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Background: Vancomycin-resistant Enterococcus faecium (VREfm) has emerged as a nosocomial pathogen worldwide. The dissemination of VREfm is due to both clonal spread and spread of mobile genetic elements (MGEs) such as transposons.Objectives: We aimed to combine vanB-carrying transposon data with core-genome MLST (cgMLST) typing and epidemiological data to understand the pathways of transmission in nosocomial outbreaks.Methods: Retrospectively, 36 VREfm isolates obtained from 34 patients from seven VREfm outbreak investigations in 2014 were analysed. Isolates were sequenced on a MiSeq and a MinION instrument. De novo assembly was performed in CLC Genomics Workbench and the hybrid assemblies were obtained through Unicycler v0.4.1. Ridom SeqSphere+ was used to extract MLST and cgMLST data. Detailed analysis of each transposon and their integration points was performed using the Artemis Comparison Tool (ACT) and multiple blast analyses.Results: Four different vanB transposons were found among the isolates. cgMLST divided ST80 isolates into three cluster types (CTs); CT16, CT104 and CT106. ST117 isolates were divided into CT24, CT103 and CT105. Within VREfm isolates belonging to CT103, two different vanB transposons were found. In contrast, VREfm isolates belonging to CT104 and CT106 harboured an identical vanB transposon.Conclusions: cgMLST provides a high discriminatory power for the epidemiological analysis of VREfm. However, additional transposon analysis is needed to detect horizontal gene transfer. Combining these two methods allows investigation of both clonal spread as well as the spread of MGEs. This leads to new insights and thereby better understanding of the complex transmission routes in VREfm outbreaks.

Preventing the spread of multidrug-resistant Gram-negative bacteria (MDRGNB) is a public health priority. However, the definition of MDRGNB applied for planning infection prevention measures such as barrier precautions differs depending on national guidelines. This is particularly relevant in the Dutch-German border region, where patients are transferred between healthcare facilities located in the two different countries, because clinicians and infection control personnel must understand antibiograms indicating MDRGNB from both sides of the border and using both national guidelines. This retrospective study aimed to compare antibiograms of Gram-negative bacteria and classify them using the Dutch and German national standards for MDRGNB definition. A total of 31,787 antibiograms from six Dutch and four German hospitals were classified. Overall, 73.7% were no MDRGNB according to both guidelines. According to the Dutch and German guideline, 7772/31,787 (24.5%) and 4586/31,787 (12.9%) were MDRGNB, respectively (p

Objectives: To reveal the prevalence and epidemiology of extended-spectrum β-lactamase (ESBL)- and/or plasmid AmpC (pAmpC)- and carbapenemase (CP) producing Enterobacteriaceae and vancomycin-resistant enterococci (VRE) across the Northern Dutch-German border region. Methods: A point-prevalence study on ESBL/pAmpC/CP producing Enterobacteriaceae and VRE was carried out in hospitalized patients in the Northern Netherlands (n = 445, 2012-2013) and Germany (n = 242, 2012). Healthy individuals from the Dutch community (n = 400, 2010-2012) were also screened. In addition, a genome-wide gene-by-gene approach was applied to study the epidemiology of ESBL-Escherichia coli and VRE. Results: A total of 34 isolates from 27 patients (6.1%) admitted to Dutch hospitals were ESBL/pAmpC positive and 29 ESBL-E. coli, three pAmpC-E. coli, one ESBL-Enterobacter cloacae, and one pAmpC-Proteus mirabilis were found. In the German hospital, 18 isolates (16 E. coli and 2 Klebsiella pneumoniae) from 17 patients (7.7%) were ESBL positive. In isolates from the hospitalized patients CTX-M-15 was the most frequently detected ESBL-gene. In the Dutch community, 11 individuals (2.75%) were ESBL/pAmpC positive: 10 ESBL-E. coli (CTX-M-1 being the most prevalent gene) and one pAmpC E. coli. Six Dutch (1.3%) and four German (3.9%) hospitalized patients were colonized with VRE. Genetic relatedness by core genome multi-locus sequence typing (cgMLST) was found between two ESBL-E. coli isolates from Dutch and German cross-border hospitals and between VRE isolates from different hospitals within the same region. Conclusion: The prevalence of ESBL/pAmpC-Enterobacteriaceae was similar in hospitalized patients across the Dutch-German border region, whereas VRE prevalence was slightly higher on the German side. The overall prevalence of the studied pathogens was lower in the community than in hospitals in the Northern Netherlands. Cross-border transmission of ESBL-E. coli and VRE seems unlikely based on cgMLST analysis, however continuous monitoring is necessary to control their spread and stay informed about their epidemiology.

The study describes the transmission of a CTX-M-15-producing ST15 Klebsiella pneumoniae between patients treated in a single center and the subsequent inter-institutional spread by patient referral occurring between May 2012 and September 2013. A suspected epidemiological link between clinical K. pneumoniae isolates was supported by patient contact tracing and genomic phylogenetic analysis from May to November 2012. By May 2013, a patient treated in three institutions in two cities was involved in an expanding cluster caused by this high-risk clone (HiRiC) (local expansion, CTX-M-15 producing, and containing hypervirulence factors). A clone-specific multiplex PCR was developed for patient screening by which another patient was identified in September 2013. Genomic phylogenetic analysis including published ST15 genomes revealed a close homology with isolates previously found in the USA. Environmental contamination and lack of consistent patient screening were identified as being responsible for the clone dissemination. The investigation addresses the advantages of whole-genome sequencing in the early detection of HiRiC with a high propensity of nosocomial transmission and prolonged circulation in the regional patient population. Our study suggests the necessity for inter-institutional/regional collaboration for infection/outbreak management of K. pneumoniae HiRiCs.

Contains fulltext : 172306.pdf (Publisher’s version ) (Closed access) A survey of diagnosis and treatment of invasive aspergillosis was conducted in eight University Medical Centers (UMCs) and eight non-academic teaching hospitals in the Netherlands. Against a background of emerging azole resistance in Aspergillus fumigatus routine resistance screening of clinical isolates was performed primarily in the UMCs. Azole resistance rates at the hospital level varied between 5% and 10%, although rates up to 30% were reported in high-risk wards. Voriconazole remained first choice for invasive aspergillosis in 13 out of 16 hospitals. In documented azole resistance 14 out of 16 centres treated patients with liposomal amphotericin B.

Meningitis in adults due to infection with Mycoplasma hominis is rarely reported. Here, we document the third case of M. hominis meningitis in an adult individual, developed upon neurosurgery following a subarachnoid haemorrhage. Our findings are noteworthy, because the presence of M. hominis in cerebrospinal fluid cannot be identified by standard culturing, Gram-staining, or matrix-assisted laser desorption ionization time-of-flight mass spectrometry. Importantly, however, 16S rDNA sequencing did lead to an unambiguous diagnosis and guided successful antimicrobial therapy. Based on our present findings and a review of the respective literature, we conclude that M. hominis should be considered as a candidate causative agent of infections of the central nervous system following neurosurgical procedures, especially if there is no response to standard antimicrobial therapy, and routine culturing yields negative results. (C) 2012 Elsevier GmbH. All rights reserved.

Extended-spectrum -lactamase producing Klebsiella pneumoniae have emerged as one of the major nosocomial pathogens. Between July and September 2012, a CTX-M-15 producing K. pneumoniae caused an outbreak in a university hospital in the Netherlands. The outbreak isolates were characterized and assigned to a novel sequence type (ST1427). An epidemiological link between affected patients was supported by patient contact tracing and whole-genome phylogenetic analysis. Intra-strain polymorphism was detected among multiple isolates obtained from different body sites of the index patient, which may relate to antibiotic treatment and/or host adaptation. Environmental contamination caused by the outbreak clone was found in the patient rooms even on medical equipment. The novel clone was not closely related to any known endemic/epidemic clone, but carried a set of a plasmid-borne resistance genes [bla CTX-M-15, bla TEM-1, bla OXA-1, aac(6')-Ib-cr, qnrB1, tetA(A), aac(3)-II]. Analysis of its virulence factors revealed a previously uncharacterized capsular biosynthesis region and two uncharacterized fimbriae gene clusters, and suggested that the new clone was not hypervirulent. To our knowledge, this is the first outbreak report of K. pneumoniae ST1427, and our study could be of help to understand the features of this newly emerging clone.

Objective.To reveal the source of a nosocomial outbreak of colonization and infection with a strain ofSerratia marcescenspositive for Guiana extended-spectrumβ-lactamase 1 (GES-1) that occurred among patients in a neurosurgical intensive care unit (ICU) in a Dutch university medical center from May 2002 through March 2003.Methods.Samples from the environment and from the hands of healthcare workers (HCWs) were cultured. A retrospective case-control study was carried out.Results.Fifteen neurosurgical ICU patients who had 1 or more cultures that yielded the epidemic strain ofS. marcescensfrom May 2002 through March 2003 were defined as case patients and matched with 30 control patients. Environmental cultures did not reveal a prominent source ofS. marcescens. Cultures of specimens from the hands of 100 HCWs revealed colonization of a single HCW with the epidemic strain. Although this HCW instantly went on leave, serial cultures detected prolonged carriage of the epidemic strain on the hands of the HCW for 3 months. The skin of the HCW's hands was psoriatic. The epidemic abruptly ended after the colonized HCW went on leave. Retrospective case-control analysis showed that the patients colonized or infected withS. marcescensreceived significantly more nursing care from the colonized HCW than did control patients (PS. marcescens. In October 2004, the formerly colonized HCW appeared to have carriage of the epidemic strain on the hands again.Conclusions.A single HCW with the epidemic strain ofS. marcescenson the hands was considered the source of this outbreak.

The specificity of random primer R143 for Aspergillus fumigatus DNA was determined in order to test its usefulness in establishing the presence of A. fumigatus DNA in fungal cultures. When PCR reaction products of these cultures were compared with those of 21 other bacterial and fungal DNA samples, R143 proved to produce a 1346 bp band with only A. fumigatus. This band has been sequenced completely and the EcoRI restriction site was used for subsequent confirmation of PCR products. The specificity for A. fumigatus DNA was also confirmed by Southern blotting. Comparison of morphological typing of Aspergillus species in cultures with PCR using R143 on DNA isolated from these cultures showed concordance in 22 of 24 cases. In two cases there was discordance: both times PCR results showed correctly the presence of A. fumigatus, initially not detected by culture. R143 is an A. fumigatus specific random primer, with potential for use in detection of A. fumigatus DNA in clinical specimens.

Item does not contain fulltext BACKGROUND: Azole resistance is an emerging problem in Aspergillus fumigatus and complicates the management of patients with Aspergillus-related diseases. Selection of azole resistance may occur through exposure to azole fungicides in the environment. In the Netherlands a surveillance network was used to investigate the epidemiology of resistance selection in A. fumigatus. METHODS: Clinical A. fumigatus isolates were screened for azole resistance in 8 university hospitals using azole agar dilution plates. Patient information was collected using an online questionnaire and azole-resistant A. fumigatus isolates were analyzed using gene sequencing, susceptibility testing, and genotyping. Air sampling was performed to investigate the presence of resistant isolates in hospitals and domiciles. RESULTS: Between December 2009 and January 2011, 1315 A. fumigatus isolates from 921 patients were screened. A new cyp51A-mediated resistance mechanism (TR46/Y121F/T289A) was observed in 21 azole-resistant isolates from 15 patients in 6 hospitals. TR46/Y121F/T289A isolates were highly resistant to voriconazole (minimum inhibitory concentration >/=16 mg/L). Eight patients presented with invasive aspergillosis due to TR46/Y121F/T289A, and treatment failed in all 5 patients receiving primary therapy with voriconazole. TR46/Y121F/T289A Aspergillus fumigatus was recovered from 6 of 10 sampled environmental sites. CONCLUSIONS: We describe the emergence and geographical migration of a voriconazole highly resistant A. fumigatus that was associated with voriconazole treatment failure in patients with invasive aspergillosis. Recovery of TR46/Y121F/T289A from the environment suggests an environmental route of resistance selection. Exposure of A. fumigatus to azole fungicides may facilitate the emergence of new resistance mechanisms over time, thereby compromising the use of azoles in the management of Aspergillus-related diseases.

Contains fulltext : 89507.pdf (Publisher’s version ) (Open Access) We report the use of PCR techniques on a formalin-fixed and paraffin-embedded tissue specimen for direct detection of one dominant azole resistance mechanism in a case of disseminated invasive aspergillosis. Rapid detection of mutations associated with azole resistance directly in tissue significantly reduces diagnostic delay. 01 april 2010

Rhodococcus equi, mainly known from veterinary medicine as a pathogen in domestic animals, can also cause infections in immunocompromised humans, especially in those with defects in cellular immunity. Alemtuzumab, an anti-CD52 monoclonal antibody, causes lymphocytopenia by eliminating CD52-positive cells. We report a patient in whom Rhodococcus equi infection developed after alemtuzumab therapy.

SUMMARY The gram-positive bacterium Staphylococcus aureus is a frequent component of the human microbial flora that can turn into a dangerous pathogen. As such, this organism is capable of infecting almost every tissue and organ system in the human body. It does so by actively exporting a variety of virulence factors to the cell surface and extracellular milieu. Upon reaching their respective destinations, these virulence factors have pivotal roles in the colonization and subversion of the human host. It is therefore of major importance to obtain a clear understanding of the protein transport pathways that are active in S. aureus . The present review aims to provide a state-of-the-art roadmap of staphylococcal secretomes, which include both protein transport pathways and the extracytoplasmic proteins of these organisms. Specifically, an overview is presented of the exported virulence factors, pathways for protein transport, signals for cellular protein retention or secretion, and the exoproteomes of different S. aureus isolates. The focus is on S. aureus , but comparisons with Staphylococcus epidermidis and other gram-positive bacteria, such as Bacillus subtilis , are included where appropriate. Importantly, the results of genomic and proteomic studies on S. aureus secretomes are integrated through a comparative “secretomics” approach, resulting in the first definition of the core and variant secretomes of this bacterium. While the core secretome seems to be largely employed for general housekeeping functions which are necessary to thrive in particular niches provided by the human host, the variant secretome seems to contain the “gadgets” that S. aureus needs to conquer these well-protected niches.

Pantoea infections are rare in humans, especially in neonates. Infections are usually associated with plant thom injury or outbreaks traced to contaminated parenteral nutrition, intravenous anesthetics or packed er throcytes. Between I st of January 1994 and Ist of June 2005, 125 cf 6383 patients (2%) in a 24-bed level III NICU became colonized with Pantoea agglomerans. Three new-borns exhibited late-onset Pantoea agglomerans septicemia and died. Sporadic cases of Pantoea agglomerans septicemia have not been reported in neonatal intensive care so far.

A case of bacteremia due to Ochrobactrum intermedium , with concomitant liver abscesses, in an orthotopic liver transplant recipient is presented. Identical microorganisms were isolated from fecal specimens and from an aspirate of a liver abscess that was indicative of invasion of the graft by gastrointestinal spread. 16S DNA sequence analysis of the blood isolate revealed the recovery of the recently proposed new species O. intermedium , closely related to Ochrobactrum anthropi and Brucella spp.

INTRODUCTION: Nowadays Enterococcus faecium has become one of the most emerging and challenging nosocomial pathogens. The aim of this study was to determine risk factors in haematology patients who are at risk of an Enterococcus faecium bloodstream infection (BSI) and should be considered for pre-emptive glycopeptide treatment. With these identified risk factors a prediction model can be developed for clinical use.METHODS: Retrospectively clinical and microbiological data in 33 patients with an E. faecium BSI were compared to 66 control patients during a 5-year period at the haematology ward. Multivariate logistic regression was used to explore the independent risk factors and a prediction model was developed to determine the risk of an E. faecium BSI.RESULTS: E. faecium BSIs were found to be associated with high mortality rates. Independent risk factors for E. faecium BSI were colonization with E. faecium 30 days prior to blood culture (OR 5.71; CI 1.7-18.7), combination of neutropenia and abdominal focus (4.37; 1.4-13.4), age > 58 years (4.01; 1.3-12.5), hospital stay prior to blood culture > 14 days (3.55; 0.98-12.9) and CRP (C-reactive protein) level >125 mg/L (4.37; 1.1-10.2).CONCLUSION: Using data from this study, risk stratification for the development of an E. faecium BSI in patients with haematological malignancies is possible. Pre-emptive treatment should be considered in those patients who are at high risk. Using a prediction model as designed in this study, antibiotic stewardship in terms of prudent use of glycopeptides can be improved and might be helpful in controlling further spread of VRE (vancomycin resistant enterococci).

Enterococcus faecium has become a major cause of nosocomial infections especially in patients with hematologic malignancies. The aim of this study was to determine risk factors in those patients who are at risk of Enterococcus faecium blood stream infection (BSI) and should be considered for empirical treatment.

We describe the introduction of various multi-drug resistant bacterial strains, including an NDM-1-producing Klebsiella pneumoniae, through a traveller returning from Egypt, where they had been admitted to a private hospital. All family members of the patient were colonised with one or more extended-spectrum beta-lactamase producing strains. These findings emphasise the importance of adherence to isolation precautions for returning patients and suggest the need for inclusion of Enterobacteriaceae in admission screening. We here report of a patient who had been hospitalised in Egypt for appendicitis in July 2013, and was colonised with various multiresistant Enterobacteriaceae including strains producing NDM-1, oxacillinase-48 (OXA-48) and extended spectrum beta-lactamase (ESBL). Explorative screening for multiresistant microorganisms among the patient's family members also yielded several ESBL-producing microorganisms. This report addresses the need for heightened awareness of patients and family members who have recently been exposed to healthcare environments in countries with high levels of antibiotic resistance. Patients repatriated after hospitalisation abroad are a risk for introducing multiresistant microorganisms into hospitals in their home countries. In 2008, New Delhi metallo-beta-lactamase (NDM), which hydrolyses last-line carbapenem antibiotics, has been for the first time described in a Swedish patient returning from India [1]. Most reports on NDM are related to travellers returning from Pakistan and India. However, the global dispersal of NDM is of growing concern [2]. In the past two years, NDM-producing strains have been reported in patients returning from the African continent without obvious links to the Indian subcontinent [3,4].