Hepatitis B is highly endemic in Papua New Guinea (PNG). Vaccination at birth is a key mother-to-child transmission prevention strategy. Despite recommendations for newborns to be vaccinated within 24 hours of delivery, a 2005 survey showed 23% coverage among children born in health facilities. Our study examined hepatitis B birth-dose coverage and knowledge, practices and barriers to vaccine delivery in five major PNG hospitals. Data on births and vaccines administered were sourced from the National Department of Health (NDoH) and directly from the five hospitals. A maternity unit audit and staff survey were undertaken. Across the five hospitals, the hospital-level data of hepatitis B birth-dose coverage was 79% (range: 40-96%) compared to 19% from national data (range: 0-106%). Despite hospitals having adequate vaccine supply, access to appropriately stored vaccine in maternity units was compromised with only one unit having a vaccine-specific temperature-monitored refrigerator. In interviews of 25 staff, incorrect reasons given for delaying vaccination were prematurity (60%), low birthweight (48%) and difficult birth (36%). This study found encouraging birth-dose coverage rates in five major hospitals but 20% of babies still missed receiving the recommended vaccine. The NDoH Immunization Unit will use the results of this study to inform strategies to improve hepatitis B birth-dose coverage in hospitals.