[Show abstract][Hide abstract] ABSTRACT: De novo genome assembly can be challenging due to inherent properties of the reads, even when using current state-of-the-art assembly tools based on de Bruijn graphs. Often users are not bio-informaticians and, in a black box approach, utilise assembly parameters such as contig length and N50 to generate whole genome sequences, potentially resulting in mis-assemblies.
[Show abstract][Hide abstract] ABSTRACT: Rhinovirus infections occur frequently throughout life and have been reported in about one-third of asymptomatic cases. The clinical significance of sequential rhinovirus infections remains unclear. To determine the incidence and clinical relevance of sequential rhinovirus detections, nasopharyngeal samples from 2485 adults with acute cough/lower respiratory illness were analysed. Patients were enrolled prospectively by general practitioners from 12 European Union countries during three consecutive years (2007-2010). Nasopharyngeal samples were collected at the initial general practitioner consultation and 28 days thereafter and symptom scores were recorded by patients over that period. Rhinovirus RNA was detected in 444 (18%) out of 2485 visit one samples and in 110 (4.4%) out of 2485 visit two respiratory samples. 21 (5%) of the 444 patients had both samples positive for rhinovirus. Genotyping of both virus detections was successful for 17 (81%) out of 21 of these patients. Prolonged rhinovirus shedding occurred in six (35%) out of 21 and re-infection with a different rhinovirus in 11 (65%) out of 21. Rhinovirus re-infections were significantly associated with chronic obstructive pulmonary disease (p=0.04) and asthma (p=0.02) and appeared to be more severe than prolonged infections. Our findings indicate that in immunocompetent adults rhinovirus re-infections are more common than prolonged infections, and chronic airway comorbidities might predispose to more frequent rhinovirus re-infections.
The European respiratory journal. 07/2014; 44(1):169-77.
[Show abstract][Hide abstract] ABSTRACT: Infection with human cytomegalovirus (CMV) is a significant cause of morbidity and mortality in solid organ and hematopoietic stem cell transplant (HSCT) recipients.
[Show abstract][Hide abstract] ABSTRACT: Canuti and Martin Deijs have contributed equally to this work. Previously unknown or unexpected pathogens may be responsible for that proportion of respiratory diseases in which a causative agent cannot be identified. The application of broad-spectrum, sequence independent virus discovery techniques may be useful to reduce this proportion and widen our knowledge about respiratory pathogens. Thanks to the availability of high-throughput sequencing (HTS) technology, it became today possible to detect viruses which are present at a very low load, but the clinical relevance of those viruses must be investigated. In this study we used VIDISCA-454, a restriction enzyme based virus discovery method that utilizes Roche 454 HTS system, on a nasal swab collected from a subject with respiratory complaints. A γ-papillomavirus was detected (complete genome: 7142 bp) and its role in disease was investigated. Respiratory samples collected both during the acute phase of the illness and 2 weeks after full recovery contained the virus. The patient presented antibodies directed against the virus but there was no difference between IgG levels in blood samples collected during the acute phase and 2 weeks after full recovery. We therefore concluded that the detected γ-papillomavirus is unlikely to be the causative agent of the respiratory complaints and its presence in the nose of the patient is not related to the disease. Although HTS based virus discovery techniques proved their great potential as a tool to clarify the etiology of some infectious diseases, the obtained information must be subjected to cautious interpretations. This study underlines the crucial importance of performing careful investigations on viruses identified when applying sensitive virus discovery techniques, since the mere identification of a virus and its presence in a clinical sample are not satisfactory proofs to establish a causative link with a disease.
Frontiers in Microbiology 07/2014; 5:374. · 3.90 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The purposes of this study were to investigate the intestinal carriage of extended-spectrum β-lactamase-harbouring Enterobacteriaceae (ESBL-EN) and associated fluoroquinolone resistance (FQ-R) in 120 hospitalised patients with antibiotic-associated diarrhoea, and to investigate a correlation between Clostridium difficile (C. difficile) infection and intestinal colonisation with ESBL-EN in these patients. Stool samples were screened for C. difficile infection by toxin A/B enzyme-linked immunosorbent assay (ELISA) and for the presence of enterobacterial isolates producing β-lactamases by plating on β-lactamase screening (BLSE) agar. Recovered isolates were confirmed pheno- and genotypically for the presence of ESBL genes (bla CTX-M, bla TEM, bla SHV) by the double-disc synergy test and polymerase chain reaction (PCR) sequencing, and tested for the presence of topoisomerase mutations (gyrA, parC) and plasmid-mediated quinolone resistance (PMQR) determinants [qnrA, qnrB, qnrS, qepA, aac(6')-Ib-cr] by PCR sequencing. ESBL-EN were detected in 44/120 (37 %) stool samples. C. difficile-infected patients showed a significantly higher frequency of intestinal colonisation with ESBL-EN compared to C. difficile non-infected patients (62 % vs. 31 %, p = 0.008). Of the 73 ESBL-EN recovered, 46 (63 %) showed high-level FQ-R [ciprofloxacin minimum inhibitory concentration (MIC) ≥32 mg/L]. The largest group consisted of 21 bla CTX-M-15-harbouring Enterobacteriaceae (ciprofloxacin MIC ≥64 mg/L) with multiple topoisomerase mutations in gyrA and parC, in combination with co-carriage of aac(6')-Ib-cr. Most of them were Escherichia coli isolates belonging to sequence types ST131 and ST410. We found remarkably high rates of intestinal colonisation with high-level FQ-R ESBL-EN in hospitalised patients with antibiotic-associated diarrhoea, especially among C. difficile-infected patients. These data underscore the need for stringent infection control to contain this potentially infectious and multidrug-resistant reservoir.
[Show abstract][Hide abstract] ABSTRACT: Although implantation of cellular material in the central nervous system (CNS) is a key direction in CNS regenerative medicine, this approach is currently limited by the occurrence of strong endogenous immune cell responses. In a model of mesenchymal stem cell (MSC) grafting in the CNS of immune-competent mice, we previously described that MSC grafts become highly surrounded and invaded by Iba1(+) myeloid cells (microglia and/or macrophages). Here, following grafting of blue fluorescent protein (BFP)-expressing MSC in the CNS of CX3CR1(+/-) and CX3CR1(-/-) mice, our results indicate: (1) that the observed inflammatory response is independent of the fractalkine signalling axis, and (2) that a significant spatial distribution of Iba1(+) inflammatory cells occurs, in which Iba1(+) CX3CR1(+) myeloid cells mainly surround the MSC graft and Iba1(+) CX3CR1(-) myeloid cells mainly invade the graft at 10 days post transplantation. Although Iba1(+) CX3CR1(+) myeloid cells are considered to be of resident microglial origin, Iba1(+) CX3CR1(-) myeloid cells are most likely of peripheral monocyte/macrophage origin. In order to confirm the latter, we performed MSC-BFP grafting experiments in the CNS of eGFP(+) bone marrow chimeric C57BL/6 mice. Analysis of MSC-BFP grafts in the CNS of these mice confirmed our observation that peripheral monocytes/macrophages invade the MSC graft and that resident microglia surround the MSC graft site. Furthermore, analysis of major histocompatibility complex class II (MHCII) expression revealed that mainly macrophages, but not microglia, express this M1 pro-inflammatory marker in the context of MSC grafting in the CNS. These results again highlight the complexity of cell implantation immunology in the CNS.Immunology and Cell Biology advance online publication, 1 July 2014; doi:10.1038/icb.2014.49.
Immunology and Cell Biology 07/2014; · 3.93 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Cervarix™ is approved as a preventive vaccine against infection with the human papillomavirus (HPV) strains 16 and 18, which are causally related to the development of cervical cancer. We are the first to investigate in vitro the effects of this HPV vaccine on interleukin (IL)-15 dendritic cells (DC) as proxy of a naturally occurring subset of blood DC, and natural killer (NK) cells, two innate immune cell types that play an important role in antitumour immunity. Our results show that exposure of IL-15 DC to the HPV vaccine results in increased expression of phenotypic maturation markers, pro-inflammatory cytokine production and cytotoxic activity against HPV-positive tumour cells. These effects are mediated by the vaccine adjuvant, partly through Toll-like receptor 4 activation. Next, we demonstrate that vaccine-exposed IL-15 DC in turn induce phenotypic activation of NK cells, resulting in a synergistic cytotoxic action against HPV-infected tumour cells. Our study thus identifies a novel mode of action of the HPV vaccine in boosting innate immunity, including killing of HPV-infected cells by DC and NK cells.
Journal of Cellular and Molecular Medicine 06/2014; · 4.75 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Infections remain a leading cause of morbidity and mortality in patients with reduced immunity caused by haematological disease and chemotherapy-induced neutropenia. We evaluated the clinical and microbiological impact of discontinuing fluoroquinolone prophylaxis in these patients.
We analysed 154 admissions in three sequential periods of 8 months: long-standing use, discontinuation and reintroduction of prophylaxis. Clinical endpoints were occurrence of febrile neutropenia, bacteraemia, severe sepsis, septic shock, response to antibiotic therapy, total antibiotic consumption and duration of hospital stay. Microbiological analysis included bacterial isolates from stool and blood cultures and their resistance pattern.
No significant increase in serious infectious complications was seen with the discontinuation of prophylaxis. The overall incidence of bacteraemia did not change, but a higher proportion of bacterial isolates were Gram-negative (22.2% versus 5.9% & 8.6%; p=0.030), more often multi-susceptible (50% versus 0%) and less fluoroquinolone-resistant (10% versus 100%). Screening of stools showed a higher prevalence of organisms in the discontinuation period (86.7% versus 37.3% & 55.2%; p=<0.001), but they were more frequently multi-susceptible (53.8% versus 10.5% & 6.3%; p=<0.001). After discontinuation of prophylaxis, fluoroquinolone resistance decreased rapidly from 73.7% to 7.7%, in association with a significant decrease in extended spectrum beta-lactamase (ESBL)-producing isolates from 42.1% to 10.3%. Resistance figures immediately returned to pre-discontinuation values after reinstitution of prophylaxis.
No clinically relevant short-term drawbacks were observed with the discontinuation of fluoroquinolone prophylaxis in patients with chemotherapy-induced prolonged profound neutropenia, which led to a significant decrease in fluoroquinolone resistance as well as occurrence of ESBL-producing isolates. This article is protected by copyright. All rights reserved.
European Journal Of Haematology 04/2014; · 2.55 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: There are no reliable data on antibiotic use in non-European Union (EU) southern and eastern European countries and newly independent states. We aimed to collect valid, representative, comparable data on systemic antimicrobial use in these non-EU countries of the WHO European region.
Validated 2011 total national wholesale antibiotic-use data of six southern and eastern European countries and regions and seven newly independent states were analysed in accordance with the WHO anatomical therapeutic chemical (ATC)/defined daily doses (DDD) method and expressed in DDD/1000 inhabitants per day (DID).
Total (outpatients and hospital care) antibiotic use ranged from 15·3 DID for Armenia to 42·3 DID for Turkey. Co-amoxiclav was mainly used in Georgia (42·9% of total antibiotic use) and Turkey (30·7%). Newly independent states used substantial quantities of ampicillin and amoxicillin (up to 55·9% of total antibiotic use in Azerbaijan). Montenegro and Serbia were the highest consumers of macrolides (15·8% and 19·5% of total antibiotic use, respectively), mainly azithromycin. Parenteral antibiotic treatment is common practice: 46·4% of total antibiotic use in Azerbaijan (mainly ampicillin; 5·3 DID) and 31·1% of total antibiotic use in Tajikistan (mainly ceftriaxone; 4·7 DID).
This study provides publicly available total antibiotic-use data for 13 non-EU countries and areas of the WHO European region. These data will raise awareness of inappropriate antibiotic use and stimulate policy makers to develop action plans. The established surveillance system provides a method to develop quality indicators of antibiotic use and to assess the effect of policy and regulatory actions.
Netherlands Ministry of Health, Welfare, and Sport, and EU.
The Lancet Infectious Diseases 03/2014; · 19.97 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: To complement analyses of the linear trend and seasonal fluctuation of European outpatient antibiotic use expressed in defined daily doses (DDD) by analyses of data in packages, to assess the agreement between both measures and to study changes in the number of DDD per package over time.
Data on outpatient antibiotic use, aggregated at the level of the active substance (WHO version 2011) were collected from 2000 to 2007 for 31 countries and expressed in DDD and packages per 1000 inhabitants per day (DID and PID, respectively). Data expressed in DID and PID were analysed separately using non-linear mixed models while the agreement between these measurements was analysed through a joint non-linear mixed model. The change in DDD per package over time was studied with a linear mixed model.
Total outpatient antibiotic and penicillin use in Europe and their seasonal fluctuation significantly increased in DID, but not in PID. The use of combinations of penicillins significantly increased in DID and in PID. Broad-spectrum penicillin use did not increase significantly in DID and decreased significantly in PID. For all but one subgroup, country-specific deviations moved in the same direction whether measured in DID or PID. The correlations are not perfect. The DDD per package increased significantly over time for all but one subgroup.
Outpatient antibiotic use in Europe shows contrasting trends, depending on whether DID or PID is used as the measure. The increase of the DDD per package corroborates the recommendation to adopt PID to monitor outpatient antibiotic use in Europe.
Journal of Antimicrobial Chemotherapy 03/2014; · 5.34 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: BACKGROUND Antibiotics are of limited overall clinical benefit for uncomplicated lower respiratory tract infection (LRTI) but there is uncertainty about their effectiveness for patients with features associated with higher levels of antibiotic prescribing. AIM To estimate the benefits and harms of antibiotics for acute LRTI among those producing coloured sputum, smokers, those with fever or prior comorbidities, and longer duration of prior illness. DESIGN AND SETTING Secondary analysis of a randomised controlled trial of antibiotic placebo for acute LRTI in primary care. METHOD Two thousand and sixty-one adults with acute LRTI, where pneumonia was not suspected clinically, were given amoxicillin or matching placebo. The duration of symptoms, rated moderately bad or worse (primary outcome), symptom severity on days 2-4 (0-6 scale), and the development of new or worsening symptoms were analysed in pre-specified subgroups of interest. Evidence of differential treatment effectiveness was assessed in prespecified subgroups by interaction terms. RESULTS No subgroups were identified that were significantly more likely to benefit from antibiotics in terms of symptom duration or the development of new or worsening symptoms. Those with a history of significant comorbidities experienced a significantly greater reduction in symptom severity between days 2 and 4 (interaction term -0.28, P = 0.003; estimated effect of antibiotics among those with a past history -0.28 [95% confidence interval = -0.44 to -0.11], P = 0.001), equivalent to three people in 10 rating symptoms as a slight rather than a moderately bad problem. For subgroups not specified in advance antibiotics provided a modest reduction in symptom severity for non-smokers and for those with short prior illness duration (<7 days), and a modest reduction in symptom duration for those with short prior illness duration. CONCLUSION There is no clear evidence of clinically meaningful benefit from antibiotics in the studied high-risk groups of patients presenting in general practice with uncomplicated LRTIs where prescribing is highest. Any possible benefit must be balanced against the side-effects and longer-term effects on antibiotic resistance.
British Journal of General Practice 02/2014; 64(619):e75-80. · 1.83 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The study aims were: 1) to define the prevalence of and risk factors for colonization by ESBL-producing Enterobacteriaceae (ESBL-Ent) among healthcare workers (HCWs) and family members (FMs) of ESBL-Ent-colonized patients in rehabilitation units (RUs) and 2) to compare ESBL-Ent isolates from these three groups. The study included 286 FMs of 194 ESBL-Ent-carrying patients identified in 5 RUs located in Israel, Italy, France and Spain. ESBL-Ent were detected by rectal swabs in 26 (9%) of 286 FMs screened. In multivariate analyses, older age of the FM, longer mean number of hours spent with the patient, being a daughter or a female spouse of a patient, and chronic lung disease of the patient were significantly associated with carriage in the FM. Escherichia coli was the most common organism (76%), followed by Klebsiella pneumoniae (19%). Isolates were typed by PFGE and MLST, and ESBLs were identified by PCR-sequencing. A comparison of paired species isolates from FMs and their respective patient showed that 17/23 strains were indistinguishable. ESBL-Ent was detected in 35 (3.5%, E. coli=34) of the 1001 HCWs screened. Feeding patients was associated with ESBL-Ent carriage of HCWs. Only 7 of 23 E. coli subclones cultured from HCWs were also represented among 376 patient-derived ESBL-producing E. coli isolates from the same RUs. In Spain, a higher proportion of HCW and FM were ESBL carriers than elsewhere (p<0.05). In conclusion, the molecular and epidemiological data suggest that FMs are at higher risk of ESBL-Ent acquisition from their relative patients than HCW workers. This article is protected by copyright. All rights reserved.
Clinical Microbiology and Infection 01/2014; · 4.58 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: BACKGROUND: Intensive care units (ICUs) are high-risk areas for transmission of antimicrobial-resistant bacteria, but no controlled study has tested the effect of rapid screening and isolation of carriers on transmission in settings with best-standard precautions. We assessed interventions to reduce colonisation and transmission of antimicrobial-resistant bacteria in European ICUs. METHODS: We did this study in three phases at 13 ICUs. After a 6 month baseline period (phase 1), we did an interrupted time series study of universal chlorhexidine body-washing combined with hand hygiene improvement for 6 months (phase 2), followed by a 12-15 month cluster randomised trial (phase 3). ICUs were randomly assigned by computer generated randomisation schedule to either conventional screening (chromogenic screening for meticillin-resistant Staphylococcus aureus [MRSA] and vancomycin-resistant enterococci [VRE]) or rapid screening (PCR testing for MRSA and VRE and chromogenic screening for highly resistant Enterobacteriaceae [HRE]); with contact precautions for identified carriers. The primary outcome was acquisition of resistant bacteria per 100 patient-days at risk, for which we calculated step changes and changes in trends after the introduction of each intervention. We assessed acquisition by microbiological surveillance and analysed it with a multilevel Poisson segmented regression model. We compared screening groups with a likelihood ratio test that combined step changes and changes to trend. This study is registered with ClinicalTrials.gov, number NCT00976638. FINDINGS: Seven ICUs were assigned to rapid screening and six to conventional screening. Mean hand hygiene compliance improved from 52% in phase 1 to 69% in phase 2, and 77% in phase 3. Median proportions of patients receiving chlorhexidine body-washing increased from 0% to 100% at the start of phase 2. For trends in acquisition of antimicrobial-resistant bacteria, weekly incidence rate ratio (IRR) was 0.976 (0.954-0.999) for phase 2 and 1.015 (0.998-1.032) for phase 3. For step changes, weekly IRR was 0.955 (0.676-1.348) for phase 2 and 0.634 (0.349-1.153) for phase 3. The decrease in trend in phase 2 was largely caused by changes in acquisition of MRSA (weekly IRR 0.925, 95% CI 0.890-0.962). Acquisition was lower in the conventional screening group than in the rapid screening group, but did not differ significantly (p=0.06). INTERPRETATION: Improved hand hygiene plus unit-wide chlorhexidine body-washing reduced acquisition of antimicrobial-resistant bacteria, particularly MRSA. In the context of a sustained high level of compliance to hand hygiene and chlorhexidine bathings, screening and isolation of carriers do not reduce acquisition rates of multidrug-resistant bacteria, whether or not screening is done with rapid testing or conventional testing. FUNDING: European Commission.
[Show abstract][Hide abstract] ABSTRACT: Greater use of antibiotics during the past 50 years has exerted selective pressure on susceptible bacteria and may have favoured the survival of resistant strains. Existing information on antibiotic resistance patterns from pathogens circulating among community-based patients is substantially less than from hospitalized patients on whom guidelines are often based. We therefore chose to assess the relationship between the antibiotic resistance pattern of bacteria circulating in the community and the consumption of antibiotics in the community.
Both gray literature and published scientific literature in English and other European languages was examined. Multiple regression analysis was used to analyse whether studies found a positive relationship between antibiotic consumption and resistance. A subsequent meta-analysis and meta-regression was conducted for studies for which a common effect size measure (odds ratio) could be calculated.
Electronic searches identified 974 studies but only 243 studies were considered eligible for inclusion by the two independent reviewers who extracted the data. A binomial test revealed a positive relationship between antibiotic consumption and resistance (p < .001) but multiple regression modelling did not produce any significant predictors of study outcome. The meta-analysis generated a significant pooled odds ratio of 2.3 (95% confidence interval 2.2 to 2.5) with a meta-regression producing several significant predictors (F(10,77) = 5.82, p < .01). Countries in southern Europe produced a stronger link between consumption and resistance than other regions.
Using a large set of studies we found that antibiotic consumption is associated with the development of antibiotic resistance. A subsequent meta-analysis, with a subsample of the studies, generated several significant predictors. Countries in southern Europe produced a stronger link between consumption and resistance than other regions so efforts at reducing antibiotic consumption may need to be strengthened in this area. Increased consumption of antibiotics may not only produce greater resistance at the individual patient level but may also produce greater resistance at the community, country, and regional levels, which can harm individual patients.
[Show abstract][Hide abstract] ABSTRACT: Background aims
Dendritic cell (DC)-based immunotherapy has shown potential to counteract autoimmunity in multiple sclerosis (MS).
We compared the phenotype and T-cell stimulatory capacity of in vitro generated monocyte-derived DC from MS patients with those from healthy controls.
Except for an increase in the number of C-C chemokine receptor 7–expressing DC from MS patients, no major differences were found between groups in the expression of maturation-associated membrane markers or in the in vitro capacity to stimulate autologous T cells.
Our observations may pave the way for the development of patient-tailored DC-based vaccination strategies to treat MS.
[Show abstract][Hide abstract] ABSTRACT: Methicillin-resistant Staphylococcus aureus (MRSA) causes serious infections that are even more difficult to treat when associated with a biofilm phenotype that facilitates evasion of the host immune system and antibiotics. As a first step toward understanding the mechanisms underlying biofilm formation, we sequenced the genomes of two prolific biofilm-forming strains belonging to the two most important globally disseminated clonal lineages, USA300 and EMRSA-15.
[Show abstract][Hide abstract] ABSTRACT: Epidemic methicillin-resistant S. aureus (MRSA) clones cause infections in both hospital and community settings. As a biofilm phenotype further facilitates evasion of the host immune system and antibiotics, we compared the biofilm-forming capacities of various MRSA clones.
PLoS ONE 01/2014; 9(8):e104561. · 3.73 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Widespread multidrug resistance in Escherichia coli has necessitated the reintroduction of older antibiotics, such as nitrofurantoin. However, mechanisms by which resistance to nitrofurantoin emerges in E. coli are not well elucidated. Toward this aim, we sequenced two nitrofurantoin-sensitive E. coli sequence types (ST540 and ST2747) and their four nitrofurantoin-resistant derivatives generated in vitro under aerobic and anaerobic growth conditions.
[Show abstract][Hide abstract] ABSTRACT: Background:In a pan-European randomised controlled trial (GRACE INTRO) of two interventions, (i) a point-of-care C-reactive protein test and/or (ii) training in communication skills and use of an interactive patient booklet, both interventions resulted in large reductions in antibiotic prescribing for acute cough.Aims:This process evaluation explored patients' views of primary care consultations using the two interventions in six European countries.Methods:Sixty-two interviews were conducted with patients who had participated in the GRACE INTRO trial. Interviews were transcribed verbatim and translated into English where necessary. Analysis used techniques from thematic and framework analysis.Results:Most patients were satisfied with their consultation despite many not receiving an antibiotic. Patients appeared to accept the use of both intervention approaches. A minority, but particularly in the trial arm with both interventions, reported that they would wait longer before consulting for cough in future.Conclusions:Patients perceived that both interventions supported the general practitioner's (GP's) prescribing decisions by helping them understand when an antibiotic was, and was not, needed. Patients consulting with acute cough had largely positive views about the GP's enhanced communication skills, which included understanding their concerns, and the use of a near-patient test as an additional investigation.
NPJ primary care respiratory medicine. 01/2014; 24:14026.