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This is the first study of the highest elevation cyanobacteria-dominated microbial mat yet described. The desiccated mat was sampled in 2010 from an ephemeral rock pool at 5500 m above sea level in the Cordillera Vilcanota of southern Perú. After being frozen for 6 years at -20 °C in the lab, pieces of the mat were sequenced to fully characterize both the 16 and 18S microbial communities and experiments were conducted to determine if organisms in the mat could revive and become active under the extreme freeze-thaw conditions that these mats experience in the field. Sequencing revealed an unexpectedly diverse, multi-trophic microbial community with 16S OTU richness comparable to similar, seasonally desiccated mats from the Dry Valleys of Antarctica and low elevation sites in the Atacama Desert region. The bacterial community of the mat was dominated by phototrophs in the Cyanobacteria (Nostoc) and the Rhodospirillales, whereas the eukaryotic community was dominated by predators such as bdelloid rotifers (Philodinidae). Microcosm experiments showed that bdelloid rotifers in the mat were able to come out of dormancy and actively forage even under realistic field conditions (diurnal temperature fluctuations of -12 °C at night to + 27 °C during the day), and after being frozen for 6 years. Our results broaden our understanding of the diversity of life in periodically desiccated, high-elevation habitats and demonstrate that extreme freeze-thaw cycles per se are not a major factor limiting the development of at least some members of these unique microbial mat systems.