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Introduction: Bacteriological profile and antimicrobial susceptibility patterns of urine culture isolates were determined among patients in the Ndjamena General Hospital, a National Reference centre. Methods: A cross-sectional study was carried out from July to November 2014. Six hundred and sixty patients were enrolled, to whom a cytobacteriological examination of urine was prescribed. Urine was collected and cultured. Bacterial identification and antimicrobial susceptibility patterns were performed using Vitek 2 compact automated system. Results: 216 isolates were recovered from patients (age range: 10-90 years). E. coli was the pathogen frequently cultured 128 (59.3%) followed by K. pneumonia 28 (13.0%). Bacteriuria was more present in inpatients (70.4%) compared to outpatients (29.6%). High antibiotic-resistance rate (> 60%) of the total isolates was observed with ampicillin, ciprofloxacin and cephalosporins. Imipeneme (94.9%) displayed satisfactory activity against bacteria isolates. ESBLs phenotype was present in 68/105 (64.7%) of betalactamine resistant isolates. AAC(3)-I and AAC(6’)-I enzymes were found respectively in 16/36 (44.4%) and 20/36 (55.6%) of aminoglycosides resistant isolates. Resistance of isolates to quinolones was mainly due to an association of target modification (gyrA and parC), porin reduction and/or efflux mechanisms and was present in 107/213 (49%) of quinolones resistant isolates. Conclusion: E. coli is the predominant uropathogen isolated in our setting and there are antibiotic-resistant uropathogens among the studied population. Therefore, routine surveillance of bacterial uropathogens to common used antibiotics must be a continuous process so as to provide physicians with up to date information about the local data of antimicrobial resistance.