Recreational bathing waters are complex systems with diverse inputs from multiple anthropogenic and zoogenic sources of faecal contamination. Faecal contamination is a substantial threat to water ...quality and public health. Here we present a comprehensive strategy to estimate the contribution of faecal indicator bacteria (FIB) from different biological sources on two at-risk beaches in Dublin, Ireland. The daily FIB loading rate was determined for three sources of contamination: a sewage-impacted urban stream, dog and wild bird fouling. This comparative analysis determined that the stream contributed the highest daily levels of FIB, followed by dog fouling. Dog fouling may be a significant source of FIB, contributing approximately 20 % of E. coli under certain conditions, whereas wild bird fouling contributed a negligible proportion of FIB (<3 %). This study demonstrates that source-specific quantitative microbial source apportionment (QMSA) strategies are vital to identify primary public health risks and target interventions to mitigate faecal contamination.
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•Source-specific QMSA is vital to quantify diverse biological sources of faeces.•Sewage-impacted stream contributed the highest daily FIB loading onto beaches.•Dogs were estimated to load 0.6 –h 2.2 kg of faeces and 20 % of E. coli daily.•Wild bird fouling contributed a negligible proportion of FIB.
Numerical health risk assessment models have been developed to describe faecal contamination of water using Escherichia coli as an indicator bacterium. Although many previously established numerical ...models for E. coli in aquatic environments have only considered the effects of one or two water quality parameters such as temperature and sunlight, it is difficult to simulate E. coli survival with only one or two parameters because the aquatic environment is a complex system. This study conducted a series of comparative experiments to select water quality parameters that should be preferentially considered in a numerical model for E. coli survival in lakes. The parameters considered were temperature, pH, dissolved oxygen (DO), total dissolved solids (TDS), suspended solids (SS), coexisting microbes, and light intensity. In the laboratory experiments, the survival of E. coli was observed by controlling two of these seven parameters, and the effects of these parameters on the rate of E. coli population change were statistically compared. Consequently, light intensity affected the survival of E. coli most significantly, followed by the presence of coexisting microbes, temperature, pH, and TDS. However, DO and SS had smaller effects on survival than other parameters. High-impact interactions on E. coli survival were observed between temperature and TDS and temperature and coexisting microbes. These results suggest that existing numerical models for simulating E. coli survival in lakes should be modified to consider the independent and interactive effects of multiple parameters such as sunlight, coexisting microbes, temperature, pH, and TDS.
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•To refine models, we determined parameters that highly impact E. coli survival.•Light affected E. coli survival most significantly followed by coexisting microbes.•Temperature, pH, and total dissolved solids also affected E. coli survival.•Dissolved oxygen and suspended solids affected E. coli survival less.•Temperature and total dissolved solids/coexisting microbes strongly interacted.
Faecal pollution in aquatic environments is a worldwide public health concern, yet the reliability and comprehensiveness of the methods used to assess faecal contamination are still debated. We ...compared three approaches, namely a culture-based method to enumerate Faecal Indicator Bacteria (FIB), a FIB-targeting qPCR assay, and High-Throughput Sequencing (HTS) to detect faeces- and sewage-associated taxa in water and sediment samples of an impacted model lagoon and its adjacent sea across one year. Despite at different levels, all approaches agreed in showing a higher contamination in the lagoon than in the sea, and higher in sediments than water. FIB significantly correlated when considering separately sediment and water, and when using both cultivation and qPCR. Similarly, FIB correlated between cultivation and qPCR, but qPCR provided consistently higher estimates of FIB. Faeces-associated bacteria positively correlated with cultivated FIB in both compartments, whereas sewage-associated bacteria did only in water. Considering their benefits and limitations, we conclude that, in our study site, improved quali-quantitative information on contamination is provided when at least two approaches are combined (e.g., cultivation and qPCR or HTS data). Our results provide insights to move beyond the use of FIB to improve faecal pollution management in aquatic environments and to incorporate HTS analysis into routine monitoring.
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•Three approaches to assess faecal pollution were compared.•Each approach exhibited benefits and limitations.•Combining at least two approaches provides exhaustive information on contamination.
Many wastewater treatment technologies have been shown to remove bacterial pathogens more effectively than viral pathogens and, in aquatic environments, levels of traditional faecal indicator ...bacteria (FIB) do not appear to correlate consistently with levels of human viral pathogens. There is, therefore, a need for novel viral indicators of faecal pollution and surrogates of viral pathogens, especially given the increasing importance of indirect and direct wastewater reuse. Potential candidates include bacteriophages (phages) and the study described here sought to elucidate the relationship between three groups of phages (somatic coliphages (SOMPH), F-RNA coliphages (F-RNAPH) and human-specific phages infecting B. fragilis (Bf124PH) – enumeration using double layer agar technique) and viral pathogens (human adenovirus (HuAdV) and norovirus (NoV) – enumeration using molecular methods) through full-scale municipal wastewater treatment processes. FIB (faecal coliforms (FC) and intestinal enterococci (ENT) – enumeration using membrane filtration) were also monitored. Samples were collected every fortnight, during a twelve-month period, at each stage of four full-scale wastewater treatment plants (WWTP) in southern England (two activated sludge (AS) and two trickling filter (TF) plants) (n = 360 samples). FIB and SOMPH were consistently found in all samples tested, whereas F-RNAPH, Bf124PH and HuAdV were less frequently detected, especially following AS treatment. The detection rate of NoV was low and consequently discussion of this group of viruses is limited. Concentrations of SOMPH and FIB were statistically higher (p value < 0.05) than concentrations of F-RNAPH, Bf124PH and HuAdV in raw wastewater. FIB were more effectively removed than phages in both systems. Removal rates of HuAdV were similar to those of phages at the secondary treatment stage of both systems. In TF systems, HuAdV were removed at the same rate as F-RNAPH, but at lower rates than SOMPH and Bf124PH. The findings suggest that phages (in particular SOMPH) are better indicators of the fate of viral pathogens in WWTP than existing FIB and that these organisms may have a useful role to play in future sanitation safety planning.
•FIB were more readily removed than phages and viral pathogens in all WWTP monitored.•Removal rates of phages were shown to be similar to those of human adenovirus.•Phages likely to better indicate the removal of viral pathogens in WWTP than FIB.•Phages as surrogates of viral pathogens in WWTP may support safe wastewater reuse.
Quantitative PCR (qPCR) and next generation sequencing (NGS) are nucleic acid based microbiology techniques that provide new insights into drinking water quality, but considerable uncertainty remains ...around their correct interpretation. We noticed the presence of bacterial DNA from various putative pathogens, including from faecal indicator bacteria (FIB), in disinfected water, when culturable FIB were absent. To understand these observations better we studied the effect of chlorination on conventional and DNA based microbial water quality assessments. Surface water chlorination reduced plate counts for various FIB by up to >6 log units, intact cell counts by flow cytometry by 3.3 log units, and 16S rRNA gene copies by qPCR by 1.5 and 1.6 log units for total bacteria and total coliforms, respectively. Nanopore sequencing of 16S rRNA amplicons with the portable MinION device revealed the DNA from several families containing putative pathogens appeared to be more resistant than that of other bacteria to degradation by chlorine disinfection. For instance, 16S rRNA genes assigned to the Enterobacteriaceae family, members of which are mostly the target of coliform tests, increased in relative abundance from 0.001 ± 0.0002% to 0.0036 ± 0.003% after chlorine treatment. Hence, metagenomic drinking water data needs to be interpreted with caution. Plate counts and flow cytometry in combination with DNA based analysis provide more robust insight than NGS or qPCR alone.
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•The effects of chlorination on water microbiomes were studied.•The DNA from the inactivated cells can persist in disinfected water.•16S rRNA amplicon sequencing outcomes were altered by chlorination.•Multiple lines of evidence are required for drinking water assessment.•Frequent water quality monitoring is key element of the water security plan.
•Faecal pollution data from the past 11 years in the Adriatic sea were analysed.•FIB contamination changed over space and time.•A general reduction of structural zeros over time was found.River ...discharge and rainfall did not explain faecal pollution•CSOs events were highly correlated with peaks in FIB.
Faecal contamination of the coastal sea poses widespread hazard to human and environmental health and is predicted to rise in response to global change and human pressure. For better management and risk reduction it is thus imperative to clarify and predict trends of faecal pollution over spatial and temporal scales, and to assess links with climate and other variables. Here, we investigated the spatio-temporal variation in the Faecal Indicator Bacteria (FIB) Escherichia coli and enterococci, over a time frame spanning 11 years (2011–2021) along a coastal area covering approximately 40 km and 59 bathing sites in the Marche region (Adriatic Sea, Italy), characterized by intense beach tourism, high riverine inputs, resident population, maritime traffic and industrial activities. Our analysis, that considers 5,183 measurements during the bathing season (April to October), shows that FIB abundance varied significantly among years. A general, although not significant, increase over time of both FIB was observed, mainly due to a general reduction of structural zeros (i.e., zeros originated from the actual absence of the response variable) over the examined time period. FIB abundances displayed their maxima and minima in different years according to the municipality, with overall peaks recorded in different months (May-June or September), whereas the lowest values were always observed in October. FIB levels were not significantly related neither to rainfalls nor to river discharge, but the activation of combined sewer overflows (CSOs), typically occurring after intense rainfall events, appeared as a necessary condition for the high faecal contamination levels. Considering climate change scenarios predicting significant increases in extreme weather events, our findings support the usefulness of analysing long-term trends to identify pollution sources, and the prioritization of control strategies to better manage the release of microbial pollutants from combined sewer overflows in coastal waters to reduce human risks.
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•Faecal indicators should be based on the hydrodynamic properties of water systems.•FIB are appropriate in advection dominant waterbodies.•crAssphage and PMMoV are appropriate in dispersion/advection ...dominant waterbodies.
The EU Bathing Water Directive (BWD) requires member states to assess bathing water quality according to the levels of faecal indicator bacteria (FIB) in designated bathing areas. However, this criterion has two significant limitations given that the BWD does not; (i) account for differences in hydrodynamic properties of bathing waters and, (ii) assumes that all faecal pathogens decay equally in aquatic environments. This study simulated sewage discharge events in three hypothetical aquatic environments characterised by different advection and dispersion parameters in the solute transport equation. Temporal changes in the downstream concentration of six faecal indicators were determined in simulations that utilised measured decay rates of each faecal indicator from a programme of controlled microcosm experiments in fresh and seawater environments. The results showed that the decay rates of faecal indicators are not a critical parameter in advection dominant water bodies, such as in fast-flowing rivers. Therefore, faecal indicator selection is less important in such systems and for these, FIB remains the most cost-effective faecal indicator to monitor the public health impacts of faecal contamination. In contrast, consideration of faecal indicator decay is important when assessing dispersion and advection/dispersion dominant systems, which would pertain to transitional (estuarine) and coastal waterbodies. Results suggest that the inclusion of viral indicators, such as crAssphage and PMMoV, could improve the reliability of water quality modelling and minimise the risk of waterborne illnesses from faecal contamination.
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Faecal pollution of water and the resulting potential presence of human enteric pathogens is a predominant threat to public health. Microbiological water quality can be assessed by the detection of ...standard faecal indicator bacteria (SFIB) such as E. coli or certain Enterococcus species. In recent years, isothermal amplification methods have become a useful alternative to polymerase chain reaction (PCR), allowing molecular diagnostics with simple or no instrumentation. In this study, a novel screening method for the molecular detection of Enterococcus spp. by loop-mediated isothermal amplification (LAMP) is described. A set of six specific LAMP primers was designed to amplify a diagnostic fragment of the Enterococcus 23S rRNA gene, which is present in several enterococcal species targeted by quantitative PCR (qPCR), which is the standard technique recommended by the US Environmental Protection Agency. Sensitivity and specificity tests were performed using a set of 30 Enterococcus and non-target bacterial reference strains. It is shown that LAMP is equally sensitive and even more specific than the qPCR assay. A dilution series of Enterococcus faecalis DNA revealed that the LAMP method can reliably detect 130 DNA target copies per reaction within 45 min. Additionally, enterococci isolated from Austrian surface waterbodies, as well as a set of DNA extracts from environmental waters, were tested. Contingency analysis demonstrated a highly significant correlation between the results of the developed LAMP assay and the reference qPCR method. Furthermore, a simple staining procedure with a fluorescence dye demonstrated the identification of amplified products by eye. In conclusion, this method is an important component for the efficient screening and testing of water samples in low-resource settings lacking sophisticated laboratory equipment and highly trained personnel, requiring only a simple heating block.
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•A rapid method for the molecular detection of enterococci in water is presented.•The developed LAMP assay is equally sensitive and specific as the reference qPCR.•The reaction takes place at a constant temperature provided by a heating block.•Results can be visualised within 1 min.
Monitoring of a constructed wetland receiving episodic pulses of subsurface drainage waters from grazed dairy pastures showed good nitrogen reductions, but moderate increases in median Escherichia ...coli concentrations. This unexpected increase of E. coli was initially postulated to be an artefact of the routine sampling approach, which was biased towards receding flows and likely missed high inflows of contaminated drainage. Intensive monitoring during storm events was therefore instigated using auto-samplers to quantify responses during rainfall-generated flow events. Determination of inflow and outflow E. coli fluxes and loads for ten storm events concluded that the wetland was consistently a net exporter of E. coli with increases ranging from 1.5 to 26-fold (median 10.2-fold). Whilst wildlife defecation in the wetland may account for a proportion of the rise in E. coli, it is likely that (consistent with associated genetic profiling studies) much of the increase is due to mobilisation of environmentally adapted strains of E. coli persisting and growing within the sediment, decomposing plant litter and organic detritus of the wetland. This raises interesting questions regarding the microbial ecology of E. coli and its utility as an indicator of faecal pathogens, and potential health risks from diffuse agricultural drainage after passage through wetlands.
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•Unexpected E. coli increase in a constructed wetland treating agricultural drainage.•In and out-flows, concentrations and loads intensively monitored over 10 stormflows.•Median 10-fold increase in exported E. coli load after passage through wetland.•Credible evidence for persistence and generation of E. coli within the wetland.•Implications for the utility of E. coli as a faecal indicator for human health risk.
•Machine-learning is applied to mimic a process-based hydrodynamic and water quality model.•Process-based models forced by highly variable inputs are well emulated by the LSTM.•Spatio-temporal ...variations of water quality in reservoirs can be efficiently predicted by the LSTM.•Compared to process-based models, shorter simulation time is required with the LSTM.•Real-time hydrodynamic and water quality simulations can be made easier with machine-learning.
Process-based models are very efficient in simulating hydrodynamics and water quality in surface water bodies. However, their complex characteristics in terms of implementation, data requirements, and simulation time limit their application in regular drinking water source management. This study demonstrates the potential of a ML model (Long Short-Term Memory (LSTM)) as viable alternatives to process-based hydrodynamic and water quality models in water source management. Using meteorological and hydrological measurements, a hydrodynamic and water quality model was first calibrated to predict time series, profiles, and contours of water variables namely Escherichia coli (E. coli), faecal coliforms, zinc, and lead concentrations in the Brusdalsvatnet lake, which is the drinking water source for the city of Ålesund in Norway. The results obtained were combined with the input data to train a suite of LSTM models to emulate the results achieved with the process-based modelling. The results indicate that ML models can conveniently reproduce the spatio-temporal evolution of water quality in the lake that is achievable with the process-based model, particularly when specific locations within the lake are of interest. Compared to R2, NS and MSE ranges of 0.72–0.87, 0.68–0.85, and 0.21–0.44 achieved with the process-based model in the prediction of temperature in the lake, 0.78–0.95, 0.75–0.89, and 0.011–0.028 were respectively achieved in testing the LSTM models. Similar performance levels were achieved with the LSTM model in the prediction of Escherichia coli (E. coli), faecal coliforms, Zinc, and Lead concentrations at different depths in the lake. While setting up and training the LSTM model to emulate the process-based model simulations was very time-consuming, a validated model as developed in this study can offer an opportunity for real-time simulation of water quality in drinking water sources when integrated with cloud data transmission from field sensors.
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