Th e present study was conducted to determine the phenotypic and genotypic characterization of thermophilic bacteria isolated from Pasinler hot spring, Erzurum, Turkey. Fatty acid profi les, BOX PCR fi ngerprints, and 16S rDNA sequence data were used for the phenotypic and genotypic characterization of thermophilic bacteria. Totally 9 diff erent bacterial strains were selected based on morphological, physiological, and biochemical tests. Th ese strains were characterized by molecular tests including fatty acid and BOX profi les, and 16S rDNA sequence. Th e data of fatty acid analysis showed the presence of 14 diff erent fatty acids in the 9 bacterial strains examined. Additionally, 5 of these fatty acids, 15:0 iso, 15:0 anteiso, 16:0 iso, 17:0 iso, and 17:0 anteiso fatty acids, were found in all isolates. Based on fatty acid profi les, it was determined that the bacterial strains were classifi ed into 2 phenotypic groups, and these data were confi rmed by BOX PCR genomic fi ngerprint profi les and 16S rDNA sequence analysis. Th e fi rst group, identifi ed as Bacillus licheniformis, was represented by 4 strains, and the second group, identifi ed as Aeribacillus pallidus, was represented by 5 strains.