The hypothesis that catheter-related sepsis (CRS) may be preceded by contamination of the catheter hub was tested in neonates with central venous catheters. Cultures of the catheter hub were obtained three times per week. One hundred thirteen catheters were placed in 88 patients. Of 35 episodes of sepsis, 28 were catheter-related, for a catheter sepsis rate of 1.03/100 catheter-days. CRS occurred in 26 (23%) of 113 catheters. In 10 of 28 episodes, the infecting microorganism was cultured from the hub before its culture from blood obtained at the time of clinical sepsis. In an additional 5 cases, a culture of the catheter hub at the time of clinical sepsis yielded the same isolate as the blood culture. Thus, 54% of episodes of CRS were preceded by or coincided with contamination of the hub. The catheter hub may be a major portal of entry for microorganisms causing sepsis in a neonatal intensive care unit.