Phylogenetic systematics of the superfamily Cryptoplacoidea (Mollusca: Polyplacophora: Chitonida)

Abstract

Chitons (class Polyplacophora) represent an early-diverging molluscan lineage, with approximately 1,090 currently recognized living species. Their basic body plan, with eight dorsal shell plates surrounded by a fleshy girdle, has remained largely unchanged for hundreds of millions of years. Nonetheless, chitons exhibit remarkable innovations, which include an intricate sensory system embedded within the shell plates (the aesthetes), the unique Schwabe organ, radular teeth reinforced with iron magnetite, and unusually high rates of chromosomal rearrangements. They inhabit marine environments from intertidal shores to the deep sea and play important ecological role as grazers. They possess a wide range of morphological characters, such as shell sculpture, gill arrangement, radular morphology, and valve insertion slits, which form the basis of their taxonomic classification. This thesis integrates morphological taxonomy, molecular systematics, and museomics to advance understanding of chiton diversity and evolution, with a particular focus on the clade Cryptoplacoidea. The first aim was to accelerate marine species discovery by providing high-quality species descriptions that combine morphology and, when possible, molecular data within an integrative taxonomic framework. The second aim was to revise the phylogenetic systematics of Cryptoplacoidea through the implementation of museomic techniques, which required testing multiple approaches. Chapter II presents detailed descriptions of new chiton species based on morphological and molecular evidence. It includes four species descriptions: Placiphorella methanophila Vončina, 2024 (SOSA et al. 2024), Craspedochiton zefranki Vončina, 2025 (SOSA et al. 2025), and two additional species, Notoplax sp. nov. and Placiphorella sp. nov. Chapter III focuses on the North-East Atlantic representatives of the genus Acanthochitona. Results from this chapter confirmed the taxonomic status of Acanthochitona discrepans as valid species. It also provided detailed redescriptions of two closely related taxa, A. crinita and A. fascicularis. Phylogenetic analyses indicated the presence of undescribed species of Acanthochitona in the Mediterranean Sea, and suggested that A. crinita may represent a “wastebasket taxon” comprising multiple unrecognized species. This chapter underscores the importance of comprehensive taxonomic reassessment in clarifying species boundaries even within a well-studied genus. Chapter IV explores the genetic potential of historical chiton specimens for museomic analysis. Optimized, low-contamination extraction protocols and newly developed chiton-specific mitochondrial primers successfully recovered DNA fragments even from specimens over a century old. These methods provide a framework for species identification in global museum collections and enable the incorporation of previously inaccessible material into genetic research. Chapter V applied ultraconserved elements (UCEs) to chiton phylogenomics for the first time, using a chiton-specific probe-set (Chen et al. 2025, Appendix D). The resulting dataset contained thousands of loci recovered from both recent and historical specimens, enabling the reconstruction of the first comprehensive phylogeny of Cryptoplacoidea. The analysis confirmed the monophyly of the superfamily, and revealed that the family Acanthochitonidae is paraphyletic, comprising three distinct families. These results carry major systematic implications, supporting the proposal of a new superfamily and family, as well as the reinstatement of a family and genus. The study also demonstrated that the genus Choneplax is paraphyletic, with its three species distributed across different genera and families. Ancestral-state reconstruction of key morphological traits highlights convergence and morphological plasticity, providing new insights into chiton evolution and classification. In conclusion, this thesis advances the understanding of chiton diversity and evolution by integrating complementary morphological and molecular approaches, thereby laying a strong foundation for future systematic revisions. The findings prompt major taxonomic reassessments, including the revision and clarification of higher-level classifications, and position chitons as a valuable model system for studying phenotypic evolution in morphologically challenging groups. More broadly, they demonstrate the utility of chitons as case studies for addressing fundamental questions about genetic and morphological evolution.