Hospital-acquired infection (HAI) results in an enormous burden of excess morbidity, mortality and cost in both adults and children. Monitoring HAI is difficult, especially with limited resources, but it is vital if infection control measures are to be appropriately implemented and assessed. Cross-sectional prevalence surveys, repeated every six months, have been used effectively to monitor HAI in adults, but this technique has not been previously employed in the paediatric population. We performed prevalence surveys of HAI on a single day once every six months for five years, using a standardized questionnaire. Of the 1623 inpatients surveyed, 125 (7.7%) had HAI and 352 (21%) had community-acquired infection. In those with HAI, central-line infections, pneumonia, and wound infections predominated. A hospital stay of greater than seven days was associated with a sixfold increase in the risk of HAI. In addition, admission to a paediatric or neonatal intensive care unit, the presence of a urinary or vascular catheter, the presence of an endotracheal tube, immunosuppression and recent surgery were all associated with a significantly increased risk of HAI. In contrast to other studies, younger children were not at increased risk of HAI; admission to the neonatal unit, rather than age per se, was associated with increased risk. We conclude that repeated prevalence surveys enable simple and cost-effective assessment of HAI, facilitating appropriate infection control interventions. They should be used more widely in the paediatric setting.