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The phylogenetic diversity of picocyanobacteria in seven alkaline lakes on the Tibetan Plateau was analyzed using the molecular marker 16S-23S rRNA internal transcribed spacer sequence. A total of 1,077 environmental sequences retrieved from the seven lakes were grouped into seven picocyanobacterial clusters, with two clusters newly described here. Each of the lakes was dominated by only one or two clusters, while different lakes could have disparate communities, suggesting low alpha diversity but high beta diversity of picocyanobacteria in these high-altitude freshwater and saline lakes. Several globally distributed clusters were found in these Tibetan lakes, such as subalpine cluster I and the Cyanobium gracile cluster. Although other clusters likely exhibit geographic restriction to the plateau temporally, reflecting endemicity, they can indeed be distributed widely on the plateau. Lakes with similar salinities may have similar genetic populations despite a large geographic distance. Canonical correspondence analysis identified salinity as the only environmental factor that may in part explain the diversity variations among lakes. Mantel tests suggested that the community similarities among lakes are independent of geographic distance. A portion of the picocyanobacterial clusters appear to be restricted to a narrow salinity range, while others are likely adapted to a broad range. A seasonal survey of Lake Namucuo across 3 years did not show season-related variations in diversity, and depth-related population partitioning was observed along a vertical profile of the lake. Our study emphasizes the high dispersive potential of picocyanobacteria and suggests that the regional distribution may result from adaptation to specified environments.