Daniel Stec

An integrative description of Macrobiotus paulinae sp. nov. (Tardigrada: Eutardigrada: Macrobiotidae: hufelandi group) from Kenya.

In this paper we describe Macrobiotus paulinae, a new species of the hufelandi group from the Kenyan highlands. In addition to the traditional taxonomic description, aided with morphometrics as well as light and scanning microscopy imaging, we also provide nucleotide sequences of three nuclear (18S rRNA, 28S rRNA, ITS-2) and one mitochondrial (COI) DNA fragment of the new species. The sequences allowed not only a more accurate description but also provided an independent verification of the taxonomic status of Ma. paulinae sp. nov. Such integrative approach requires a considerable number of individuals and eggs, which we have partially subsidised by employing an in vitro culture of the new species. Our analyses revealed that Ma. paulinae sp. nov. is most similar to Macrobiotus madegassus Maucci, 1993 and Macrobiotus modestus Pilato & Lisi, 2009, however it differs from these species, as well as from all other known species of the hufelandi group, by the presence of seven paired dorso-lateral patches of cuticular granulation and the presence of chorionic filaments growing out of terminal discs of egg processes. Macrobiotus paulinae sp. nov. is an example of a species with a miniaturised buccal apparatus (i.e. with reduced peribuccal lamellae and oral cavity armature, and stylet supports inserted on the buccal tube more anteriorly than in typical Ma. hufelandi group species), and it therefore resembles two recently described two-macroplacoided Minibiotus species: Mi. acadianus Meyer & Domingue, 2011 and Mi. julianae Meyer, 2012. The re-examination of the type material for these two species confirmed that they are equipped with peribuccal lamellae and therefore we transfer them to the genus Macrobiotus, specifically to the hufelandi group.

Determinants and taxonomic consequences of extreme egg shell variability in Ramazzottius subanomalus (Biserov, 1985) (Tardigrada)

Nearly a half of known eutardigrade species lay ornamented eggs. The ornamentation is thought to provide attachment of the egg to the substrate and protection for the developing embryo, but from the taxonomic point of view chorion morphology may also provide key characters for species differentiation and identification, especially between closely related taxa. Nonetheless, despite the evolutionary and taxonomic importance of the egg shell, the determinants of its morphology are very poorly, if at all, understood. Here, we combine morphological, molecular and experimental approaches in an attempt to separate the genetic and environmental factors that shape egg chorion morphology in Ramazzottius subanomalus (Biserov, 1985). Our integrative study, based on a population of R. subanomalus isolated from a single moss sample, revealed (1) remarkable variation in egg shell morphology, but (2) relatively little variation in animal morphometric traits, and (3) genetic differentiation, expressed as two ITS-2 haplotypes, but no parallel polymorphism in COI. Although animals did not differ morphometrically between the haplotypes, eggs laid by haplotype 1 and 2 females exhibited highly statistically significant differences in all measured traits. The study demonstrates, for the first time, a correlation between phenotypic and genetic variability within a tardigrade species. The revealed congruence between genetic and morphological traits might be viewed as an example of incipient speciation that illustrates early evolutionary steps leading to species complexes that differ primarily in terms of egg shell morphology. Moreover, our data confirm the value of the ITS-2 fragment in distinguishing very closely related tardigrade lineages.

An integrative revision of Mesocrista Pilato, 1987 (Tardigrada: Eutardigrada: Hypsibiidae)

ABSTRACT Mesocrista is a Holarctic tardigrade genus currently classified within the subfamily Itaquasconinae (Eutardigrada: Parachela: Hypsibiidae). The position of the genus has been so far inferred solely on the basis of light microscopy observations. Here, we present the first-ever scanning electron microscopy images of a dissected bucco-pharyngeal apparatus and DNA sequences for Mesocrista from Spitsbergen, mainland Norway and Poland. The new data allowed an integrative redescription of the nominal M. spitzbergensis from the locus typicus and uncovered a new species, M. revelata, from the European localities. The two species differ phenotypically by the oral cavity armature and claw morphology as well as by a number of morphometric traits, and they are also distinct genetically in all sequenced DNA markers, three nuclear (18S rRNA, 28S rRNA, ITS-2) and one mitochondrial (COI). Both molecular and morphometric data presented in this paper suggest that previous records of M. spitzbergensis should be treated with caution, since most probably some of them signify different Mesocrista species. Our phylogenetic analysis confirmed that Mesocrista is indeed a member of the Itaquasconinae. We also found that the most closely related, among genera for which there are molecular data available, is Adropion, from which Mesocrista differs by the shape of the apophyses for insertion of stylet muscles and the width of the bucco-pharyngeal tube. Finally, we advocate that Diphascon marcusi, a rare species hypothesised to represent either Mesocrista or Adropion, should be transferred to the latter genus and that the species should be redescribed to confirm its validity and systematic position. http://zoobank.org/urn:lsid:zoobank.org:pub:6515D58D-8469-4854-8687-177232112BDA

Unmasking hidden species diversity within the Ramazzottius oberhaeuseri complex, with an integrative redescription of the nominal species for the family Ramazzottiidae (Tardigrada : Eutardigrada : Parachela)

Outdated and inaccurate original descriptions of species nominal for genera or species complexes give birth to notorious difficulties in species identification and often translate into erroneous classifications of multiple species as one. This, in turn, results in severe overestimations of specific geographic ranges and ultimately in underestimations of the diversity of organisms living on our planet. Tardigrades, ubiquitous microscopic metazoans exhibiting relatively few taxonomically useful traits, bear many such problematic species. In this paper, we investigated, morphologically and molecularly, 10 European populations of Ramazzottius cf. oberhaeuseri (Doyère, 1840), a putatively cosmopolitan nominal species for the family Ramazzottiidae. The analysis of COI sequences revealed eight potential species, including one from Paris, the type locality of R. oberhaeuseri, which we used to integratively redescribe the nominal taxon. The identification of seven further species in a handful of samples collected from a single continent explicitly shows the considerable undescribed species diversity within this group and puts in question the cosmopolitan distribution of R. oberhaeuseri. The integrative redescription of R. oberhaeuseri opens the opportunity for descriptions of other species in the complex. http://zoobank.org/urn:lsid:zoobank.org:pub:50B99B53-C317-4DBD-8CF1-16297B85C4F3

High Mitochondrial Diversity in a New Water Bear Species (Tardigrada: Eutardigrada) from Mountain Glaciers in Central Asia, with the Erection of a New Genus Cryoconicus

Abstract. Glaciers and ice sheets are considered a biome with unique organism assemblages. Tardigrada (water bears) are micrometazoans that play the function of apex consumers on glaciers. Cryoconite samples with the dark-pigmented tardigrade Cryoconicus gen. nov. kaczmareki sp. nov. were collected from four locations on glaciers in China and Kyrgyzstan. The erection of the new genus is based on a unique combination of morphological traits as well as on phylogenetic analyses. The analysis of COI sequences in the new species revealed high genetic differentiation with 9 haplotypes shared among 13 sequenced individuals from three sequenced populations. There was no apparent geographic structure in COI haplotype diversity, which might indicate effective dispersal abilities of the new species. A recovery of numerous live individuals from a sample that was frozen for 11 years suggests high survival rates in the natural environment. The ability to withstand low temperatures, combined with dark pigmentation that is hypothesised to protect from intense UV radiation, could explain how the new taxon is able to dwell in an extreme glacial habitat. We also found that a rare mountain tardigrade Ramazzottius cataphractus (Maucci, 1974) is morphologically similar to the new species, therefore we propose to transfer it to the new genus. Our study indicates that glacier invertebrate fauna is still poorly known and requires intense research.

Description of Macrobiotus scoticus sp. nov. (Tardigrada : Macrobiotidae : hufelandi group) from Scotland by means of integrative taxonomy

Abstract. A new species of the Macrobiotus hufelandi group is described from Scotland. Both light and scanning electron microscopy were used for collecting morphological and morphometric data whereas genotyping allowed sequences for three nuclear (18S rRNA, 28S rRNA and ITS-2) and one mitochondrial (COI) DNA fragment to be obtained. By undertaking in vitro culturing, we were able to obtain a number of eggs and provide a detailed description. Macrobiotus scoticus sp. nov. exhibits a reduced oral cavity armature, with only the ridges of the third band of teeth visible under light microscopy, therefore the animals of the new species are most similar to individuals of M. almadai Fontoura et al., 2008; M. diversus Biserov, 1990; M. madegassus Maucci, 1993; M. martini Bartels et al., 2009; M. modestus Pilato and Lisi, 2009; and M. paulinae Stec et al., 2015. However, M. scoticus sp. nov. lays eggs with strongly modified processes, which makes this species unique not only among the species listed above but from all hufelandi group taxa. In contrast to mushroom-shaped egg processes found in the majority of the hufelandi complex species, processes in M. scoticus sp. nov. are in the shape of spatulas, with longitudinally flattened terminal portions. Superficially, the eggs of the new species resemble those of M. kristenseni Guidetti et al., 2013, which exhibits elongated, spike- or filament-like processes. Nonetheless, the eggs of M. kristenseni have a smooth inter-process surface whereas in the new species the surface is covered with a very dense reticulum. Moreover, M. kristenseni has a better developed oral cavity armature, with all three bands of teeth detectable under light microscopy.

The tardigrade fauna of Tunisia, with an integrative description of Bryodelphax maculatus sp. nov. (Heterotardigrada: Echiniscidae)

The current knowledge of the diversity and distribution of tardigrade species in northern Africa is scarce and far from satisfactory. In this paper we review old and provide new faunistic records for various regions of Tunisia. Among the newly recorded taxa, we found a new heterotardigrade Bryodelphax maculatus sp. nov. The new species belongs to the weglarskae group, characterised by the presence of ventral plates, but differs from its congeners by a different arrangement of ventral plates and by conspicuous groups of cuticular tubercles on the ventral cuticle. The analysis of larval, juvenile and mature instars of the new species revealed, for the first time in heterotardigrades, ontogenetic variability in the development of ventral armature.

An integrative description of Macrobiotus shonaicus sp. nov. (Tardigrada: Macrobiotidae) from Japan with notes on its phylogenetic position within the hufelandi group

Tardigrade research in Japan dates back over 100 years, and to date, 167 species of this ecdysozoan phylum have been reported from the country. Of these species, the Macrobiotus hufelandi complex has been represented only by the nominal taxon of this group, Macrobiotus hufelandi. In this article, a new species of the hufelandi group from Japan, Macrobiotus shonaicus sp. nov., is described using integrative taxonomy. In addition to the detailed morphological and morphometric data, obtained using phase contrast light microscopy (PCM) and scanning electron microscopy (SEM), we provide DNA sequences of four molecular markers (both nuclear and mitochondrial). The new species belongs to the persimilis subgroup and is most similar to M. anemone from USA, M. naskreckii from Mozambique, and M. patagonicus from Argentina, but it can be easily distinguished from these species by the presence of thin flexible filaments on terminal discs of the egg process. By the latter character, the new species is most similar to M. paulinae and M. polypiformis, but it can be easily distinguished from them by having a solid egg surface between egg processes (i.e., without pores or reticulum). A phylogenetic analysis of available DNA sequences of the COI marker for the hufelandi group revealed that the new species clusters with the two other species that exhibit filaments on egg process discs (M. paulinae and M. polypiformis) and with two species that have entire egg processes modified into filaments (M. kristenseni and M. scoticus). All five species form a clade distinct from all other sequenced species of the hufelandi group with typical mushroom- or inverted goblet-shaped egg processes, which may suggest that the ancestor of the five species with atypical egg processes had a mutation allowing derivations from the mushroom or inverted chalice-like shape of egg processes.

An integrative description of a new tardigrade species Mesobiotus romani sp. nov. (Macrobiotidae: harmsworthi group) from the Ecuadorian Pacific coast

In a mixed moss and lichen sample collected in Esmeraldas Province in north-western Ecuador, 20 tardigrades and 11 eggs, belonging to a new species of the genus Mesobiotus, were found. In addition to the traditional taxonomic description with morphometrics, light and scanning microscopy imaging, we also provide nucleotide sequences of three nuclear (18S rRNA, 28S rRNA, ITS-2) and one mitochondrial (COI) DNA fragments of the new species. Based on the egg chorion morphology, Mesobiotus romani sp. nov. is the most similar to: M. binieki, M. coronatus, M. dimentmani, M. patiens, M. perfidus, M. philippinicusi, M. pseudoblocki, M. pseudocoronatus, M. pseudopatiens, M. radiatus, M. rigidus, M. simulans and M. wuzhishanensis, but differs mainly by some specific characters of both egg and adult morphology, and morphometrics.

Paramacrobiotus lachowskae, a new species of Tardigrada from Colombia (Eutardigrada : Parachela : Macrobiotidae)

ABSTRACT In this article we describe a new Paramacrobiotus species of the areolatus group by means of integrative taxonomy. Together with the detailed morphological and morphometric data (obtained from light and scanning electron microscopy) we also provide DNA sequences of four universal molecular markers used in tardigrade taxonomy (three nDNA fragments: 18S rRNA, 28S rRNA, ITS-2, and one mtDNA fragment: COI). The animals of Paramacrobiotus lachowskae sp. nov. are similar to several species of the areolatus complex and the eggshell ornamentation is similar to that of two species of the richtersi group. Therefore, the new species can be easily distinguished from the areolatus group species by egg morphology (dome-like, wrinkled processes with long flexible spines/filaments covered by fine short hairs) and from species of the richtersi complex by the absence of the microplacoid. Paramacrobiotus lachowskae sp. nov. is the 43rd species reported from Colombia and fifth Paramacrobiotus species from this South American country. http://zoobank.org/urn:lsid:zoobank.org:pub:FEDC11D3-DCCF-4699-A704-CE6DEA1CDED4