Perennially cold habitats are diminishing as a result of climate change; however, little is known of the diversity or biogeography of microbes that thrive in such environments. Here we use targeted 16S rRNA gene surveys to evaluate the global affinities of cold-dwelling cyanobacteria from lake, stream and ice communities living at the northern limit of High Arctic Canada. Pigment signature analysis by HPLC confirmed the dominance of cyanobacteria in the phototrophic communities of these High Arctic microbial mats, with associated populations of chlorophytes and chromophytes. Microscopic analysis of the cyanobacteria revealed a diverse assemblage of morphospecies grouping into orders Oscillatoriales, Nostocales and Chroococcales. The 16S rRNA gene sequences from six clone libraries grouped into a total of 24 ribotypes, with a diversity in each mat ranging from five ribotypes in ice-based communities to 14 in land-based pond communities. However, no significant differences in composition were observed between these two microbial mat systems. Based on clone-library and phylogenetic analysis, several of the High Arctic ribotypes were found to be >99% similar to Antarctic and alpine sequences, including to taxa previously considered endemic to Antarctica. Among the latter, one High Arctic sequence was found 99.8% similar to Leptolyngbya antarctica sequenced from the Larsemann Hills, Antarctica. More than 68% of all identified ribotypes at each site matched only cyanobacterial sequences from perennially cold terrestrial ecosystems, and were <97.5% similar to sequences from warmer environments. These results imply the global distribution of low-temperature cyanobacterial ecotypes throughout the cold terrestrial biosphere.