Neonatal sepsis still remains a major cause of morbidity and mortality in neonatal intensive care unit (NICU). Recently, soluble CD14 subtype (sDC14-ST) also named presepsin, was proposed as an effective biomarker for diagnosing, monitoring, and assessing the risk of neonatal sepsis and septic shock. The aim of this study was to investigate the diagnostic accuracy of sCD14-ST presepsin in diagnosing neonatal bacterial sepsis and in discriminating non-bacterial systemic inflammatory response syndrome (SIRS) from bacterial sepsis. This study involved 65 critically ill full-term and preterm newborns admitted to the neonatal intensive care unit (NICU), divided into three groups: 25 newborns with bacterial neonatal sepsis (group A); 15 newborns with a diagnosis of non-bacterial SIRS and with no localizing source of bacterial infection (group B); and 25 babies with no clinical or bacteriological signs of systemic or local infection receiving routine NICU care, most of them treated with phototherapy for neonatal jaundice (group C). A total of 102 whole blood samples were collected, 40 in group A, 30 in group B and 32 in group C. In 10 babies included in group A, sCD14-ST presepsin was also measured in an additional second blood sample collected 3 days after the start of antibiotic treatment. sCD14-ST presepsin was measured by a commercially available chemiluminescent enzyme immunoassay (CLEIA) optimized on an automated immunoassay analyzer. Statistical analysis was performed by means of MedCalc® statistical package; receiver operating characteristic (ROC) analysis was computed, and the area under the ROC curve (AUC) was used to evaluate the ability of sCD14-ST to discriminate neonatal bacterial sepsis from non-bacterial SIRS. Blood sCD14-ST presepsin levels were found significantly higher in bacterial sepsis when compared with controls (p<0.0001); similarly, they were higher in non-bacterial SIRS when compared with controls (p<0.0001). However, no statistically significant difference was found between bacterial sepsis and non-bacterial SIRS (p=0.730). In our population, CRP and sCD14-ST did not correlate with each other. ROC analysis revealed that sCD14-ST presepsin has an area under the curve (AUC) of 0.995 (95% C.I.: 0.941-1.00) greater than that of CRP (0.827; 95% C.I.: 0.72-0.906). Similarly, in the group of babies with non-infectious SIRS, sCD14-ST AUC was greater than CRP AUC (0.979; 95% C.I.: 0.906-0.999 versus 0.771; 95% C.I.: 0.647-0.868). In controls, preliminary reference intervals for sCD14-ST ranged 223.4-599.7 ng/L, being significantly different from those previously published elsewhere. In conclusion, sCD14-ST presepsin could be introduced in clinical practice as a diagnostic tool for improving the management of neonatal sepsis and non-bacterial SIRS.