In 2014 to 2017, the prevalence of leaf blight disease was surveyed in various rice fields located in northern Thailand with 40% disease incidence. Rice leaves with leaf blight symptoms (30 to 50% of the lesion area) ranging from yellow, light brown, to slightly reddish spots were collected and used for bacterial isolation, identification, and storage. Pieces of rice leaf were cut, surface sterilized, and then placed on nutrient glucose agar (NGA). After incubation at 28°C for 24 to 48 h, a single colony of each isolate was picked and restreaked on the NGA plate two additional times. Round, yellow, smooth, nonflat, mucoid colonies were observed on the NGA plate. Bacterial isolates that were gram-negative and rod-shaped were selected for molecular identification. The positive bacterial isolates were used for DNA extraction and PCR amplification of the 16S-23S rDNA spacer region of Xanthomonas oryzae pv. oryzae (∼470 bp) (Adachi and Oku 2000). An unexpected PCR product (∼700 bp) using the XOR primer pair was observed. The PCR product was gel purified for DNA sequencing analysis. The sequencing data were subjected to a BLASTn search of the NCBI database. Results showed 694/700 (99%) identity of our suspected sequences to the gyrB gene of Pantoea ananatis (GenBank no. KT203549.1). Additionally, four pairs of primers specific to housekeeping genes of P. ananatis were designed from P. ananatis strain LMG 20103 (GenBank no. CP001875.2), which are gyrB, rpoB, infB, and atpD. The result showed all four primers successfully amplified all four amplicons in 11 bacterial isolates, and all the sequencing data after BLASTn showed 99 to 100% similarity to P. ananatis (GenBank no. KT203549.1 with 666/670 [99%] identity [gyrB], KT735144.1 with 564/564 [100%] identity [rpoB], KT429228.1 with 499/499 [100%] identity [infB], and KT203523.1 with 593/594 [99%] identity [atpD]). Seven bacterial isolates were shown to be positive only for gyrB and rpoB gene amplifications, and the sequencing data indicated that they belonged to P. stewartii (GenBank no. KT203549.1 with 670/670 [100%] identity [gyrB] and KT735140.1 with 564/564 [100%] identity [rpoB]). Moreover, to validate the species identification, biochemical characterization was performed using the API 20E system, and the result showed all 18 bacterial isolates were identified as Pantoea spp. 2 (urease negative and indole positive). The partial sequencing data of gyrB gene and rpoB genes of all bacterial isolates conducted in this study were deposited in the NCBI database with accession numbers MH244161 to MH244200. Two bacterial isolates from each species were used to confirm for the infectivity of these pathogens in rice via the clipping method. The bacterial suspension (10⁸ CFU/ml) in sterile distilled water was inoculated into rice plants. Inoculation of rice plants with sterile distilled water was used as a negative control (five individual plants per group). The yellow, orange to brown stripes were observed at day 7 postinoculation as expected. To fulfill Koch’s postulates, the bacteria were reisolated from symptomatic leaves for DNA extraction and PCR amplification. The sequence alignment of rpoB gene showed 100% identity between the reisolated sample and the original one. This study is the first report of leaf blight in rice caused by P. ananatis and P. stewartii in Thailand. Toward this end, it has become increasingly evident that P. ananatis and P. stewartii are among the most important and newly emerged leaf blight disease causing pathogens in rice.