BACKGROUND: Acute respiratory infection (ARI) is a leading cause of morbidity and mortality globally, with 83% of ARI mortality occurring in low-income and middle-income countries (LMICs) before the COVID-19 pandemic. We aimed to estimate the effect of interventions promoting handwashing with soap on ARI in LMICs. METHODS: In our systematic review and meta-analysis, we searched MEDLINE, Embase, Web of Science, Scopus, Cochrane Library, Global Health, and Global Index Medicus for studies of handwashing with soap interventions in LMICs from inception to May 25, 2021. We included randomised and non-randomised controlled studies of interventions conducted in domestic, school, or childcare settings. Interventions promoting hand hygiene methods other than handwashing with soap were excluded, as were interventions in health-care facilities or the workplace. The primary outcome was ARI morbidity arising from any pathogen for participants of any age. Secondary outcomes were lower respiratory infection, upper respiratory infection, influenza confirmed by diagnostic test, COVID-19 confirmed by diagnostic test, and all-cause mortality. We extracted relative risks (RRs), using random-effects meta-analysis to analyse study results, and metaregression to evaluate heterogeneity. We assessed risk of bias in individual studies using an adapted Newcastle-Ottawa scale, and assessed the overall body of evidence using a Grading of Recommendations, Assessment, Development, and Evaluation (GRADE) approach. The study is registered with PROSPERO, CRD42021231414. FINDINGS: 26 studies with 161 659 participants met inclusion criteria, providing 27 comparisons (21 randomised). Interventions promoting handwashing with soap reduced any ARI compared with no handwashing intervention (RR 0·83 [95% CI 0·76-0·90], I2 88%; 27 comparisons). Interventions also reduced lower respiratory infections (0·78 [0·64-0·94], I2 64%; 12 comparisons) and upper respiratory infections (0·74 [0·59-0·93], I2 91%; seven comparisons), but not test-confirmed influenza (0·94 [0·42-2·11], I2 90%; three comparisons), test-confirmed COVID-19 (no comparisons), or all-cause mortality (prevalence ratio 0·95 [95% CI 0·71-1·27]; one comparison). For ARI, no heterogeneity covariates were significant at p<0·1 and the GRADE rating was moderate certainty evidence. INTERPRETATION: Interventions promoting handwashing with soap can reduce ARI in LMICs, and could help to prevent the large burden of respiratory disease. FUNDING: Bill & Melinda Gates Foundation, Reckitt Global Hygiene Institute, and UK FCDO.