Despite detailed study of selected thermophilic taxa, overall community diversity of bacteria in thermophilic mats remains relatively poorly understood. A sequence-based survey of bacterial communities from several hot spring locations in central Tibet was undertaken. Diversity and frequency of occurrence for 140 unique 16S rRNA gene phylotypes were identified in clone libraries constructed from environmental samples. A lineage-per-time plot revealed that individual locations have evolved to support relatively large numbers of phylogenetically closely related phylotypes. Application of the F ( ST ) statistic and P test to community data was used to demonstrate that phylogenetic divergence between locations was significant, thus emphasizing the status of hot springs as isolated habitats. Among phylotypes, only the Chlorobi were ubiquitous to all mats, other phototrophs (Cyanobacteria and Chloroflexi) occurred in most but not all samples and generally accounted for a large number of recovered phylotypes. Phylogenetic analyses of phototrophic phylotypes revealed support for location-specific lineages. The alpha, beta and gamma proteobacteria were also frequently recovered phyla, suggesting they may be abundant phylotypes in mats, a hitherto unappreciated aspect of thermophilic mat biodiversity. Samples from one location indicated that where phototrophic bacteria were rare or absent due to niche disturbance, the relative frequency of proteobacterial phylotypes increased.