Horizontal transmission of hepatitis B virus (HBV) is responsible for about half of the hepatitis B surface antigen (HBsAg) carriers in Taiwan. To investigate the routes and risk factors of horizontal HBV transmission, 131 HBsAg carrier children of HBsAg seronegative mothers were compared with 98 age-matched children seronegative for all hepatitis B markers. Possible factors influencing transmission of HBV, including family size, age at attending kindergarten, frequency of intramuscular injection during infancy and at the toddler stage, contact with HBsAg carriers, and HBV status of parents, siblings and caretakers were studied by questionnaire survey. Intramuscular injection frequency during infancy and at the toddler stage and prevalence of HBsAg positivity among siblings were significantly higher in carrier children than in the control group. Other factors studied were not different in these two groups. The results indicated that unsterilized, multiple intramuscular injections and intrafamilial spread among siblings might be the major routes of horizontal HBV transmission in children in Taiwan.