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Four strains of obligately anaerobic spirochetes were isolated from marine intertidal mud. These strains grew to yields of 2 X lo8 to 6 x 10' cells per ml in chemically defined media containing glucose, amino acids or NH4Cl, sulfide, vitamins, and inorganic salts. These spirochetes required relatively high levels of Na' for growth and utilized carbohydrates but not amino acids as growth substrates. Glucose was fermented to COz, H2, ethanol, and acetate. These four strains also formed the branched-chain fatty acids isovalerate , 2-methylbutyrate, and isobutyrate as fermentation products of L-leucine, L-isoleucine, and L-valine, respectively. The adenosine 5'-triphosphate generated via fermentation of these amino acids was expended by the spirochetes to prolong survival during periods of growth substrate starvation. The deoxyribonucleic acids of the four strains contained 63.6 to 65.6 mol% guanine plus cytosine. We concluded that these four strains are representatives of a new species, for which we propose the name Spirochaeta isovalerica; the type strain of this species is strain MA-2 (= DSM 2461). Spirochetes are frequently observed in sam- ples of anoxic mud collected from freshwater and saltwater environments. The obligately and facultatively anaerobic spirochetes present in salt marsh habitats have been enumerated by using rifampin as a selective agent, and these organisms have been found to be present at densities ranging from lo4 to lo6 colony-forming units per g (wet weight) of sediment (19). Al- though significant numbers of spirochetes are present in marine sediments, few marine species have been described. We isolated 14 strains of spirochetes from intertidal muds collected in Pacific and Atlantic coastal regions and exam- ined these isolates for selected ultrastructural and metabolic properties. In the course of these studies we identified four strains of obligately anaerobic spirochetes that formed the four- and five-carbon branched-chain fatty acids isovaler- ate, 2-methylbutyrate, and isobutyrate as minor fermentation products. These fermentation products are not formed by the other marine spirochetes that have been described and have not been reported for any of the recognized species of free-living terrestrial spirochetes (4). In previous papers we have described the results of studies carried out to elucidate the significance of branched-chain fatty acid forma- tion by one of the four spirochete strains (strain MA-2T (type strain)) (9-11). We determined that

Spirochaeta isovalerica sp. nov., a Marine Anaerobe That Forms Branched-Chain Fatty Acids as Fermentation Products