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Background: The increasing and rapid spread of metallo-beta-lactamase (MBL) producing Enterobacteriaceae, particularly Escherichia coli and Klebsiella pneumoniae represents an emerging public health threat. However, limited data is available on MBL production in clinical isolates of E. coli and K. pneumoniae from Nepal. We have therefore undertaken this study to ascertain the incidence of MBL production in clinical isolates of E. coli and K. pneumoniae at a tertiary care teaching hospital in central Nepal. Methods: A total of 401 consecutive, non-duplicate isolates of E. coli (n = 216) and K. pneumoniae (n = 185) were recovered from various clinical samples between July and December, 2012. These isolates were screened for the detection of carbapenemase production on the basis of their reduced susceptibility to meropenem or ertapenem by the disc diffusion method. The screened isolates were further phenotypically studied for carbapenemase production by modified Hodge test (MHT). MBL production was detected by performing combined disc test by using imipenem discs with and without ethylenediaminetetraacetic acid (EDTA), which chelates zinc required for MBL activity. Results: Out of 216 E. coli isolates, a total of 41 isolates (18.98%) and out of 185 K. pneumoniae isolates, a total of 39 isolates (21.08%) were suspected to be carbapenemase- producers on the basis of their reduced susceptibility to meropenem or ertapenem. Interestingly, all the initially suspected isolates of E. coli and K. pneumoniae for carbapenemase production were found to be positive in both MHT and combined disc test. However, few weakly positive reactions were observed in MHT. All the MBL producing isolates were multidrug-resistant (MDR). In addition, 75.60% E. coli and 71.79% of K. pneumoniae isolates producing MBL were found to be “pandrug- resistant”. Conclusions: Our findings showed MBL production in a considerable number of E. coli and K. pneumoniae isolates with MDR and pandrug-resistant phenotypes. Combined disc method can provide a sensible choice for phenotypic detection of MBL production in clinical microbiology laboratories as detection of MBL in bacterial isolates is indispensable for establishing the effective antibiotic policies and infection control strategies in the hospital setting. [ABSTRACT FROM AUTHOR]