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Background: Production of metallo-β-lactamase, an enzyme that hydrolyze a variety of β-lactams including carbapenems leaving little therapeutic option is increasing. To manage patients effectively, it is important to know the local prevalence of MBLs in the hospital. Present study was undertaken to determine prevalence of metallo-β-lactamase production along with the clinical profile of Acinetobacter baumannii and Pseudomonas aeruginosa. Methods: Prospective cross sectional study was carried out during December-2010 to November-2011. Relevant demographic and clinical details were collected. Acinetobacter baumannii and Pseudomonas aeruginosa were subjected to antimicrobial susceptibility testing by Kirby-bauer disc diffusion method and mini API system, (bioMerieux-France). Imipenem/meropenem resistant isolates were tested for metallo-β-lactamase production by imipenem-EDTA combined disc test. Results: 81 strains of Acinetobacter baumannii and 28 strains of Pseudomonas aeruginosa were isolated. Most common specimens from which Acinetobacter baumanni and Pseudomonas aeruginosa isolated were from respiratory tract i.e. 47 (58%) and 12 (42.9%) respectively. Majority of the strains of Acinetobacter baumannii 44 (54.3%) and Pseudomonas aeruginosa 17 (60.7%) were isolated from non critical areas. Both organisms showed high prevalence of multidrug resistance with MBLs production of 29.6% in Acinetobacter baumannii and 42.9% in Pseudomonas aeruginosa. Conclusion: Metallo-β-lactamase-mediated carbapenem resistance is a significant threat in hospitalized patients. It should be addressed with rapid detection and stringent infection control measures. [ABSTRACT FROM AUTHOR]