Metallo-β-lactamases (MBLs) hydrolyze all classes of β-lactams except monobactams and are not inhibited by classic serine β-lactamase inhibitors. Gram-negative pathogens isolated from patient infections were collected from 202 medical centers in 40 countries as part of a global surveillance study in 2012-2014. Carbapenem non-susceptible Enterobacteriaceae and Pseudomonas aeruginosa were characterized for bla genes encoding VIM, IMP, NDM, SPM, and GIM variants using PCR and sequencing. 471 MBL-positive isolates included the following species (number): P. aeruginosa (308), Klebsiella spp. (85), Enterobacter spp. (39), Proteeae (16), Citrobacter freundii (12), Escherichia coli (6), and Serratia marcescens (5) and were submitted by sites from 34 countries. Of these, 69.6% were collected in 9 countries (number): Russia (72), Greece (61), the Philippines (54), Venezuela (29), and 20-25 isolates each from Kuwait, Nigeria, Romania, South Africa, and Thailand. Thirty-two different MBL variants were detected (14 VIM, 14 IMP, 4 NDM). Seven novel MBL variants were encountered in the study, each differing from a previously reported variant by one amino acid substitution: VIM-42 (VIM-1 [V223I]), VIM-43 (VIM-4 [A24V]), VIM-44 (VIM-2 [K257N]), VIM-45 (VIM-2 [T35I]), IMP-48 (IMP-14 [I69T]), IMP-49 (IMP-18 [V49F]) and NDM-16 (NDM-1 [R264H]). The in vitro activities of all tested antibiotics against MBL-positive Enterobacteriaceae were significantly reduced with the exception of aztreonam-avibactam (MIC90 0.5-1 μg/mL), whereas colistin was the most effective agent against MBL-positive P. aeruginosa (>97% susceptible). Although the global percentage of isolates encoding MBLs remains relatively low, their detection in twelve species, 34 countries, and all regions participating in this surveillance study is concerning.