WP1 System Sampling & Characterization (DTU-ENV, NCU, USC, UoB)
Objectives:
- Describe the three sewer catchments in Denmark, the United Kingdom, and Spain
- Design and execute sampling campaigns (preliminary, phase 1, phase 2) in these catchments
- Construct, distribute and curate samples
- Perform chemical analyses of samples including heavy metal and antibiotic content
- Perform meta-data collection on antibiotic consumption in the three catchments
WP2 Inventory and Quantification of the AMRG and AMR gene-carrying MGEs (UCPH, USC, CHUS, DTU-CBS, NCL)
Objectives:
- Deeply characterize microbial community diversity in each UWS ecosystem, including phageomes and plasmidomes (all sequences associated with phages and plasmids, respectively)
- Quantify clinically relevant AMR genes and compare their occurrence between and across UWSs.
- Characterize clinically-relevant AMRGs from core locations in the UWS to:
- Reveal the diversity of β-lactamases and carbapenemases genes
- Identify their location (i.e. presence on a bacterial chromosome, a plasmid or a phage) and describe their context (i.e. flanking genes indicating co-selection or potential for mobility)
WP3 Host Identification and Realized Transfer (NCU, DTU-CBS, DTU-ENV, CHUS)
Objectives:
- Isolate and obtain carbapenem- and β-lactam-resistant strains from urban water systems (UWS)
- Characterize resistant hosts physiologically and genomically, and determine their relative pathogenicity
- Identify the most probable host(s) of key MGEs (from WP2) by datamining the WP2 metagenomes
- Infer the transfer history of AMR within and between each UWS ecosystem compartment, using AMR isolates, and AMR and MGE gene data from across each system
WP4 Transfer Potential (DTU-ENV, USC, UCPH)
Objectives:
- Establish the potential of the UWS communities to acquire and mobilize plasmids and phages similar to those identified in WP2;
- Establish potential transfer host ranges (diversity of bacteria that can take up mobile genetic elements) of these plasmids and phages;
- Compare these potential transfer host ranges to the realized host ranges (from WP3)
WP5 Fitness Effects of the AMR determinants (USC, NCU)
Objectives:
- Information on genomic context (WP2), and identified hosts (WP3) will be used to screen the most abundant or persistent plasmids or phages to quantify:
- effects of plasmid or phage carriage on bacterial physiology (metabolic burden and environmental fitness
- effects of plasmids and phages on biofilm formation
- plasmid and phage stability
WP6 Data integration, Modelling and Microbial Risk Assessment (UoB 24, RHCC 15, CHUS 3, all partners)
Objectives:
- Build a modelling framework, conduct parameter and structural sensitivity analyses (how important are HGT, ARG selection, competition and dilution in determining system behaviour?)
- Infer parameters and confidence intervals to answer quantitative questions (do virulence genes decline more rapidly than resistance genes? how strong is co-selection for multiple resistances?)
- Evaluate potential mitigation strategies (how likely would separate treatment of hospital sewage reduce co-selection, how much would replacing the activated sludge stage at the WWTP (DK, ES) with trickling filters (UK) reduce pathogen/ARG transmission risks?)
- Conduct risk analysis for public health (with what probability would HGT of ARGs between pathogens and environmental strains occur? how high are the risks of resistance genes and/or pathogens leaving the WWTP to enter sources of drinking water? what are the consequences to public health? can UWS sampling serve to monitor resistance in the population? can novel AMR detected in UWS precede and predict new AMR trends in the population?)