Simultaneous detection of pathogens and antimicrobial resistance genes with the open source, cloud-based, CZ ID pipeline.

Antimicrobial resistant (AMR) pathogens represent urgent threats to human health, and their surveillance is of paramount importance. Metagenomic next generation sequencing (mNGS) has revolutionized such efforts, but remains challenging due to the lack of open-access bioinformatics tools capable of simultaneously analyzing both microbial and AMR gene sequences. To address this need, we developed the CZ ID AMR module, an open-access, cloud-based workflow designed to integrate detection of both microbes and AMR genes in mNGS and whole-genome sequencing (WGS) data. It leverages the Comprehensive Antibiotic Resistance Database and associated Resistance Gene Identifier software, and works synergistically with the CZ ID short-read mNGS module to enable broad detection of both microbes and AMR genes. We highlight diverse applications of the AMR module through analysis of both publicly available and newly generated mNGS and WGS data from four clinical cohort studies and an environmental surveillance project. Through genomic investigations of bacterial sepsis and pneumonia cases, hospital outbreaks, and wastewater surveillance data, we gain a deeper understanding of infectious agents and their resistomes, highlighting the value of integrating microbial identification and AMR profiling for both research and public health. We leverage additional functionalities of the CZ ID mNGS platform to couple resistome profiling with the assessment of phylogenetic relationships between nosocomial pathogens, and further demonstrate the potential to capture the longitudinal dynamics of pathogen and AMR genes in hospital acquired bacterial infections. In sum, the new AMR module advances the capabilities of the open-access CZ ID microbial bioinformatics platform by integrating pathogen detection and AMR profiling from mNGS and WGS data. Its development represents a critical step toward democratizing pathogen genomic analysis and supporting collaborative efforts to combat the growing threat of AMR.

Addressing Fecal Contamination in Rural Kenyan Households: The Roles of Environmental Interventions and Animal Ownership.

Combined water, sanitation, and handwashing (WSH) interventions could reduce fecal contamination 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 2-year cluster-randomized controlled trial evaluating single and combined WSH interventions. Water treatment with chlorine reduced E. coli in drinking water. A combined WSH intervention improved water quality by the same magnitude but did not affect E. coli levels on hands or toys. One potential explanation for the limited impact of the sanitation intervention (upgraded latrines) is failure to address dog and livestock fecal contamination. Small ruminant (goat or sheep) ownership was associated with increased E. coli levels in stored water and on child hands. Cattle and poultry ownership was protective against child stunting, and domesticated animal ownership was not associated with child diarrhea. Our findings do not support restricting household animal ownership to prevent child diarrheal disease or stunting but do support calls for WSH infrastructure that can more effectively reduce household fecal contamination.

Experimental evolution of Staphylococcus aureus in macrophages: dissection of a conditional adaptive trait promoting intracellular survival.

Staphylococcus aureus is a major pathogen associated with important diseases in humans and animals. Macrophages are a key component of the innate immune response to S. aureus infection and play a major role in disease outcomes. To investigate the adaptive evolution of S. aureus in response to macrophages, we developed an experimental infection assay. S. aureus strains representing major human epidemic clones were passaged many times in a macrophage cell line, accumulating mutations in an array of genomic loci. Phenotypic analysis revealed the emergence of a lineage exhibiting increased survival in macrophages and human blood, and resistance to vancomycin. The evolved lineage exhibited a previously undescribed small colony variant (SCV) phenotype characterized by hyper-pigmentation, which resulted from a missense mutation in rsbW. Notably, the novel SCV was a conditional adaptive trait that was unstable in nutrient-replete conditions in vitro, rapidly converting from hyper-pigmented SCV to a non-pigmented large colony variant via spontaneous sigB deletion events. Importantly, we identified similar deletions in the genome sequences of a limited number of clinical S. aureus isolates from public databases, indicating that related events may occur during clinical infection. Experimental infection of zebrafish did not reveal a difference in virulence between parent and novel SCV but demonstrated an in vivo fitness cost for the compensatory sigB deletion events. Taken together, we report an experimental evolutionary approach for investigating bacterial innate immune cell interactions, revealing a conditional adaptation that promotes S. aureus survival in macrophages and resistance to vancomycin. IMPORTANCE: Staphylococcus aureus is an important human bacterial pathogen. The host response to S. aureus involves the production of innate immune cells such as macrophages which are important for fighting infection. Here we report a new model of experimental evolution for studying how S. aureus can evade killing by macrophages. We identified a novel adaptive phenotype that promotes survival in macrophages and blood and resistance to antibiotics. The phenotype is lost rapidly upon growth in nutrient-rich conditions via disruption of the alternative sigma factor sigB, revealing a conditional niche-specific fitness advantage. Genomic analysis of clinical isolates suggests similar adaptations may occur during human infections. Our model may be used broadly to identify adaptations of S. aureus to the innate immune response.

Simultaneous detection of pathogens and antimicrobial resistance genes with the open source, cloud-based, CZ ID pipeline.

Antimicrobial resistant (AMR) pathogens represent urgent threats to human health, and their surveillance is of paramount importance. Metagenomic next generation sequencing (mNGS) has revolutionized such efforts, but remains challenging due to the lack of open-access bioinformatics tools capable of simultaneously analyzing both microbial and AMR gene sequences. To address this need, we developed the Chan Zuckerberg ID (CZ ID) AMR module, an open-access, cloud-based workflow designed to integrate detection of both microbes and AMR genes in mNGS and whole-genome sequencing (WGS) data. It leverages the Comprehensive Antibiotic Resistance Database and associated Resistance Gene Identifier software, and works synergistically with the CZ ID short-read mNGS module to enable broad detection of both microbes and AMR genes. We highlight diverse applications of the AMR module through analysis of both publicly available and newly generated mNGS and WGS data from four clinical cohort studies and an environmental surveillance project. Through genomic investigations of bacterial sepsis and pneumonia cases, hospital outbreaks, and wastewater surveillance data, we gain a deeper understanding of infectious agents and their resistomes, highlighting the value of integrating microbial identification and AMR profiling for both research and public health. We leverage additional functionalities of the CZ ID mNGS platform to couple resistome profiling with the assessment of phylogenetic relationships between nosocomial pathogens, and further demonstrate the potential to capture the longitudinal dynamics of pathogen and AMR genes in hospital acquired bacterial infections. In sum, the new AMR module advances the capabilities of the open-access CZ ID microbial bioinformatics platform by integrating pathogen detection and AMR profiling from mNGS and WGS data. Its development represents a critical step toward democratizing pathogen genomic analysis and supporting collaborative efforts to combat the growing threat of AMR.

Improved Child Feces Management Mediates Reductions in Childhood Diarrhea from an On-Site Sanitation Intervention: Causal Mediation Analysis of a Cluster-Randomized Trial in Rural Bangladesh.

BACKGROUND: The WASH benefits Bangladesh trial multi-component sanitation intervention reduced diarrheal disease among children < 5 years. Intervention components included latrine upgrades, child feces management tools, and behavioral promotion. It remains unclear which components most impacted diarrhea. METHODS: We conducted mediation analysis within a subset of households (n = 720) from the sanitation and control arms. Potential mediators were categorized into indicators of latrine quality, latrine use practices, and feces management practices. We estimated average causal mediation effects (ACME) as prevalence differences (PD), defined as the intervention's effect on diarrhea through its effect on the mediator. RESULTS: The intervention improved all indicators compared to controls. We found significant mediation through multiple latrine use and feces management practice indicators. The strongest mediators during monsoon seasons were reduced open defecation among children aged < 3 and 3-8 years, and increased disposal of child feces into latrines. The strongest mediators during dry seasons were access to a flush/pour-flush latrine, reduced open defecation among children aged 3-8 years, and increased disposal of child feces into latrines. Individual mediation effects were small (PD = 0.5-2 percentage points) compared to the overall intervention effect but collectively describe significant mediation pathways. DISCUSSION: The effect of the WASH Benefits Bangladesh sanitation intervention on diarrheal disease was mediated through improved child feces management and reduced child open defecation. Although the intervention significantly improved latrine quality, relatively high latrine quality at baseline may have limited benefits from additional improvements. Targeting safe child feces management may increase the health benefits of rural sanitation interventions.

Geographic pair matching in large-scale cluster randomized trials.

Cluster randomized trials are often used to study large-scale public health interventions. In large trials, even small improvements in statistical efficiency can have profound impacts on the required sample size and cost. Location integrates many socio-demographic and environmental characteristics into a single, readily available feature. Here we show that pair matching by geographic location leads to substantial gains in statistical efficiency for 14 child health outcomes that span growth, development, and infectious disease through a re-analysis of two large-scale trials of nutritional and environmental interventions in Bangladesh and Kenya. Relative efficiencies from pair matching are ≥1.1 for all outcomes and regularly exceed 2.0, meaning an unmatched trial would need to enroll at least twice as many clusters to achieve the same level of precision as the geographically pair matched design. We also show that geographically pair matched designs enable estimation of fine-scale, spatially varying effect heterogeneity under minimal assumptions. Our results demonstrate broad, substantial benefits of geographic pair matching in large-scale, cluster randomized trials.

Effects of breastfeeding on children's gut colonization with multidrug-resistant Enterobacterales in peri-urban Lima, Peru.

Children living in low-resource settings are frequently gut-colonized with multidrug-resistant bacteria. We explored whether breastfeeding may protect against children's incident gut colonization with extended-spectrum beta-lactamase-producing Escherichia coli (ESBL-Ec) and Klebsiella, Enterobacter, or Citrobacter spp. (ESBL-KEC). We screened 937 monthly stool samples collected from 112 children aged 1-16 months during a 2016-19 prospective cohort study of enteric infections in peri-urban Lima. We used 52,816 daily surveys to examine how exposures to breastfeeding in the 30 days prior to a stool sample were associated with children's risks of incident gut-colonization, controlling for antibiotic use and other covariates. We sequenced 78 ESBL-Ec from 47 children to explore their diversity. Gut-colonization with ESBL-Ec was increasingly prevalent as children aged, approaching 75% by 16 months, while ESBL-KEC prevalence fluctuated between 18% and 36%. Through 6 months of age, exclusively providing human milk in the 30 days prior to a stool sample did not reduce children's risk of incident gut-colonization with ESBL-Ec or ESBL-KEC. From 6 to 16 months of age, every 3 additional days of breastfeeding in the prior 30 days was associated with 6% lower risk of incident ESBL-Ec gut-colonization (95% CI: 0.90, 0.98, p = .003). No effects were observed on incident ESBL-KEC colonization. We detected highly diverse ESBL-Ec among children and few differences between children who were predominantly breastfed (mean age: 4.1 months) versus older children (10.8 months). Continued breastfeeding after 6 months conferred protection against children's incident gut colonization with ESBL-Ec in this setting. Policies supporting continued breastfeeding should be considered in efforts to combat antibiotic resistance.

Is detection of enteropathogens and human or animal faecal markers in the environment associated with subsequent child enteric infections and growth: an individual participant data meta-analysis.

BACKGROUND: Quantifying contributions of environmental faecal contamination to child diarrhoea and growth faltering can illuminate causal mechanisms behind modest health benefits in recent water, sanitation, and hygiene (WASH) trials. We aimed to assess associations between environmental detection of enteropathogens and human or animal microbial source tracking markers (MSTM) and subsequent child health outcomes. METHODS: In this individual participant data meta-analysis we searched we searched PubMed, Embase, CAB Direct Global Health, Agricultural and Environmental Science Database, Web of Science, and Scopus for WASH intervention studies with a prospective design and concurrent control that measured enteropathogens or MSTM in environmental samples, or both, and subsequently measured enteric infections, diarrhoea, or height-for-age Z-scores (HAZ) in children younger than 5 years. We excluded studies that only measured faecal indicator bacteria. The initial search was done on Jan 19, 2021, and updated on March 22, 2023. One reviewer (AM) screened abstracts, and two independent reviewers (AM and RT) examined the full texts of short-listed articles. All included studies include at least one author that also contributed as an author to the present Article. Our primary outcomes were the 7-day prevalence of caregiver-reported diarrhoea and HAZ in children. For specific enteropathogens in the environment, primary outcomes also included subsequent child infection with the same pathogen ascertained by stool testing. We estimated associations using covariate-adjusted regressions and pooled estimates across studies. FINDINGS: Data from nine published reports from five interventions studies, which included 8603 children (4302 girls and 4301 boys), were included in the meta-analysis. Environmental pathogen detection was associated with increased infection prevalence with the same pathogen and lower HAZ (ΔHAZ -0·09 [95% CI -0·17 to -0·01]) but not diarrhoea (prevalence ratio 1·22 [95% CI 0·95 to 1·58]), except during wet seasons. Detection of MSTM was not associated with diarrhoea (no pooled estimate) or HAZ (ΔHAZ -0·01 [-0·13 to 0·11] for human markers and ΔHAZ -0·02 [-0·24 to 0·21] for animal markers). Soil, children's hands, and stored drinking water were major transmission pathways. INTERPRETATION: Our findings support a causal chain from pathogens in the environment to infection to growth faltering, indicating that the lack of WASH intervention effects on child growth might stem from insufficient reductions in environmental pathogen prevalence. Studies measuring enteropathogens in the environment should subsequently measure the same pathogens in stool to further examine theories of change between WASH, faecal contamination, and health. Given that environmental pathogen detection was predictive of infection, programmes targeting specific pathogens (eg, vaccinations and elimination efforts) can environmentally monitor the pathogens of interest for population-level surveillance instead of collecting individual biospecimens. FUNDING: The Bill & Melinda Gates Foundation and the UK Foreign and Commonwealth Development Office.

Safe Today, Unsafe Tomorrow: Tanzanian Households Experience Variability in Drinking Water Quality.

Measuring Escherichia coli in a single-grab sample of stored drinking water is often used to characterize drinking water quality. However, if water quality exhibits variability temporally, then one-time measurement schemes may be insufficient to adequately characterize the quality of water that people consume. This study uses longitudinal data collected from 193 households in peri-urban Tanzania to assess variability in stored water quality and to characterize uncertainty with different data collection schemes. Households were visited 5 times over the course of a year. At each visit, information was collected on water management practices, and a sample of stored drinking water was collected for E. coli enumeration. Water quality was poor for households, with 80% having highly contaminated (>100 CFU per 100 mL) water during at least one visit. There was substantial variability of water quality for households, with only 3% of households having the same category (low, medium, or high) of water quality for all five visits. These data suggest a single sample would inaccurately characterize a household's drinking water quality over the course of a year and lead to misestimates of population level access to safe drinking water.

Risk factors for extended-spectrum beta-lactamase (ESBL)-producing E. coli carriage among children in a food animal-producing region of Ecuador: A repeated measures observational study.

BACKGROUND: The spread of antibiotic-resistant bacteria may be driven by human-animal-environment interactions, especially in regions with limited restrictions on antibiotic use, widespread food animal production, and free-roaming domestic animals. In this study, we aimed to identify risk factors related to commercial food animal production, small-scale or "backyard" food animal production, domestic animal ownership, and practices related to animal handling, waste disposal, and antibiotic use in Ecuadorian communities. METHODS AND FINDINGS: We conducted a repeated measures study from 2018 to 2021 in 7 semirural parishes of Quito, Ecuador to identify determinants of third-generation cephalosporin-resistant E. coli (3GCR-EC) and extended-spectrum beta-lactamase E. coli (ESBL-EC) in children. We collected 1,699 fecal samples from 600 children and 1,871 domestic animal fecal samples from 376 of the same households at up to 5 time points per household over the 3-year study period. We used multivariable log-binomial regression models to estimate relative risks (RR) of 3GCR-EC and ESBL-EC carriage, adjusting for child sex and age, caregiver education, household wealth, and recent child antibiotic use. Risk factors for 3GCR-EC included living within 5 km of more than 5 commercial food animal operations (RR: 1.26; 95% confidence interval (CI): 1.10, 1.45; p-value: 0.001), household pig ownership (RR: 1.23; 95% CI: 1.02, 1.48; p-value: 0.030) and child pet contact (RR: 1.23; 95% CI: 1.09, 1.39; p-value: 0.001). Risk factors for ESBL-EC were dog ownership (RR: 1.35; 95% CI: 1.00, 1.83; p-value: 0.053), child pet contact (RR: 1.54; 95% CI: 1.10, 2.16; p-value: 0.012), and placing animal feces on household land/crops (RR: 1.63; 95% CI: 1.09, 2.46; p-value: 0.019). The primary limitations of this study are the use of proxy and self-reported exposure measures and the use of a single beta-lactamase drug (ceftazidime with clavulanic acid) in combination disk diffusion tests for ESBL confirmation, potentially underestimating phenotypic ESBL production among cephalosporin-resistant E. coli isolates. To improve ESBL determination, it is recommended to use 2 combination disk diffusion tests (ceftazidime with clavulanic acid and cefotaxime with clavulanic acid) for ESBL confirmatory testing. Future studies should also characterize transmission pathways by assessing antibiotic resistance in commercial food animals and environmental reservoirs. CONCLUSIONS: In this study, we observed an increase in enteric colonization of antibiotic-resistant bacteria among children with exposures to domestic animals and their waste in the household environment and children living in areas with a higher density of commercial food animal production operations.

Leveraging the COVID-19 pandemic as a natural experiment to assess changes in antibiotic use and antibiotic-resistant E. coli carriage in semi-rural Ecuador.

The coronavirus 2019 (COVID-19) pandemic has had significant impacts on health systems, population dynamics, public health awareness, and antibiotic stewardship, which could affect antibiotic resistant bacteria (ARB) emergence and transmission. In this study, we aimed to compare knowledge, attitudes, and practices (KAP) of antibiotic use and ARB carriage in Ecuadorian communities before versus after the COVID-19 pandemic began. We leveraged data collected for a repeated measures observational study of third-generation cephalosporin-resistant E. coli (3GCR-EC) carriage among children in semi-rural communities in Quito, Ecuador between July 2018 and September 2021. We included 241 households that participated in surveys and child stool sample collection in 2019, before the pandemic, and in 2021, after the pandemic began. We estimated adjusted Prevalence Ratios (aPR) and 95% Confidence Intervals (CI) using logistic and Poisson regression models. Child antibiotic use in the last 3 months declined from 17% pre-pandemic to 5% in 2021 (aPR: 0.30; 95% CI 0.15, 0.61) and 3GCR-EC carriage among children declined from 40 to 23% (aPR: 0.48; 95% CI 0.32, 0.73). Multi-drug resistance declined from 86 to 70% (aPR: 0.32; 95% CI 0.13; 0.79), the average number of antibiotic resistance genes (ARGs) per 3GCR-EC isolate declined from 9.9 to 7.8 (aPR of 0.79; 95% CI 0.65, 0.96), and the diversity of ARGs was lower in 2021. In the context of Ecuador, where COVID-19 prevention and control measures were strictly enforced after its major cities experienced some of the world's the highest mortality rates from SARS-CoV-2 infections, antibiotic use and ARB carriage declined in semi-rural communities of Quito from 2019 to 2021.

Evaluating the relationship between community water and sanitation access and the global burden of antibiotic resistance: an ecological study.

BACKGROUND: Antibiotic resistance is a leading cause of death, with the highest burden occurring in low-resource settings. There is little evidence on the potential for water, sanitation, and hygiene (WASH) access to reduce antibiotic resistance in humans. We aimed to determine the relationship between the burden of antibiotic resistance in humans and community access to drinking water and sanitation. METHODS: In this ecological study, we linked publicly available, geospatially tagged human faecal metagenomes (from the US National Center for Biotechnology Information Sequence Read Archive) with georeferenced household survey datasets that reported access to drinking water sources and sanitation facility types. We used generalised linear models with robust SEs to estimate the relationship between the abundance of antibiotic resistance genes (ARGs) in human faecal metagenomes and community-level coverage of improved drinking water and sanitation within a defined radii of faecal metagenome coordinates. FINDINGS: We identified 1589 metagenomes from 26 countries. The mean abundance of ARGs, in units of log10 ARG fragments per kilobase per million mapped reads classified as bacteria, was highest in Africa compared with Europe (p=0·014), North America (p=0·0032), and the Western Pacific (p=0·011), and second highest in South-East Asia compared with Europe (p=0·047) and North America (p=0·014). Increased access to improved water and sanitation was associated with lower ARG abundance (effect estimate -0·22, [95% CI -0·39 to -0·05]) and the association was stronger in urban (-0·32 [-0·63 to 0·00]) than in rural (-0·16 [-0·38 to 0·07]) areas. INTERPRETATION: Although additional studies to investigate causal effects are needed, increasing access to water and sanitation could be an effective strategy to curb the proliferation of antibiotic resistance in low-income and middle-income countries. FUNDING: Bill & Melinda Gates Foundation.

Biosecurity and water, sanitation, and hygiene (WASH) interventions in animal agricultural settings for reducing infection burden, antibiotic use, and antibiotic resistance: a One Health systematic review.

Prevention and control of infections across the One Health spectrum is essential for improving antibiotic use and addressing the emergence and spread of antibiotic resistance. Evidence for how best to manage these risks in agricultural communities-45% of households globally-has not been systematically assembled. This systematic review identifies and summarises evidence from on-farm biosecurity and water, sanitation, and hygiene (WASH) interventions with the potential to directly or indirectly reduce infections and antibiotic resistance in animal agricultural settings. We searched 17 scientific databases (including Web of Science, PubMed, and regional databases) and grey literature from database inception to Dec 31, 2019 for articles that assessed biosecurity or WASH interventions measuring our outcomes of interest; namely, infection burden, microbial loads, antibiotic use, and antibiotic resistance in animals, humans, or the environment. Risk of bias was assessed with the Systematic Review Centre for Laboratory Animal Experimentation tool, Risk of Bias in Non-Randomized Studies of Interventions, and the Appraisal tool for Cross-Sectional Studies, although no studies were excluded as a result. Due to the heterogeneity of interventions found, we conducted a narrative synthesis. The protocol was pre-registered with PROSPERO (CRD42020162345). Of the 20 672 publications screened, 104 were included in this systematic review. 64 studies were conducted in high-income countries, 24 studies in upper-middle-income countries, 13 studies in lower-middle-income countries, two in low-income countries, and one included both upper-middle-income countries and lower-middle-income countries. 48 interventions focused on livestock (mainly pigs), 43 poultry (mainly chickens), one on livestock and poultry, and 12 on aquaculture farms. 68 of 104 interventions took place on intensive farms, 22 in experimental settings, and ten in smallholder or subsistence farms. Positive outcomes were reported for ten of 23 water studies, 17 of 35 hygiene studies, 15 of 24 sanitation studies, all three air-quality studies, and 11 of 17 other biosecurity-related interventions. In total, 18 of 26 studies reported reduced infection or diseases, 37 of 71 studies reported reduced microbial loads, four of five studies reported reduced antibiotic use, and seven of 20 studies reported reduced antibiotic resistance. Overall, risk of bias was high in 28 of 57 studies with positive interventions and 17 of 30 studies with negative or neutral interventions. Farm-management interventions successfully reduced antibiotic use by up to 57%. Manure-oriented interventions reduced antibiotic resistance genes or antibiotic-resistant bacteria in animal waste by up to 99%. This systematic review highlights the challenges of preventing and controlling infections and antimicrobial resistance, even in well resourced agricultural settings. Most of the evidence emerges from studies that focus on the farm itself, rather than targeting agricultural communities or the broader social, economic, and policy environment that could affect their outcomes. WASH and biosecurity interventions could complement each other when addressing antimicrobial resistance in the human, animal, and environmental interface.

Geographic pair-matching in large-scale cluster randomized trials.

Custer randomized trials are often used to study large-scale public health interventions. In large trials, even small improvements in statistical efficiency can have profound impacts on the required sample size and cost. Pair matched randomization is one strategy with potential to increase trial efficiency, but to our knowledge there have been no empirical evaluations of pair-matching in large-scale, epidemiologic field trials. Location integrates many socio-demographic and environmental characteristics into a single feature. Here, we show that geographic pair-matching leads to substantial gains in statistical efficiency for 14 child health outcomes that span growth, development, and infectious disease through a re-analysis of two large-scale trials of nutritional and environmental interventions in Bangladesh and Kenya. We estimate relative efficiencies ≥1.1 for all outcomes assessed and relative efficiencies regularly exceed 2.0, meaning an unmatched trial would have needed to enroll at least twice as many clusters to achieve the same level of precision as the geographically pair-matched design. We also show that geographically pair-matched designs enable estimation of fine-scale, spatially varying effect heterogeneity under minimal assumptions. Our results demonstrate broad, substantial benefits of geographic pair-matching in large-scale, cluster randomized trials.

Effects of High Temperature and Heavy Precipitation on Drinking Water Quality and Child Hand Contamination Levels in Rural Kenya.

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 have not been tested and validated. We linked measurements of Escherichia coli in source water (n = 1673), stored drinking water (n = 9692), and hand rinses from children <2 years old (n = 2634) with publicly available gridded temperature and precipitation data (at ≤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, 95% CI: 0.07, 0.24), while heavy 7-day total precipitation was associated with a 0.29 increase in log10 E. coli levels (p < 0.001, 95% CI: 0.13, 0.44). In household stored drinking water, heavy 7-day precipitation was associated with a 0.079 increase in log10 E. coli levels (p = 0.042, 95% CI: 0.07, 0.24). Heavy precipitation did not increase E. coli levels among respondents who treated their water, suggesting that water treatment can mitigate effects on water quality. On child hands, high 7-day temperature was associated with a 0.39 decrease in log10 E. coli levels (p < 0.001, 95% CI: -0.52, -0.27). Our findings provide insight on how climate change could impact environmental transmission of bacterial pathogens in Kenya. We suggest water treatment is especially important after heavy precipitation (particularly when preceded by dry periods) and high temperatures.

Effects of water, sanitation, and hygiene interventions on detection of enteropathogens and host-specific faecal markers in the environment: a systematic review and individual participant data meta-analysis.

BACKGROUND: Water, sanitation, and hygiene (WASH) improvements are promoted to reduce diarrhoea in low-income countries. However, trials from the past 5 years have found mixed effects of household-level and community-level WASH interventions on child health. Measuring pathogens and host-specific faecal markers in the environment can help investigate causal pathways between WASH and health by quantifying whether and by how much interventions reduce environmental exposure to enteric pathogens and faecal contamination from human and different animal sources. We aimed to assess the effects of WASH interventions on enteropathogens and microbial source tracking (MST) markers in environmental samples. METHODS: We did a systematic review and individual participant data meta-analysis, which included searches from Jan 1, 2000, to Jan 5, 2023, from PubMed, Embase, CAB Direct Global Health, Agricultural and Environmental Science Database, Web of Science, and Scopus, of prospective studies with water, sanitation, or hygiene interventions and concurrent control group that measured pathogens or MST markers in environmental samples and measured child anthropometry, diarrhoea, or pathogen-specific infections. We used covariate-adjusted regression models with robust standard errors to estimate study-specific intervention effects and pooled effect estimates across studies using random-effects models. FINDINGS: Few trials have measured the effect of sanitation interventions on pathogens and MST markers in the environment and they mostly focused on onsite sanitation. We extracted individual participant data on nine environmental assessments from five eligible trials. Environmental sampling included drinking water, hand rinses, soil, and flies. Interventions were consistently associated with reduced pathogen detection in the environment but effect estimates in most individual studies could not be distinguished from chance. Pooled across studies, we found a small reduction in the prevalence of any pathogen in any sample type (pooled prevalence ratio [PR] 0·94 [95% CI 0·90-0·99]). Interventions had no effect on the prevalence of MST markers from humans (pooled PR 1·00 [95% CI 0·88-1·13]) or animals (pooled PR 1·00 [95% CI 0·97-1·03]). INTERPRETATION: The small effect of these sanitation interventions on pathogen detection and absence of effects on human or animal faecal markers are consistent with the small or null health effects previously reported in these trials. Our findings suggest that the basic sanitation interventions implemented in these studies did not contain human waste and did not adequately reduce exposure to enteropathogens in the environment. FUNDING: Bill and Melinda Gates Foundation and the UK Foreign and Commonwealth Development Office.

Learning and STEM identity gains from an online module on sequencing-based surveillance of antimicrobial resistance in the environment: An analysis of the PARE-Seq curriculum.

COVID-19 necessitated the rapid transition to online learning, challenging the ability of Science, Technology, Engineering, and Math (STEM) professors to offer laboratory experiences to their students. As a result, many instructors sought online alternatives. In addition, recent literature supports the capacity of online curricula to empower students of historically underrepresented identities in STEM fields. Here, we present PARE-Seq, a virtual bioinformatics activity highlighting approaches to antimicrobial resistance (AMR) research. Following curricular development and assessment tool validation, pre- and post-assessments of 101 undergraduates from 4 institutions revealed that students experienced both significant learning gains and increases in STEM identity, but with small effect sizes. Learning gains were marginally modified by gender, race/ethnicity, and number of extracurricular work hours per week. Students with more extracurricular work hours had significantly lower increase in STEM identity score after course completion. Female-identifying students saw greater learning gains than male-identifying, and though not statistically significant, students identifying as an underrepresented minority reported larger increases in STEM identity score. These findings demonstrate that even short course-based interventions have potential to yield learning gains and improve STEM identity. Online curricula like PARE-Seq can equip STEM instructors to utilize research-driven resources that improve outcomes for all students, but support must be prioritized for students working outside of school.

Adoption of Point-of-Use Chlorination for Household Drinking Water Treatment: A Systematic Review.

BACKGROUND: Centralized chlorination of urban piped water supplies has historically contributed to major reductions in waterborne illness. In locations without effective centralized water treatment, point-of-use (POU) chlorination for households is widely promoted to improve drinking water quality and health. Realizing these health benefits requires correct, consistent, and sustained product use, but real-world evaluations have often observed low levels of use. To our knowledge, no prior reviews exist on adoption of chlorine POU products. OBJECTIVES: Our objectives were to identify which indicators of adoption are most often used in chlorine POU studies, summarize levels of adoption observed, understand how adoption changes over time, and determine how adoption is affected by frequency of contact between participants and study staff. METHODS: We conducted a systematic review of household POU chlorination interventions or programs from 1990 through 2021 that reported a quantitative measure of adoption, were conducted in low- and middle-income countries, included data collection at households, and reported the intervention start date. RESULTS: We identified 36 studies of household drinking water chlorination products that met prespecified eligibility criteria and extracted data from 46 chlorine intervention groups with a variety of chlorine POU products and locations. There was no consensus definition of adoption of household water treatment; the most common indicator was the proportion of household stored water samples with free chlorine residual >0.1 or 0.2mg/L. Among studies that reported either free or total chlorine-confirmed adoption of chlorine POU products, use was highly variable (across all chlorine intervention groups at the last time point measured in each study; range: 1.5%-100%; sample size-weighted median=47%; unweighted median=58%). The median follow-up duration among intervention groups was 3 months. On average, adoption declined over time and was positively associated with frequency of contact between respondents and study staff. DISCUSSION: Although prior research has shown that POU chlorine products improve health when correctly and consistently used, a reliance on individual adoption for effective treatment is unlikely to lead to the widespread public health benefits historically associated with pressurized, centralized treatment of piped water supplies. https://doi.org/10.1289/EHP10839.