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Collembola, commonly known as springtails, are important detritivores, abundant in leaf litter and soil globally. Springtails are wingless hexapods with many North American species having wide distributions ranging from as far as Alaska to Mexico. Here, we analyze the occurrence and intraspecific diversity of springtails with a globular body shape (Symphypleona and Neelipleona), in southern high Appalachia, a significant biodiversity hotspot. The peaks of high Appalachia represent ‘sky islands’ due to their physical isolation, and they host numerous endemic species in other taxa. We surveyed globular Collembola through COI metabarcoding, assessing geographic and genetic diversity across localities and species. Intraspecific diversity in globular Collembola was extremely high, suggesting that considerable cryptic speciation has occurred. While we were able to associate morphospecies with described species in most of the major families in the region (Dicyrtomidae, Katiannidae, Sminthuridae, and Sminthurididae), other families (Neelidae, and Arrhopalitidae) are in more pressing need of taxonomic revision before species identities can be confirmed. Due to poor representation in databases, and high intraspecific variability, no identifications were accomplished through comparison with available DNA barcodes.