The antimicrobial resistance profiles of Escherichia coli and Pseudomonas aeruginosa isolated from private groundwater wells in the Republic of Ireland.

The role of the natural environment in the dissemination of antimicrobial resistant bacteria has been increasingly recognised in the literature. However, knowledge surrounding the critical factors and mechanisms mediating their occurrence is still limited, particularly in relatively 'pristine' groundwater environments. In the Republic of Ireland (RoI), a country characterised by high groundwater reliance, household-based (unregulated) wells provide drinking water to 11% of the population. These private wells are generally located in rural areas, where the risk of microbiological contamination is high due to intensive agricultural practices and high reliance on domestic wastewater treatment systems; both of which are also potential sources of antimicrobials and antimicrobial resistant bacteria. Accordingly, the current research sought to elucidate current rates of antimicrobial resistant bacteria and the principal factors associated with their presence in private wells in the RoI. A total of 250 samples (from 132 wells nationwide) were assessed for the presence of faecal (Escherichia coli) and environmental (Pseudomonas aeruginosa) bacteria, with single isolates from each contaminated sample tested phenotypically against 18 and 9 antimicrobials, respectively. Findings show that while 16.7% of E. coli (n = 8/48) were categorically resistant to ≥1 antimicrobial, with a further 79.2% classified as intermediately resistant, no categorical resistance was found among P. aeruginosa isolates (n = 0/6), with just one intermediately resistant isolate detected. Multivariate regression modelling indicates significantly higher odds of resistant E. coli detection in concurrence with elevated cattle density (OR = 1.028, p = 0.032), aligning with findings of highest resistance rates to veterinary antimicrobials (e.g., streptomycin = 14.6%, tetracycline = 12.5%, and ampicillin = 12.5%). Multivariate model results also suggest overland flow culminating in direct wellhead ingress as a primary ingress mechanism for resistant E. coli. Study findings may inform groundwater source protection initiatives and antimicrobial resistance surveillance moving forward.

Characterization of Shiga toxin-producing Escherichia coli presence, serogroups and risk factors from private groundwater sources in western Ireland.

Over recent years, Ireland has reported the highest crude incidence rates of Shiga toxin-producing Escherichia coli (STEC) enteritis in Europe. Unregulated private groundwater sources have emerged as an important potential transmission route for STEC, with up to 750,000 Irish residents reliant on these sources for domestic waters. This study aimed to investigate the prevalence and serogroup profile of STEC contamination from domestic private wells in western Ireland. Fifty-two groundwater sources were analysed during two sampling campaigns in the autumn (September/October) of 2019 (n = 21) and 2021 (n = 31). Untreated groundwater samples (30 L) were collected and analysed using the "CapE" (capture, amplify, extract) method. Extracted DNA was tested using multiplex real-time PCR for Shiga toxin stx1 and/or stx2 and eae genes. STEC positive DNA samples were tested for clinically relevant serogroups by real-time PCR. Data relating to 27 potential groundwater contamination risk factors were geospatially linked to each well and assessed for association with E. coli, stx1 and/or stx2 and eae presence/absence. Overall, 20/52 wells (38.4 %) were positive for E. coli (median concentration 8.5 MPN/100 mL as assessed by Colilert-18 method). Stx1 and/or stx2 was detected in 10/52 (19.2 %) wells overall and 8/20 E. coli positive wells, equating to a STEC to "generic" E. coli detection ratio of 40 %. Six of these wells (30 %) were also positive for eae. One or more serogroup-specific gene targets were identified in all but one stx1 and/or stx2 positive sample, with O145 (n = 6), O157 (n = 5) and O103 (n = 4) most prevalent. STEC presence was significantly associated with decreasing well depth (U = -2.243; p = 0.024) and increasing 30-day mean antecedent rainfall (U = 2.126; p = 0.034). Serogroup O104 was associated with increased sheep density (U = 2.089; p = 0.044) and detection of stx1 and/or stx2 + eae with increased septic tank density (U = 2.246 p = 0.023). Findings indicate high detection rates of clinically relevant STEC in E. coli contaminated groundwater sources in Ireland.

Spatiotemporal dynamics of Escherichia coli presence and magnitude across a national groundwater monitoring network, Republic of Ireland, 2011-2020.

Groundwater is a vital drinking water resource and its protection from microbiological contamination is paramount to safeguard public health. The Republic of Ireland (RoI) is characterised by the highest incidence of verocytotoxigenic Escherichia coli (VTEC) enteritis in the European Union (EU), linked to high reliance on unregulated groundwater sources (~16% of the population). Yet, the spatio-temporal factors influencing the frequency and magnitude of microbial contamination remain largely unknown, with past studies typically constrained to spatio-temporally 'limited' sampling campaigns. Accordingly, the current investigation sought to analyse an extensive spatially distributed time-series (2011-2020) of groundwater monitoring data in the RoI. The dataset, compiled by the Environmental Protection Agency (EPA), showed 'high' contamination rates, with 66.7% (88/132) of supplies testing positive for E. coli, and 29.5% (39/132) exceeding concentrations of 10MPN/100 ml (i.e. gross contamination) at least once during the 10-year monitoring period. Seasonal decomposition analyses indicate that E. coli detection rates peak during late autumn/early winter, coinciding with increases in annual rainfall, while gross contamination peaks in spring (May) and late-summer (August), likely reflecting seasonal shifts in agricultural practices. Mixed effects logistic regression modelling indicates that monitoring sources located in karst limestone are statistically associated with E. coli presence (OR = 2.76, p = 0.03) and gross contamination (OR = 2.54, p = 0.037) when compared to poorly productive aquifers (i.e., transmissivity below 10m2/d). Moreover, 5-day and 30-day antecedent rainfall increased the likelihood of E. coli contamination (OR = 1.027, p < 0.001 and OR = 1.005, p = 0.016, respectively), with the former also being associated with gross contamination (OR = 1.042, p < 0.001). As such, it is inferred that preferential flow and direct ingress of surface runoff are the most likely ingress mechanisms associated with E. coli groundwater supply contamination. The results presented are expected to inform policy change around groundwater source protection and provide insight for the development of groundwater monitoring programmes in geologically heterogeneous regions.

The role of the natural aquatic environment in the dissemination of extended spectrum beta-lactamase and carbapenemase encoding genes: A scoping review.

The natural aquatic environment is a significant contributor to the development and circulation of clinically significant antibiotic resistance genes (ARGs). The potential for the aquatic environment to act as a reservoir for ARG accumulation in areas receiving anthropogenic contamination has been thoroughly researched. However, the emergence of novel ARGs in the absence of external influences, as well as the capacity of environmental bacteria to disseminate ARGs via mobile genetic elements remain relatively unchallenged. In order to address these knowledge gaps, this scoping literature review was established focusing on the detection of two important and readily mobile ARGs, namely, extended spectrum beta-lactamase (ESBL) and carbapenemase genes. This review included 41 studies from 19 different countries. A range of different water bodies including rivers (n = 26), seawaters (n = 6) and lakes (n = 3), amongst others, were analysed in the included studies. ESBL genes were reported in 29/41 (70.7%) studies, while carbapenemase genes were reported in 13/41 (31.7%), including joint reporting in 9 studies. The occurrence of mobile genetic elements was evaluated, which included the detection of integrons (n = 22), plasmids (n = 18), insertion sequences (n = 4) and transposons (n = 3). The ability of environmental bacteria to successfully transfer resistance genes via conjugation was also examined in 11 of the included studies. The findings of this scoping review expose the presence of clinically significant ARGs in the natural aquatic environment and highlights the potential ability of environmental isolates to disseminate these genes among different bacterial species. As such, the results presented demonstrate how anthropogenic point discharges may not act as the sole contributor to the development and spread of clinically significant antibiotic resistances. A number of critical knowledge gaps in current research were also identified. Key highlights include the limited number of studies focusing on antibiotic resistance in uncontaminated aquatic environments as well as the lack of standardisation among methodologies of reviewed investigations.

Mapping and Analysing Potential Sources and Transmission Routes of Antimicrobial Resistant Organisms in the Environment using Geographic Information Systems-An Exploratory Study.

Antimicrobial resistance (AMR) is one of the leading threats to human health worldwide. The identification of potential sources of antimicrobial resistant organisms (AROs) and their transmission routes in the environment is important for improving our understanding of AMR and to inform and improve policy and monitoring systems, as well as the identification of suitable sampling locations and potential intervention points. This exploratory study uses geographic information systems (GIS) to analyse the spatial distribution of likely ARO sources and transmission routes in four local authority areas (LAAs) in Ireland. A review of relevant spatial data in each LAA, grouped into themes, and categorised into sources and transmission routes, was undertaken. A range of GIS techniques was used to extract, organise, and collate the spatial data into final products in the form of thematic maps for visual and spatial analysis. The results highlight the location of 'clusters' at increased risk of harbouring AMR in each LAA. They also demonstrate the relevance of aquatic transmission routes for ARO mobility and risk of human exposure. The integration of a GIS approach with expert knowledge of AMR is shown to be a useful tool to gain insights into the spatial dimension of AMR and to guide sampling campaigns and intervention points.