PLOS Water

In-line chlorination automatically treats drinking water in piped water systems or at points-of-collection, substantially reducing morbidity and mortality while averting the burden of water treatment on individuals. Approximately 2.3 billion people globally use fecally contaminated drinking water infrastructure that is potentially compatible with in-line chlorination. In India, the Ministry of Jal Shaktis Jal Jeevan Mission has increased access to piped drinking water among rural households, however, water is often supplied intermittently without treatment. The aims of this study were to 1) test installation and operational procedures for in-line chlorination in rural drinking water systems, 2) evaluate acceptability and adoption of chlorinated water, and 3) assess reductions in drinking water fecal contamination, including with antibiotic resistant bacteria. We conducted a randomized controlled implementation trial of in-line chlorination in 20 villages in Rayagada district, Odisha, India over a 1-year period. Data collection included a baseline and post-intervention census (N=914) and surveys with a random subset of households over six timepoints (N=1,041) to assess water quality. We discuss operational challenges, including maintaining consistent chlorine dosing and addressing taste and odor preferences. In the treatment group, we detected free chlorine residual in 51% of tap water samples; this prevalence rose to 80% after increasing the target dose. Despite taste complaints in treatment communities, participants continued using piped water as their primary drinking water source. Individuals in treatment communities reported reduced time spent manually treating their drinking water, a responsibility borne by women in 92% of households. Treatment reduced E. coli prevalence by 70% in household tap water and by 47% in stored water, as well as reduced the presence of antibiotic resistant E. coli. In-line chlorination can lead to better quality water, lower the time burden of water treatment, and increase user adoption of chlorinated drinking water, but achieving this requires dedicated implementation and monitoring.

BackgroundThere is increasing evidence of the efficacy of point-of-use water filters on diarrhea prevalence in numerous global settings, in both observational studies and randomized experiments. Most studies, however, are focused on rural locations. Methods We use self-report household surveys to monitor a set of approximately 10,000 households receiving point-of-use water filters and WASH training in Kibera, Kenya. Twenty-five drinking water sources throughout the 7 neighborhoods were also selected for testing of E. coli, total coliform, bacterial 16S rRNA community sequencing and metals. Albendazole was provided to all households at distribution as part of the standard filter distribution protocol, with a subset of 2,642 households not receiving Albendazole at distribution, instead receiving it at the second follow-up (approximately 5 weeks after filter distribution). ResultsAfter data cleaning, a sample of 6,795 households were analyzed using mixed effects generalized linear models to account for repeated household measurements, geospatial and temporal effects, interviewer and other household covariates. Models predicted self-reported, 2-week prevalence of diarrhea. After accounting for confounding factors, self-reported diarrhea rates dropped from 52.7% to 2.2% after approximately 70 days of filter use. Field testing characterized most water sources (18 out of 25) as unsafe for Total coliforms, many for E. coli (6 out of 25), and one source above WHO health guidelines for arsenic. There was no evidence of a difference in self-reported diarrhea prevalence between households receiving Albendazole at distribution vs. those that didnt (p>0.05). ConclusionsThe introduction of Sawyer filters to households in a densely populated informal settlement reduced diarrhea and other health related problems. Representative water quality testing indicates a high frequency of drinking water source contamination with E. coli and Total Coliforms but a very low frequency of dissolved metals present, above WHO guidelines for drinking water. Anti-parasitic medication distribution had little to no impact on the results. Future randomized controlled studies with objective health measures are needed to ensure cause-effect impact of the filters, and study of filter longevity in the field continues to be a critical need.

BackgroundAchieving universal access to safe and affordable drinking water remains a critical global challenge, particularly in low- and middle-income countries where intermittencies in water supply are common. We aimed to examine relationships between water intermittency and multiple outcomes, including microbial water contamination, child health, and caregiver stress. MethodsWe conducted a cross-sectional study with 237 households with a child <5 years old in a peri-urban neighborhood of Blantyre, Malawi. We conducted a structured questionnaire with primary caregivers to record water intermittencies, water handling and hygiene practices, caregiver-reported child diarrhea and acute respiratory infection (ARI) symptoms, and stress among caregivers. Drinking water samples were tested for E. coli and cefotaxime-resistant E. coli using IDEXX Quanti-Tray/2000. We used generalized linear models to evaluate how intermittency occurrence, frequency, and duration influenced outcomes, adjusting for sociodemographic and WASH factors. FindingsOf 237 households, 32.5% reported [&ge;]1 water intermittency in the past month. These households were more likely to experience water insecurity, skip bathing and laundry, and report less handwashing after animal contact or outdoor work. E. coli was detected in 65.7% and cefotaxime-resistant E. coli in 8.4% of water samples. Intermittency was not associated with impaired water quality. Children in intermittent households had higher prevalence of diarrhea (PR=1.94, 95% CI: 1.11-3.39) and ARI with fever (PR=2.00, 95% CI: 1.11-3.60). Rare/short intermittencies were more strongly associated with diarrhea; frequent/long intermittencies were more strongly associated with ARI. Caregivers in households with short and frequent intermittencies reported higher stress. InterpretationWater intermittencies were associated with impaired hygiene, child illness, and caregiver stress. Frequency and duration modified the associations, suggesting that short- vs. long-term behavioral adaptations may differently influence exposure to enteric and respiratory pathogens and stress responses. Interventions like low-flow handwashing stations and water reuse may help reduce health risks in intermittent water settings.

Communities relying on drinking-water storage routinely document containers as a reservoir for pathogens when they are not properly cleaned. Although lab-based research provides guidance on cleaning protocols, gaps remain in understanding: 1) what methods are practiced, and 2) method effectiveness at reducing biofilm growth. In partnership with AFMAC, this pilot explored biofilm contamination in household water storage containers of four Zambian communities engaged in WASH-promoting community health clubs and health promoters. A mixed-methods design was used: 15-20 households per community completed a knowledge, attitudes, and practices survey and provided whole-container biofilm samples. Four focus group discussions with program members and four interviews with AFMAC staff explored container cleaning sensitizations. Findings revealed household awareness of container cleaning, inconsistent cleaning practices, and high levels of biofilm in 100% of storage containers. The absence of a standardized container cleaning module, reliance on visual cues as a proxy for microbial safety, and disproportionate burden placed on women underscore critical gaps in current programming and highlight container cleaning as a critical pathway to interrupt recontamination. This pilot highlights the need for locally adapted, practical protocols using accessible materials. Advancing container hygiene through integrated, adaptive, and scalable approaches is essential for reducing public health risks from unsafe water storage and improving point-of-use water quality. SynopsisField evidence on household water storage container cleaning is limited. The study found high biofilm levels and inconsistent cleaning of household water storage containers, revealing a gap between knowledge and practice in maintaining safe water quality. Table of Content Graphic O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=142 SRC="FIGDIR/small/25339087v1_ufig1.gif" ALT="Figure 1000"> View larger version (49K): org.highwire.dtl.DTLVardef@1ccb1ddorg.highwire.dtl.DTLVardef@1b15447org.highwire.dtl.DTLVardef@11416b3org.highwire.dtl.DTLVardef@1de5f0b_HPS_FORMAT_FIGEXP M_FIG C_FIG

Weather events associated with climate change can influence the environmental spread and survival of fecal pathogens, potentially impacting the efficacy of water, sanitation, and hygiene (WASH) interventions. We used longitudinal data from a randomized controlled trial in Bangladesh to assess whether rainfall and temperature modified the effect of an on-site sanitation intervention on fecal contamination. Over 3.5 years, we enumerated E. coli in household samples along eight fecal-oral pathways (n=23,238 samples) and obtained daily weather data. The intervention included the provision of double-pit latrines, child potties, and scoops for removing child and animal feces, along with behavior change promotion. The intervention was associated with larger reductions in E. coli in/on mother hands, child hands, ponds, and flies (0.10-log to 0.91-log) following higher rainfall and in/on food, mother hands, child hands, soil, and ponds (0.11-log to 0.40-log) following higher temperatures, compared to drier and colder periods. The intervention slightly reduced E. coli in stored drinking water and had no consistent effect on E. coli in tubewell water, regardless of weather. Our findings suggest that sanitation interventions can help mitigate the effects of increased rainfall and temperature on environmental fecal contamination. Previous analyses of these data without stratification by daily weather only found small (approximately 0.10-log) reductions in E. coli in/on stored drinking water and child hands. Future WASH trials should incorporate weather data to identify periods of differential intervention effectiveness to understand how weather variability influences the outcomes of public health interventions and develop strategies to enhance resilience against climate change impacts in vulnerable communities. Research in ContextO_ST_ABSEvidence before this studyC_ST_ABSHigher temperatures and increased rainfall linked to climate change have been associated with increased infectious disease transmission. Effective water, sanitation, and hygiene (WASH) interventions, such as access to improved sanitation facilities, are pivotal in reducing these risks. However, the efficacy of WASH interventions in interrupting fecal-oral transmission can be influenced by weather fluctuations, which can compromise service delivery and/or exacerbate environmental dissemination of fecal waste. We conducted a literature review on the impact of weather events and climate change on the environmental spread of fecal pathogens and their influence on WASH interventions. We used Google Scholar to identify articles published from 2000 to 2024 using the following search terms: ("climate change" OR weather OR weather events OR temperature OR heatwave OR rainfall OR precipitation) AND (sanitation OR "WASH intervention" OR "sanitation intervention" OR "intervention efficacy") AND (pathogen OR enteropathogen OR "Escherichia coli" OR "E. coli" OR "fecal indicator" OR "fecal contamination" OR "fecal pathogens). Studies have documented the influence of weather conditions, particularly rainfall and temperature, on the effectiveness of WASH interventions in reducing fecal contamination and diarrhea. A recent meta-analysis found that lack of access to improved latrines and piped drinking water modified the association between rainfall/temperature and diarrhea incidence in children under the age of five but this analysis only focused on health outcomes and did not include measures of environmental contamination. In Bangladesh, WASH interventions, including water treatment and handwashing, were more effective in reducing E. coli contamination in stored drinking water and food during dry seasons compared to wet seasons, however, in a different longitudinal analysis focused on a sanitation intervention, seasonality did not significantly modify the relationship between the intervention and fecal contamination. However, these studies used seasonal definitions (wet vs. dry) which can miss transient weather fluctuations within a season. One study in Kenya used daily rainfall and temperature data to show that increases in drinking water contamination associated with increased rainfall were mitigated if households treated their water. No study has evaluated the influence of daily weather parameters on the effect of WASH improvements on a comprehensive set of fecal-oral pathways. Added value of this studyThis study utilizes longitudinal data from a randomized controlled trial that implemented water treatment, handwashing and sanitation interventions in Bangladesh (WASH Benefits), with over 23,000 E. coli measurements collected over 3.5 years. The measurements were taken along eight fecal-oral transmission pathways, including mother and child hands, stored food, stored drinking water, groundwater from tubewells, ponds, courtyard soil, and captured flies. Previous analyses of these data with no stratification by daily weather found only small reductions in environmental fecal contamination associated with the WASH interventions. In the present analysis, we re-analyzed these data using daily weather measurements to investigate whether rainfall and temperature modified the efficacy of the improved on-site sanitation intervention in reducing E. coli contamination in the domestic environment. Unlike previous analyses that did not consider daily weather variations, stratifying by daily weather conditions revealed differential intervention impacts on E. coli contamination, with larger reductions observed following periods of higher rainfall and temperature across multiple fecal-oral pathways (mother hands, child hands, food, soil, ponds, and flies). Implications of all the available evidenceUnderstanding how interventions perform under varying weather conditions is crucial, particularly in low- and middle-income countries susceptible to extreme weather events. Integrating fine-grained weather data into sanitation intervention assessments can inform climate adaptation strategies. Our findings, combined with evidence from previous studies, indicate that on-site sanitation improvements, water treatment and broader WASH approaches can help protect against environmental fecal contamination associated with increased rainfall and temperatures. These insights are pivotal for designing resilient WASH interventions capable of mitigating climate-related health risks in vulnerable populations, supporting the Sustainable Development Goals in low- and middle-income countries. The WASH Benefits sanitation interventions implemented in rural Bangladesh demonstrated enhanced efficacy during periods of higher rainfall and temperature. These effects were not discernible in previous analyses that did not stratify by daily weather, where average reductions in E. coli counts were minimal, underscoring the importance of capturing weather variability in assessing intervention impacts. This approach also contrasts with earlier studies using calendar-based definitions of seasons, which may not fully capture the nuanced effects of climate variables on intervention efficacy. Collectively, these findings underscore the importance of incorporating fine-grained weather data in WASH trials to enhance the understanding of intervention efficacy under variable environmental conditions.

BackgroundWeather extremes are predicted to influence pathogen exposure but their effects on specific fecal-oral transmission pathways are not well investigated. We evaluated effects of extreme rain and temperature during different antecedent periods (0-14 days) on E. coli along eight fecal-oral transmission pathways in rural Bangladeshi households. MethodsE. coli was enumerated in mother and child hand rinses, food, stored drinking water, tubewells, flies, ponds, and courtyard soil using IDEXX Quanti-Tray/2000 in nine rounds over 3{middle dot}5 years (n=26,659 samples) and spatiotemporally matched to daily weather data. We used generalized linear models with robust standard errors to estimate E. coli count ratios (ECRs) associated with extreme rain and temperature, defined as >90th percentile of daily values during the study period. FindingsControlling for temperature, extreme rain on the sampling day was associated with increased E. coli in food (ECR=3{middle dot}13 (1{middle dot}63, 5{middle dot}99), p=0{middle dot}001), stored drinking water (ECR=1{middle dot}98 (1{middle dot}36, 2{middle dot}88), p<0{middle dot}0005) and ponds (ECR=3{middle dot}46 (2{middle dot}34, 5{middle dot}11), p<0{middle dot}0005), and reduced E. coli in soil (ECR=0{middle dot}36 (0{middle dot}24, 0{middle dot}53), p<0{middle dot}0005). Extreme rain the day before sampling was associated with reduced E. coli in tubewells (ECR=0{middle dot}10 (0{middle dot}02, 0{middle dot}62), p=0{middle dot}014). Effects were similar for rainfall 1-7 days before sampling and slightly attenuated for rainfall 14 days before sampling. Controlling for rainfall, extreme temperature on the sampling day was associated increased E. coli in stored drinking water (ECR=1{middle dot}49 (1{middle dot}05, 2{middle dot}12), p=0{middle dot}025) and food (ECR=3{middle dot}01 (1{middle dot}51, 6{middle dot}01), p=0{middle dot}002). Rainfall/temperature was not consistently associated with E. coli on hands and flies. InterpretationIn rural Bangladesh, measures to control enteric infections following weather extremes should focus on reducing contamination of drinking water and food stored at home and reducing exposure to surface waters. FundingBill & Melinda Gates Foundation, National Institutes of Health, World Bank. Research in Context Evidence before this studyHigher temperatures and levels of rainfall are associated with increased waterborne and vector-borne disease incidence, including child diarrhea. However, the specific pathways that facilitate increased transmission of diarrheagenic pathogens under these weather conditions have not been well investigated. We searched Google Scholar for articles published since 2000 using the following search terms: ("climate change" OR weather OR temperature OR heatwave OR rainfall OR precipitation) AND (pathogen OR enteropathogen OR "Escherichia coli" OR "E. coli" OR "fecal indicator" OR "fecal contamination") AND (environment OR water OR hands OR food OR soil OR flies). A large body of literature has evaluated the effect of rainfall or temperature on water quality and generally found that higher temperatures and magnitudes of rainfall were associated with higher levels of fecal indicator bacteria, such as Escherichia coli (E. coli), in surface and groundwaters, public and private drinking water sources and drinking water stored at homes. However, studies on the impact of rainfall and temperature on fecal contamination along non-waterborne fecal-oral transmission pathways are limited. Contamination of food stored at home has been linked to storage temperature. We found only one study on hand contamination with respect to weather, which found lower E. coli counts on child hands when daily temperatures were higher but no effect from rain. No studies have simultaneously assessed the effects of weather events on a comprehensive set of fecal-oral transmission pathways, which are typically described with the F-diagram and can include drinking water (at the source or stored), surface waters, caregiver and child hands, food, soil and flies. Added value of this studyWe spatiotemporally matched historical meteorological data to over 26,000 E. coli measurements collected in nine rounds over 3{middle dot}5 years in a randomized controlled trial in rural Bangladesh. E. coli was measured across eight different pathogen transmission pathways in the domestic environment. To our knowledge, this study is the first to utilize a large longitudinal dataset of environmental measurements collected over multiple years to investigate how increased rainfall and temperature affect fecal contamination across the full span of pathways described by the F-diagram. Our findings can help identify fecal-oral transmission pathways that are the most susceptible to extreme weather events and should be prioritized for intervention in their wake, as well as offer guidance on the time windows when interventions should be implemented with respect to weather events to interrupt these pathways in the context of climate change. This study can inform the effective delivery of WASH interventions, supporting climate change adaptation to reduce the enteric disease burden associated with weather extremes. Implications of all the available evidenceExtreme rainfall within two weeks of sampling was associated with increased E. coli contamination in stored food, stored drinking water and ponds, and reduced contamination of tubewell water and courtyard soil. Extreme temperature during the same timeframe was associated with increased contamination of stored food and stored drinking water. These findings illuminate environmental mechanisms behind previously reported increases in diarrheal diseases associated with extreme rainfall and temperature. Our findings suggest that, as extreme weather events become more common with climate change, intervention efforts to control exposure to fecal contamination in rural Bangladesh should prioritize reducing contamination of stored food and drinking water as well as reducing exposure to contaminated surface waters.

Universal access to safely managed water, defined as water from an improved source that is accessible on premises, available when needed, and free from contamination, remains far off in many low-resource settings. Progress toward safely managed services may include intermediate steps focused on improving some service attributes over others. However, knowledge surrounding households service preferences remains limited. This study explored satisfaction with existing water service characteristics and willingness-to-pay for hypothetical improvements in water supply. We conducted surveys with 1,748 households among 120 rural communities in northern Ghana. While few households (<2%) had safely managed water services, approximately half (52%) had at least basic access (i.e., an improved source where collection time is within 30 minutes). Satisfaction with existing water services was associated with source type, accessibility, availability, and perceived safety. For example, respondents who always had drinking water available in sufficient quantities had almost three times the odds of being satisfied (odds ratio [OR]: 2.66, 95% CI: 1.94-3.66), similar to respondents able to collect water within 30 minutes (OR: 3.09, 95% CI: 2.28-4.19) and those with on-premises water access (OR: 2.90, 95% CI: 1.26-6.66). Accessibility and water quality were particularly high priorities. Using discrete choice experiments, we estimated participants were willing to pay 3.6-4.9 times more for on-premises piped connections (compared with handpumps 30 minutes away) and chlorination (compared with no treatment). To a lesser extent, households were willing to pay for improved reliability, increased availability, and shorter collection times (10 vs. 30 minutes). Accordingly, we considered the possibility of introducing an intermediate service level: "proximate access", defined as using an improved source where water is collected within 10 minutes, available when needed, and (possibly) free from contamination. Such a category may provide feasible intermediate improvements to advance progress toward more highly valued safely managed services in rural, low-resource settings.

Background Bacteriological contamination of drinking water remains a major public health burden in sub-Saharan Africa, yet the full contamination chain from source to household has rarely been quantified at national scale. This study analyses water quality at both levels using the 2021 Cote d'Ivoire Demographic and Health Survey (DHS-CI 2021). Methods Cross-sectional secondary analysis of DHS-CI 2021 data. Households with paired bacteriological tests at the source (SH3227) and at the household (SH3225) were included (n = 2,541 for determinants; n = 2,528 for chain analysis). Contamination was defined as >0 CFU/100 ml. Determinants of source contamination were assessed by weighted logistic regression accounting for complex survey design. The contamination chain was described across four categories: safe throughout, recontaminated during transport/storage, decontaminated at home, and contaminated throughout. Results Weighted prevalence of source contamination was 63.6% [95% CI: 60.7-66.5%] and 77.0% [74.1-79.9%] at the household. Only 15.0% of households had safe water throughout the chain; 21.2% showed domestic recontamination and 60.8% consumed water contaminated at both levels. Key determinants of source contamination were use of an unimproved source (aOR = 8.15; 95% CI: 4.54-14.66), administrative region, travel time [&le;]30 minutes (aOR = 1.92; 95% CI: 1.41-2.62), and higher wealth quintiles (protective; aOR = 0.25 for richest). Model discrimination was good (AUC = 0.809). Conclusions The vast majority of Ivorian households consume bacteriologically unsafe water, with domestic recontamination representing a distinct and significant degradation pathway even among users of improved sources. Dual interventions targeting source protection and safe household water storage are urgently needed to advance progress toward SDG 6 in Cote d'Ivoire.

BackgroundFrom April 25, 2014, to October 15, 2015, residents of Flint, Michigan, were unknowingly exposed to contaminated water after a drinking water source switch. This public health disaster, the Flint water crisis (FWC), resulted in population-wide lead-in-water exposure for 18 months. A potent neurotoxin, lead is associated with multiple long-term adverse health conditions. In response to the FWC, the Flint Registry (FR) was formed to mitigate the impact of the crisis by proactively identifying the health and development concerns of participants and providing secondary prevention resources to address their needs. MethodsDesigned as a public health intervention with robust community engagement in every aspect, the FR enrollment is non-end-dated with ongoing evaluation and screening. Eligibility requirements include individuals who lived, worked, or went to school at an address serviced by the City of Flint water system anytime from 4/25/2014 to 10/15/2015, including those exposed prenatally. Recruitment is list-based, community-outreach-based, and marketing-based. Participants enroll primarily online or by phone and complete a survey to evaluate health impacts of the FWC and screen for service needs. Data collected at enrollment includes demographics, physical/mental health, child development, prior utilization of services, and environmental/lead-exposure risks. Based on responses, enrollees are referred within a community referral network to services in the categories of lead elimination, health, nutrition, and child development via an automated process. Frequencies of reported diagnoses, health symptoms, food access/insecurity problems, and educational support needs are compared to city, county, state, and national measures to identify ongoing disparities. Referrals identify the unmet needs for secondary prevention services. DiscussionThe FR protocol is unique because it not only conducts longitudinal surveillance, but it also improves public health through a community-wide referral process, allowing many people to be connected to health-promoting resources. This protocol is applicable to other public health crises due to its broad, city-wide surveillance and mitigation efforts, reduction of barriers to maximize participation, and incorporation of community voice. The FR protocol was designed in concert with the community, and community engagement remains a priority. Future work includes a diversified community engagement approach, ongoing enrollment, referrals, surveillance, and mitigation efforts.

Measurement of Enterococcus spp. levels with qPCR allows for same-day advisory notification of recreational water quality conditions, representing a major advance over traditional culture-based methods that require 18 or more hours to obtain results. In 2015, the United States Environmental Protection Agency released an Enterococcus qPCR protocol for recreational water quality testing. Over the past decade, there have been multiple advances in qPCR-based environmental testing, affording the opportunity to update the current methodology. A streamlined Enterococcus qPCR protocol is introduced that simplifies the mathematical model to estimate target sequence concentrations (TSC), reduces sample testing time by 20 min, incorporates a certified control material for standard curve generation, and introduces an inactivated E. faecalis whole cell DNA standard (WCDS) control material. A series of experiments were conducted 1) to compare results of the two Enterococcus qPCR protocols in analysis of marine, estuarine, and freshwater samples (n = 60), 2) to investigate alternative practices to adjust results due to potential water sample matrix interference, control material degradation, and/or analyst inconsistencies, and 3) to evaluate the performance, homogeneity, and stability of an inactivated E. faecalis cell preparation as a WCDS control material. Findings indicate a strong correlation between water sample mean log10 TSC per reaction results (R2 = 0.980) and 100% agreement in amplification and sample processing control tests. A Bayesian approach that accounts for uncertainty in qPCR measurements confirmed statistical equivalence for all water samples yielding paired measurements in the range of quantification, with 72.7% of samples exhibiting reduced error with the new streamlined protocol. Evaluation of three alternative practices to adjust for variation in Enterococcus qPCR measurements indicated no significant difference in water sample log10 TSC per reaction results with varying concentrations of treated sewage influent. Systematic testing of an inactivated WCDS control material yielded statistically equivalent performance compared to viable E. faecalis cell preparations. Homogeneity and stability experiments indicated that Enterococcus qPCR measurements of inactivated WCDS are reproducible across multiple preparations and that the material is stable at -20{degrees}C for at least 38 weeks. Together, experiments demonstrate that the streamlined protocol and alternative practices should make Enterococcus qPCR faster, easier to implement, safer, and more reproducible.

IntroductionSalmonella Typhi and Salmonella Paratyphi, fecal-oral transmitted bacterium have temporally and geographically heterogeneous pathways of transmission. Previous work in Kathmandu Nepal implicated stone waterspouts as a dominant transmission pathway after 77% of samples tested positive for S. Typhi and 70% for S. Paratyphi. Due to a falling water table, these spouts no longer provide drinking water, but typhoid fever persists, and the question of the diseases dominant pathway of transmission remains unanswered. MethodsWe used environmental surveillance to detect S. Typhi and Paratyphi DNA from potential sources of transmission. We collected 1L drinking water samples from a population-based random sample of households between February and October 2019 Between November 2019 and July 2021, we collected monthly 50 mL river water samples from 19 sites along the rivers leading through the Kathmandu and Kavre Districts of Nepal. We processed drinking water samples using a single qpcr and processed river water samples using differential centrifugation and qPCR at time 0 and after 16 hours of culture enrichment. A 3-cycle threshold (Ct) decrease of S. Typhi or S. Paratyphi, pre- and post-enrichment, was used as evidence of growth. We also performed structured observations of human-environment interactions to understand pathways of potential exposure. ResultsAmong 370 drinking water samples, S. Typhi was detected in 7 samples (1.8%) and S. Paratyphi A was detected in 4 (1.0%) samples. Among 381 river water samples, S. Typhi was detected in 171 (45%) and S. Paratyphi A was detected in 152 (42%) samples. Samples located upstream of the Kathmandu city center were positive for S. Typhi 12% of the time while samples from locations in and downstream the city had bacterial DNA detected 58% and 67% of the time respectively. Individuals were observed bathing in the rivers, washing clothes, and washing vegetables for sale in Kathmandu markets. ImplicationsThese results suggest that drinking water was not the dominant pathway of transmission of S. Typhi and S. Paratyphi A in the Kathmandu Valley in 2019. The high degree of river water contamination and its use for washing vegetables raises the possibility that river systems srepresent an important source of typhoid exposure in Kathmandu. Author SummaryUnderstanding the dominant route of transmission of a pathogen is important for designing and implementing effective control strategies. Salmonella Typhi and Paratyphi which cause typhoid and paratyphoid fever infect an estimated 10 million people and kill more than 100,000 annually. In Kathmandu prior work suggested that stone spouts where people collected drinking water were contaminated and driving transmission of the diseases. However, many of these spouts no longer function, and people are still getting sick. We tested drinking water from households in this area as well as local river water and found that 13 drinking water samples were positive for S.Typhi and 15 were positive for S. Paratyphi and many river samples tested positive for these bacterium. River water samples were not often positive upstream of Kathmandu city center (12% positive for S.Typhi) but were often positive within the city center (58% positive for S.Typhi) and in rural areas up to 10 km downstream of the city (67% positive for S.Typhi). During sample collection, individuals were observed interacting with rivers by walking in them, washing clothes and washing vegetables for sale in markets. This study shows that drinking water may not be primary driver of typhoid transmission in the Kathmandu valley, but that sewage contaminated river water may be a foci of transmission into the wider population.

Climate change may impact human health through the influence of weather on environmental transmission of diarrhea. Previous studies have found that high temperatures and heavy precipitation are associated with increased diarrhea prevalence, but the underlying causal mechanisms are not clear. We linked measurements of Escherichia coli in source water (n=1,673), stored drinking water (n=8,924), and hand rinses from children <2 years old (n=2,660) with publicly available gridded temperature and precipitation data (at [&le;]0.2 degree spatial resolution and daily temporal resolution) by the GPS coordinates and date of sample collection. Measurements were collected over a 3-year period across a 2500 km2 area in rural Kenya. In drinking water sources, high 7-day temperature was associated with a 0.16 increase in log10 E. coli levels (p<0.001), while heavy 7-day total precipitation was associated with a 0.29 increase in log10 E. coli levels (p<0.001). In household stored drinking water, heavy 7-day precipitation was associated with a 0.079 increase in log10 E. coli levels (p=0.042). On child hands, high 7-day temperature was associated with a 0.39 decrease in log10 E. coli levels (p<0.001). Our findings provide insight on how climate change could impact environmental transmission of bacterial pathogens in Kenya, and suggest water treatment could be a mitigation strategy.

Women shoulder the majority of water collection labor globally, yet how their water collection and water-related work experiences may change over time or by water source type remains insufficiently understood. We conducted a longitudinal, mixed-methods study in rural Kenya and Honduras to understand how women's experiences collecting water and performing water-related work varied between (a) two time points, (b) improved and unimproved water source types, and (c) water source location. Data were collected in 2023 and 2024 using interviews, observation, GPS-enabled watches, and scales to measure time and distance traveled, water weight and volume carried, and calories expended. 133 women participated in data collection (66 Kenya, 67 Honduras). We compared women's experience data by time point (2023 vs. 2024), source type (improved vs. unimproved), and source location (off-premises vs. on-premises) (t-test, Mann-Whitney U test). We also mapped participants' routes and activities to show which sources were visited, when, and for what activities. In Kenya, mean water collection time, distance, and caloric expenditure were significantly lower and water volume was significantly higher in 2024 when there were unexpected rains compared to 2023 when there was a persistent drought. When comparing source types during the 2023 drought, journeys to improved sources took significantly less time and energy and covered less distance than journeys to unimproved sources. These differences were not observed during the rainy conditions of 2024 when unimproved sources were closer and more accessible. In Honduras, water collection and water work burdens did not differ significantly by time point or source type. We found women with on-premises water access to still expend considerable time and caloric expenditure engaging in water work within their household compounds. Findings from Kenya suggest that water infrastructure improvements can reduce women's water collection burdens, though benefits may depend on and vary by season and source location. Findings from Honduras show that water labor does not end once water is in the household. Rather, substantial time and energy are expended carrying out water-related work even when sources are on premises, suggesting that efforts to assess water labor need to extend beyond collection alone. To meaningfully reduce burdens and ensure improved water sources are utilized during all seasons, initiatives need to consider source location, seasonal variability, and work beyond collection. Evaluations to assess infrastructure impacts on women's labor and well-being are needed and long overdue.

Managed aquifer recharge (MAR), a hydro-geological intervention designed to dilute groundwater salinity, pumps pond water treated through a slow sand filter into the underground aquifers. We evaluated the microbiological safety of the resulting MAR water at sites from three districts in southwest coastal Bangladesh. We collected monthly paired pond-MAR water samples from July 2016-June 2017 and enumerated fecal coliforms and E. coli using the IDEXX quanti-tray technique, by the most probable number (MPN) method. We used WHO risk categories for microbiological quality; no risk (<1 MPN), low risk (1-10 MPN) and moderate to high risk (>10 MPN per 100 mL water). We estimated the difference in mean log10 MPN in pond and MAR water using linear mixed effect models with random intercepts and cluster adjusted robust standard error. Almost all pond water samples (292/299, 98%) had moderate- to high-risk level (>10 MPN) fecal coliforms and E. coli (283/299, 95%). In contrast, 81% (242/300) of MAR water samples had no or low risk level fecal coliforms (0-10 MPN), of which 60% (179/300) had no fecal coliforms. We detected no or low risk level E. coli in 94% (283/300) of MAR water samples of these 80% (240/300) had no E. coli. MAR samples had lower mean log10 MPN fecal coliforms (-2.37; 95% CI: -2.56, -2.19) and E. coli (-2.26; 95% CI: - 2.43, -2.09) than pond water; microbial reductions remained consistent during the wet (May-October) and dry seasons. MAR-systems provided water with reduced fecal indicator bacteria compared to infiltered pond water.

Environmental exposure to Burkholderia pseudomallei, the causative agent of melioidosis, remains poorly characterised due to the limited sensitivity of conventional detection methods. This hinders accurate risk mapping and delays public health responses. Here, we developed CRISPR-BEEPs - a sensitive, equipment-light CRISPR-based assay - that demonstrated substantially improved sensitivity (93.5% vs 19.4%) and specificity (100% vs 98.0%) compared to conventional culture-based plate inspection techniques. We applied CRISPR-BEEPs to water samples collected from both natural and piped sources across 15,118 km{superscript 2} in northeast Thailand, including households of confirmed melioidosis patients and controls. B. pseudomallei was detected in 73.3% of groundwater samples, 32.4% of surface water, and 28.3% of piped water, with peak detection during the flood season. Importantly, the assays improved sensitivity enabled detection of a significant association between environmental B. pseudomallei detection within a 10 km radius of participants households and melioidosis risk (OR 2.74; 95% CI: 1.38- 5.48) - an association undetectable by conventional methods. These findings highlight major gaps in water treatment and distribution infrastructure and demonstrate the value of high-resolution environmental diagnostics. Strengthening water sanitation and surveillance systems is essential for mitigating melioidosis transmission and addressing the broader burden of waterborne diseases in vulnerable settings.

Combined water, sanitation, and handwashing (WSH) interventions have the potential to reduce fecal pathogens along more transmission pathways than single interventions alone. We measured Escherichia coli levels in 3909 drinking water samples, 2691 child hand rinses, and 2422 toy ball rinses collected from households enrolled in a two-year cluster-randomized controlled trial evaluating single and combined WSH interventions. Water treatment alone reduced E. coli in drinking water, while a combined WSH intervention improved water quality by the same magnitude but did not affect levels of fecal indicator bacteria on child hands or toy balls. The failure of the WSH interventions to reduce E. coli along important child exposure pathways is consistent with the lack of a protective effect from the interventions on child diarrhea or child growth during the trial. Our results have important implications for WSH program design; the sanitation and handwashing interventions implemented in this trial should not be expected to reduce child exposure to fecal contamination in other similar settings.

Refugees living in camps are particularly vulnerable to infectious disease outbreaks because of overcrowding, inadequate preventative health care, and limited water, sanitation, and hygiene services. As a result of the high disease prevalence, surveillance of cholera in Malawi provides a strong rationale for site and pathogen selection for refugee camp wastewater and environmental surveillance feasibility studies. We conducted a study in Dzaleka camp, Malawi, during a nationwide cholera outbreak. Incentivized refugee volunteers took samples for 19 weeks from seven high-use public pit latrines and a vacuum pump truck used for desludging fecal sludge. The National Microbiological Reference Laboratory of the Public Health Institute of Malawi used culture methods with confirmation via VITEK(R) MS or Analytical Profile Index to detect Vibrio cholerae. Academic partners provided technical input, training, and quality assurance through the study. The results were reviewed weekly at partner-coordination meetings. No V. cholerae was detected in samples, but one quality assurance sample tested positive during the study period. Here, we discuss the unique challenges of conducting wastewater and environmental surveillance in refugee camp settings, including our operational framework. This study provides a framework for up-scaling wastewater and environmental surveillance efforts in other humanitarian contexts through robust partnerships, processes and in-country tools. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=68 SRC="FIGDIR/small/25321306v1_ufig1.gif" ALT="Figure 1"> View larger version (34K): org.highwire.dtl.DTLVardef@1f7d609org.highwire.dtl.DTLVardef@4f630org.highwire.dtl.DTLVardef@1bef989org.highwire.dtl.DTLVardef@f09d8c_HPS_FORMAT_FIGEXP M_FIG C_FIG HighlightsO_LIA refugee camp in Malawi was investigated during a nationwide cholera outbreak C_LIO_LISamples were collected for 19 weeks from seven pit latrines and a pump truck C_LIO_LIVibrio cholerae was largely absent using culture methods C_LIO_LISurveillance of non-sewered sanitation in a camp setting is achievable through partnerships C_LI

IntroductionWastewater-based epidemiology (WBE) was implemented at the 2024 Republican and Democratic National Conventions (RNC and DNC, respectively)--two prominent large-scale events, each with estimated attendances of >50,000 persons. In preparation for event monitoring, the Wisconsin and Chicago WBE programs (associated with the RNC and DNC public health response, respectively) developed site-specific monitoring strategies and response plans, prioritized additional pathogens for event surveillance, and further optimized laboratory workflows to ensure rapid daily data reporting to public health. The Chicago program expanded the sewer sampling network to include new locations closer to event venues than previously available. Sampling was also conducted before the events, to establish baselines for endemic pathogens, as well as after each event to monitor for residual community transmission. MethodsSurveillance was expanded from the four respiratory pathogens regularly assessed by both WBE programs (SARS-CoV-2, influenza A, influenza B, respiratory syncytial virus) to include 3 gastrointestinal pathogens (norovirus, Salmonella enterica, Shiga toxin-producing E. coli). The Wisconsin program also conducted monitoring for the measles, mumps, rubella, and hepatitis A viruses. Wastewater sampling for the RNC was conducted at the community water reclamation facility level, while at the DNC samples were collected from manholes located downstream of the event venues. For both events, WBE data were summarized and contextualized alongside traditional public health surveillance data in daily situation reports. ResultsBetween the RNC and DNC response, a total of 112 wastewater samples were collected and assayed to provide concentration data on as many as 11 distinct pathogens of interest. Concentration results for the suite of pathogens were available within 12 to 36 hours of sample collection. In each instance when wastewater concentrations exceeded pre-established thresholds for action and flagged as an alert, other sources of contemporaneous public health surveillance information (e.g., clinical data) did not corroborate the WBE findings. ConclusionExisting WBE infrastructure in two U.S. cities was readily adapted for public health surveillance at two high-profile, large-scale events. Assays for additional event-relevant pathogens were quickly incorporated into routine laboratory workflows and data from wastewater samples were generated and reported with rapid turnaround-time. In considering the unique benefits of wastewater data, WBE results were a valuable supplement to other public health surveillance data in monitoring potential public health threats during these two large-scale events.

BackgroundSchools are high-risk settings for SARS-CoV-2 transmission, but necessary for childrens educational and social-emotional wellbeing. Previous research suggests that wastewater monitoring can detect SARS-CoV-2 infections in controlled residential settings with high levels of accuracy. However, its effective accuracy, cost, and feasibility in non-residential community settings is unknown. MethodsThe objective of this study was to determine the effectiveness and accuracy of community-based passive wastewater and surface (environmental) surveillance to detect SARS-CoV-2 infection in neighborhood schools compared to weekly diagnostic (PCR) testing. We implemented an environmental surveillance system in nine elementary schools with 1700 regularly present staff and students in southern California. The system was validated from November 2020 - March 2021. FindingsIn 447 data collection days across the nine sites 89 individuals tested positive for COVID-19, and SARS-CoV-2 was detected in 374 surface samples and 133 wastewater samples. Ninety-three percent of identified cases were associated with an environmental sample (95% CI: 88% - 98%); 67% were associated with a positive wastewater sample (95% CI: 57% - 77%), and 40% were associated with a positive surface sample (95% CI: 29% - 52%). The techniques we utilized allowed for near-complete genomic sequencing of wastewater and surface samples. InterpretationPassive environmental surveillance can detect the presence of COVID-19 cases in non-residential community school settings with a high degree of accuracy. FundingCounty of San Diego, Health and Human Services Agency, National Institutes of Health, National Science Foundation, Centers for Disease Control

BackgroundAntibiotic resistance is a leading cause of death, with the highest burden in low-resource settings. There is limited evidence on the potential for water, sanitation, and hygiene (WASH) infrastructure to reduce the burden of antibiotic resistance in humans. MethodsWe used geospatially tagged human gut metagenomes and household survey datasets to determine the association between antibiotic resistance gene (ARG) abundance and community-level coverage of improved drinking water points and improved sanitation facilities. Adjusted general linearized models with robust standard errors were used to estimate the relationship between ARG abundance in the human gut and access to water and sanitation. FindingsWe identified 1589 publicly available metagenomes from 26 countries. The average abundance of ARGs, in units of log10 ARG reads per kilobase per million (RPKM) mapped reads classified as bacteria, was highest in Africa compared to other World Health Organization (WHO) regions (one-way ANOVA p<0.001, post hoc Tukey HSD p<0.05). Increased access to both improved water and sanitation was associated with lower ARG abundance (effect estimate: -0.26, 95% CI [-0.44, -0.08]); the association was stronger in urban (-0.37 [-0.68, -0.07]) compared to rural areas (-0.16 [-0.38, 0.07]). Improved sanitation alone was associated with reduced ARG abundance (-0.16 [-0.32, 0.00]) while improved drinking water was not (-0.09 [-0.35, 0.16]). InterpretationWhile additional studies to investigate casual effects are needed, increasing access to water and sanitation could be an effective strategy to curb the proliferation of antibiotic resistance in low- and middle-income countries. FundingBill & Melinda Gates Foundation Research in ContextO_ST_ABSEvidence before this studyC_ST_ABSAntibiotic resistance is a growing global health threat that disproportionately affects low- and middle-income countries (LMICs). In 2019, an estimated 5 million deaths were associated with antibiotic resistance, with the highest death rate in western sub-Saharan Africa. Water, sanitation, and hygiene (WASH) interventions (e.g., household drinking water treatment, flush toilet, hand washing facilities with soap) can reduce diarrheal and respiratory infections, as reported in previous meta-analyses. Estimates, based on probability modeling, suggest improvements in water and sanitation could decrease antibiotic use for diarrheal disease treatment by 47-50% and 69-72%, respectively. Improving WASH infrastructure could theoretically contribute to the control of antibiotic resistance by preventing the release of antibiotics, resistant organisms, or antibiotic resistance genes (ARGs) into the environment, thus decreasing the burden of antibiotic-resistant infections. One global analysis across 73 countries suggested that improved infrastructure, including WASH services, was associated with reduced antibiotic resistance prevalence in isolates, however the independent effect of WASH access was not assessed. We searched PubMed for evidence on the impact of WASH interventions (excluding those related to animals and agriculture) on antibiotic resistance using the following keyword chain: (water OR sanitation OR hygiene OR WASH) AND (antimicrobial OR antibiotic) AND resistance) NOT ("OneHealth" OR "One Health" OR animal OR livestock). We selected reviews and systematic reviews (n=1420) to be screened for relevance to WASH and antibiotic resistance. The reference lists of included reviews were then searched for individual studies. We also consulted international agency guidelines and online resources from the Joint Programming Initiative on Antimicrobial Resistance, the International Scientific Forum on Hygiene, ReAct, Resistomap, and the London School of Hygiene and Tropical Medicine AMR Centre. Studies focusing on centralized water or wastewater treatment technologies in high income countries (HICs) reported variable removal of antibiotics (53 to >90%), antibiotic-resistant bacteria (90-99.9%) and ARGs (90-99.9%) from waste streams. Other studies were conducted on hand hygiene, which has proven effective at reducing human infections and antibiotic use. No studies were identified on the effect of on-site sanitation systems (e.g., pour-flush toilets, pit latrines), which serve an estimated 2.7 billion people globally, or fecal sludge management interventions on antibiotic resistance. Added value of this studyIn this study, we used 1589 publicly available human gut metagenomes from around the world to assess the abundance of ARGs as a function of access to improved drinking water and sanitation infrastructure. This analysis provides new evidence of differences in the abundance of antibiotic resistance in the human gut across the world and finds that decreased gut abundance of ARGs is associated with increased access to improved drinking water and sanitation. Implications of all the available evidenceCurrent approaches to controlling antibiotic resistance in humans predominantly focus on antibiotic stewardship; however, this approach is challenging in LMICs where infectious illnesses are generally more prevalent and unregulated antibiotic usage is common. Along with efforts to provide other known social benefits, such as reducing infectious disease and improving gender equality, improving access to safe drinking water and sanitation could contribute to reducing the burden of antibiotic resistance. This work highlights improving access to adequate water and sanitation as a potentially effective strategy, although additional studies designed to rigorously investigate the casual relationship between WASH and antibiotic resistance are needed.