Three morphologically similar thermo-acidophilic strains, USBA-GBX-501, USBA-GBX-502 and USBA-GBX-503(T), were isolated from acidic thermal springs at the National Natural Park Los Nevados (Colombia). All isolates were spore-forming, Gram-stain-positive and motile, growing aerobically at 25-55 degrees C (optimum ~45 degrees C) and at pH 1.5-4.5 (optimum pH ~3.0). Phylogenetic analysis of the 16S rRNA gene sequences of these isolates showed an almost identical sequence (99.0 % similarity) and they formed a robust cluster with the closest relative Alicyclobacillus tolerans DSM 16297(T) with a sequence similarity of 99.0 %. Average similarity to other species of the genus Alicyclobacillus was 93.0 % and average similarity to species of the genus Effusibacillus was 90 %. In addition, the level of DNA-DNA hybridization between strain USBA-GBX-503(T) and Alicyclobacillus tolerans DSM 16297(T) was 31.7 %. The genomic DNA G+C content of strain USBA-GBX-503(T) was 44.6 mol%. The only menaquinone was MK-7 (100.0 %). No omega-alicyclic fatty acids were detected in strain USBA-GBX-503(T), and the major cellular fatty acids were C18 : 1omega7c, anteiso-C17 : 0 and iso-C17 : 0. Based on phenotypic and chemotaxonomic characteristics, phylogenetic analysis and DNA-DNA relatedness values, along with low levels of identity at the whole genome level (ANIb and ANIm values of <67.0 and <91.0 %, respectively), it can be concluded that strain USBA-GBX-503(T) represents a novel species of the genus Alicyclobacillus, for which the name Alicyclobacillus montanus sp. nov. is proposed. The type strain is USBA-GBX-503(T) (=CMPUJ UGB U503(T)=CBMAI1927(T)).