Neonatal septicaemia is the leading cause of death in neonates admitted in the neonatal intensive care unit in India. Furthermore this situation is nowadays worsened by the beta lactamase producing bacteria occurring as a causative agent of neonatal septicaemia. These ESBL producing gram negative organisms with increased resistance should be monitored regularly and antibiogram prepared so as to plan effective empiric therapy.  Aim of this study was to determine the bacteriological spectrum and resistance pattern of these gram negative isolates from neonatal intensive care unit. ESBL production among these gram negative organisms was also determined. A study was done in the department of Microbiology of our medical college where 282 neonates admitted in the neonatal intensive care unit between june 2016 to august 2016 with suspicion of septicaemia were included in the study. About 2ml of blood was collected under aseptic condition and put in Bact T Alert PF plus bottles (paediatric bottles) for culture. Gram negative isolates recovered from these septicemic patients were further tested for the presence of ESBL and resistance pattern. Among the 282 suspected patients of neonatal septicaemia, blood culture was positive in 89 patients. Gram negative isolates were seen in 48 cases whereas gram positive isolates were seen in 22 cases. Candida species were isolated from 19 cases of neonatal septicaemia. Most common gram negative organism isolated was Klebsiella species (22.2%). ESBL production was there in 39.6% of the gram negative isolates. Multidrug resistance was more common in ESBL producers than Non ESBL producers. Septicaemia in neonates is increasing rapidly in India.  ESBL  producing gram negative organisms are also increasing with alarming proportion throughout the world. As the gold standard for the diagnosis of septicemia is culture which takes about 48-72 hours so precious time is lost in the diagnosis. Clinician has to rely on the empirical therapy till the result of the culture is obtained. Empiric therapy should be based on the knowledge of prevalence of beta lactamases in that hospital. Bacteriological profile of neonatal septicaemia also varies in different regions of the vast country like India.

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