Ivo de Sena Oliveira

A world checklist of Onychophora (velvet worms), with notes on nomenclature and status of names.

Abstract Currently, the number of valid species of Onychophora is uncertain. To facilitate taxonomic work on this understudied animal group, we present an updated checklist for the two extant onychophoran subgroups, Peripatidae and Peripatopsidae, along with an assessment of the status of each species. According to our study, 82 species of Peripatidae and 115 species of Peripatopsidae have been described thus far. However, among these 197 species, 20 are nomina dubia due to major taxonomic inconsistencies. Apart from nomina dubia, many of the valid species also require revision, in particular representatives of Paraperipatus within the Peripatopsidae, and nearly all species of Peripatidae. In addition to extant representatives, the record of unambiguous fossils includes three species with uncertain relationship to the extant taxa. For all species, we provide a list of synonyms, information on types and type localities, as well as remarks on taxonomic and nomenclatural problems and misspellings. According to recent evidence of high endemism and cryptic speciation among the Peripatidae and Peripatopsidae, previous synonyms are revised. Putative mutations, subspecies and variations are either raised to the species status or synonymised with corresponding taxa. In our revised checklist, we follow the rules and recommendations of the International Code of Zoological Nomenclature to clarify previous inconsistencies.

Unexplored Character Diversity in Onychophora (Velvet Worms): A Comparative Study of Three Peripatid Species

Low character variation among onychophoran species has been an obstacle for taxonomic and phylogenetic studies in the past, however we have identified a number of new and informative characters using morphological, molecular, and chromosomal techniques. Our analyses involved a detailed examination of Epiperipatus biolleyi from Costa Rica, Eoperipatus sp. from Thailand, and a new onychophoran species and genus from Costa Rica, Principapillatus hitoyensis gen. et sp. nov.. Scanning electron microscopy on embryos and specimens of varying age revealed novel morphological characters and character states, including the distribution of different receptor types along the antennae, the arrangement and form of papillae on the head, body and legs, the presence and shape of interpedal structures and fields of modified scales on the ventral body surface, the arrangement of lips around the mouth, the number, position and structure of crural tubercles and anal gland openings, and the presence and shape of embryonic foot projections. Karyotypic analyses revealed differences in the number and size of chromosomes among the species studied. The results of our phylogenetic analyses using mitochondrial COI and 12S rRNA gene sequences are in line with morphological and karyotype data. However, our data show a large number of unexplored, albeit informative, characters in the Peripatidae. We suggest that analysing these characters in additional species would help unravel species diversity and phylogeny in the Onychophora, and that inconsistencies among most diagnostic features used for the peripatid genera in the literature could be addressed by identifying a suite of characters common to all peripatids.

Cryptic Speciation in Brazilian Epiperipatus (Onychophora: Peripatidae) Reveals an Underestimated Diversity among the Peripatid Velvet Worms

BACKGROUND Taxonomical studies of the neotropical Peripatidae (Onychophora, velvet worms) have proven difficult, due to intraspecific variation and uniformity of morphological characters across this onychophoran subgroup. We therefore used molecular approaches, in addition to morphological methods, to explore the diversity of Epiperipatus from the Minas Gerais State of Brazil. METHODOLOGY/PRINCIPAL FINDINGS Our analyses revealed three new species. While Epiperipatus diadenoproctussp. nov. can be distinguished from E. adenocryptussp. nov. and E. paurognostussp. nov. based on morphology and specific nucleotide positions in the mitochondrial cytochrome c oxidase subunit I (COI) and small ribosomal subunit RNA gene sequences (12S rRNA), anatomical differences between the two latter species are not evident. However, our phylogenetic analyses of molecular data suggest that they are cryptic species, with high Bayesian posterior probabilities and bootstrap and Bremer support values for each species clade. The sister group relationship of E. adenocryptussp. nov. and E. paurognostussp. nov. in our analyses correlates with the remarkable morphological similarity of these two species. To assess the species status of the new species, we performed a statistical parsimony network analysis based on 582 base pairs of the COI gene in our specimens, with the connection probability set to 95%. Our findings revealed no connections between groups of haplotypes, which have been recognized as allopatric lineages in our phylogenetic analyses, thus supporting our suggestion that they are separate species. CONCLUSIONS/SIGNIFICANCE Our findings suggest high cryptic species diversity and endemism among the neotropical Peripatidae and demonstrate that the combination of morphological and molecular approaches is helpful for clarifying the taxonomy and species diversity of this apparently large and diverse onychophoran group.

Apodemes associated with limbs support serial homology of claws and jaws in onychophora (velvet worms)

Although the onychophoran jaw blades are believed to be derivatives of foot claws, serial homology of these structures has not been demonstrated. To shed light on the evolutionary origin of the onychophoran jaws, we searched for morphological landmarks and compared the internal and external anatomy of jaws and distal leg portions in representatives of the two major onychophoran subgroups, the Peripatidae and Peripatopsidae. Our data revealed hitherto unknown structures associated with the onychophoran limbs, such as a soft diastemal membrane separating the anterior and posterior portions of the inner jaw blade (present only in Peripatidae), apodemes associated with feet, an eversible dorsal sac at the basis of each foot claw, and a specific arrangement of musculature associated with the sclerotised claws, jaws and their apodemes. Specific correspondences in structure and position of apodemes support serial homology of claws and jaws, suggesting that the onychophoran jaw evolved from the distal portion rather than the entire limb in the last common ancestor of Onychophora. J. Morphol. 274:1180–1190, 2013.

The role of ventral and preventral organs as attachment sites for segmental limb muscles in Onychophora

BackgroundThe so-called ventral organs are amongst the most enigmatic structures in Onychophora (velvet worms). They were described as segmental, ectodermal thickenings in the onychophoran embryo, but the same term has also been applied to mid-ventral, cuticular structures in adults, although the relationship between the embryonic and adult ventral organs is controversial. In the embryo, these structures have been regarded as anlagen of segmental ganglia, but recent studies suggest that they are not associated with neural development. Hence, their function remains obscure. Moreover, their relationship to the anteriorly located preventral organs, described from several onychophoran species, is also unclear. To clarify these issues, we studied the anatomy and development of the ventral and preventral organs in several species of Onychophora.ResultsOur anatomical data, based on histology, and light, confocal and scanning electron microscopy in five species of Peripatidae and three species of Peripatopsidae, revealed that the ventral and preventral organs are present in all species studied. These structures are covered externally with cuticle that forms an internal, longitudinal, apodeme-like ridge. Moreover, phalloidin-rhodamine labelling for f-actin revealed that the anterior and posterior limb depressor muscles in each trunk and the slime papilla segment attach to the preventral and ventral organs, respectively. During embryonic development, the ventral and preventral organs arise as large segmental, paired ectodermal thickenings that decrease in size and are subdivided into the smaller, anterior anlagen of the preventral organs and the larger, posterior anlagen of the ventral organs, both of which persist as paired, medially-fused structures in adults. Our expression data of the genes Delta and Notch from embryos of Euperipatoides rowelli revealed that these genes are expressed in two, paired domains in each body segment, corresponding in number, position and size with the anlagen of the ventral and preventral organs.ConclusionsOur findings suggest that the ventral and preventral organs are a common feature of onychophorans that serve as attachment sites for segmental limb depressor muscles. The origin of these structures can be traced back in the embryo as latero-ventral segmental, ectodermal thickenings, previously suggested to be associated with the development of the nervous system.

Myoanatomy of the velvet worm leg revealed by laboratory-based nanofocus X-ray source tomography

Significance X-ray computed tomography (CT) imaging has become popular for investigating, nondestructively and three-dimensionally, both external and internal structures of various specimens. However, the limited resolution of conventional laboratory-based CT systems (≥500 nm) still hampers the detailed visualization of features on the low nanometer level. We present a laboratory CT device and data processing pipeline to routinely and efficiently generate high-resolution 3D data (≈100 nm) without requiring synchrotron radiation facilities. Our setup is especially relevant for conducting detailed analysis of very small biological samples, as demonstrated for a walking appendage of a velvet worm. Comparative analyses of our CT data with those obtained from other popular imaging methods highlight the advantages and future applicability of the nanoCT setup. X-ray computed tomography (CT) is a powerful noninvasive technique for investigating the inner structure of objects and organisms. However, the resolution of laboratory CT systems is typically limited to the micrometer range. In this paper, we present a table-top nanoCT system in conjunction with standard processing tools that is able to routinely reach resolutions down to 100 nm without using X-ray optics. We demonstrate its potential for biological investigations by imaging a walking appendage of Euperipatoides rowelli, a representative of Onychophora—an invertebrate group pivotal for understanding animal evolution. Comparative analyses proved that the nanoCT can depict the external morphology of the limb with an image quality similar to scanning electron microscopy, while simultaneously visualizing internal muscular structures at higher resolutions than confocal laser scanning microscopy. The obtained nanoCT data revealed hitherto unknown aspects of the onychophoran limb musculature, enabling the 3D reconstruction of individual muscle fibers, which was previously impossible using any laboratory-based imaging technique.

A new and critically endangered species and genus of Onychophora (Peripatidae) from the Brazilian savannah – a vulnerable biodiversity hotspot

In recent years, a high number of endemic species might have gone extinct silently in the Brazilian savannah (=Cerrado), a biodiversity hotspot, as researchers are not able to describe them as fast as their natural habitats are being destroyed. This especially holds true for onychophorans, or velvet worms, which include many point endemic species and, in this biome, only occur in small gallery forests susceptible to disturbances. In the present work, we use a combined morphological and molecular approach to describe a relict and critically endangered onychophoran species from the Cerrado. Our morphological analyses using light and scanning electron microscopy revealed novel characters and character states for Peripatidae, such as six interpedal structures per segment, modified scales on the genital and posterior body region of females and a third class of dermal papillae, herein named peripheral accessory papillae. Since Cerradopatus sucuriuensis gen. et sp. nov. showed a unique set of morphological characters and could not be assigned to any described genus of Peripatidae, a new genus is raised for this species herein. In addition to morphological data, the new taxon is supported by karyotypic, slime protein profiling and molecular data, including sequences of the mitochondrial genes COI and 12S rRNA. Unfortunately, Cerradopatus sucuriuensis gen. et sp. nov. is prone to an extremely high risk of extinction in the wild, as it only occurs in specific humid patches within three small, unprotected fragments of forest that are being progressively affected by human activities. According to the IUCN Red List categories and criteria, this species is classified as Critically Endangered and requires urgent conservation efforts. http://zoobank.org/urn:lsid:zoobank.org:pub:10D0762E-7842-428B-8ABB-9096BED10F65

Capture of Prey, Feeding, and Functional Anatomy of the Jaws in Velvet Worms (Onychophora)

Onychophorans are carnivorous, terrestrial invertebrates that occur in tropical and temperate forests of the Southern Hemisphere and around the Equator. Together with tardigrades, onychophorans are regarded as one of the closest relatives of arthropods. One of the most peculiar features of onychophorans is their hunting and feeding behavior. These animals secrete a sticky slime, which is ejected via a pair of slime-papillae, to entangle the prey. After the prey has been immobilized, its cuticle is punctured using a pair of jaws located within the mouth. These jaws constitute internalized appendages of the second body segment and are innervated by the deutocerebrum; thus, they are homologous to the chelicerae of chelicerates, and to the (first) antennae of myriapods, crustaceans, and insects. The jaws are also serial homologs of the paired claws associated with each walking limb of the trunk. The structure of the jaws is similar in representatives of the two major onychophoran subgroups, the Peripatidae and Peripatopsidae. Each jaw is characterized by an outer and an inner blade; while the outer blade consists only of a large principal tooth and up to three accessory teeth, the inner blade bears numerous additional denticles. These denticles are separated from the remaining part of the inner jaw by a diastema and a soft membrane only in peripatids. The onychophoran jaws are associated with large apodemes and specialized muscles that enable their movement. In contrast to the mandibles of arthropods, the onychophoran jaws are moved along, rather than perpendicular to, the main axis of the body. Our elemental analysis reveals an increased incorporation of calcium at the tip of each blade, which might provide rigidity, whereas there is no evidence for incorporation of metal or prominent mineralization. Stability of the jaw might be further facilitated by the cone-in-cone organization of its cuticle, as each blade consists of several stacked, cuticular elements. In this work, we summarize current knowledge on the jaws of onychophorans, which are a characteristic feature of these animals.

Evaluation of contrasting techniques for X-ray imaging of velvet worms (Onychophora)

Non‐invasive imaging techniques like X‐ray computed tomography have become very popular in zoology, as they allow for simultaneous imaging of the internal and external morphology of organisms. Nevertheless, the effect of different staining approaches required for this method on samples lacking mineralized tissues, such as soft‐bodied invertebrates, remains understudied. Herein, we used synchrotron radiation‐based X‐ray micro‐computed tomography to compare the effects of commonly used contrasting approaches on onychophorans – soft‐bodied invertebrates important for studying animal evolution. Representatives of Euperipatoides rowelli were stained with osmium tetroxide (vapour or solution), ruthenium red, phosphotungstic acid, or iodine. Unstained specimens were imaged using both standard attenuation‐based and differential phase‐contrast setups to simulate analyses with museum material. Our comparative qualitative analyses of several tissue types demonstrate that osmium tetroxide provides the best overall tissue contrast in onychophorans, whereas the remaining staining agents rather favour the visualisation of specific tissues and/or structures. Quantitative analyses using signal‐to‐noise ratio measurements show that the level of image noise may vary according to the staining agent and scanning medium selected. Furthermore, box‐and‐whisker plots revealed substantial overlap in grey values among structures in all datasets, suggesting that a combination of semiautomatic and manual segmentation of structures is required for comprehensive 3D reconstructions of Onychophora, irrespective of the approach selected. Our results show that X‐ray micro‐computed tomography is a promising technique for studying onychophorans and, despite the benefits and disadvantages of different staining agents for specific tissues/structures, this method retrieves informative data that may eventually help address evolutionary questions long associated with Onychophora.

Revision of Tasmanian viviparous velvet worms (Onychophora : Peripatopsidae) with descriptions of two new species

Abstract. The restricted distribution of viviparous onychophorans in Tasmania has long been a subject of discussion, but their evolutionary history remains unclear. We applied morphological, molecular and karyological methods to assess the taxonomy and phylogenetic relationships of the four viviparous species reported from Tasmania, including Tasmanipatus barretti, T. anophthalmus and two undescribed species previously referred to as ‘Tasmania’ sp. 1 and sp. 2. We demonstrate that all four species can be unambiguously distinguished based on independent character sets. The two ‘Tasmania’ species, which were previously thought to be cryptic, proved to exhibit a set of distinct morphological characters. Molecular phylogenetic analyses revealed that the four species belong to a major clade that includes Peripatoides from New Zealand, and that species from the two landmasses show reciprocal monophyly within this clade. Within the Tasmanian clade, T. anophthalmus is more closely related to the two ‘Tasmania’ species than to T. barretti. Based on this relationship and the lack of morphological and/or karyological characters supporting the Tasmanian viviparous clade, we erect two new genera to accommodate the two ‘Tasmania’ species (Diemenipatus, gen. nov.) and T. anophthalmus (Leucopatus, gen. nov.). An emended diagnosis followed by a redescription of T. barretti is provided and ‘Tasmania’ sp. 1 and sp. 2 are formally described as D. taiti, gen. et sp. nov. and D. mesibovi, gen. et sp. nov., respectively.