Three New Freshwater Cochliopodium Species (Himatismenida, Amoebozoa) from the Southeastern United States.

Cochliopodium is a lens-shaped genus of Amoebozoa characterized by a flexible layer of microscopic dorsal scales. Recent taxonomic and molecular studies reported cryptic diversity in this group and suggested that the often-used scale morphology is not a reliable character for species delineation in the genus. Here, we described three freshwater Cochliopodium spp. from the southeastern United States based on morphological, immunocytochemistry (ICC), and molecular data. A maximum-likelihood phylogenetic analysis and pairwise comparison of COI sequences of Cochliopodium species showed that each of these monoclonal cultures were genetically distinct from each other and any described species with molecular data. Two of the new isolates, "crystal UK-YT2" (Cochliopodium crystalli n. sp.) and "crystal-like UK-YT3" (C. jaguari n. sp.), formed a clade with C. larifeili, which all share a prominent microtubule organizing center (MTOC) and have cubical-shaped crystals. The "Marrs Spring UK-YT4" isolate, C. marrii n. sp., was 100% identical to "Cochliopodium sp. SG-2014 ." These sequences formed a clade with C. actinophorum and C. arabianum. While the new isolates can be separated morphologically, most of the taxonomic features used in the group show plasticity; therefore, Cochliopodium species can only be reliably identified with the help of molecular data.