Suctioning of newborns immediately after birth, as part of delivery room resuscitation, is only recommended if the airway is obstructed. The aim of this study was to describe the use of suctioning during newborn resuscitation among survivors versus those who died within 3 days and potential suction-related heart rate responses and associations to newborn characteristics. This was a retrospective observational study from July 2013 to July 2016 in a referral hospital in rural Tanzania. Research assistants observed and documented all deliveries, newborn resuscitations were video-recorded, and newborn heart rates were captured with a dry-electrode electrocardiogram. Liveborn infants ≥34 weeks gestation who received ventilation and with complete datasets were eligible. All 30 newborns who died were included, and a total of 46 survivors were selected as controls. Videos were annotated and heart rate patterns were observed before and after the suction events. Suctioning was performed more frequently than recommended. No differences were found in suctioning characteristics between newborns who died versus those who survived. In 13% of suction events, a significant heart rate change (i.e., arrhythmia or brief/sustained >15% fall in heart rate) was observed in relation to suctioning. This represents a potential additional harm to already depressed newborns undergoing resuscitation.