Taxonomic studies were performed on seven strains isolated from microbial mats in Antarctic lakes of the McMurdo Dry Valleys. Phylogenetic analysis based on 16S rRNA gene sequences indicated that these strains are related to the genus Flavobacterium; sequence similarity values with their nearest phylogenetic neighbours ranged from 97?0 to 98?7 %. Results of DNA-DNA hybridization and comparison of repetitive extragenic palindromic DNA-PCR fingerprinting patterns revealed that these strains are members of two distinct species. Genotypic results, together with phenotypic characteristics, allowed these species to be differentiated from related recognized Flavobacterium species. The isolates are psychrophilic, Gram-negative, chemoheterotrophic, rod-shaped cells. Their whole-cell fatty acid profiles are similar and include C 15 : 0 , C 15 : 0 anteiso, C 15 : 0 iso, C 15 : 1 v6c, C 16 : 0 iso, C 16 : 0 iso 3-OH, C 16 : 1 iso and summed feature 3 (which comprises C 15 : 0 iso 2-OH and/or C 16 : 1 v7c) as the major fatty acid components. On the basis of these results, two novel species are proposed: Flavobacterium fryxellicola sp. nov., consisting of three strains with LMG 22022 T (=CIP 108325 T ) as the type strain; and Flavobacterium psychrolimnae sp. nov., consisting of four strains with LMG 22018 T (=CIP 108326 T ) as the type strain. DNA G+C contents of F. fryxellicola and F. psychrolimnae are 35?2-35?9 and 33?8-34?5 mol%, respectively. Members of the phylum Bacteroidetes are abundant in freshwater and marine ecosystems and these bacteria may have a specialized role in the uptake and degradation of organic matter in cold, aquatic environments (Kirchman, 2002). Several novel species, added to the genus Flavobacterium since 1996, have originated from Antarctic habitats (Van Trappen et al., 2003, 2004; and references cited therein).During the MICROMAT project (November 1998-February 2001, 746 bacterial strains were isolated under heterotrophic conditions from microbial mat samples that were collected from ten Antarctic lakes (Van Trappen et al., 2002). Numerical analysis of the fatty acid composition of the isolates and 16S rRNA gene sequence analysis, performed on representative strains, showed that the diversity of heterotrophic bacteria in microbial mats from Antarctic lakes is very high. Moreover, many fatty acid clusters were shown to contain multiple taxa when tested by repetitive extragenic palindromic DNA-PCR (rep-PCR) fingerprinting, a technique used to investigate the genomic diversity of each fatty acid cluster in more detail (Van Trappen et al., 2003, 2004. Several strains belonging to fatty acid clusters 5, 6 and 10 have previously been described as novel Flavobacterium species: Flavobacterium gelidilacus, Flavobacterium degerlachei, Flavobacterium frigoris and Flavobacterium micromati (Van Trappen et al., 2003, 2004.In the present work, the taxonomic relationships of 22 strains from fatty acid cluster 5 (as delineated by Van Trappen et al., 2002) were studied. A group of 11 of these strains was identif...