On-line detection of Escherichia coli intrusion in a pilot-scale drinking water distribution system.

Improvements in microbial drinking water quality monitoring are needed for the better control of drinking water distribution systems and for public health protection. Conventional water quality monitoring programmes are not always able to detect a microbial contamination of drinking water. In the drinking water production chain, in addition to the vulnerability of source waters, the distribution networks are prone to contamination. In this study, a pilot-scale drinking-water distribution network with an on-line monitoring system was utilized for detecting bacterial intrusion. During the experimental Escherichia coli intrusions, the contaminant was measured by applying a set of on-line sensors for electric conductivity (EC), pH, temperature (T), turbidity, UV-absorbance at 254 nm (UVAS SC) and with a device for particle counting. Monitored parameters were compared with the measured E. coli counts using the integral calculations of the detected peaks. EC measurement gave the strongest signal compared with the measured baseline during the E. coli intrusion. Integral calculations showed that the peaks in the EC, pH, T, turbidity and UVAS SC data were detected corresponding to the time predicted. However, the pH and temperature peaks detected were barely above the measured baseline and could easily be mixed with the background noise. The results indicate that on-line monitoring can be utilized for the rapid detection of microbial contaminants in the drinking water distribution system although the peak interpretation has to be performed carefully to avoid being mixed up with normal variations in the measurement data.

Sand filters for removal of microbes and nutrients from wastewater during a one-year pilot study in a cold temperate climate.

Onsite wastewater treatment systems (OWTS) are recognised as potential threats to groundwater or other water environments subject to discharged effluents. In this study, the microbiological and nutrient removal properties of three different pilot-scale sand filters (SFs) were followed over a one-year period. Moreover, a separate phosphorus removal unit was tested for six months. For the best treatment system, the average log removals were 2.2-3.5 for pathogenic human noro- and adenoviruses and 4.3-5.2 and 4.6-5.4 for indicator viruses and bacteria, respectively. The system that effectively removed microbes was also efficient at removing nutrients. However, the poorest treatment system yielded substantially lower removals. The remarkable differences noted between the studied SFs highlights the importance of construction materials and the careful planning of the filters. Moreover, seasonal conditions appear to have a clear effect on purification efficiencies, emphasising the vulnerability of these systems especially in cold climates.

Composition of active bacterial communities and presence of opportunistic pathogens in disinfected and non-disinfected drinking water distribution systems in Finland.

Many factors, including microbiome structure and activity in the drinking water distribution system (DWDS), affect the colonization potential of opportunistic pathogens. The present study aims to describe the dynamics of active bacterial communities in DWDS and identify the factors that shape the community structures and activity in the selected DWDSs. Large-volume drinking water and hot water, biofilm, and water meter deposit samples were collected from five DWDSs. Total nucleic acids were extracted, and RNA was further purified and transcribed into its cDNA from a total of 181 water and biofilm samples originating from the DWDS of two surface water supplies (disinfected with UV and chlorine), two artificially recharged groundwater supplies (non-disinfected), and a groundwater supply (disinfected with UV and chlorine). In chlorinated DWDSs, concentrations of <0.02-0.97 mg/l free chlorine were measured. Bacterial communities in the RNA and DNA fractions were analysed using Illumina MiSeq sequencing with primer pair 341F-785R targeted to the 16S rRNA gene. The sequence libraries were analysed using QIIME pipeline, Program R, and MicrobiomeAnalyst. Not all bacterial cells were active based on their 16S rRNA content, and species richness was lower in the RNA fraction (Chao1 mean value 490) than in the DNA fraction (710). Species richness was higher in the two DWDSs distributing non-disinfected artificial groundwater (Chao1 mean values of 990 and 1 000) as compared to the two disinfected DWDSs using surface water (Chao1 mean values 190 and 460) and disinfected DWDS using ground water as source water (170). The difference in community structures between non-disinfected and disinfected water was clear in the beta-diversity analysis. Distance from the waterworks also affected the beta diversity of community structures, especially in disinfected distribution systems. The two most abundant bacteria in the active part of the community (RNA) and total bacterial community (DNA) belonged to the classes Alphaproteobacteria (RNA 28 %, DNA 44 %) and Gammaproteobacteria (RNA 32 %, DNA 30 %). The third most abundant and active bacteria class was Vampirovibrionia (RNA 15 %), whereas in the total community it was Paceibacteria (DNA 11 %). Class Nitrospiria was more abundant and active in both cold and hot water in DWDS that used chloramine disinfection compared to non-chlorinated or chlorine-using DWDSs. Thirty-eight operational taxonomic units (OTU) of Legionella, 30 of Mycobacterium, and 10 of Pseudomonas were detected among the sequences. The (RT)-qPCR confirmed the presence of opportunistic pathogens in the DWDSs studied as Legionella spp. was detected in 85 % (mean value 4.5 × 104 gene copies/100 ml), Mycobacterium spp. in 95 % (mean value 8.3 × 106 gene copies/100 ml), and Pseudomonas spp. in 78 % (mean value 1.6 × 105 gene copies/100 ml) of the water and biofilm samples. Sampling point inside the system (distance from the waterworks and cold/hot system) affected the active bacterial community composition. Chloramine as a chlorination method resulted in a recognizable community composition, with high abundance of bacteria that benefit from the excess presence of nitrogen. The results presented here confirm that each DWDS is unique and that opportunistic pathogens are present even in conditions when water quality is considered excellent.

Campylobacter species, Salmonella serotypes and ribosomal RNA-based fecal source tracking in the Kokemäki River watershed.

Fecal contamination of surface water compromises the usability of surface water for drinking water production due to an increase in human health risks. In this study, we collected surface water samples for two years from the Kokemäki River (Finland). The downstream river stretch is used for feeding production of artificial ground water for a major drinking water treatment plant. The prevalence of Campylobacter species and Salmonella serotypes together with fecal source identifiers targeting general, human, gull, swine, and ruminant were evaluated at 16 sampling sites throughout the studied watershed. We detected Campylobacter spp. from all 16 sampling sites with Campylobacter jejuni and Campylobacter lari as the most detected species. Salmonella spp. was detected in 10 out of 16 sampling sites, with Salmonella Typhimurium being the most common serovar. Regarding spatial variation in the hygienic quality of surface water, the upstream area (urban proximity) and downstream area (agricultural proximity) had higher microbial loads than the middle section of the study area. Samples taken in fall and spring had higher microbial loads than summer and winter samples. The lower ratio of rRNA to rRNA-gene (rDNA) of studied microbes in the winter than in other seasons may indicate low metabolic activity of bacterial targets during winter. The number of gulls, swine, and cattle in the catchment area concorded with the number of fecal source identifiers in the surface water. Further, the prevalence of gull-specific source identifier agreed with the detection of C. coli, C. lari, and S. Typhimurim, whereas the prevalence of swine- and ruminant-specific source identifiers agreed with the detection of C. jejuni and C. coli. Thus, fecal source identifiers are shown to be important tools for monitoring zoonotic pathogens affecting microbial quality of surface water. Further, variation in fecal loads indicates such variation in health risks related to surface water use.

Comparison of culture and qPCR methods in detection of mycobacteria from drinking waters.

Environmental mycobacteria are common bacteria in man-made water systems and may cause infections and hypersensitivity pneumonitis via exposure to water. We compared a generally used cultivation method and a quantitative polymerase chain reaction (qPCR) method to detect mycobacteria in 3 types of drinking waters: surface water, ozone-treated surface water, and groundwater. There was a correlation between the numbers of mycobacteria obtained by cultivation and qPCR methods, but the ratio of the counts obtained by the 2 methods varied among the types of water. The qPCR counts in the drinking waters produced from surface or groundwater were 5 to 34 times higher than culturable counts. In ozone-treated surface waters, both methods gave similar counts. The ozone-treated drinking waters had the highest concentration of assimilable organic carbon, which may explain the good culturability. In warm tap waters, qPCR gave 43 times higher counts than cultivation, but both qPCR counts and culturable counts were lower than those in the drinking waters collected from the same sites. The TaqMan qPCR method is a rapid and sensitive tool for total quantitation of mycobacteria in different types of clean waters. The raw water source and treatments affect both culturability and total numbers of mycobacteria in drinking waters.

Manual faucets induce more biofilms than electronic faucets.

Electronic faucets (types E1 and E2) and manual (M) faucets were studied for microbial quality, i.e., biomass and pathogenic microbes of biofilms in the faucet aerator, the water, and the outer surface of faucet in a hospital in Finland. Heterotrophic plate count content reflecting culturable microbial biomass and adenosine triphosphate content representing viable microbial biomass were smaller in the biofilms of E1-type electronic faucets than E2-type electronic faucets or M faucets. The likely explanation is the mixing point of cold and hot water (E1 and M: in the faucet; E2: in a separate box 50 cm before the actual faucet part). The highest amounts of Legionella (serogroups 2-15 of Legionella pneumophila) in a water sample (5000 cfu/L) and in biofilm samples (May-June 2008 sampling: 240 cfu/mL; November 2008: 1100 cfu/mL) were found in one E1-type faucet, which was lacking a back pressure valve due to faulty installation. This study reveals that certain types of electronic faucets seem to promote hospital hygiene, as they were associated with less microbial growth in biofilms in the faucet aerator, than some other types of electronic faucets or manual faucets, likely owing to the mixing point of cold and hot water. However, the faucet type had no direct effect on the presence of Legionella spp. Also correct installation is crucial.

Occurrence of thermotolerant Campylobacter spp. and adenoviruses in Finnish bathing waters and purified sewage effluents.

A total of 50 Finnish bathing water samples and 34 sewage effluent samples originating from 17 locations were studied in the summers of 2006 and 2007. Campylobacter were present in 58% and adenoviruses in 12% of all bathing water samples; 53% of all sewage effluent samples were positive for Campylobacter spp. and 59% for adenoviruses. C. jejuni was the most common Campylobacter species found and human adenovirus serotype 41 was the most common identified adenovirus type. Bathing water temperature displayed a significant negative relationship with the occurrence of Campylobacter. One location had identical pulsed-field gel electrophoresis patterns of C. coli isolates in the bathing water and in sewage effluent, suggesting that sewage effluent was the source of C. coli at this bathing site. The counts of faecal indicator bacteria were not able to predict the presence of Campylobacter spp. or adenoviruses in the bathing waters. Thus the observed common presence of these pathogens in Finnish sewage effluents and bathing waters may represent a public health risk. The low water temperature in Finland may enhance the prevalence of Campylobacter in bathing waters. More attention needs to be paid to minimizing the concentrations of intestinal pathogens in bathing waters.

Microbial reduction in wastewater treatment using Fe(3+) and Al(3+) coagulants and PAA disinfectant.

Wastewater is an important source of pathogenic enteric microorganisms in surface water and a major contaminating agent of drinking water. Although primary and secondary wastewater treatments reduce the numbers of microorganisms in wastewater, significant numbers of microbes can still be present in the effluent. The aim of this study was to test the feasibility of tertiary treatment for municipal wastewater treatment plants (WWTPs) using PIX (FeCl3) or PAX (AlCl3) coagulants and peracetic acid (PAA) the disinfectant to reduce microbial load in effluent. Our study showed that both PIX and PAX efficiently reduced microbial numbers. PAA disinfection greatly reduced the numbers of culturable indicator microorganisms (Escherichia coli, intestinal enterococci, F-specific RNA coliphages and somatic DNA coliphages). In addition, pathogenic microorganisms, thermotolerant Campylobacter, Salmonella and norovirus GI, were successfully reduced using the tertiary treatments. In contrast, clostridia, Legionella, rotavirus, norovirus GII and adenovirus showed better resistance against PAA compared to the other microorganisms. However, interpretation of polymerase chain reaction (PCR) analysis results will need further studies to clarify the infectivity of the pathogenic microbes. In conclusion, PIX and PAX flocculants followed by PAA disinfectant can be used as a tertiary treatment for municipal WWTP effluents to reduce the numbers of indicator and pathogenic microorganisms.

Clostridium difficile contamination of public tap water distribution system during a waterborne outbreak in Finland.

AIMS: In November through December 2007, the drinking water distribution system in the town of Nokia, Finland, was contaminated with treated sewage effluent that resulted in a large gastroenteritis outbreak in the community. The aim of the present study was to investigate if the contaminated water in this outbreak was also a potential source of Clostridium difficile infections. METHODS: Samples from the contaminated tap water and treated sewage effluent were collected. Stool samples from a portion of patients that fell ill during the outbreak were examined for C. difficile. PCR ribotyping was performed on toxin positive C. difficile isolates and the genetic profiles of the water and patient isolates were compared. RESULTS: Twelve toxin-positive C. difficile isolates were found in water samples: five from contaminated tap water and seven from treated sewage effluent. Among these, four and five distinct PCR ribotype profiles were identified, respectively. Four PCR ribotype profiles were found among nine human faecal C. difficile isolates. Two isolates, one from tap water and one from a patient, had an indistinguishable PCR ribotype profile. CONCLUSIONS: Our findings demonstrate for the first time C. difficile contamination of a tap water distribution system and waterborne transmission of toxigenic C. difficile seems possible.

Effects of temperature and light exposure on the decay characteristics of fecal indicators, norovirus, and Legionella in mesocosms simulating subarctic river water.

Knowledge of the decay characteristics of health-related microbes in surface waters is important for modeling the transportation of waterborne pathogens and for assessing their public health risks. Although water temperature and light exposure are major factors determining the decay characteristics of enteric microbes in surface waters, such effects have not been well studied in subarctic surface waters. This study comprehensively evaluated the effect of temperature and light on the decay characteristics of health-related microbes [Escherichia coli, enterococci, microbial source tracking markers (GenBac3 & HF183 assays), coliphages (F-specific and somatic), noroviruses GII and Legionella spp.] under simulated subarctic river water conditions. The experiments were conducted in four different laboratory settings (4 °C/dark, 15 °C/dark, 15 °C/light, and 22 °C/light). The T90 values (time required for a 90 % reduction in the population of a target) of all targets were higher under cold and dark (2.6-51.3 days depending upon targets) than under warm and light conditions (0.6-3.5 days). Under 4 °C/dark (simulated winter) water conditions, F-specific coliphages had 27.2 times higher, and coliform bacteria had 3.3 times higher T90 value than under 22 °C/light (simulated summer) water conditions. Bacterial molecular markers also displayed high variation in T90 values, with the greatest difference between 4 °C/dark and 22 °C/light recorded for HF183 DNA (20.6 times) and the lowest difference for EC23S857 RNA (6.6 times). E. coli, intestinal enterococci, and somatic coliphages were relatively more sensitive to light than water temperature, but F-specific coliphages, norovirus, and all bacterial rDNA and rRNA markers were relatively more sensitive to temperature than light exposure. Due to the slow microbial decay in winter under subarctic conditions, the microbial quality of river water might remain low for a long time after a sewage spill. This increased risk associated with fecal pollution during winter may deserve more attention, especially when river waters are used for drinking water production.

Monitoring groundwater quality with real-time data, stable water isotopes, and microbial community analysis: A comparison with conventional methods.

Groundwater provides much of the world's potable water. Nevertheless, groundwater quality monitoring programmes often rely on a sporadic, slow, and narrowly focused combination of periodic manual sampling and laboratory analyses, such that some water quality deficiencies go undetected, or are detected too late to prevent adverse consequences. In an effort to address this shortcoming, we conducted enhanced monitoring of untreated groundwater quality over 12 months (February 2019-February 2020) in four shallow wells supplying potable water in Finland. We supplemented periodic manual sampling and laboratory analyses with (i) real-time online monitoring of physicochemical and hydrological parameters, (ii) analysis of stable water isotopes from groundwater and nearby surface waters, and (iii) microbial community analysis of groundwater via amplicon sequencing of the 16S rRNA gene and 16S rRNA. We also developed an early warning system (EWS) for detecting water quality anomalies by automating real-time online monitoring data collection, transfer, and analysis - using electrical conductivity (EC) and turbidity as indirect water quality indicators. Real-time online monitoring measurements were largely in fair agreement with periodic manual measurements, demonstrating their usefulness for monitoring water quality; and the findings of conventional monitoring, stable water isotopes, and microbial community analysis revealed indications of surface water intrusion and faecal contamination at some of the studied sites. With further advances in technology and affordability expected into the future, the supplementary methods used here could be more widely implemented to enhance groundwater quality monitoring - by contributing new insights and/or corroborating the findings of conventional analyses.

A comparative analysis employing a gene- and genome-centric metagenomic approach reveals changes in composition, function, and activity in waterworks with different treatment processes and source water in Finland.

The emergence and development of next-generation sequencing technologies (NGS) has made the analysis of the water microbiome in drinking water distribution systems (DWDSs) more accessible and opened new perspectives in microbial ecology studies. The current study focused on the characterization of the water microbiome employing a gene- and genome-centric metagenomic approach to five waterworks in Finland with different raw water sources, treatment methods, and disinfectant. The microbial communities exhibit a distribution pattern of a few dominant taxa and a large representation of low-abundance bacterial species. Changes in the community structure may correspond to the presence or absence and type of disinfectant residual which indicates that these conditions exert selective pressure on the microbial community. The Archaea domain represented a small fraction (up to 2.5%) and seemed to be effectively controlled by the disinfection of water. Their role particularly in non-disinfected DWDS may be more important than previously considered. In general, non-disinfected DWDSs harbor higher microbial richness and maintaining disinfectant residual is significantly important for ensuring low microbial numbers and diversity. Metagenomic binning recovered 139 (138 bacterial and 1 archaeal) metagenome-assembled genomes (MAGs) that had a >50% completeness and <10% contamination consisting of 20 class representatives in 12 phyla. The presence and occurrence of nitrite-oxidizing bacteria (NOB)-like microorganisms have significant implications for nitrogen biotransformation in drinking water systems. The metabolic and functional complexity of the microbiome is evident in DWDSs ecosystems. A comparative analysis found a set of differentially abundant taxonomic groups and functional traits in the active community. The broader set of transcribed genes may indicate an active and diverse community regardless of the treatment methods applied to water. The results indicate a highly dynamic and diverse microbial community and confirm that every DWDS is unique, and the community reflects the selection pressures exerted at the community structure, but also at the levels of functional properties and metabolic potential.

Decontamination of a drinking water pipeline system contaminated with adenovirus and Escherichia coli utilizing peracetic acid and chlorine.

A contaminated drinking water distribution network can be responsible for major outbreaks of infections. In this study, two chemical decontaminants, peracetic acid (PAA) and chlorine, were used to test how a laboratory-scale pipeline system can be cleaned after simultaneous contamination with human adenovirus 40 (AdV40) and Escherichia coli. In addition, the effect of the decontaminants on biofilms was followed as heterotrophic plate counts (HPC) and total cell counts (TCC). Real-time quantitative polymerase chain reaction (qPCR) was used to determine AdV40 and plate counting was used to enumerate E. coli. PAA and chlorine proved to be effective decontaminants since they decreased the levels of AdV40 and E. coli to below method detection limits in both water and biofilms. However, without decontamination, AdV40 remained present in the pipelines for up to 4 days. In contrast, the concentration of cultivable E. coli decreased rapidly in the control pipelines, implying that E. coli may be an inadequate indicator for the presence of viral pathogens. Biofilms responded to the decontaminants by decreased HPCs while TCC remained stable. This indicates that the mechanism of pipeline decontamination by chlorine and PAA is inactivation rather than physical removal of microbes.

Detection and quantification of SARS-CoV-2 RNA in wastewater influent in relation to reported COVID-19 incidence in Finland.

Wastewater-based surveillance is a cost-effective concept for monitoring COVID-19 pandemics at a population level. Here, SARS-CoV-2 RNA was monitored from a total of 693 wastewater (WW) influent samples from 28 wastewater treatment plants (WWTP, N = 21-42 samples per WWTP) in Finland from August 2020 to May 2021, covering WW of ca. 3.3 million inhabitants (∼ 60% of the Finnish population). Quantity of SARS-CoV-2 RNA fragments in 24 h-composite samples was determined by using the ultrafiltration method followed by nucleic acid extraction and CDC N2 RT-qPCR assay. SARS-CoV-2 RNA signals at each WWTP were compared over time to the numbers of confirmed COVID-19 cases (14-day case incidence rate) in the sewer network area. Over the 10-month surveillance period with an extensive total number of samples, the detection rate of SARS-CoV-2 RNA in WW was 79% (including 6% uncertain results, i.e., amplified only in one out of four, two original and two ten-fold diluted replicates), while only 24% of all samples exhibited gene copy numbers above the quantification limit. The range of the SARS-CoV-2 detection rate in WW varied from 33% (including 10% uncertain results) in Pietarsaari to 100% in Espoo. Only six out of 693 WW samples were positive with SARS-COV-2 RNA when the reported COVID-19 case number from the preceding 14 days was zero. Overall, the 14-day COVID-19 incidence was 7.0, 18, and 36 cases per 100 000 persons within the sewer network area when the probability to detect SARS-CoV-2 RNA in wastewater samples was 50%, 75% and 95%, respectively. The quantification of SARS-CoV-2 RNA required significantly more COVID-19 cases: the quantification rate was 50%, 75%, and 95% when the 14-day incidence was 110, 152, and 223 COVID-19 cases, respectively, per 100 000 persons. Multiple linear regression confirmed the relationship between the COVID-19 incidence and the SARS-CoV-2 RNA quantified in WW at 15 out of 28 WWTPs (overall R2 = 0.36, p < 0.001). At four of the 13 WWTPs where a significant relationship was not found, the SARS-CoV-2 RNA remained below the quantification limit during the whole study period. In the five other WWTPs, the sewer coverage was less than 80% of the total population in the area and thus the COVID-19 cases may have been inhabitants from the areas not covered. Based on the results obtained, WW-based surveillance of SARS-CoV-2 could be used as an indicator for local and national COVID-19 incidence trends. Importantly, the determination of SARS-CoV-2 RNA fragments from WW is a powerful and non-invasive public health surveillance measure, independent of possible changes in the clinical testing strategies or in the willingness of individuals to be tested for COVID-19.

Increased information on waterborne outbreaks through efficient notification system enforces actions towards safe drinking water.

In 1997, a compulsory notification system for waterborne outbreaks was introduced in Finland. The main aim of this notification is to obtain immediate information on suspected waterborne outbreaks in order to restrict and manage the outbreak promptly. During the past ten years, there have been 67 waterborne outbreaks in Finland, mainly associated with small groundwater supplies or private wells. The number of reported waterborne outbreaks has increased since the launch of the notification system indicating that the threshold limit of outbreak detection has most probably decreased. The number of cases of illness has fulfilled the national health target, which is below 0.01% of the population, but more action is still needed to ensure the production of safe drinking water under all circumstances. Ten years accumulation of knowledge on outbreaks has revealed that a compulsory notification system is an effective tool to gather information on waterborne outbreaks. The system has also increased awareness of possible problems related to the quality of drinking water. This article summarises management and legislative actions and policy measures taken so far in Finland to reduce the number of outbreaks and cases of illness related to them.

Microbial contamination of groundwater at small community water supplies in Finland.

The raw water quality and associations between the factors considered as threats to water safety were studied in 20 groundwater supplies in central Finland in 2002-2004. Faecal contaminations indicated by the appearance of Escherichia coli or intestinal enterococci were present in five small community water supplies, all these managed by local water cooperatives. Elevated concentrations of nutrients in raw water were linked with the presence of faecal bacteria. The presence of on-site technical hazards to water safety, such as inadequate well construction and maintenance enabling surface water to enter into the well and the insufficient depth of protective soil layers above the groundwater table, showed the vulnerability of the quality of groundwater used for drinking purposes. To minimize the risk of waterborne illnesses, the vulnerable water supplies need to be identified and appropriate prevention measures such as disinfection should be applied.

Phage Biocontrol of Pseudomonas aeruginosa in Water.

Bacteriophage control of harmful or pathogenic bacteria has aroused growing interest, largely due to the rise of antibiotic resistance. The objective of this study was to test phages as potential agents for the biocontrol of an opportunistic pathogen Pseudomonas aeruginosa in water. Two P. aeruginosa bacteriophages (vB_PaeM_V523 and vB_PaeM_V524) were isolated from wastewater and characterized physically and functionally. Genomic and morphological characterization showed that both were myoviruses within the Pbunavirus genus. Both had a similar latent period (50-55 min) and burst size (124-134 PFU/infected cell), whereas there was variation in the host range. In addition to these environmental phages, a commercial Pseudomonas phage, JG003 (DSM 19870), was also used in the biocontrol experiments. The biocontrol potential of the three phages in water was tested separately and together as a cocktail against two P. aeruginosa strains; PAO1 and the environmental strain 17V1507. With PAO1, all phages initially reduced the numbers of the bacterial host, with phage V523 being the most efficient (>2.4 log10 reduction). For the environmental P. aeruginosa strain (17V1507), only the phage JG003 caused a reduction (1.2 log10) compared to the control. The cocktail of three phages showed a slightly higher decrease in the level of the hosts compared to the use of individual phages. Although no synergistic effect was observed in the host reduction with the use of the phage cocktail, the cocktail-treated hosts did not appear to acquire resistance as rapidly as hosts treated with a single phage. The results of this study provide a significant step in the development of bacteriophage preparations for the control of pathogens and harmful microbes in water environments.

Bacterial Genes Encoding Resistance Against Antibiotics and Metals in Well-Maintained Drinking Water Distribution Systems in Finland.

Information on the co-occurrence of antibiotic resistance genes (ARGs) and metal resistance genes (MRGs) among bacterial communities in drinking water distribution systems (DWDSs) is scarce. This study characterized ARGs and MRGs in five well-maintained DWDSs in Finland. The studied DWDSs had different raw water sources and treatment methods. Two of the waterworks employed artificially recharged groundwater (ARGW) and used no disinfection in the treatment process. The other three waterworks (two surface and one groundwater source) used UV light and chlorine during the treatment process. Ten bulk water samples (two from each DWDS) were collected, and environmental DNA was extracted and then sequenced using the Illumina HiSeq platform for high-throughput shotgun metagenome sequencing. A total of 430 ARGs were characterized among all samples with the highest diversity of ARGs identified from samples collected from non-disinfected DWDSs. Furthermore, non-disinfected DWDSs contained the highest diversity of bacterial communities. However, samples from DWDSs using disinfectants contained over double the ratio of ARG reads to 16S rRNA gene reads and most of the MRG (namely mercury and arsenic resistance genes). The total reads and types of ARGs conferring genes associated with antibiotic groups namely multidrug resistance, and bacitracin, beta-lactam, and aminoglycoside and mercury resistance genes increased in waterworks treating surface water with disinfection. The findings of this study contribute toward a comprehensive understanding of ARGs and MRGs in DWDSs. The occurrence of bacteria carrying antibiotic or metal resistance genes in drinking water causes direct exposure to people, and thus, more systematic investigation is needed to decipher the potential effect of these resistomes on human health.

Surface Water Intrusion, Land Use Impacts, and Bacterial Community Composition in Shallow Groundwater Wells Supplying Potable Water in Sparsely Populated Areas of a Boreal Region.

Rural communities often rely on groundwater for potable water supply. In this study, untreated groundwater samples from 28 shallow groundwater wells in Finland (<10 m deep and mostly supplying untreated groundwater to <200 users in rural areas) were assessed for physicochemical water quality, stable water isotopes, microbial water quality indicators, host-specific microbial source tracking (MST) markers, and bacterial community composition, activity, and diversity (using amplicon sequencing of the 16S rRNA gene and 16S rRNA). Indications of surface water intrusion were identified in five wells, and these indications were found to be negatively correlated, overall, with bacterial alpha diversity (based on amplicon sequencing of the 16S rRNA gene). High levels of turbidity, heterotrophs, and iron compromised water quality in two wells, with values up to 2.98 nephelometric turbidity units (NTU), 16,000 CFU/ml, and 2,300 µg/liter, respectively. Coliform bacteria and general fecal indicator Bacteroidales bacteria (GenBac3) were detected in 14 and 10 wells, respectively (albeit mostly at low levels), and correlations were identified between microbial, physicochemical, and environmental parameters, which may indicate impacts from nearby land use (e.g., agriculture, surface water, road salt used for deicing). Our results show that although water quality was generally adequate in most of the studied wells, the continued safe use of these wells should not be taken for granted. IMPORTANCE Standard physicochemical water quality analyses and microbial indicator analyses leave much of the (largely uncultured) complexity of groundwater microbial communities unexplored. This study combined these standard methods with additional analyses of stable water isotopes, bacterial community data, and environmental data about the surrounding areas to investigate the associations between physicochemical and microbial properties of 28 shallow groundwater wells in Finland. We detected impaired groundwater quality in some wells, identified potential land use impacts, and revealed indications of surface water intrusion which were negatively correlated with bacterial alpha diversity. The potential influence of surface water intrusion on groundwater wells and their bacterial communities is of particular interest and warrants further investigation because surface water intrusion has previously been linked to groundwater contamination, which is the primary cause of waterborne outbreaks in the Nordic region and one of the major causes in the United States and Canada.

Diverse and active archaea communities occur in non-disinfected drinking water systems-Less activity revealed in disinfected and hot water systems.

The knowledge about the members of active archaea communities in DWDS is limited. The current understanding is based on high-throughput 16S ribosomal RNA gene (DNA-based) amplicon sequencing that reveals the diversity of active, dormant, and dead members of the prokaryote (bacteria, archaea) communities. The sequencing primers optimized for bacteria community analysis may underestimate the share of the archaea community. This study characterized archaea communities at five full-scale drinking water distribution systems (DWDS), representing a variety of drinking water production units (A-E); A&B use artificially recharged non-disinfected groundwater (ARG), the other DWDS's supplied water disinfected by using ultraviolet (UV) light and chlorine compounds, C&D were surface waterworks and E was a ground waterworks. For the first time for archaea community analyses, this study employed the archaea-specific high-throughput sequencing primers for 16S ribosomal RNA (rRNA) as a target (reverse-transcribed cDNA; an RNA-based approach) in addition to the previously used 16S rRNA gene target (rDNA; a DNA-based approach) to reveal the active fraction of the archaea present in DWDS. The archaea community structure in varying environmental conditions in the water and biofilm of the five DWDSs were investigated by taking into consideration the system properties (cold or hot water system) and water age (distance from the treatment plants) in samples from each season of one year. The RNA-based archaea amplicon reads were obtained mostly from cold water samples from DWDSs (A-B) distributing water without disinfection where the DNA-based and RNA-based analysis created separate clusters in a weighted beta-diversity analysis. The season and location in DWDS A further affected the diversity of these archaea communities as was seen by different clusters in beta-diversity plots. The recovery of archaea reads was not adequate for analysis in any of the disinfected samples in DWDSs C-E or non-disinfected hot water in DWDSs A-B when utilizing RNA-based template. The metabolically active archaea community of DWDSs thus seemed to be effectively controlled by disinfection of water and in the hot water systems by the temperature. All biofilms regardless of DWDS showed lower species richness values (mainly Nitrososphaeria class) than non-disinfected water from DWDSs A-B where several archaea classes occurred (e.g. Woesearchaeia, Nitrososphaeria, Micrarchaeia, Methanomicrobia, Iairchaeia, Bathyarchaeia) indicating only part of the archaea members were able to survive in biofilms. Thus, Archaea has been shown as a significant part of normal DWDS biota, and their role especially in non-disinfected DWDS may be more important than previously considered.