2 days versus 5 days of postoperative antibiotics for complex appendicitis: a pragmatic, open-label, multicentre, non-inferiority randomised trial.

BACKGROUND: The appropriate duration of postoperative antibiotics for complex appendicitis is unclear. The increasing global threat of antimicrobial resistance warrants restrictive antibiotic use, which could also reduce side-effects, length of hospital stay, and costs. METHODS: In this pragmatic, open-label, non-inferiority trial in 15 hospitals in the Netherlands, patients with complex appendicitis (aged ≥8 years) were randomly assigned (1:1) to receive 2 days or 5 days of intravenous antibiotics after appendicectomy. Randomisation was stratified by centre, and treating physicians and patients were not masked to treatment allocation. The primary endpoint was a composite endpoint of infectious complications and mortality within 90 days. The main outcome was the absolute risk difference (95% CI) in the primary endpoint, adjusted for age and severity of appendicitis, with a non-inferiority margin of 7·5%. Outcome assessment was based on electronic patient records and a telephone consultation 90 days after appendicectomy. Efficacy was analysed in the intention-to-treat and per-protocol populations. Safety outcomes were analysed in the intention-to-treat population. This trial was registered with the Netherlands Trial Register, NL5946. FINDINGS: Between April 12, 2017, and June 3, 2021, 13 267 patients were screened and 1066 were randomly assigned, 533 to each group. 31 were excluded from intention-to-treat analysis of the 2-day group and 30 from the 5-day group owing to errors in recruitment or consent. Appendicectomy was done laparoscopically in 955 (95%) of 1005 patients. The telephone follow-up was completed in 664 (66%) of 1005 patients. The primary endpoint occurred in 51 (10%) of 502 patients analysed in the 2-day group and 41 (8%) of 503 patients analysed in the 5-day group (adjusted absolute risk difference 2·0%, 95% CI -1·6 to 5·6). Rates of complications and re-interventions were similar between trial groups. Fewer patients had adverse effects of antibiotics in the 2-day group (45 [9%] of 502 patients) than in the 5-day group (112 [22%] of 503 patients; odds ratio [OR] 0·344, 95% CI 0·237 to 0·498). Re-admission to hospital was more frequent in the 2-day group (58 [12%] of 502 patients) than in the 5-day group (29 [6%] of 503 patients; OR 2·135, 1·342 to 3·396). There were no treatment-related deaths. INTERPRETATION: 2 days of postoperative intravenous antibiotics for complex appendicitis is non-inferior to 5 days in terms of infectious complications and mortality within 90 days, based on a non-inferiority margin of 7·5%. These findings apply to laparoscopic appendicectomy conducted in a well resourced health-care setting. Adopting this strategy will reduce adverse effects of antibiotics and length of hospital stay. FUNDING: The Netherlands Organization for Health Research and Development.

The 2021 Dutch Working Party on Antibiotic Policy (SWAB) guidelines for empirical antibacterial therapy of sepsis in adults.

BACKGROUND: The Dutch Working Party on Antibiotic Policy (SWAB) in collaboration with relevant professional societies, has updated their evidence-based guidelines on empiric antibacterial therapy of sepsis in adults. METHODS: Our multidisciplinary guideline committee generated ten population, intervention, comparison, and outcome (PICO) questions relevant for adult patients with sepsis. For each question, a literature search was performed to obtain the best available evidence and assessed using the Grading of Recommendations Assessment, Development, and Evaluation (GRADE) system. The quality of evidence for clinically relevant outcomes was graded from high to very low. In structured consensus meetings, the committee formulated recommendations as strong or weak. When evidence could not be obtained, recommendations were provided based on expert opinion and experience (good practice statements). RESULTS: Fifty-five recommendations on the antibacterial therapy of sepsis were generated. Recommendations on empiric antibacterial therapy choices were differentiated for sepsis according to the source of infection, the potential causative pathogen and its resistance pattern. One important revision was the distinction between low, increased and high risk of infection with Enterobacterales resistant to third generation cephalosporins (3GRC-E) to guide the choice of empirical therapy. Other new topics included empirical antibacterial therapy in patients with a reported penicillin allergy and the role of pharmacokinetics and pharmacodynamics to guide dosing in sepsis. We also established recommendations on timing and duration of antibacterial treatment. CONCLUSIONS: Our multidisciplinary committee formulated evidence-based recommendations for the empiric antibacterial therapy of adults with sepsis in The Netherlands.

Enterobacteriaceae and Bacteroidaceae provide resistance to travel-associated intestinal colonization by multi-drug resistant Escherichia coli.

Previous studies have shown high acquisition risks of extended-spectrum beta-lactamase-producing Enterobacteriaceae (ESBL-E) among international travelers visiting antimicrobial resistance (AMR) hotspots. Although antibiotic use and travelers' diarrhea have shown to influence the ESBL-E acquisition risk, it remains largely unknown whether successful colonization of ESBL-E during travel is associated with the composition, functional capacity and resilience of the traveler's microbiome. The microbiome of pre- and post-travel fecal samples from 190 international travelers visiting Africa or Asia was profiled using whole metagenome shotgun sequencing. A metagenomics species concept approach was used to determine the microbial composition, population diversity and functional capacity before travel and how it is altered longitudinally. Eleven travelers were positive for ESBL-E before travel and removed from the analysis. Neither the microbial richness (Chao1), diversity (effective Shannon) and community structure (Bray-Curtis dissimilarity) in pretravel samples nor the longitudinal change of these metrics during travel were predictive for ESBL-E acquisition. A zero-inflated two-step beta-regression model was used to determine how the longitudinal change in both prevalence and abundance of each taxon was related to ESBL acquisition. There were detected increases in both the prevalence and abundance of Citrobacter freundii and two members of the genus Bacteroides, in association with remaining uncolonized by ESBL-E. These results highlight the potential of these individual microbes as a microbial consortium to prevent the acquisition of ESBL-E. The ability to alter a person's colonization resistance to a bacterium could be key to intervention strategies that aim to minimize the spread of MDR bacteria.

[A new definition of the I-category in the antibiogram: I-category antibiotics no longer an option to avoid]. / Nieuwe definitie van I-categorie in het antibiogram.

Currently, the meaning of the I-category of the antibiogram in culture results is changing in The Netherlands. Before, the I-category was called 'intermediate' and included combinations of antibiotics and micro-organisms in which the chance of therapeutic success was doubtful. Therefore, in clinical practice this category was mostly avoided and considered as 'in this case not clear'. From now on, the definition of I has changed to 'susceptible, increased exposure' and can be considered as a valid treatment option when adequate (higher) dosing is applied. Because of the large-scale implementation, not all microbiological laboratories may be able to implement the new I at the same moment and the way they may execute this may differ in detail. When considering treating a patient with 'susceptible, increased exposure' and you are in doubt whether your microbiological laboratory already has implemented the new definition of I or about the correct dosage, consult your clinical microbiologist or Antimicrobial Stewardship Team.

Destination shapes antibiotic resistance gene acquisitions, abundance increases, and diversity changes in Dutch travelers.

BACKGROUND: Antimicrobial-resistant bacteria and their antimicrobial resistance (AMR) genes can spread by hitchhiking in human guts. International travel can exacerbate this public health threat when travelers acquire AMR genes endemic to their destinations and bring them back to their home countries. Prior studies have demonstrated travel-related acquisition of specific opportunistic pathogens and AMR genes, but the extent and magnitude of travel's effects on the gut resistome remain largely unknown. METHODS: Using whole metagenomic shotgun sequencing, functional metagenomics, and Dirichlet multinomial mixture models, we investigated the abundance, diversity, function, resistome architecture, and context of AMR genes in the fecal microbiomes of 190 Dutch individuals, before and after travel to diverse international locations. RESULTS: Travel markedly increased the abundance and α-diversity of AMR genes in the travelers' gut resistome, and we determined that 56 unique AMR genes showed significant acquisition following international travel. These acquisition events were biased towards AMR genes with efflux, inactivation, and target replacement resistance mechanisms. Travel-induced shaping of the gut resistome had distinct correlations with geographical destination, so individuals returning to The Netherlands from the same destination country were more likely to have similar resistome features. Finally, we identified and detailed specific acquisition events of high-risk, mobile genetic element-associated AMR genes including qnr fluoroquinolone resistance genes, blaCTX-M family extended-spectrum ß-lactamases, and the plasmid-borne mcr-1 colistin resistance gene. CONCLUSIONS: Our results show that travel shapes the architecture of the human gut resistome and results in AMR gene acquisition against a variety of antimicrobial drug classes. These broad acquisitions highlight the putative risks that international travel poses to public health by gut resistome perturbation and the global spread of locally endemic AMR genes.

Non-adherence to antimicrobial guidelines in patients with bloodstream infection visiting the emergency department.

OBJECTIVE: Non-adherence to antimicrobial guidelines in patients with bloodstream infection can result in undertreatment, overtreatment, or equivalent treatment, and could lead to suboptimal care. Our aim was to examine the association between non-adherence and appropriate coverage as well as to assess the impact of non-adherence on 30-day mortality. METHODS: We conducted a retrospective cohort study between 2012 and 2017 at a tertiary university hospital. Adult patients attending the emergency department with a bloodstream infection were included. Adherence was defined as guideline-recommended antibiotic therapy. Non-adherence was either undertreatment (too narrow-spectrum), overtreatment (too broad-spectrum), or equivalent treatment. Outcomes were appropriate coverage (i.e. antibiotic therapy that matches in vitro susceptibility of the isolated bacteria) and 30-day mortality. RESULTS: We included 909 patients of whom 395 (43.5%) were treated adherently, 355 (39.1%) were undertreated, 87 (9.6%) were overtreated, and 72 (7.9%) received an equivalent treatment. Overtreated patients were more severely ill, whilst undertreated patients had more favorable patient characteristics. Overtreatment did not result in higher appropriate coverage, whereas undertreatment was associated with lower coverage (OR[95%CI]: 0.18 [0.12; 0.26]). Overtreatment and undertreatment were not associated with 30-day mortality. CONCLUSIONS: Guideline adherence likely depends on disease severity, because overtreatment was more often observed in patients with high disease severity and undertreatment in less severely ill patients. Undertreatment was associated lower appropriate coverage but not with higher mortality. However, this can be the result of residual confounding . Overtreatment did not result in higher appropriate antibiotic coverage nor a survival benefit . Therefore, overtreatment seems not justifiable.

Author Correction: Global phylogenetic analysis of Escherichia coli and plasmids carrying the mcr-1 gene indicates bacterial diversity but plasmid restriction.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Low antimicrobial resistance in general practice patients in Rotterdam, the city with the largest proportion of immigrants in the Netherlands.

Antimicrobial resistance (AMR) is an increasing problem. The prevalence of antimicrobial resistance in general practice patients is expected to be relatively high in Rotterdam, the Dutch city with the largest proportion non-Western immigrants. The aim of this study was to assess the prevalence of antibiotic-resistant uropathogens (Escherichia coli, Klebsiella pneumoniae, and Proteus mirabilis) in general practices in Rotterdam, and to find a possible association between the prevalence of antibiotic-resistant E. coli and age, gender, and socioeconomic status (SES). A retrospective analysis was performed of urine samples from general practice patients in 2016. The prevalence of AMR in uropathogens was compared with national resistance data, as was the prevalence of highly and multidrug resistant and extended spectrum ß-lactamase (ESBL) producing E. coli and K. pneumoniae. Univariate logistic regression was used to study associations between antibiotic-resistant E. coli and age, gender, and SES area score. No clinically relevant differences were observed in the prevalence of antibiotic-resistant uropathogens in Rotterdam compared with the national prevalence. For E. coli and K. pneumoniae, the prevalence was 3.6% for ESBL production (both pathogens together), while the prevalence ranged between 4.2%-5.0% for high resistance and between 1.2%-3.3% for multidrug resistance. Ciprofloxacin-resistant E. coli was significantly associated with higher age. Although Rotterdam has a high percentage of non-western immigrants and a low SES, AMR is low among general practice patients. This indicates that adherence to national guidelines in general practice enables maintenance of low AMR, even in high-risk populations.

Prevalence and risk factors for carriage of ESBL-producing Enterobacteriaceae in a population of Dutch travellers: A cross-sectional study.

BACKGROUND: We investigated prevalence and predictive factors for ESBL-E carriage in a population of mostly travellers prior to their travel (n = 2216). In addition, we examined ESBL genotype before travel and compared these to returning travellers. METHOD: A questionnaire and faecal sample were collected before travel, and a second faecal sample was collected immediately after travel. Faecal samples were analysed for ESBL-E, with genotypic characterization by PCR and sequencing. Risk factors for ESBL-E carriage prior to travel were identified by logistic regression analyses. RESULTS: Before travel, 136 participants (6.1%) were colonized with ESBL-E. Antibiotic use in the past three months (ORadjusted 2.57; 95% CI 1.59-4.16) and travel outside of Europe in the past year (1.92, 1.28-2.87) were risk factors for ESBL-E colonisation prior to travel. Travel outside of Europe carried the largest attributable risk (39.8%). Prior to travel 31.3% (40/128) of participants carried blaCTX-M 15 and 21.9% (28/128) blaCTX-M 14/18. In returning travellers 633 acquired ESBL-E of who 53.4% (338/633) acquired blaCTX-M 15 and 17.7% (112/633) blaCTX-M 14/18. CONCLUSION: In our population of Dutch travellers we found a pre-travel ESBL-E prevalence of 6.1%. Prior to travel, previous antibiotic use and travel outside of Europe were the strongest independent predictors for ESBL-E carriage, with travel outside of Europe carrying the largest attributable risk. Our molecular results suggest ESBL genes found in our study population prior to travel were in large part travel related.

Appropriate empirical antibiotic therapy and mortality: Conflicting data explained by residual confounding.

OBJECTIVE: Clinical practice universally assumes that appropriate empirical antibiotic therapy improves survival in patients with bloodstream infection. However, this is not generally supported by previous studies. We examined the association between appropriate therapy and 30-day mortality, while minimizing bias due to confounding by indication. METHODS: We conducted a retrospective cohort study between 2012 and 2017 at a tertiary university hospital in the Netherlands. Adult patients with bloodstream infection attending the emergency department were included. Based on in vitro susceptibility, antibiotic therapy was scored as appropriate or inappropriate. Primary outcome was 30-day mortality. To control for confounding, we performed conventional multivariable logistic regression and propensity score methods. Additionally, we performed an analysis in a more homogeneous subgroup (i.e. antibiotic monotherapy). RESULTS: We included 1.039 patients, 729 (70.2%) received appropriate therapy. Overall 30-day mortality was 10.4%. Appropriately treated patients had more unfavorable characteristics, indicating more severe illness. Despite adjustments, we found no association between appropriate therapy and mortality. For the antibiotic monotherapy subgroup (n = 449), patient characteristics were more homogeneous. Within this subgroup, appropriate therapy was associated with lower mortality (Odds Ratios [95% Confidence Intervals] ranging from: 0.31 [0.14; 0.67] to 0.40 [0.19; 0.85]). CONCLUSIONS: Comparing heterogeneous treatment groups distorts associations despite use of common methods to prevent bias. Consequently, conclusions of such observational studies should be interpreted with care. If possible, future investigators should use our method of attempting to identify and analyze the most homogeneous treatment groups nested within their study objective, because this minimizes residual confounding.

Risk of acquisition of human diarrhoeagenic Escherichia coli virulence genes in intercontinental travellers: A prospective, multi-centre study.

BACKGROUND: We studied geographic distribution of diarrhoeagenic Escherichia coli virulence genes (DEC VGs) acquisition in travellers and investigated if they acquired highly virulent EAEC/STEC hybrid strains. METHODS: From the prospective, multicentre COMBAT study among 2001 Dutch travellers, 491 travellers were selected based on travel destination to 7 subregions. Faecal samples taken directly before and after travel were screened for nine DEC VGs with real-time PCR. Incidence proportions and rates were calculated for each gene and subregion. RESULTS: 479 travellers were analysed. 21.8% acquired aggR (EAEC), with highest acquisition rates in Northern and Western Africa and 15.3% acquired eae (STEC/EPEC) with highest rates in travellers to Western and Eastern Africa. ETEC (elt or est gene) was acquired by 4.2% of travellers and acquisition of est was associated with traveller's diarrhoea. Overall, the risk of acquiring DEC VGs was low in Southern Africa and South America. Although the combination of aggR (EAEC) and stx1/2 (STEC) was acquired by 3 travellers, these genes could not be detected together in a single E. coli strain. CONCLUSIONS: The risk of acquisition of DEC VGs strongly depends on the travel destination, with those travelling to Africa - except Southern Africa - having a higher risk.

Two versus five days of antibiotics after appendectomy for complex acute appendicitis (APPIC): study protocol for a randomized controlled trial.

BACKGROUND: Acute appendicitis is one of the most common indications for emergency surgery. In patients with a complex appendicitis, prolonged antibiotic prophylaxis is recommended after appendectomy. There is no consensus regarding the optimum duration of antibiotics. Guidelines propose 3 to 7 days of treatment, but shorter courses may be as effective in the prevention of infectious complications. At the same time, the global issue of increasing antimicrobial resistance urges for optimization of antibiotic strategies. The aim of this study is to determine whether a short course (48 h) of postoperative antibiotics is non-inferior to current standard practice of 5 days. METHODS: Patients of 8 years and older undergoing appendectomy for acute complex appendicitis - defined as a gangrenous and/or perforated appendicitis or appendicitis in presence of an abscess - are eligible for inclusion. Immunocompromised or pregnant patients are excluded, as well as patients with a contraindication to the study antibiotics. In total, 1066 patients will be randomly allocated in a 1:1 ratio to the experimental treatment arm (48 h of postoperative intravenously administered (IV) antibiotics) or the control arm (5 days of postoperative IV antibiotics). After discharge from the hospital, patients participate in a productivity-cost-questionnaire at 4 weeks and a standardized telephone follow-up at 90 days after appendectomy. The primary outcome is a composite endpoint of infectious complications, including intra-abdominal abscess (IAA) and surgical site infection (SSI), and mortality within 90 days after appendectomy. Secondary outcomes include IAA, SSI, restart of antibiotics, length of hospital stay (LOS), reoperation, percutaneous drainage, readmission rate, and cost-effectiveness. The non-inferiority margin for the difference in the primary endpoint rate is set at 7.5% (one-sided test at ɑ 0.025). Both per-protocol and intention-to-treat analyses will be performed. DISCUSSION: This trial will provide evidence on whether 48 h of postoperative antibiotics is non-inferior to a standard course of 5 days of antibiotics. If non-inferiority is established, longer intravenous administration following appendectomy for complex appendicitis can be abandoned, and guidelines need to be adjusted accordingly. TRIAL REGISTRATION: Dutch Trial Register, NTR6128 . Registered on 20 December 2016.

Global phylogenetic analysis of Escherichia coli and plasmids carrying the mcr-1 gene indicates bacterial diversity but plasmid restriction.

To understand the dynamics behind the worldwide spread of the mcr-1 gene, we determined the population structure of Escherichia coli and of mobile genetic elements (MGEs) carrying the mcr-1 gene. After a systematic review of the literature we included 65 E. coli whole genome sequences (WGS), adding 6 recently sequenced travel related isolates, and 312 MLST profiles. We included 219 MGEs described in 7 Enterobacteriaceae species isolated from human, animal and environmental samples. Despite a high overall diversity, 2 lineages were observed in the E. coli population that may function as reservoirs of the mcr-1 gene, the largest of which was linked to ST10, a sequence type known for its ubiquity in human faecal samples and in food samples. No genotypic clustering by geographical origin or isolation source was observed. Amongst a total of 13 plasmid incompatibility types, the IncI2, IncX4 and IncHI2 plasmids accounted for more than 90% of MGEs carrying the mcr-1 gene. We observed significant geographical clustering with regional spread of IncHI2 plasmids in Europe and IncI2 in Asia. These findings point towards promiscuous spread of the mcr-1 gene by efficient horizontal gene transfer dominated by a limited number of plasmid incompatibility types.

Travel-related acquisition of diarrhoeagenic bacteria, enteral viruses and parasites in a prospective cohort of 98 Dutch travellers.

BACKGROUND: Limited prospective data are available on the acquisition of viral, bacterial and parasitic diarrhoeagenic agents by healthy individuals during travel. METHODS: To determine the frequency of travel associated acquisition of 19 pathogens in 98 intercontinental travellers, qPCR was used to detect 8 viral pathogens, 6 bacterial enteric pathogens and 5 parasite species in faecal samples collected immediately before and after travel. RESULTS: We found high pre-travel carriage rates of Blastocystis spp. and Dientamoeba fragilis of 32% and 19% respectively. Pre-travel prevalences of all other tested pathogens were below 3%. Blastocystis spp. (10%), Plesiomonas shigelloides (7%), D. fragilis (6%) and Shigella spp. (5%) were the most frequently acquired pathogens and acquisition of enteral viruses and hepatitis E virus in this relatively small group of travellers was rare or non-existent. CONCLUSIONS: Our findings suggest that the role of viruses as the cause of persisting traveller's diarrhoea is limited and bacterial pathogens are more likely as a cause of traveller's diarrhoea. The substantial proportion of travellers carrying Blastocystis spp. and D. fragilis before travel warrants cautious interpretation of positive samples in returning travellers with gastrointestinal complaints.

Import and spread of extended-spectrum ß-lactamase-producing Enterobacteriaceae by international travellers (COMBAT study): a prospective, multicentre cohort study.

BACKGROUND: International travel contributes to the dissemination of antimicrobial resistance. We investigated the acquisition of extended-spectrum ß-lactamase-producing Enterobacteriaceae (ESBL-E) during international travel, with a focus on predictive factors for acquisition, duration of colonisation, and probability of onward transmission. METHODS: Within the prospective, multicentre COMBAT study, 2001 Dutch travellers and 215 non-travelling household members were enrolled. Faecal samples and questionnaires on demographics, illnesses, and behaviour were collected before travel and immediately and 1, 3, 6, and 12 months after return. Samples were screened for the presence of ESBL-E. In post-travel samples, ESBL genes were sequenced and PCR with specific primers for plasmid-encoded ß-lactamase enzymes TEM, SHV, and CTX-M group 1, 2, 8, 9, and 25 was used to confirm the presence of ESBL genes in follow-up samples. Multivariable regression analyses and mathematical modelling were used to identify predictors for acquisition and sustained carriage, and to determine household transmission rates. This study is registered with ClinicalTrials.gov, number NCT01676974. FINDINGS: 633 (34·3%) of 1847 travellers who were ESBL negative before travel and had available samples after return had acquired ESBL-E during international travel (95% CI 32·1-36·5), with the highest number of acquisitions being among those who travelled to southern Asia in 136 of 181 (75·1%, 95% CI 68·4-80·9). Important predictors for acquisition of ESBL-E were antibiotic use during travel (adjusted odds ratio 2·69, 95% CI 1·79-4·05), traveller's diarrhoea that persisted after return (2·31, 1·42-3·76), and pre-existing chronic bowel disease (2·10, 1·13-3·90). The median duration of colonisation after travel was 30 days (95% CI 29-33). 65 (11·3%) of 577 remained colonised at 12 months. CTX-M enzyme group 9 ESBLs were associated with a significantly increased risk of sustained carriage (median duration 75 days, 95% CI 48-102, p=0·0001). Onward transmission was found in 13 (7·7%) of 168 household members. The probability of transmitting ESBL-E to another household member was 12% (95% CI 5-18). INTERPRETATION: Acquisition and spread of ESBL-E during and after international travel was substantial and worrisome. Travellers to areas with a high risk of ESBL-E acquisition should be viewed as potential carriers of ESBL-E for up to 12 months after return. FUNDING: Netherlands Organisation for Health Research and Development (ZonMw).

Prolonged carriage and potential onward transmission of carbapenemase-producing Enterobacteriaceae in Dutch travelers.

AIM: The aim was to study acquisition and persistence of carbapenemase-producing Enterobacteriaceae (CPE) among travelers. MATERIALS & METHODS: Stools from 2001 travelers and 215 nontraveling household members, collected before and immediately post-travel as well as 1, 3, 6 and 12 months upon return, were screened for CPE. RESULTS: Five travelers, all visiting Asia outside the Indian subcontinent, acquired CPE. One traveler persistently carried the same OXA-244 CPE up to 6 months post-travel. Three months after travel, her co-traveling spouse also became positive for this OXA-244 CPE strain, suggesting clonal transmission within this household. CONCLUSION: Acquisition of CPE is not restricted to travelers to the Indian subcontinent and/or to travelers seeking healthcare during travel and can persist up to at least 6 months post-travel.

The Carriage Of Multiresistant Bacteria After Travel (COMBAT) prospective cohort study: methodology and design.

BACKGROUND: Antimicrobial resistance (AMR) is one of the major threats to public health around the world. Besides the intense use and misuse of antimicrobial agents as the major force behind the increase in antimicrobial resistance, the exponential increase of international travel may also substantially contribute to the emergence and spread of AMR. However, knowledge on the extent to which international travel contributes to this is still limited. The Carriage Of Multiresistant Bacteria After Travel (COMBAT) study aims to 1. determine the acquisition rate of multiresistant Enterobacteriaceae during foreign travel 2. ascertain the duration of carriage of these micro-organisms 3. determine the transmission rate within households 4. identify risk factors for acquisition, persistence of carriage and transmission of multiresistant Enterobacteriaceae. METHODS/DESIGN: The COMBAT-study is a large-scale multicenter longitudinal cohort study among travellers (n = 2001) and their non-travelling household members (n = 215). Faecal samples are collected before and immediately after travel and 1 month after return from all participants. Follow-up faecal samples are collected 3, 6 and 12 months after return from travellers (and their non-travelling household members) who acquired multiresistant Enterobacteriaceae. Questionnaires are collected from all participants at each time-point. Faecal samples are screened phenotypically for the presence of extended-spectrum beta-lactamase (ESBL) or carbapenemase-producing Enterobacteriaceae. Positive post-travel isolates from travellers with negative pre-travel samples are genotypically analysed for ESBL and carbapenemase genes with microarray and gene sequencing. DISCUSSION: The design and scale of the COMBAT-study will enable us to provide much needed detailed insights into the risks and dynamics of introduction and spread of ESBL- and carbapenemase-producing Enterobacteriaceae by healthy travellers and the potential need and measures to monitor or manage these risks. TRIAL REGISTRATION: The study is registered at clinicaltrials.gov under accession number NCT01676974.

Antibiotic trapping by plasmid-encoded CMY-2 ß-lactamase combined with reduced outer membrane permeability as a mechanism of carbapenem resistance in Escherichia coli.

A liver transplant patient was admitted with cholangitis, for which meropenem therapy was started. Initial cultures showed a carbapenem-susceptible (CS) Escherichia coli strain, but during admission, a carbapenem-resistant (CR) E. coli strain was isolated. Analysis of the outer membrane protein profiles showed that both CS and CR E. coli lacked the porins OmpF and OmpC. Furthermore, PCR and sequence analysis revealed that both CS and CR E. coli possessed bla(CTX-M-15) and bla(OXA-1). The CR E. coli strain additionally harbored bla(CMY-2) and demonstrated a >15-fold increase in ß-lactamase activity against nitrocefin, but no hydrolysis of meropenem was detected. However, nitrocefin hydrolysis appeared strongly inhibited by meropenem. Furthermore, the CMY-2 enzyme demonstrated lower electrophoretic mobility after its incubation either in vitro or in vivo with meropenem, indicative of its covalent modification with meropenem. The presence of the acyl-enzyme complex was confirmed by mass spectrometry. By transformation of the CMY-2-encoding plasmid into various E. coli strains, it was established that both porin deficiency and high-level expression of the enzyme were needed to confer meropenem resistance. In conclusion, carbapenem resistance emerged by a combination of elevated ß-lactamase production and lack of porin expression. Due to the reduced outer membrane permeability, only small amounts of meropenem can enter the periplasm, where they are trapped but not degraded by the large amount of the ß-lactamase. This study, therefore, provides evidence that the mechanism of "trapping" by CMY-2 ß-lactamase plays a role in carbapenem resistance.