BackgroundOrphaned children are often deprived of quality care, making them more susceptible to diseases due to inadequate hand hygiene. ObjectiveThe present study aimed to assess the prevalence of hand hygiene practices and detect bacterial loads on children's hands before and after hygiene interventions in an orphanage school. Method: The study enrolled all the orphan children registered with the Save Our Souls (SOS) children’s orphanage School in Pakistan. The WHO standard checklist for assessing handwashing practices and swab samples from the hand were collected to evaluate the impact of hand-hygiene practices on bacterial load before and after the intervention. The Quantitative Microbial Risk Assessment (QMRA) model was used to predict the health risk. ResultThe study identified the two most common bacteria: S. aureus and E. coli. Before exposure to the intervention, S. aureus contamination was observed in both groups: intervention (1261 CFU/Hand) and control (1008 CFU/Hand) while E. coli in children's hands were prevalent in the intervention (1042 CFU/Hand) and control (1798 CFU/Hand) groups. The bacterial contamination was significantly reduced after the intervention (S. aureus 166 CFU/ml and E. coli 185 CFU/ml). The higher bacterial ingestion rate was attributed to hand contamination and increased bacteria transfer from hand to mouth. ConclusionThe implementation of the multicomponent hand hygiene intervention showed improvement in accessibility to hand hygiene resources and practices. The findings underscore the need for hygiene interventions in orphanage schools to improve health and educational outcomes.