Neonatal septicemia is still a leading cause of mortality and morbidity in developing countries like India. This prospective study involved 419 clinically suspected cases of neonatal septicemia over a period of one year from April 2013 to March 2014. Of the total 419 cases studied 197 (47%) were blood culture positive. Among the culture positive cases, septicemia was more common in male neonates, seen in 52.79%, compared to 47.2% of female neonates. Of the 197 culture positive cases, early onset septicemia (EOS) was more common, seen in 78.68% compared to late onset septicemia (LOS) in 21.31% of the cases. Gram negative organisms were the predominant causative agents of septicemia in(56.85%), compared to Gram positive organisms in (43.14%) of the cases. Klebsiella pneumoniae was the commonest gram negative isolate in 23.91% of cases, followed by Staphylococcus aureus in 25.37% of cases. All the Gram positive isolates were 100% susceptible to Vancomycin and linezolid followed by Ciprofloxacin and 100% resistant to Penicillin and Tetracycline. Majority of Gram negative isolates were susceptible to Meropenem followed by Ceftazidime while maximum resistance was noted with Ampicillin, Amikacin, Cefotaxime. To conclude, Blood culture is still the “Gold standard” for the diagnosis of septicemia in neonates and should be done in all cases of suspected septicemia prior to starting the antibiotics.