• The antimicrobial resistome in relation to antimicrobial use and biosecurity in pig farming, a metagenome-wide association study in nine European countries.

      Van Gompel, Liese; Luiken, Roosmarijn E C; Sarrazin, Steven; Munk, Patrick; Knudsen, Berith E; Hansen, Rasmus B; Bossers, Alex; Aarestrup, Frank M; Dewulf, Jeroen; Wagenaar, Jaap A; et al. (2019-04-01)
      Previous studies in food-producing animals have shown associations between antimicrobial use (AMU) and resistance (AMR) in specifically isolated bacterial species. Multi-country data are scarce and only describe between-country differences. Here we investigate associations between the pig faecal mobile resistome and characteristics at the farm-level across Europe. A cross-sectional study was conducted among 176 conventional pig farms from nine European countries. Twenty-five faecal samples from fattening pigs were pooled per farm and acquired resistomes were determined using shotgun metagenomics and the Resfinder reference database, i.e. the full collection of horizontally acquired AMR genes (ARGs). Normalized fragments resistance genes per kilobase reference per million bacterial fragments (FPKM) were calculated. Specific farm-level data (AMU, biosecurity) were collected. Random-effects meta-analyses were performed by country, relating farm-level data to relative ARG abundances (FPKM). Total AMU during fattening was positively associated with total ARG (total FPKM). Positive associations were particularly observed between widely used macrolides and tetracyclines, and ARGs corresponding to the respective antimicrobial classes. Significant AMU-ARG associations were not found for β-lactams and only few colistin ARGs were found, despite high use of these antimicrobial classes in younger pigs. Increased internal biosecurity was directly related to higher abundances of ARGs mainly encoding macrolide resistance. These effects of biosecurity were independent of AMU in mutually adjusted models. Using resistome data in association studies is unprecedented and adds accuracy and new insights to previously observed AMU-AMR associations. Major components of the pig resistome are positively and independently associated with on-farm AMU and biosecurity conditions.
    • Author Correction: Setting a baseline for global urban virome surveillance in sewage.

      Nieuwenhuijse, David F; Oude Munnink, Bas B; Phan, My V T; Munk, Patrick; Venkatakrishnan, Shweta; Aarestrup, Frank M; Cotten, Matthew; Koopmans, Marion P G (2021-08-25)
    • Description and determinants of the faecal resistome and microbiome of farmers and slaughterhouse workers: A metagenome-wide cross-sectional study.

      Van Gompel, Liese; Luiken, Roosmarijn E C; Hansen, Rasmus B; Munk, Patrick; Bouwknegt, Martijn; Heres, Lourens; Greve, Gerdit D; Scherpenisse, Peter; Jongerius-Gortemaker, Betty G M; Tersteeg-Zijderveld, Monique H G; et al. (2020-07-14)
    • Farm dust resistomes and bacterial microbiomes in European poultry and pig farms.

      Luiken, Roosmarijn E C; Van Gompel, Liese; Bossers, Alex; Munk, Patrick; Joosten, Philip; Hansen, Rasmus Borup; Knudsen, Berith E; García-Cobos, Silvia; Dewulf, Jeroen; Aarestrup, Frank M; et al. (2020-07-29)
    • Global monitoring of antimicrobial resistance based on metagenomics analyses of urban sewage.

      Hendriksen, Rene S; Munk, Patrick; Njage, Patrick; van Bunnik, Bram; McNally, Luke; Lukjancenko, Oksana; Röder, Timo; Nieuwenhuijse, David; Pedersen, Susanne Karlsmose; Kjeldgaard, Jette; et al. (2019-03-08)
      Antimicrobial resistance (AMR) is a serious threat to global public health, but obtaining representative data on AMR for healthy human populations is difficult. Here, we use metagenomic analysis of untreated sewage to characterize the bacterial resistome from 79 sites in 60 countries. We find systematic differences in abundance and diversity of AMR genes between Europe/North-America/Oceania and Africa/Asia/South-America. Antimicrobial use data and bacterial taxonomy only explains a minor part of the AMR variation that we observe. We find no evidence for cross-selection between antimicrobial classes, or for effect of air travel between sites. However, AMR gene abundance strongly correlates with socio-economic, health and environmental factors, which we use to predict AMR gene abundances in all countries in the world. Our findings suggest that global AMR gene diversity and abundance vary by region, and that improving sanitation and health could potentially limit the global burden of AMR. We propose metagenomic analysis of sewage as an ethically acceptable and economically feasible approach for continuous global surveillance and prediction of AMR.
    • Proficiency Testing of Virus Diagnostics Based on Bioinformatics Analysis of Simulated High-Throughput Sequencing Data Sets.

      Brinkmann, Annika; Andrusch, Andreas; Belka, Ariane; Wylezich, Claudia; Höper, Dirk; Pohlmann, Anne; Nordahl Petersen, Thomas; Lucas, Pierrick; Blanchard, Yannick; Papa, Anna; et al. (2019-08-01)
      Quality management and independent assessment of high-throughput sequencing-based virus diagnostics have not yet been established as a mandatory approach for ensuring comparable results. The sensitivity and specificity of viral high-throughput sequence data analysis are highly affected by bioinformatics processing using publicly available and custom tools and databases and thus differ widely between individuals and institutions. Here we present the results of the COMPARE [Collaborative Management Platform for Detection and Analyses of (Re-)emerging and Foodborne Outbreaks in Europe] in silico virus proficiency test. An artificial, simulated in silico data set of Illumina HiSeq sequences was provided to 13 different European institutes for bioinformatics analysis to identify viral pathogens in high-throughput sequence data. Comparison of the participants' analyses shows that the use of different tools, programs, and databases for bioinformatics analyses can impact the correct identification of viral sequences from a simple data set. The identification of slightly mutated and highly divergent virus genomes has been shown to be most challenging. Furthermore, the interpretation of the results, together with a fictitious case report, by the participants showed that in addition to the bioinformatics analysis, the virological evaluation of the results can be important in clinical settings. External quality assessment and proficiency testing should become an important part of validating high-throughput sequencing-based virus diagnostics and could improve the harmonization, comparability, and reproducibility of results. There is a need for the establishment of international proficiency testing, like that established for conventional laboratory tests such as PCR, for bioinformatics pipelines and the interpretation of such results.
    • Secrets of the Hospital Underbelly: Patterns of Abundance of Antimicrobial Resistance Genes in Hospital Wastewater Vary by Specific Antimicrobial and Bacterial Family.

      Perry, Meghan R; Lepper, Hannah C; McNally, Luke; Wee, Bryan A; Munk, Patrick; Warr, Amanda; Moore, Barbara; Kalima, Pota; Philip, Carol; de Roda Husman, Ana Maria; et al. (2021-09-10)
      Background: Hospital wastewater is a major source of antimicrobial resistance (AMR) outflow into the environment. This study uses metagenomics to study how hospital clinical activity impacts antimicrobial resistance genes (ARGs) abundances in hospital wastewater. Methods: Sewage was collected over a 24-h period from multiple wastewater collection points (CPs) representing different specialties within a tertiary hospital site and simultaneously from community sewage works. High throughput shotgun sequencing was performed using Illumina HiSeq4000. ARG abundances were correlated to hospital antimicrobial usage (AMU), data on clinical activity and resistance prevalence in clinical isolates. Results: Microbiota and ARG composition varied between CPs and overall ARG abundance was higher in hospital wastewater than in community influent. ARG and microbiota compositions were correlated (Procrustes analysis, p=0.014). Total antimicrobial usage was not associated with higher ARG abundance in wastewater. However, there was a small positive association between resistance genes and antimicrobial usage matched to ARG phenotype (IRR 1.11, CI 1.06-1.16, p<0.001). Furthermore, analyzing carbapenem and vancomycin resistance separately indicated that counts of ARGs to these antimicrobials were positively associated with their increased usage [carbapenem rate ratio (RR) 1.91, 95% CI 1.01-3.72, p=0.07, and vancomycin RR 10.25, CI 2.32-49.10, p<0.01]. Overall, ARG abundance within hospital wastewater did not reflect resistance patterns in clinical isolates from concurrent hospital inpatients. However, for clinical isolates of the family Enterococcaceae and Staphylococcaceae, there was a positive relationship with wastewater ARG abundance [odds ratio (OR) 1.62, CI 1.33-2.00, p<0.001, and OR 1.65, CI 1.21-2.30, p=0.006 respectively]. Conclusion: We found that the relationship between hospital wastewater ARGs and antimicrobial usage or clinical isolate resistance varies by specific antimicrobial and bacterial family studied. One explanation, we consider is that relationships observed from multiple departments within a single hospital site will be detectable only for ARGs against parenteral antimicrobials uniquely used in the hospital setting. Our work highlights that using metagenomics to identify the full range of ARGs in hospital wastewater is a useful surveillance tool to monitor hospital ARG carriage and outflow and guide environmental policy on AMR.
    • Setting a baseline for global urban virome surveillance in sewage.

      Nieuwenhuijse, David F; Oude Munnink, Bas B; Phan, My V T; Munk, Patrick; Venkatakrishnan, Shweta; Aarestrup, Frank M; Cotten, Matthew; Koopmans, Marion P G (2020-08-13)