Cassava (Manihot esculenta Crantz) is one of the most important crops in Vietnam, providing food, starch sources, and raw materials for production of bio-fuel. Cassava bacterial blight disease caused by Xanthonomas axonopodis pv. manihotis (Xam) has affected cassava production in Vietnam. In the present study, PCR technique applied the primers rpoD_17F/rpoD_1005R and XgyrconpcrF1/Xgyrconrpcr1 specific for rpoD and gyrB genes, respectively with genomic DNAs extracted directly from the symptomatic leaves without isolation and pure culture of the causal bacterium. The two primer pairs amplified amplicons of about 900 bp from the 30 symptomatic leaf samples collected from Krong Pac, M’Drak, and Ea Kar districts of Dak Lak province. The amplified DNA sequences were identical in each gene and shared 100% similarity to that of Xam. Phylogenetic trees were constructed with the sequences of rpoD and gyrB genes of different Xanthomonas species from GenBank. The results indicated that the bacterial species analysed by either rpoD or gyrB as the target genes showed 100% identity with that of Xanthomonas axonopodis pv. manihotis causing cassava bacterial blight disease in Dong Nai, Vietnam (Accession numbers: MF774491 and MF774490, respectively) and in some other countries. DNA and phylogenetic analysis based on the sequences of rpoD and gyrB genes have confirmed that X. axonopodis pv. manihotis is the causal agent of cassava bacterial blight in Krong Pak, M’Drak, and Ea Kar districts of Dak Lak province, Vietnam. The protocol in this study is rapid and sensitive for detection and identification of X. axonopodis pv. manihotis without isolation and pure culture of the causal bacterium.