Spiralian phylogeny informs the evolution of microscopic lineages.

Despite rapid advances in the study of metazoan evolutionary history [1], phylogenomic analyses have so far neglected a number of microscopic lineages that possess a unique combination of characters and are thus informative for our understanding of morphological evolution. Chief among these lineages are the recently described animal groups Micrognathozoa and Loricifera, as well as the two interstitial "Problematica" Diurodrilus and Lobatocerebrum [2]. These genera show a certain resemblance to Annelida in their cuticle and gut [3, 4]; however, both lack primary annelid characters such as segmentation and chaetae [5]. Moreover, they show unique features such as an inverted body-wall musculature or a novel pharyngeal organ. This and their ciliated epidermis have led some to propose relationships with other microscopic spiralians, namely Platyhelminthes, Gastrotricha, and in the case of Diurodrilus, with Micrognathozoa [6, 7]-lineages that are grouped by some analyses into "Platyzoa," a clade whose status remains uncertain [1, 8-11]. Here, we assess the interrelationships among the meiofaunal and macrofaunal members of Spiralia using 402 orthologs mined from genome and transcriptome assemblies of 90 taxa. Lobatocerebrum and Diurodrilus are found to be deeply nested members of Annelida, and unequivocal support is found for Micrognathozoa as the sister group of Rotifera. Analyses using site-heterogeneous substitution models further recover a lophophorate clade and position Loricifera + Priapulida as sister group to the remaining Ecdysozoa. Finally, with several meiofaunal lineages branching off early in the diversification of Spiralia, the emerging concept of a microscopic, acoelomate, direct-developing ancestor of Spiralia is reviewed.

Comparative myoanatomy of Echinoderes (Kinorhyncha): a comprehensive investigation by CLSM and 3D reconstruction.

INTRODUCTION: Kinorhyncha is a clade of marine invertebrate meiofauna. Their body plan includes a retractable introvert bearing rings of cuticular spines, and a limbless trunk with distinct segmentation of nervous, muscular and epidermal organ systems. As derived members within the basal branch of Ecdysozoa, kinorhynchs may provide an important example of convergence on the evolution of segmentation within one of three bilaterian superclades. We describe the myoanatomy of Echinoderes, the most specious kinorhynch genus, and build upon historical studies of kinorhynch ultrastructure and gross morphology. This is the first multi-species comparison of a complete organ system by confocal microscopy and three-dimensional reconstruction within Kinorhyncha. RESULTS: Myoanatomy of adult Echinoderes is composed of the following: Head with two mouth cone circular muscles, nine pairs of oral style muscles, ten introvert retractors, one introvert circular muscle, and fourteen introvert circular muscle retractors; Neck with one circular muscle; Trunk showing distinct pairs of ventral and dorsal muscles within segments 1-10, dorsoventral muscles within segments 3-10, diagonal muscles within segments 1-8, longitudinal fibers spanning segments 1-9, three pairs of terminal spine muscles, and one pair of male penile spine muscles; Gut showing a pharynx with ten alternating rings of radial and circular muscle fibers enclosed in a complex sheath of protractors and retractors, an orthogonal grid of longitudinal and circular fibers surrounding the intestine, and paired hindgut dilators. CONCLUSIONS: Myoanatomy is highly conserved between species of Echinoderes. Interspecific variation is observed in the arrangement and number of introvert fibers and the composition of pharyngeal muscles. Segmented trunk musculature facilitates the movements of articulated cuticular plates along the anterior-posterior axis. Intersegmental muscle fibers assist with dorsoventral and lateral trunk movements. Protractors, retractors and circular muscles coordinate eversion and retraction of the introvert and mouth cone, and relocation of the pharynx during locomotion and feeding behaviors. Pairs of posterior fibers suggest independent movements of terminal spines, and male penile spines. Within Scalidophora, myoanatomy is more similar between Kinorhyncha and Loricifera, than either group is to Priapulida. Kinorhynch myoanatomy may reflect a convergent transition from vermiform to segmented body plans during the early radiation of Ecdysozoa.

A complete three-dimensional reconstruction of the myoanatomy of Loricifera: comparative morphology of an adult and a Higgins larva stage.

INTRODUCTION: Loricifera is a group of small, marine animals, with undetermined phylogenetic relationships within Ecdysozoa (molting protostome animals). Despite their well-known external morphology, data on the internal anatomy of loriciferans are still incomplete. Aiming to increase the knowledge of this enigmatic phylum, we reconstruct for the first time the three-dimensional myoanatomy of loriciferans. Adult Nanaloricus sp. and the Higgins larva of Armorloricus elegans were investigated with cytochemical labeling techniques and CLSM. We discuss our findings with reference to other loriciferan species and recently established phylogenies. RESULTS: The somatic musculature of both adult and larval stages is very complex and includes several muscles arranged in three orientations: circular, transverse and longitudinal. In adult Nanaloricus sp., the introvert is characterized by a net-like muscular arrangement, which is composed of five thin circular fibers crossed by several (up to 30) thin longitudinal fibers with bifurcated anterior ends. Two sets of muscles surround the pre-pharyngeal armature: 6 buccal tube retractors arranged 3 × 2 in a conical shaped structure, and 8 mouth cone retractors. Additionally, a thick, circular muscle marks the neck region and a putative anal sphincter is the posteriormost myoanatomical feature. In the Higgins larva of A. elegans, two circular muscles are distinguished anteriorly in the introvert: a dorsal semicircular fiber and a thin ring muscle. The posteriormost region of the body is characterized by an anal sphincter and a triangular muscle. CONCLUSIONS: Based on the currently available knowledge, the myoanatomical bodyplan of adult loriciferans includes: (i) 8 mouth cone retractors, (ii) a pharynx bulb composed of transversal fibers arranged radially, (iii) circular muscles of the head and neck, (iv) internal muscles of the spinoscalids, (v) longitudinal muscles spanning all body regions, and (vi) transverse (circular) muscles in the abdomen. Concerning the Higgins larva, the muscle subsets assigned to its myoanatomical ground pattern are the (i) longitudinal retractors of the mouth cone, introvert, and abdomen, (ii) abdominal transverse muscles, and (iii) a pharynx bulb composed of transverse, radial fibers. In a comparison with phyla traditionally regarded as phylogenetically close, our data show that the overall myoanatomy of Loricifera is more similar to Kinorhyncha and Nematomorpha than to Priapulida. However, the head musculature of all these groups is very similar, which supports homology of their introverts and head morphology.

Comparative myoanatomy of cycliophoran life cycle stages.

The metazoan phylum Cycliophora includes small cryptic epibionts that live attached to the mouthparts of clawed lobsters. The life cycle is complex, with alternating sexual and asexual generations, and involves several sessile and free-living stages. So far, the morphological and genetic characterization of cycliophorans has been unable to clarify the phylogenetic position of the phylum. In this study, we add new details on the muscular anatomy of the feeding stage, the attached Prometheus larva, the dwarf male, and the female of one of the two hitherto described species, Symbion pandora. The musculature of the feeding stage is composed of myofibers that run longitudinally in the buccal funnel (two fibers) and in the trunk (variable number of fibers). The mouth opening is lined by a myoepithelial ring musculature. A complex myoepithelial sphincter is situated proximal to the anus. In the attached Prometheus larva, three longitudinal sets of myofilaments run dorsally, laterally, and ventrally along the entire anterior-posterior body axis. The muscular architecture of the dwarf male is complex, especially close to the penis, in the posterior part of the body. An X-shaped muscle structure is found on the dorsal side, whereas on the ventral side, longitudinal muscles and a V-shaped muscle structure are present. These muscles are complemented by additional dorsoventral muscles. The mesodermal muscle fibers attach to the cuticle via the epidermis in all life cycle stages studied herein. The musculature of the female is similar to that of the Pandora larva of Symbion americanus and includes dorsoventral muscles and longitudinal muscles that run in the dorsal and ventral body region. Overall, our results reveal striking similarities in the muscular arrangement of the life cycle stages of both Symbion species.