Five strains of filamentous, sulfur-oxidizing bacteria were isolated from sulfur mats of different sulfide springs from various regions of the Northern Caucasus, Russia. A phylogenetic analysis based on 16S rRNA gene sequence comparison showed that all of the isolates are affiliated with the filamentous, colourless, sulfur-oxidizing bacteria of the genus Thiothrix within the Gammaproteobacteria and are closely related to Thiothrix fructosivorans. All strains are capable of growing heterotrophically, lithoautotrophically with thiosulfate or sulfide as the sole energy source and mixotrophically. Strains G1(T), G2, P and K2 are able to fix molecular nitrogen, but strain BL(T) is not. Randomly amplified polymorphic DNA (RAPD)-PCR analysis was used to assess the level of genetic relationships among the Thiothrix isolates. The Nei and Li similarity index revealed high genetic similarity among strains G1(T), G2, P and K2 (above 75 %), indicating that they are closely related. In combination with physiological and morphological data, strains G1(T), G2, P and K2 can be considered as members of the same species. The lowest genetic similarity (approx. 20 %) was reached between strain BL(T) and the other isolated Thiothrix strains. Strains BL(T) and G1(T) shared 35 % DNA-DNA relatedness and showed 51 and 53 % relatedness, respectively, to Thiothrix fructosivorans ATCC 49749. On the basis of this polyphasic analysis, strains G1(T), G2, P and K2 represent a novel species within the genus Thiothrix, for which the name Thiothrix caldifontis sp. nov. is proposed, with strain G1(T) (=DSM 21228(T) =VKM B-2520(T)) as the type strain. In addition, strain BL(T) represents a second novel species, Thiothrix lacustris sp. nov., with strain BL(T) (=DSM 21227(T) =VKM B-2521(T)) as the type strain.
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