence factors revealed the presence of the same genes in all strains studied, the results demonstrated spa type crossing alterations in adhesion abilities in addition to a strongly enhanced lysis activity directly linked to impaired clearance attributable to polymorphonuclear leukocytes (PMNs). This change in virulence pattern indicates high heterogenicity in the ST398 pool that is not based on a different genetic make-up, but probably on variations in the genetic regulation systems. These modifications, which are tightly connected to pathogenicity, cannot be detected by conventional diagnostic methods.

Klebsiella pneumoniae carbapenemases (KPCs) are considered a serious threat to antibiotic therapy, as they confer resistance to carbapenems, which are used to treat extended-spectrum beta-lactamase (ESBL)-producing bacteria. Here, we describe the development and evaluation of a DNA microarray for the detection and genotyping of KPC genes ( bla KPC ) within a 5-h period. To test the whole assay procedure (DNA extraction plus a DNA microarray assay) directly from clinical specimens, we compared two commercial DNA extraction kits (the QIAprep Spin miniprep kit [Qiagen] and the urine bacterial DNA isolation kit [Norgen]) for the direct DNA extraction from urine samples (dilution series spiked in human urine). Reliable single nucleotide polymorphism (SNP) typing was demonstrated using 1 × 10 5 CFU/ml urine for Escherichia coli (Qiagen and Norgen) and 80 CFU/ml urine, on average, for K. pneumoniae (Norgen). This study presents, for the first time, the combination of a new KPC microarray with commercial sample preparation for detecting and genotyping microbial pathogens directly from clinical specimens; this paves the way toward tests providing epidemiological and diagnostic data, enabling better antimicrobial stewardship.

Staphylococcus aureus (S. aureus) is a Gram-positive opportunistic pathogen that colonizes frequently and asymptomatically the anterior nares of humans and animals. It can cause different kinds of infections and is considered to be an important nosocomial pathogen. Nasal carriage of S. aureus can be permanent or intermittent and may build the reservoir for autogenous infections and cross-transmission to other individuals. Most of the studies on the epidemiology of S. aureus performed in the past were focused on the emergence and dissemination of methicillin-resistant Staphylococcus aureus (MRSA) in healthcare settings. There are, however, a number of more recent epidemiological studies have aimed at analysing carriage patterns over time in the community settings providing new insights on risk factors for colonization and important data for the development of strategies to prevent infections. This chapter aims to give a review of current epidemiological studies on S. aureus carriage patterns in the general community and put them into perspective with recent, yet unpublished, investigations on the S. aureus epidemiology in the general population in northern Germany.

The implementation of routine whole-genome sequencing (WGS) promises to transform our ability to monitor the emergence and spread of bacterial pathogens. Here we combined WGS data from 308 invasive Staphylococcus aureus isolates corresponding to a pan-European population snapshot, with epidemiological and resistance data. Geospatial visualization of the data is made possible by a generic software tool designed for public health purposes that is available at the project URL (http:// www.microreact.org/project/EkUvg9uY?tt=rc). Our analysis demonstrates that high-risk clones can be identified on the basis of population level properties such as clonal relatedness, abundance, and spatial structuring and by inferring virulence and resistance properties on the basis of gene content. We also show that in silico predictions of antibiotic resistance profiles are at least as reliable as phenotypic testing. We argue that this work provides a comprehensive road map illustrating the three vital components for future molecular epidemiological surveillance: (i) large-scale structured surveys, (ii) WGS, and (iii) communityoriented database infrastructure and analysis tools. IMPORTANCE The spread of antibiotic-resistant bacteria is a public health emergency of global concern, threatening medical intervention at every level of health care delivery. Several recent studies have demonstrated the promise of routine wholegenome sequencing (WGS) of bacterial pathogens for epidemiological surveillance, outbreak detection, and infection control. However, as this technology becomes more widely adopted, the key challenges of generating representative national and international data sets and the development of bioinformatic tools to manage and interpret the data become increasingly pertinent. This study provides a road map for the integration of WGS data into routine pathogen surveillance. We emphasize the importance of large-scale routine surveys to provide the population context for more targeted or localized investigation and the development of open-access bioinformatic tools to provide the means to combine and compare independently generated data with publicly available data sets., 0, SCOPUS: ar.j, info:eu-repo/semantics/published

Staphylococcus aureus (S. aureus) is a Gram-positive opportunistic pathogen that colonizes frequently and asymptomatically the anterior nares of humans and animals. It can cause different kinds of infections and is considered to be an important nosocomial pathogen. Nasal carriage of S. aureus can be permanent or intermittent and may build the reservoir for autogenous infections and cross-transmission to other individuals. Most of the studies on the epidemiology of S. aureus performed in the past were focused on the emergence and dissemination of methicillin-resistant Staphylococcus aureus (MRSA) in healthcare settings. There are, however, a number of more recent epidemiological studies have aimed at analysing carriage patterns over time in the community settings providing new insights on risk factors for colonization and important data for the development of strategies to prevent infections. This chapter aims to give a review of current epidemiological studies on S. aureus carriage patterns in the general community and put them into perspective with recent, yet unpublished, investigations on the S. aureus epidemiology in the general population in northern Germany.

Enterococci (mainly E. faecalis, E. faecium) are important nosocomial pathogens predominantly affecting older and/or immunocompromised patients. The bacteria possess a broad spectrum of intrinsic and acquired antibiotic resistance properties. Among these, the transferrable glycopeptide resistance of the vanA and vanB genotypes in vancomycin-resistant enterococci (VRE; reservoir: E. faecium) as well as resistance to last resort antibiotics (e.g. linezolid and tigecycline) are of special concern. Enterococci (including VRE) are easily transferred in hospitals; however, colonizations are far more frequent than infections. Resistance frequencies for vancomycin in clinical E. faecium isolates have remained at a relatively constant level of 8-15% (but with local or regional variations) in recent years whereas frequencies for teicoplanin resistance have shown a slight decrease. Glycopeptide resistance trends correlate with a spread of hospital-associated E. faecium strains carrying the vanA and, with rising frequency in recent years, the vanB gene cluster, the latter being associated with teicoplanin susceptibility. This increased occurrence of vanB-positive E. faecium strains may be caused by an increased use of antibiotics selecting enterococci and VRE as well as due to methodological reasons (e.g. reduced EUCAST MIC-breakpoints for glycopeptides; increased use and sensitive performance of chromogenic VRE agars, increased use of molecular diagnostic assays).

Frequencies of vanB -type Enterococcus faecium increased in Europe during the last years. VanB enterococci show various levels of vancomycin MICs even below the susceptible breakpoint challenging a reliable diagnostics. The performance of 3 chromogenic vancomycin-resistant enterococci (VRE) screening agars, 2 Etest® vancomycin protocols, and different microdilution methods to detect 129 clinical vanB E. faecium strains was investigated. Altogether, 112 (87%) were correctly identified as VanB-type Enterococcus by microdilution MICs. An Etest® macromethod protocol was more sensitive than the standard protocol while keeping sufficient specificity in identifying 15 vanA/vanB -negative strains. Three chromogenic VRE agars performed similarly with 121 (94%), 123 (95%), and 124 (96%) vanB isolates that grew on Brilliance TM VRE Agar, CHROMagar TM VRE, and chromID TM VRE agar, respectively. Using identical media and conditions, we did not identify different growth behaviour on agar and in broth. A few vanB strains showed growth of microcolonies inside the Etest® vancomycin inhibition zones, suggesting a VanB heteroresistance phenotype.

To determine the prevalence of extended-spectrum β-lactamase (ESBL) production in Enterobacteriaceae in retail chicken meat in Germany.A total of 399 chicken meat samples from nine supermarket chains, four organic food stores and one butcher's shop in two geographically distinct regions (Berlin and Greifswald) were screened for ESBL production using selective agar. Phenotypic ESBL isolates were tested for bla(TEM), bla(CTX-M) and bla(SHV) genes using PCR and DNA sequencing. Antibiotic coresistances were determined and strain typing was performed using PCR-based phylogenetic grouping and XbaI-PFGE.A total of 185 confirmed ESBL isolates were obtained from 175 samples (43.9%) from all tested sources. The majority of isolates were Escherichia coli producing ESBL types SHV-12 (n = 82), CTX-M-1 (n = 77) and TEM-52 (n = 16). No differences could be observed in the prevalence of ESBL producers between organic and conventional samples. 73.0% of the ESBL producers showed coresistance to tetracycline, 35.7% to co-trimoxazole and 7.6% to ciprofloxacin. Strain typing of selected E. coli isolates from Berlin revealed identical macrorestriction patterns for several isolates from samples taken from the same stores.This is the first comprehensive study from Germany showing a high prevalence of TEM-, CTX-M- and SHV-type ESBLs in Enterobacteriaceae isolated from retail chicken meat. The high rate of coresistance to different classes of antibiotics in the ESBL producers might reflect the common veterinary usage of these and related substances. There is an urgent need to further evaluate the role of poultry in the transmission of highly resistant ESBL-producing bacteria in humans.

While flagella-independent motility has long been described in representatives of the genus Acinetobacter, the mechanism of motility remains ambiguous. Acinetobacter baumannii, a nosocomial pathogen appearing increasingly multidrug-resistant, may profit from motility during infection or while persisting in the hospital environment. However, data on the frequency of motility skills among clinical A. baumannii isolates is scarce. We have screened a collection of 83 clinical A. baumannii isolates of different origin and found that, with the exception of one isolate, all were motile on wet surfaces albeit to varying degrees and exhibiting differing morphologies. Screening a collection of transposon mutants of strain ATCC 17978 for motility defects, we identified 2 akinetic mutants carrying transposon insertions in the dat and ddc gene, respectively. These neighbouring genes contribute to synthesis of 1,3-diaminopropane (DAP), a polyamine ubiquitously produced in Acinetobacter. Supplementing semi-solid media with DAP cured the motility defect of both mutants. HPLC analyses confirmed that DAP synthesis was abolished in ddc and dat mutants of different A. baumannii isolates and was re-established after genetic complementation. Both, the dat and ddc mutant of ATCC 17978 were attenuated in the Galleria mellonella caterpillar infection model. Taken together, surface-associated motility is a common trait of clinical A. baumannii isolates that requires DAP and may play a role in its virulence.

Nine carbapenem-resistant Enterobacteriaceae isolates collected from eight patients in five German hospitals were investigated. Six isolates produced the OXA-48 carbapenemase, and three isolates produced OXA-162, which is a point mutant form of OXA-48. Both carbapenemase genes were located on IncL/M-type conjugative plasmids. Insertion sequence IS 1999 (truncated or not by IS 1R ) was located upstream of the bla OXA-48 and bla OXA-162 genes in all of the isolates. Pulsed-field gel electrophoresis typing indicated the clonal transmission of an OXA-48-producing Klebsiella pneumoniae strain in two hospitals.

This study reports on the emergence of linezolid-resistant coagulase-negative staphylococci (CoNS) containing the multiresistance gene cfr in veal calves and pigs, as well as in humans exposed to these animals. CoNS (Staphylococcus auricularis, Staphylococcus cohnii, Staphylococcus lentus, Staphylococcus kloosii, Staphylococcus sciuri, Staphylococcus simulans), but not Staphylococcus aureus, carrying the gene cfr were detected in samples of 12 out of 52 calves at three farms which had a history of florfenicol use. Nasal swabs from 10 humans living on these farms were negative for cfr-carrying staphylococci. Nasal swabs taken from 142 calves at 16 farms in the same area that did not use florfenicol were also negative for cfr-carrying staphylococci. 14 cfr-carrying CoNS (S. kloosii, S. saprophyticus, S. simulans) were detected in three of eight conventional pig farms investigated. One of 12 humans living on these farms harboured a cfr-carrying S. cohnii. Among the nasal swabs taken from 169 veterinarians from all over Germany, four (2.3%) were positive for cfr-carrying CoNS (three S. epidermidis, one S. saprophyticus), and three (1.1%) of 263 contact persons of this group also harboured cfr-carrying CoNS (one S. epidermidis, two S. saprophyticus). In vitro conjugation of cfr by filter mating to S. aureus 8325-4 was possible for 10 of 34CoNS and the cfr gene was associated with plasmids of 38-40kb. Moreover, a total of 363 humans of a German municipal community were investigated for nasal carriage of cfr-carrying staphylococci to get an idea whether such isolates are disseminated as nasal colonizers in non-hospitalized humans in the community, were all negative.

Tracing the spatial spread of pathogens is a key objective of molecular infectious disease epidemiology. Accordingly, a wide range of genotyping approaches have been used to monitor the dissemination of Staphylococcus aureus strains, from localized outbreaks to global spread. We provide a critical review of available methods, revealing that molecular markers currently in use for typing S. aureus acquire changes so slowly that they monitor evolutionary change over timescales that are largely irrelevant to epidemiology. Moreover, the more variable markers frequently do not reflect the pathogen's evolutionary history and, hence, provide potentially misleading information about spread. More recent work has demonstrated that staphylococcal evolution proceeds sufficiently fast that the dynamics of S. aureus spatial spread can be elucidated at great detail on the basis of genome-wide single-nucleotide polymorphisms.

Fast and reliable diagnostics of vancomycin-resistant enterococci (VRE) is an important prerequisite for containing VRE transmission rates and controlling VRE outbreaks among hospital patients. The BD GeneOhm™ VanR Assay (Becton Dickinson Diagnostics, Erembodegem, Belgium) is a real-time polymerase chain reaction (PCR) assay for screening perianal/rectal samples for the presence of vanA or vanB genes that can be associated with VRE. A set of 51 reference strains (vanA-G genotypes) were correctly identified. Performance of the assay was evaluated and compared with culture-based methods and subsequent PCR analysis in 2 university hospitals with a different VRE prevalence. A total of 1786 samples were analyzed. With the use of the BD GeneOhm™ VanR Assay, 88 of 102 vanA-positive specimens, 62 of 67 vanB-positive specimens, 3 of 4 vanA- and vanB-positive specimens, and 1403 of 1613 negative specimens were correctly identified. The overall sensitivity was 93.1%; the specificity was 87.0% mainly due to false-positive vanB results. Results did not differ between study institutions.