BACKGROUND: Beta lactams are the most extensively used group of antimicrobials, however growing resistance to these invaluable drugs mediated by extended spectrum beta-lactamase (ESBL) enzymes is a major concern. The frequency of ESBL producing strains among clinical isolates has been steadily increasing over the past few years that has generated a major problem in clinical therapeutics. OBJECTIVES: Our aim was to determine the prevalence of ESBL producing Escherichia coli, study their antibiogram and to evaluate the association between ESBL production and antibiotic resistance in Escherichia coli. SETTINGS AND DESIGN: This prospective study was conducted in the Department of Microbiology, Jawaharlal Nehru Medical College, Aligarh Muslim University, Aligarh. STATISTICAL ANALYSIS USED: Chi-square test was used to analyze the data statistically. Probability values less than 0.05 were considered significant. MATERIALS AND METHODS: Two hundred and eighty six consecutive, non-repeated isolates of Escherichia coli obtained from pus, urine, blood, stool, throat swab, cervical swab, sputum, CSF and conjunctival swab samples received in our bacteriology laboratory were analyzed in this study. These bacterial isolates were identified and tested for antibiotic sensitivity by standard microbiological procedures. Subsequently, they were screened and then phenotypically confirmed for ESBL production by phenotypic confirmatory disk diffusion test (PCDDT). RESULTS : Out of 286 isolates of Escherichia coli screened for ESBL production, 65.03% (n=186) were detected to be positive using either ceftazidime or cefotaxime. In the screen positives, 91.94% (n=171) were phenotypically confirmed ESBL producers by PCDDT method. The overall prevalence of ESBL producing Escherichia coli was 59.79% (n=171/286) with 87.72% obtained from in-patients and 12.28% from out-patients. Majority of ESBL producing Escherichia coli were recovered from stool (73.33%), followed by pus (62.62%), urine (61.80%) and blood (60.61%) samples. The antibiogram revealed no resistance to imipenem, while the highest resistance rate was detected against cefoperazone, amoxicillin/clavulanic acid, ampicillin, cefotaxime, aztreonam, cefoxitin, piperacillin, ceftazidime, and ceftriaxone. CONCLUSIONS: ESBL producing strains of Escherichia coli show extremely wide spectrum of antibiotic resistance including resistance to penicillins, cephalosporins, aminoglycosides, and fluoroquinolones. This leads to significant implications in the management of patients. Advanced drug resistance surveillance and determination of molecular characteristics of ESBL isolates are necessary to formulate antibiotic prescription policies, so as to ensure appropriate and judicious use of the available antimicrobial drugs.