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Cassava (Manihot esculenta Crantz) is one of the most important crops in Vietnam for providing food, starch sources, and raw materials for the production of bio-ethanol and other purposes. Cassava bacterial blight (CBB) disease, caused by Xanthomonas axonopodis pv. manihotis (Xam), is one of the most important factors affecting cassava production in Vietnam. A rapid and sensitive molecular tool is required to support the traditional method, primarily based on biochemical reactions, that was considered less sensitive and time-consuming. In the present study, in order to detect Xam, the Polymerase Chain Reaction (PCR) technique was applied with the two primer pairs, rpoD_17F/rpoD_1005R and XgyrconpcrF1/Xgyrconrpcr1, to amplify partial sequences of rpoD and gyrB genes, respectively. The primers directed the amplification of about 900 bp DNA fragments from CBB-infected leaf and stem tissues. All the amplified DNA sequences were identical in each gene and to the Xam reference strains (accession numbers KP265376 and KP265378 for the rpoD gene, and KP265387 and KP265388 for the gyrB gene) from GenBank. The representative sequences of rpoD and gyrB genes were deposited in GenBank with accession numbers MF774491 and MF774490, respectively. DNA and phylogenetic analysis based on the sequences of rpoD and gyrB genes have confirmed that Xam is the causal agent of CBB in Dong Nai, Vietnam. This is the first confirmed identification of the causal pathogen of CBB disease in Vietnam using molecular tools, and this method is a reliable tool for the detection and identification of other plant bacterial pathogens.