The incidence of hand, foot, and mouth disease (HFMD) is increasing over the years despite current prevention and control policies in Singapore. A retrospective case-control study was conducted among parents whose children attended childcare centres in Singapore to assess the epidemiological risk factors associated with HFMD among children below 7 years old. Parents of 363 children with HFMD (as cases) and 362 children without HFMD (as controls) were enrolled from 22 childcare centres. Data of potential risk factors were collected through a standardised self-administered questionnaire from parents which include demographics and hygiene practices. Multivariate analysis were adjusted for age group, parent's education level, mother's age, HFMD-infected siblings, and preschool admission period. Child's age between 1.5 and 4.9 years, child who had been in childcare for more than 1.9years, having HFMD-infected siblings, two or more children in a family, higher educated parents, parents who had HFMD episode previously, wash toys with soap once every two to three weeks, sanitise toys once every two to three weeks, out-sourced cleaner in childcare centre, no domestic helper at home and more than 22 children in a classroom were independent risk factors of HFMD. These evidence provide crucial implications to guide more effective prevention and control of HFMD in Singapore.