Comparative effectiveness of ß-lactams for empirical treatment of methicillin-susceptible Staphylococcus aureus bacteraemia: a prospective cohort study.

OBJECTIVES: Standard once-daily dosing of ceftriaxone may not lead to adequate antibiotic exposure in all cases of Staphylococcus aureus bacteraemia (SAB). Therefore, we compared clinical effectiveness of empirical antibiotic treatment with flucloxacillin, cefuroxime and ceftriaxone in adult patients with MSSA bacteraemia. METHODS: We analysed data from the Improved Diagnostic Strategies in Staphylococcus aureus bacteraemia (IDISA) study, a multicentre prospective cohort study of adult patients with MSSA bacteraemia. Duration of bacteraemia and 30 day SAB-related mortality were compared between the three groups using multivariable mixed-effects Cox regression analyses. RESULTS: In total, 268 patients with MSSA bacteraemia were included in the analyses. Median duration of empirical antibiotic therapy was 3 (IQR 2-3) days in the total study population. Median duration of bacteraemia was 1.0 (IQR 1.0-3.0) day in the flucloxacillin, cefuroxime and ceftriaxone groups. In multivariable analyses, neither ceftriaxone nor cefuroxime was associated with increased duration of bacteraemia compared with flucloxacillin (HR 1.08, 95% CI 0.73-1.60 and HR 1.22, 95% CI 0.88-1.71). In multivariable analysis, neither cefuroxime nor ceftriaxone was associated with higher 30 day SAB-related mortality compared with flucloxacillin [subdistribution HR (sHR) 1.37, 95% CI 0.42-4.52 and sHR 1.93, 95% CI 0.67-5.60]. CONCLUSIONS: In this study, we could not demonstrate a difference in duration of bacteraemia and 30 day SAB-related mortality between patients with SAB empirically treated with flucloxacillin, cefuroxime or ceftriaxone. Since sample size was limited, it is possible the study was underpowered to find a clinically relevant effect.

Comparative genomics of ESBL-producing Escherichia coli (ESBL-Ec) reveals a similar distribution of the 10 most prevalent ESBL-Ec clones and ESBL genes among human community faecal and extra-intestinal infection isolates in the Netherlands (2014-17).

INTRODUCTION: The human gut microbiota is an important reservoir of ESBL-producing Escherichia coli (ESBL-Ec). Community surveillance studies of ESBL-Ec to monitor circulating clones and ESBL genes are logistically challenging and costly. OBJECTIVES: To evaluate if isolates obtained in routine clinical practice can be used as an alternative to monitor the distribution of clones and ESBL genes circulating in the community. METHODS: WGS was performed on 451 Dutch ESBL-Ec isolates (2014-17), including 162 community faeces and 289 urine and blood isolates. We compared proportions of 10 most frequently identified STs, PopPUNK-based sequence clusters (SCs) and ESBL gene subtypes and the degree of similarity using Czekanowski's proportional similarity index (PSI). RESULTS: Nine out of 10 most prevalent STs and SCs and 8/10 most prevalent ESBL genes in clinical ESBL-Ec were also the most common types in community faeces. The proportions of ST131 (39% versus 23%) and SC131 (40% versus 25%) were higher in clinical isolates than in community faeces (P < 0.01). Within ST131, H30Rx (C2) subclade was more prevalent among clinical isolates (55% versus 26%, P < 0.01). The proportion of ESBL gene blaCTX-M-1 was lower in clinical isolates (5% versus 18%, P < 0.01). Czekanowski's PSI confirmed that the differences in ESBL-Ec from community faeces and clinical isolates were limited. CONCLUSIONS: Distributions of the 10 most prevalent clones and ESBL genes from ESBL-Ec community gut colonization and extra-intestinal infection overlapped in majority, indicating that isolates from routine clinical practice could be used to monitor ESBL-Ec clones and ESBL genes in the community.

Faecal carriage, risk factors, acquisition and persistence of ESBL-producing Enterobacteriaceae in dogs and cats and co-carriage with humans belonging to the same household.

BACKGROUND: ESBL-producing Enterobacteriaceae (ESBL-E) are observed in many reservoirs. Pets might play an important role in the dissemination of ESBL-E to humans since they live closely together. OBJECTIVES: To identify prevalence, risk factors, molecular characteristics, persistence and acquisition of ESBL-E in dogs and cats, and co-carriage in human-pet pairs belonging to the same household. METHODS: In a nationwide study, one person per household was randomly invited to complete a questionnaire and to submit a faecal sample. Dog and cat owners were invited to also submit a faecal sample from their pet. Repeated sampling after 1 and 6 months was performed in a subset. ESBL-E were obtained through selective culture and characterized by WGS. Logistic regression analyses and random forest models were performed to identify risk factors. RESULTS: The prevalence of ESBL-E carriage in these cohorts was 3.8% (95% CI: 2.7%-5.4%) for human participants (n=550), 10.7% (95% CI: 8.3%-13.7%) for dogs (n=555) and 1.4% (95% CI: 0.5%-3.8%) for cats (n=285). Among animals, blaCTX-M-1 was most abundant, followed by blaCTX-M-15. In dogs, persistence of carriage was 57.1% at 1 month and 42.9% at 6 months. Eating raw meat [OR: 8.8, 95% CI: 4.7-16.4; population attributable risk (PAR): 46.5%, 95% CI: 41.3%-49.3%] and dry food (OR: 0.2, 95% CI: 0.1-0.5; PAR: 56.5%, 95% CI: 33.2%-66.6%) were predictors for ESBL-E carriage in dogs. Human-dog co-carriage was demonstrated in five households. Human-cat co-carriage was not observed. CONCLUSIONS: ESBL-E prevalence was higher in dogs than in humans and lowest in cats. The main risk factor for ESBL-E carriage was eating raw meat. Co-carriage in dogs and household members was uncommon.

Seasonality in carriage of extended-spectrum ß-lactamase-producing Escherichia coli and Klebsiella pneumoniae in the general population: a pooled analysis of nationwide cross-sectional studies.

Infections due to extended-spectrum ß-lactamase-producing Enterobacteriaceae (ESBL-E) are often preceded by asymptomatic carriage. Higher incidences in enteric infectious diseases during summer have been reported. Here, we assessed whether the presence of seasonality in intestinal ESBL-Escherichia coli/Klebsiella pneumoniae (ESBL-E/K) carriage in the general Dutch population exists. From 2014 to 2017, the faecal carriage of ESBL-E/K in healthy individuals was determined in three cross-sectional studies in the Netherlands, including 5985 subjects. Results were pooled to identify seasonal trends in prevalence (by month of sampling). Multivariate logistic regression analysis was used to calculate pooled odds ratios and 95% confidence intervals. Results were adjusted for age, sex, antibiotic use and travel. Overall prevalence of ESBL-E/K carriage was 4.3% (n = 260 ESBL-E/K-positive), with differences between months ranging from 2.6% to 7.4%. Compared to January, the monthly prevalence of ESBL-E carriage was highest in August (OR 1.88, 95% CI 1.02-3.49) and September (OR 2.25, 95% CI 1.30-3.89). The observed monthly differences in ESBL-E/K carriage rates suggest that there is seasonal variation in exposure to ESBL-E/K other than due to travelling and antibiotic use. This should be taken into account in designing future ESBL-E prevalence studies in temperate regions.

Intestinal carriage of ampicillin- and vancomycin-resistant Enterococcus faecium in humans, dogs and cats in the Netherlands.

Background: The prevalence of ampicillin- and/or vancomycin-resistant Enterococcus faecium (AREf and VREf) has increased in hospitalized patients in the Netherlands. Objectives: To quantify the prevalence, risk factors and co-carriage of AREf and VREf in humans, cats and dogs in the Dutch population. Methods: From 2014 to 2015, ∼2000 inhabitants of the Netherlands each month were randomly invited to complete a questionnaire and provide a faecal sample. Subjects owning pets were also asked to submit one dog or cat sample. Faecal samples were screened for AREf and VREf. The genetic relatedness of isolates was determined using core genome MLST. Logistic regression analysis was used to determine risk factors. Results: Of 25 365 subjects, 4721 (18.6%) completed the questionnaire and 1992 (42.2%) human, 277 dog and 118 cat samples were submitted. AREf was detected in 29 human (1.5%), 71 dog (25.6%) and 6 cat (5.1%) samples. VREf (vanA) was detected in one human and one dog. AREf/VREf co-carriage was not detected in 388 paired samples. The use of antibiotics (OR 4.2, 95% CI 1.7-11.2) and proton pump inhibitors (OR 2.7, 95% CI 1.1-6.3) were risk factors for AREf carriage in humans. In dogs, these were the use of antibiotics (OR 2.3, 95% CI 1.1-4.6) and eating raw meat (OR 3.2, 95% CI 1.4-6.6). Core genome MLST-based phylogenetic linkage indicated clonal relatedness for a minority of human (16.7%) and pet AREf isolates (23.8%) in three clusters. Conclusions: Intestinal carriage with AREf or VREf is rare in the Dutch general population. Although AREf carriage is high in dogs, phylogenetic linkage between human and pet AREf isolates was limited.

Carriage of antibiotic-resistant Gram-negative bacteria after discontinuation of selective decontamination of the digestive tract (SDD) or selective oropharyngeal decontamination (SOD).

BACKGROUND: Selective decontamination of the digestive tract (SDD) and selective oropharyngeal decontamination (SOD) reduce colonization with antibiotic-resistant Gram-negative bacteria (ARGNB), incidence of nosocomial infections and improve survival in ICU patients. The effect on bacterial gut colonization might be caused by growth suppression by antibiotics during SDD/SOD. We investigated intestinal colonization with ARGNB after discharge from ICU and discontinuation of SDD or SOD. METHODS: We performed a prospective, observational follow-up study in regular hospital wards of three teaching hospitals in the Netherlands in patients discharged from the ICU, who were participating in a cluster randomized trial comparing SDD with SOD. We determined rectal carriage with ARGNB at ICU discharge (time (T) = 0) and 3, 6 and 10 days after discharge. The primary endpoint was time to first colonization with ARGNB that was not present at T = 0. Bacteria that are intrinsically resistant to antibiotics were not included in the primary analysis, but were included in post-hoc analysis. RESULTS: Of 1370 patients screened for inclusion, 996 patients had samples at T = 0 (507 after SDD and 489 after SOD). At ICU discharge, the prevalence of intestinal carriage with any ARGNB was 22/507 (4.3%) after SDD and 87/489 (17.8%) after SOD (p < 0.0001): 426 (SDD) and 409 (SOD) patients had at least one follow-up sample for analysis. The hazard rate for acquiring carriage of ARGNB after discontinuation of SDD, compared to SOD, in the ICU was 0.61 (95% CI 0.40-0.91, p = 0.02), and cumulative risks of acquisition of at least one ARGNB until day 10 were 13% (SDD) and 18% (SOD). At day 10 after ICU discharge, the prevalence of intestinal carriage with ARGNB was 11.3% (26/230 patients) after SDD and 12.5% (28/224 patients) after SOD (p = 0.7). In post-hoc analysis of all ARGNB, including intrinsically resistant bacteria, colonization at ICU discharge was lower after SDD (4.9 vs. 22.3%, p < 0.0001), but acquisition rates after ICU discharge were similar in both groups. CONCLUSIONS: Intestinal carriage at ICU discharge and the acquisition rate of ARGNB after ICU discharge are lower after SDD than after SOD. The prevalence of intestinal carriage with ARGNB at 10 days after ICU discharge was comparable in both groups, suggesting rapid clearance of ARGNB from the gut after ICU discharge. TRIAL REGISTRATION: Netherlands Trial Registry, NTR3311 . Registered on 28 february 2012.

ESBL/AmpC-producing Enterobacteriaceae in households with children of preschool age: prevalence, risk factors and co-carriage.

OBJECTIVES: ESBL/AmpC-producing Enterobacteriaceae are an emerging public health concern. As households with preschool children may substantially contribute to the community burden of antimicrobial resistance, we determined the prevalence, risk factors and co-carriage of ESBL/AmpC-producing bacteria in preschool children and their parents. METHODS: From April 2013 to January 2015, each month 2000 preschool children were randomly selected from Dutch population registries. The parents were invited to complete an epidemiological questionnaire and to obtain and send a faecal sample from the selected child and from one parent. Samples were tested for ESBL/AmpC-producing bacteria. Logistic regression was used to identify risk factors for ESBL/AmpC carriage in children and parents, and findings were internally validated by bootstrapping. RESULTS: In total, 1016 families were included and ESBL/AmpC prevalence was 4.0% (95% CI 3.2%-5.0%); 3.5% (95% CI 2.5%-4.8%) in children and 4.5% (95% CI 3.4%-6.0%) in parents. Attending a daycare centre (DCC) was the only significant risk factor for children (OR 2.1, 95% CI 1.0-4.3). For parents, the only significant risk factor was having one or more children attending DCCs (OR 2.2, 95% CI 1.2-4.8). For parents of ESBL/AmpC-positive children the OR for ESBL/AmpC carriage was 19.7 (95% CI 9.2-42.4). Co-carriage of specific ESBL/AmpC genotypes in child and parent occurred more often than expected by chance (14.6% versus 1.1%, P < 0.001). CONCLUSIONS: In this study, intestinal carriage with ESBL/AmpCs was detected in ∼4% of households with preschool children. DCC attendance was a risk factor in both children and parents and co-carriage of specific genotypes frequently occurred in child-parent pairs. These findings suggest household transmission or/and family-specific exposure to common sources of ESBL/AmpC-producing bacteria.

ESBL carriage in pig slaughterhouse workers is associated with occupational exposure.

We investigated the prevalence of extended-spectrum ß-lactamase (ESBL) carriage in slaughterhouse workers and the association with occupational exposure to slaughter animals and products. Stool samples from 334 employees in a Dutch pig slaughterhouse were obtained. Presence of ESBL was determined by selective plating, microarray analysis, and gene sequencing. Questionnaires were used to collect personal and occupational information. The overall prevalence of ESBL carriage was 4·8% (16/334). All ESBL-producing isolates were Escherichia coli. The ESBL genes detected were bla CTX-M-1 (n = 8), bla CTX-M-15 (n = 3), bla CTX-M-27 (n = 2), bla CTX-M-24 (n = 1), bla CTX-M-55 (n = 1), and bla SHV-12 (n = 1). A higher prevalence of ESBL was seen in workers in jobs with as tasks 'removal of lungs, heart, liver, tongue' (33%), and 'removal of head and spinal cord' (25%). For further analysis, participants were divided in two groups based on potential exposure to ESBL as related to their job title. One group with an assumed higher exposure to ESBL (e.g. stable work, stabbing, dehairing, removal of organs) and another group with an assumed lower exposure to ESBL (e.g. refrigeration, packaging and expedition). In the 'higher exposure' group, ten out of 95 (10·5%) were carrying ESBL vs. six out of 233 (2·6%) in the 'lower exposure' group. Human ESBL carriage was significantly associated with job exposure in the slaughterhouse (OR 4·5, CI 1·6-12·6). Results suggest that ESBL carriage in slaughterhouse workers overall is comparable with the Dutch population. Within the slaughterhouse population a difference in carriage exists depending on their position along the slaughter line and tasks involved.

Gram-positive cocci in Dutch ICUs with and without selective decontamination of the oropharyngeal and digestive tract: a retrospective database analysis.

OBJECTIVES: The objective of this study was to determine time trends in the rate of Gram-positive cocci in 42 Dutch ICUs that continuously used or did not use selective oropharyngeal decontamination (SOD) or selective decontamination of the digestive tract (SDD) and ICUs that introduced SOD/SDD. METHODS: The Dutch Surveillance System on Antibiotic Resistance was used to determine monthly rates of Staphylococcus aureus, Enterococcus faecalis and Enterococcus faecium isolates, including resistant phenotypes, in blood and respiratory tract specimens from 2008 to 2013. Per patient, the last isolate per species per month was selected, and cumulative rates per 100 beds per month were determined. Time trends were analysed by multilevel Poisson regression. RESULTS: Eighteen ICUs used SOD/SDD (1296 months), 13 did not use SOD/SDD (936 months) and 11 introduced SOD/SDD (373 months before and 419 months after introduction). There was no significant increase in the rate of Gram-positive cocci in ICUs that used SOD/SDD. Introduction of SOD/SDD was associated with increased rates of S. aureus (ß = 0.018, 95% CI 0.006-0.030) and E. faecalis isolates (ß = 0.028, 95% CI 0.006-0.051) in respiratory tract specimens. Numbers of resistant phenotypes remained low, and an observed increase in E. faecium isolates (ß = 0.033, 95% CI 0.012-0.054), 97% of which were resistant to ampicillin, in the absence of SOD/SDD disappeared after the introduction of SOD/SDD. CONCLUSIONS: In ICUs with a low endemicity of resistance, introduction of SOD/SDD was associated with increased rates of S. aureus and E. faecalis isolates, but not with resistant phenotypes. Continuous 5 year use of SOD/SDD was not associated with increased isolation of Gram-positive cocci.

Risk factors for the acquisition of OXA-48-producing Enterobacteriaceae in a hospital outbreak setting: a matched case-control study.

OBJECTIVES: In the context of a large outbreak of OXA-48-producing Enterobacteriaceae (OXA-E) in a Dutch hospital we determined risk factors for acquisition of OXA-E. PATIENTS AND METHODS: A matched case-control study was performed in which cases (culture positive for OXA-E) were matched 1:3 to controls (culture negative for OXA-E) based on hospital ward, index date (±1 week) and time exposed in the hospital (best match). Stratified analyses were performed for patients with OXA-E producing and not producing ESBL. Potential risk factors included age, gender, surgery and ICU admission within 30 days preceding the index date, presence of comorbidities and in-hospital antibiotic treatment within 30 days preceding the index date. Data analysis was performed using multivariable conditional logistic regression with Firth correction. RESULTS: In total, 73 cases were matched to 211 controls. In the multivariable conditional logistic regression model, male gender (OR 2.63, 95% CI 1.25-5.53), age (per year increase, OR 1.03, 95% CI 1.00-1.05) and use of fluoroquinolones within 30 days preceding the index date (OR 2.98, 95% CI 1.06-8.41) were risk factors for acquisition of OXA-E. In the stratified multivariable conditional logistic regression model, quinolone use was a risk factor for the acquisition of ESBL-producing OXA-E and surgery was a risk factor for the acquisition of non-ESBL-producing OXA-E. CONCLUSIONS: During a large, hospital-wide OXA-E outbreak, male gender, age and previous use of fluoroquinolones were risk factors for acquisition of OXA-E. These findings may help in optimizing screening and isolation strategies in future OXA-E outbreaks.

Molecular epidemiology of MRSA in 13 ICUs from eight European countries.

OBJECTIVES: The European epidemiology of MRSA is changing with the emergence of community-associated MRSA (CA-MRSA) and livestock-associated MRSA (LA-MRSA). In this study, we investigated the molecular epidemiology of MRSA during 2 years in 13 ICUs in France, Greece, Italy, Latvia, Luxemburg, Portugal, Slovenia and Spain. METHODS: Surveillance cultures for MRSA from nose and wounds were obtained on admission and twice weekly from all patients admitted to an ICU for ≥3 days. The first MRSA isolate per patient was genotyped in a central laboratory by MLST, spa typing, agr typing and SCCmec (sub)typing. Risk factors for patients with an unknown history of MRSA colonization were identified. RESULTS: Overall, 14 390 ICU patients were screened, of whom 8519 stayed in an ICU for ≥3 days. Overall MRSA admission prevalence was 3.9% and ranged from 1.0% to 7.0% for individual ICUs. Overall MRSA acquisition rate was 2.5/1000 patient days at risk and ranged from 0.2 to 8/1000 patient days at risk per ICU. In total, 557 putative MRSA isolates were submitted to the central laboratory for typing, of which 511 (92%) were confirmed as MRSA. Each country had a distinct epidemiology, with ST8-IVc (UK-EMRSA-2/-6, USA500) being most prevalent, especially in France and Spain, and detected in ICUs in five of eight countries. Seventeen (3%) and three (<1%) isolates were categorized as CA-MRSA and LA-MRSA, respectively. Risk factors for MRSA carriage on ICU admission were age >70 years and hospitalization within 1 year prior to ICU admission. CONCLUSIONS: The molecular epidemiology of MRSA in 13 European ICUs in eight countries was homogeneous within, but heterogeneous between, countries. CA-MRSA and LA-MRSA genotypes and Panton-Valentine leucocidin-producing isolates were detected sporadically.

The Innovative Medicines Initiative's New Drugs for Bad Bugs programme: European public-private partnerships for the development of new strategies to tackle antibiotic resistance.

Antibiotic resistance (ABR) is a global public health threat. Despite the emergence of highly resistant organisms and the huge medical need for new drugs, the development of antibacterials has slowed to an unacceptable level worldwide. Numerous government and non-government agencies have called for public-private partnerships and innovative funding mechanisms to address this problem. To respond to this public health crisis, the Innovative Medicines Initiative Joint Undertaking programme has invested more than €660 million, with a goal of matched contributions from the European Commission and the European Federation of Pharmaceutical Industries and Associations, in the development of new antibacterial strategies. The New Drugs for Bad Bugs (ND4BB) programme, an Innovative Medicines Initiative, has the ultimate goal to boost the fight against ABR at every level from basic science and drug discovery, through clinical development to new business models and responsible use of antibiotics. Seven projects have been launched within the ND4BB programme to achieve this goal. Four of them will include clinical trials of new anti-infective compounds, as well as epidemiological studies on an unprecedented scale, which will increase our knowledge of ABR and specific pathogens, and improve the designs of the clinical trials with new investigational drugs. The need for rapid concerted action has driven the funding of seven topics, each of which should add significantly to progress in the fight against ABR. ND4BB unites expertise and provides a platform where the commitment and resources required by all parties are streamlined into a joint public-private partnership initiative of unprecedented scale.

KPC-Like Carbapenemase-Producing Enterobacteriaceae Colonizing Patients in Europe and Israel.

In a 2008-2011 survey, 17,945 patients in 18 hospital units in Europe and Israel were screened for carriage of Klebsiella pneumoniae carbapenemase (KPC)-producing Enterobacteriaceae, resulting in identification of 124 positive patients. The isolates were dominated by Klebsiella pneumoniae sequence type 258 (ST258) KPC-2 and ST512 KPC-3, mainly from Greece and Italy, respectively, whereas Israeli isolates were of diverse species, clones, and KPC variants. Various blaKPC platforms were observed, among which IncFIIK-FIBK plasmids with blaKPC-2/-3 genes in the Tn4401a transposon prevailed.

Rectal Carriage of Extended-Spectrum-Beta-Lactamase-Producing Enterobacteriaceae in Hospitalized Patients: Selective Preenrichment Increases Yield of Screening.

This study evaluated the added value of selective preenrichment for the detection of rectal carriage of extended-spectrum-beta-lactamase-producing Enterobacteriaceae (ESBL-E). ESBL-E rectal carriage was identified in 4.8% of hospitalized patients, and 25.9% of ESBL-E rectal carriers were identified with selective preenrichment only.

MLST reveals potentially high-risk international clones of Enterobacter cloacae.

OBJECTIVES: To perform the first multinational Enterobacter cloacae clonality study, using the MLST scheme newly developed in Japan. METHODS: The analysis included 195 rectal carriage E. cloacae isolates resistant to expanded-spectrum cephalosporins (ESCs), collected from patients in 12 hospital units across Europe and Israel. All of the isolates were typed by PFGE and 173 isolates were subjected to MLST. ESC resistance was analysed phenotypically; genes encoding ESBLs and carbapenemases were identified by PCR and sequencing. RESULTS: MLST distinguished 88 STs, which correlated with the PFGE data. PFGE was more discriminatory, producing 129 pulsotypes (169 patterns). Numerous STs were observed in several countries each. The most widespread were ST66, ST78, ST108 and ST114, each having at least 10 isolates from three to five countries, diversified into multiple pulsotypes, with clusters of related isolates in one or more centres. Analysis of the STs against the MLST database revealed several epidemic clonal complexes, such as those with central genotypes ST74 (including ST78) or ST114 (including ST66). ESC resistance was equally related to overexpression of the AmpC cephalosporinase and to ESBL production. Among ESBL producers some spreading subclones were identified, including specific ST66, ST78 and ST114 pulsotypes, associated with CTX-M-15 production. Several isolates produced carbapenemase VIM-1 or KPC-2. CONCLUSIONS: Together with the information available in the MLST database, our results suggest that, like Escherichia coli and Klebsiella pneumoniae, E. cloacae harbours clonal lineages of increased epidemic potential that may be associated with resistance spread.

Phylogenetic lineages, clones and ß-lactamases in an international collection of Klebsiella oxytoca isolates non-susceptible to expanded-spectrum cephalosporins.

OBJECTIVES: The objective of this study was to examine Klebsiella oxytoca clonal and phylogenetic diversity, based on an international collection of carriage isolates non-susceptible to expanded-spectrum cephalosporins (ESCs). METHODS: The study material comprised 68 rectal carriage K. oxytoca isolates non-susceptible to ESCs recovered in 2008-11 from patients in 14 hospitals across Europe and Israel. ESC resistance was tested phenotypically; genes encoding ESBLs, AmpC cephalosporinases and carbapenemases were amplified and sequenced. The isolates were typed by PFGE and MLST, followed by sequencing of blaOXY genes. RESULTS: MLST and PFGE distinguished 34 STs and 47 pulsotypes among the isolates, respectively. Six STs were split into several pulsotypes each. Five STs were more prevalent (n = 2-9) and occurred in several countries each, including ST2, ST9 and ST141, which belong to a growing international clonal complex (CC), CC2. Four phylogenetic lineages were distinguished, each with another type of chromosomal OXY-type ß-lactamase. Three of these, with OXY-1/-5, OXY-2 types and OXY-4, corresponded to previously described phylogroups KoI, KoII and KoIV, respectively. A single isolate from Israel represented a distinct lineage with a newly defined OXY-7 type. The phylogroups showed interesting differences in mechanisms of ESC resistance; KoI strains rarely overexpressed the OXY enzymes but commonly produced ESBLs, whereas KoII strains often were OXY hyperproducers and carried ESBLs much less frequently. AmpCs (DHA-1) and carbapenemases (VIM-1) occurred sporadically. CONCLUSIONS: The study confirmed the high genetic diversity of the collection of K. oxytoca ESC-non-susceptible isolates, composed of phylogroups with distinct types of OXY-type ß-lactamases, and revealed some STs of broad geographical distribution.

Survey of metallo-ß-lactamase-producing Enterobacteriaceae colonizing patients in European ICUs and rehabilitation units, 2008-11.

OBJECTIVES: The objective of this study was to perform a multinational survey of patients' colonization by metallo-ß-lactamase (MBL)-producing Enterobacteriaceae, including their molecular characterization. METHODS: Patients in 18 hospital units across Europe and Israel (n = 17 945) were screened between mid-2008 and mid-2011. MBL-producing isolates were typed by PFGE and MLST. MBL genes were amplified and sequenced within their integrons. Plasmids with MBL genes were analysed by nuclease S1 plus hybridization profiling, mating and transformation assays, and by PCR-based replicon typing. RESULTS: Ninety-one patients in nine centres (six countries), including 62 patients in two Greek ICUs, carried 94 non-duplicate MBL-producing organisms. Klebsiella pneumoniae isolates from Greece dominated (n = 57) and belonged mainly to ST147, ST36 and ST383. All but one of the isolates expressed VIM-1-type MBLs. Isolates of Greek origins produced five enzymes, including new VIM-39, encoded by class 1 integrons of four types. In-e541-like elements prevailed, comprising six variants located on IncR, IncFIIK, IncR + FIIK, IncR + A/C or non-typeable plasmids. The other group were new In4873 and In4863, being the first In416-like elements identified in Greece, which were present on IncA/C or non-typeable plasmids. Isolates from other countries produced only VIM-1 and the major integron was In916, identified in 16 organisms from France, Italy and Spain. In916 was carried by four plasmid types, including IncA/C, IncFIIK and IncHI2. Other integrons included a new element, In3103, in Spain and In110 identified only in Latvia. CONCLUSIONS: This study provided fully comparable data on the occurrence and molecular characteristics of VIM-producing Enterobacteriaceae in a group of hospital units across Europe, documenting recent changes in their epidemiology.

Distinct SagA from Hospital-Associated Clade A1 Enterococcus faecium Strains Contributes to Biofilm Formation.

Enterococcus faecium is an important nosocomial pathogen causing biofilm-mediated infections. Elucidation of E. faecium biofilm pathogenesis is pivotal for the development of new strategies to treat these infections. In several bacteria, extracellular DNA (eDNA) and proteins act as matrix components contributing to biofilm development. In this study, we investigated biofilm formation capacity and the roles of eDNA and secreted proteins for 83 E. faecium strains with different phylogenetic origins that clustered in clade A1 and clade B. Although there was no significant difference in biofilm formation between E. faecium strains from these two clades, the addition of DNase I or proteinase K to biofilms demonstrated that eDNA is essential for biofilm formation in most E. faecium strains, whereas proteolysis impacted primarily biofilms of E. faecium clade A1 strains. Secreted antigen A (SagA) was the most abundant protein in biofilms from E. faecium clade A1 and B strains, although its localization differed between the two groups. sagA was present in all sequenced E. faecium strains, with a consistent difference in the repeat region between the clades, which correlated with the susceptibility of biofilms to proteinase K. This indicates an association between the SagA variable repeat profile and the localization and contribution of SagA in E. faecium biofilms.

Bacteraemic and non-bacteraemic/urinary antigen-positive pneumococcal community-acquired pneumonia compared.

The diagnosis of invasive pneumococcal pneumonia is based mainly on bacteraemia. Episodes without bacteraemia, but with a positive urinary antigen test (UAT), are considered non-invasive. We determined differences in outcome between patients with bacteraemic and non-bacteraemic/UAT-positive pneumococcal community-acquired pneumonia (CAP). Adult patients with clinical and radiological evidence of CAP with blood cultures and UAT tests performed at presentation in three Dutch laboratories between June 2008 and May 2010 were included. Clinical characteristics were retrospectively extracted from hospital records. Overall, 168 patients had non-bacteraemic/UAT-positive pneumococcal CAP and 123 had bacteraemic pneumococcal CAP. The day-30 mortality was 9% and 13% for non-bacteraemic/UAT-positive and bacteraemic pneumococcal CAP patients, respectively [risk difference -4%, 95% confidence interval (CI) -11% to +3%, p = 0.28]. In a multivariable logistic regression model, age ≥ 65 years, admission to the intensive care unit/coronary care unit (ICU/CCU) and presence of an immunocompromising condition were associated with day-30 mortality. A non-significant association with mortality was found for bacteraemia [odds ratio (OR) 2.21, 95% CI 0.94-5.21, p = 0.07). No such trend was found for UAT positivity. The median lengths of hospital stay were 8 [interquartile range (IQR) 5-14] and 10 (IQR 6-18) days for non-bacteraemic/UAT-positive and bacteraemic pneumococcal CAP patients, respectively (p = 0.05). As compared to non-bacteraemic/UAT-positive pneumococcal CAP, bacteraemic pneumococcal CAP has a stronger association with day-30 mortality.