Effects of water quality, sanitation, handwashing and nutritional interventions on enteric pathogen seroconversion in rural Kenya: a cluster randomised trial

Background: We evaluated whether water, sanitation, and handwashing (WASH) and nutrition interventions reduced enteric pathogen seroprevalence during the first two years of life among children in rural Kenya. Methods: We analysed the 2x2 factorial subset of the WASH Benefits Kenya cluster-randomised trial (control, WASH, nutrition, and WASH plus nutrition). Dried blood spots were collected from all children in the birth cohort after two years of intervention and at 6 and 12 months within a longitudinal substudy. Immunoglobulin G (IgG) responses were measured using a multiplex bead assay that included 12 viral, bacterial, protozoan, and helminth pathogens. The primary outcome was IgG seroprevalence at 2 years. We estimated intention-to-treat differences, comparing WASH with no WASH and nutrition with no nutrition across the factorial design. The trial is registered with ClinicalTrials.gov ( NCT01704105 ). Findings: The primary analysis included 3030 children in 384 of 391 clusters (median age 23 months). Seroprevalence increased rapidly with age and exceeded 90% for norovirus, Campylobacter, and LT-ETEC by the final visit, with several pathogens at more moderate prevalences (24% of children were seropositive to typhoidal S. enterica HlyE). Differences between intervention groups were small: lower T. solium (−3.3 percentage points [95% CI −6.3 to −0.2]) and higher E. histolytica (3.2 [0.1 to 6.3]) with WASH versus control, and lower V. cholerae (−1.1 [−1.8 to −0.3]) and Shigella/EIEC (−2.7 [−5.5 to 0.0]) with nutrition versus no nutrition. No comparison met the 5% Benjamini-Hochberg false discovery rate threshold. Secondary analyses and age-seroprevalence curves suggested modest reductions in seroprevalence for some pathogens but an increase in norovirus at approximately 12 months when intervention adherence was highest. Interpretation: At the levels of adherence achieved, household-level WASH and nutrition interventions were insufficient to reduce widespread early-life enteric pathogen exposure in this high-transmission setting, supporting development of complementary vaccine and transmission-control strategies. Funding: National Institutes of Health (R01AI166671, R01AI106878, R01AI134814). The original WASH Benefits trial was funded by the Gates Foundation (OPPGD759).