Introduction: Resistance to higher antimicrobial agent is commonly seen in gram negative bacilli. This issue is a challenging problem to the medical practitioners in addition to it is financial impact on the health care system. Objectives: To document the prevalence of multi drug resistant gram negative bacilli isolated from urine of patients attending the Urology Department of Tertiary care Hospital of western India in year 2008. Results: Out of total 328 isolates, 118 (35.98%) E.coli, 72 (21.95 %) Klebsiella, 64 (19.51%) Pseudomonas aeruginosa, 30 (9.15%) Acinetobacter, 18 (5.49%) Proteus vulgaris, 18 (5.49%) Proteus mirabilis, 6 (1.83%) Providencia rettgerii, 2 (0.61%) Citrobacter freundii. Out of these isolates, 228 (69.51%) were b-lactamase positive, while 100 (30.51%) were β-lactamase negative. Out of 228 β-lactamase positive, 104 (45.61%) were AmpC β-lactamase positive. Conclusions: Stringent protocol such as Antibiotic policy and Hospital infection control program are mandatory to curb these microbes in a tertiary care hospital.