Objectives: The Xpert Carba-R Assay is a diagnostic test designed for the rapid detectionand differentiation of the blaKPC, blaNDM, blaVIM, blaOXA-48, and blaIMP-1 genes. We verifiedthe performance of Xpert Carba-R Assay for identification of carbapenemase genein the clinical microbiology laboratory. Methods: The analytical limit of detection was determined with two suspensions ofcarbapenemase-producing Enterobacteriaceae (CPE) isolates (KPC and NDM). A totalof 52 specimens were evaluated: 21 bacterial isolates from clinical specimens, 21 rectalswabs, and 10 contrived stool specimens. Results: In bacterial isolates, concordant results between the Xpert Carba-R Assayand PCR were found in 20 of 21; 8 KPC, 8 NDM, 1 IMP, and 2 multiple carbapenamasegenes (KPC/NDM, NDM/OXA) were detected both by Xpert Carba-R Assay and PCR.In one GES-positive isolate, Xpert Carba-R Assay showed a negative result as expected.One VIM-positive isolate tested negative by Xpert Carba-R Assay. Complete concordancewas seen in rectal swab specimens: 4 specimens with KPC and 17 specimenswith negative results both by Xpert Carba-R Assay and surveillance culture. Among the10 contrived stool specimens, Xpert Carba-R Assay detected carbapenemase genes in9 specimens as expected according to the CPE strains spiked into the contrived stool; 2KPC, 4 NDM, 1 IMP, and 2 multiple carabapenamase genes (NDM/KPC, NDM/OXA).One VIM-positive specimen tested negative by Xpert Carba-R Assay. Conclusion In conclusion, the Xpert Carba-R Assay can be used to identify carbapenemasegene in bacterial isolates cultured from clinical specimens and detect CPE carrierusing rectal swab in clinical laboratories.