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The Crystal-Wonder Cave System developed in the Western Escarpment of the southern Cumberland Plateau in the Interior Low Plateau karst region of south-central Tennessee, USA is a global hotspot of cave-limited biodiversity. We combined historical literature, museum accessions, and database occurrences with new observations from bio-inventory efforts conducted between 2005 and 2022 to compile an updated list of troglobiotic and stygobiotic biodiversity for the Crystal-Wonder Cave System. The list of cave-limited fauna includes 31 species (23 troglobionts and 8 stygobionts) with 28 and 18 species documented from the Crystal and Wonder caves, respectively, which represents five phyla, ten classes, nineteen orders, and twenty-six families (six arachnids, three springtails, two diplurans, three millipedes, six insects, three terrestrial snails, one flatworm, five crustaceans, and two vertebrates, respectively). The Crystal-Wonder Cave System is the type locality for six species—Anillinus longiceps, Pseudanophthalmus humeralis, P. intermedius, Ptomaphagus hatchi, Tolus appalachius, and Chitrella archeri. The carabid beetle Anillinus longiceps is endemic to the Crystal-Wonder Cave System. Sixteen species are of conservation concern, including twelve taxa with NatureServe conservation ranks of G1–G3. The exceptional diversity of the Crystal-Wonder Cave System has been associated with several factors, including a high dispersal potential of cave fauna associated with expansive karst exposures along the Western Escarpment of the southern Cumberland Plateau, a high surface productivity, and a favorable climate throughout the Pleistocene.