Epidermal invagination and muscle rearrangement associated with the segment addition during anamorphosis in a millipede, Niponia nodulosa

Arthropod segmentation involves not only embryonic patterning but also postembryonic segment addition through molting, a process known as anamorphosis. In millipedes (class Diplopoda), which universally undergo anamorphosis, new segmental units also known as “rings” are added anterior to the telson, yet the morphogenetic mechanisms underlying this process remain poorly understood. Here, we investigate ring addition during anamorphosis in the millipede Niponia nodulosa (Polydesmida, Cryptodesmidae), focusing on epidermal and muscular organization. Using X-ray micro-computed tomography and microscopic histological analysis, we reconstructed the sequence of tissue changes preceding molting. Our results show that a local invagination of the epidermis between the telson and the penultimate ring establishes a ring primordium, whose subsequent development gives rise to a new ring. In stadia where multiple rings are added with one molt, a corresponding number of ring primordia appeared. In addition, the muscle originally linking the penultimate ring to the telson detaches and reattaches to the newly added ring, while additional muscles differentiate within the new ring to form novel connections with the telson. These coordinated changes occur rapidly before molting, enabling ring addition within the restricted posterior body region. We provided a detailed three-dimensional description of the morphological changes accompanying ring addition. Epidermal morphogenesis and muscular rearrangement are thought to enable the addition of new rings while maintaining the functional integrity of the posterior body, including defecation. This study establishes a histological and morphological basis for understanding segmentation during postembryonic development in arthropods and paves the way for future developmental investigations.