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Two new species of heliobacteria isolated from cyanobacterial mats of two alkaline sulfidic hot springs are formally described. Strains BR4 and BG29 are assigned to anoxygenic phototrophic bacteria of the familyHeliobacteriaceae, since they possess the unique properties of this taxon: strict anaerobiosis, formation of bacteriochlorophyllg, the lack of extensive intracytoplasmic membranes and chlorosomes, an unusual cell wall structure, and phylogenetic relatedness to the low G+C gram-positive eubacteria. Based on the 16S rDNA sequence similarity, strains BR4 and BG29 are assigned to the genusHeliobacteriumand described as two new species of this genus:Heliobacterium sulfidophilumsp. nov. andHeliobacterium undosumsp. nov. The G+C content of the DNA is 51.3 mol % inHbt. sulfidophilumand 57.2-57.7 mol % inHbt. undosum.The cells ofHbt. sulfidophilumare rods, and the cells ofHbt. undosumare slightly twisted spirilla or short rods. Both new bacteria are motile by peritrichous flagella.Hbt. sulfidophilumproduces endospores. The new bacteria are strict anaerobes growing photoheterotrophically on a limited range of organic compounds. In the dark, they can switch from photosynthesis to the slow fermentation of pyruvate. Biotin is required as a growth factor. Both species are highly tolerant to sulfide (up to 2 mM at pH 7.5) and oxidize it photoheterotrophically to elemental sulfur; photoautotrophic growth was not observed. The temperature optimal for growth ofHbt. sulfidophilumandHbt undosumis 30–35‡C, and the optimal pH is 7–8.