Background: Cholestasis is a known complication of gram-negative bacterial infection, especially in infants. This syndrome is more frequent in the neonatal period and may account for as much as a third of the cases of neonatal jaundice. Objectives: to determine the prevalence of neonatal cholestasis in neonatal sepsis and to determine the prevalence of Gram negative septicemia in neonatal sepsis. Materials and Methods: This retrospective, observational study was conducted among 267 neonates attending in neonatal ward department of pediatrics at Dr. B.R.A.M. Hospital, Raipur, Chhattisgarh. Results: Only 47 infants out of 267 cases were having Cholestasis jaundice. Overwhelming majority of infection in our study were caused by gram negative organism Klebsiella pneumoniae (78.72%), E.coli (17.02%), Acinobacter (2.13%) and S.aureus (2.1%). Association of Cholestasis with infecting organism in subjects with neonatal sepsis was assessed using Chi square test. No significant association was found to exist between two parameters (p=0.07). Conclusion: It was concluded that majority of infection in our study were caused by gram negative organism Klebsiella pneumonia, E.coli, Acinobacter and S.aureus.