BACKGROUND Non-fermentative gram-negative bacilli are common causes of human infections especially nosocomial infections. These organisms are usually resistant to multiple antimicrobial agents including carbapenems. The study aimed to find out the prevalence of metallo-β-lactamase producing non-fermentative gram-negative bacilli among the samples which yielded growth of bacteria in a tertiary care hospital. METHODS This is a descriptive cross-sectional study conducted in a tertiary care hospital from February 2017 to May 2017. Convenience sampling method was used. Bacterial identification, characterization and antimicrobial susceptibility testing were done by following standard microbiological guidelines. Metallo-β-lactamase production was detected by using combined disk diffusion test and double-disc synergy test. Data were analyzed by using Statistical Package of Social Science software version 16. Point estimate at 95% confidence interval was calculated along with frequency and proportion for binary data. RESULTS Among 628 samples which yielded growth of bacteria, 118 (18.79%) at 95% Confidence Interval (15.74-21.84) were metallo-β-lactamase producing non-fermentative gram-negative bacilli. Among them, 54 (45.76%) were Pseudomonas aeruginosa and 64 (54.24%) were Acinetobacter baumannii. CONCLUSIONS A high prevalence of metallo-β-lactamase production was observed among the nonfermentative gram-negative bacilli than the study done in similar settings. It is mandatory to perform routine monitoring of metallo-β-lactamase producing isolates in clinical laboratories in order to help the clinicians prescribe proper antibiotics.