Milnesium Tardigradum Research Articles

A New Species of Bisexual Milnesium (Eutardigrada: Apochela) Having Aberrant Claws from Innhovde, Dronning Maud Land, East Antarctica.

There have been several records in the last 60 years for East Antarctica for Milnesium tardigradum Doyère, 1840 sensu lato, now considered a species complex. During the 56th Japanese Antarctic Research Expedition summer operation (2014-2015), a new tardigrade species in the genus Milnesium Doyère, 1840 was found in an ice-free Innhovde area along Lützow-Holm Bay, Dronning Maud Land, East Antarctica. The new species has aberrant claws with four to seven points on each secondary claw branch, which distinguishes it from other Milnesium species. A male specimen was found in the population and evidence showed that an isolated adult female moulted twice without oviposition. This strongly suggested bisexual reproduction for this population. The new species, Milnesium rastrum sp. nov., is described with its phylogenetic position and a discussion on the reproductive strategies for the harsh environments.

Considerations on the claws of the Apochela and a novel detail of the bucco-pharyngeal apparatus of the genus Milnesium (Tardigrada: Apochela: Milnesiidae)

This article clarifies two morphological problems, the first regarding all Apochela Schuster, Nelson, Grigarick and Christenberry 1980, and the second the genus Milnesium Doyère, 1840. Among Eutardigrada Richters, 1926, Apochela have four claws per leg, while Parachela Schuster, Nelson, Grigarick and Christenberry 1980 have two double claws per leg. Until now, the four claws per leg of Apochela have been denominated as if they had derived from two parachelan-like double claws undergoing branch separation. However, morphological and molecular data indicate the relative phylogenetic distance of Apochela from Parachela as sister groups; the ancestor of Eutardigrada was probably similar to a primitive heterotardigrade in having four separate claws per leg, which organised and differentiated in couples but without fusion in Apochela, while they fused to form proper double claws only in Parachela. If this hypothesis is correct (Apochela would have retained four separate claws per leg as plesiomorphic character), the denomination of apochelan claws must be adapted to reflect it. The second problem regards the valvular system present between buccal tube and pharyngeal bulb in the genus Milnesium. Until now, only one valvular system type was known, attributed to Milnesium tardigradum (today sensu lato), or to Milnesium cf. tardigradum. We reveal a second type recognisable in many species. The two types are described, discussing their possible correlation with life stage, sex, some morphological and morphometric traits, mounting medium, geographic distribution and currently available phylogenetic trees; the valvular system type is probably species-specific and we attribute to most species of the genus the correct type. Our conclusions are preliminary, but, in any case, the present study indicates that the exact morphology of the discussed portion of the bucco-pharyngeal apparatus in Milnesium is still not well known.

The Unique Antimicrobial Recognition and Signaling Pathways in Tardigrades with a Comparison Across Ecdysozoa

Tardigrades are microscopic animals known to withstand unfavorable abiotic conditions. These animals are also constantly exposed to biotic stresses, including parasites and internal microbiomes. However, the tardigrade immune mechanisms against these biotic stresses are largely uncharacterized. Due to the contentious phylogenetic position of tardigrades, it is not intuitive whether they possess an immune system more similar to that of arthropods (e.g., Toll, Imd, and JNK pathways of the Drosophila melanogaster antimicrobial response) or to that of nematodes (e.g., the Tir-1/Nsy-1/Sek-1/Pmk-1/Atf-7 signaling cassette [called Tir-1 pathway here]) in Caenorhabditis elegans). In this study, comparative genomic analyses were conducted to mine homologs of canonical D. melanogaster and C. elegans immune pathway genes from eight tardigrades (Echiniscoides cf. sigismundi, Echiniscus testudo, Hypsibius exemplaris, Mesobiotus philippinicus, Milnesium tardigradum, Paramacrobiotus richtersi, Richtersius cf. coronifer, and Ramazzottius varieornatus) and four non-arthropod ecdysozoans (two onychophorans: Epiperipatus sp. and Opisthopatus kwazululandi; one nematomorph: Paragordius varius; and one priapulan: Priapulus caudatus) in order to provide insights into the tardigrade antimicrobial system. No homologs of the intracellular components of the Toll pathway were detected in any of the tardigrades examined. Likewise, no homologs of most of the Imd pathway genes were detected in any of the tardigrades or any of the other non-arthropod ecdysozoans. Both the JNK and Tir-1 pathways, on the other hand, were found to be conserved across ecdysozoans. Interestingly, tardigrades had no detectable homologs of NF-κB, the major activator of antimicrobial response gene expression. Instead, tardigrades appear to possess NF-κB distantly related NFAT homologs. Overall, our results show that tardigrades have a unique gene pathway repertoire that differs from that of other ecdysozoans. Our study also provides a framework for future studies on tardigrade immune responses.

Milnesium tardigradum Doyère, 1840: The first integrative study of interpopulation variability in a tardigrade species

<i>Milnesium tardigradum</i> Doyère, 1840: The first integrative study of interpopulation variability in a tardigrade species

The Resilience of Life to Astrophysical Events

Much attention has been given in the literature to the effects of astrophysical events on human and land-based life. However, little has been discussed on the resilience of life itself. Here we instead explore the statistics of events that completely sterilise an Earth-like planet with planet radii in the range 0.5–1.5R⊕ and temperatures of ∼300 K, eradicating all forms of life. We consider the relative likelihood of complete global sterilisation events from three astrophysical sources – supernovae, gamma-ray bursts, large asteroid impacts, and passing-by stars. To assess such probabilities we consider what cataclysmic event could lead to the annihilation of not just human life, but also extremophiles, through the boiling of all water in Earth’s oceans. Surprisingly we find that although human life is somewhat fragile to nearby events, the resilience of Ecdysozoa such as Milnesium tardigradum renders global sterilisation an unlikely event.

Specimens with an Artifact Appearing as 'Three Spines' in Milnesium tardigradum var. trispinosa Rahm, 1931 (Tardigrada).

Milnesium tardigradum trispinosa Rahm, 1931 is characterized by its three spines at the posterior end of the animal, but has never been reported since its original description. Among mounted specimens of Milnesium sp. from Japan and M. tardigradum s. s. from France, several cases with these 'three spines' were observed. In these samples, the character was formed by an artifact of the fixation process. Images of these specimens show such a striking similarity to Milnesium tardigradum trispinosa that this taxon must be considered as having been erroneously established as a result of misinterpretation of an artifact and no longer valid.

Effect of ultra-high pressure on small animals, tardigrades and Artemia

AbstractThis research shows that small animals, tardigrades (Milnesium tardigradum) in tun (dehydrated) state and Artemia salina cists (dried eggs) can tolerate the very high hydrostatic pressure of 7.5 GPa. It was really surprising that living organisms can survive after exposure to such a high pressure. We extended these studies to the extremely high pressure of 20 GPa by using a Kawai-type octahedral anvil press. After exposure to this pressure for 30 min, the tardigrades were soaked in pure water and investigated under a microscope. Their bodies regained metabolic state and no serious injury could be seen. But they were not alive. A few of Artemia eggs went part of the way to hatching after soaked in sea water, but they never grew any further. Comparing with the case of blue-green alga, these animals are weaker under ultra-high pressure.

Tardigrades of the Tree Canopy: Milnesium swansoni sp. nov. (Eutardigrada: Apochela: Milnesiidae) a new species from Kansas, U.S.A.

Milnesium swansoni sp. nov. is a new species of Eutardigrada described from the tree canopy in eastern Kansas, USA. This species within the order Apochela, family Milnesiidae, genus Milnesium is distinguished by its smooth cuticle, narrow buccal tube, four peribuccal lamellae, primary claws without accessory points, and a secondary claw configuration of [3-3]-[3-3]. The buccal tube appears to be only half the width of the nominal species Milnesium tardigradum for animals of similar body length. The species adds to the available data for the phylum, and raises questions concerning species distribution.

B14-O-05Correlative imaging analysis of Tardigrada (water bears) under the active and dehydrated states by X-ray micro-computed tomography, electron microscopy and confocal microscopy

Some Tardigrades can survive in the dehydrated state called ‘tun’. However how they can achieve such a remarkable ability of anhydrobiosis has not been well understood. We studied the internal structure of the tuns of three Tardigrade species by mCT at SPring-8 (BL20XU and BL47XU) at the resolution of 83〜500nm: Milnesium tardigradum, Ramazzottius varieornatus, as well as Macrobiotus sp. collected on the Harima Science campus of the Univ. of Hyogo. Previously we identified stylets, buccal tubes, pharynx, and eggs (2010A1350, 2013A1319). Here we succeeded in visualizing organelles with less X-ray absorption (2014B1568). By comparing the mCT images with the ones by confocal microscopy, we identified them as droplets in the coelomic cells in the tun. This allowed us to map the 3D distribution of droplets in a coelomic cell as well as coelomic cells in a tun (Fig). We also visualized the structure of organs in the active state, including the brain, eyes, ganglia, muscles, and digestive organs by mCT and TEM. We are now analyzing the structural change during the recovery (rehydration) by cryo-mCT, and the nano-structure of the tun by electron microscopy.

Distribution and evolution of the serine/aspartate racemase family in invertebrates.

Free D-amino acids have been found in various invertebrate phyla, while amino acid racemase genes have been identified in few species. The purpose of this study is to elucidate the distribution, function, and evolution of amino acid racemases in invertebrate animals. We searched the GenBank databases, and found 11 homologous serine racemase genes from eight species in eight different invertebrate phyla. The cloned genes were identified based on their maximum activity as Acropora millepora (Cnidaria) serine racemase (SerR) and aspartate racemase (AspR), Caenorhabditis elegans (Nematoda) SerR, Capitella teleta (Annelida) SerR, Crassostrea gigas (Mollusca) SerR and AspR, Dugesia japonica (Platyhelminthes) SerR, Milnesium tardigradum (Tardigrada) SerR, Penaeus monodon (Arthropoda) SerR and AspR and Strongylocentrotus purpuratus (Echinodermata) AspR. We found that Acropora, Aplysia, Capitella, Crassostrea and Penaeus had two amino acid racemase paralogous genes and these paralogous genes have evolved independently by gene duplication at their recent ancestral species. The transcriptome analyses using available SRA data and enzyme kinetic data suggested that these paralogous genes are expressed in different tissues and have different functions in vivo. Phylogenetic analyses clearly indicated that animal SerR and AspR are not separated by their particular racemase functions and form a serine/aspartate racemase family cluster. Our results revealed that SerR and AspR are more widely distributed among invertebrates than previously known. Moreover, we propose that the triple serine loop motif at amino acid positions 150-152 may be responsible for the large aspartate racemase activity and the AspR evolution from SerR.

The genus Milnesium (Tardigrada: Eutardigrada: Milnesiidae) in the Great Smoky Mountains National Park (North Carolina and Tennessee, USA), with the description of Milnesium bohleberi sp. nov.

For many decades the genus Milnesium was thought to consist of a single, cosmopolitan species: Milnesium tardigradum Doyère, 1840. However, recently the genus has been re-evaluated, and numerous new species have been described. Currently, over twenty extant species and one fossil are recognised, and most appear to have very narrow geographic ranges. It is doubtful that M. tardigradum sensu stricto is truly cosmopolitan, but to evaluate this hypothesis, specimens previously identified as M. tardigradum must be re-examined using newly proposed taxonomic characters. As part of the All Taxa Biodiversity Inventory (ATBI) we collected Milnesium specimens from various locations in the Great Smoky Mountains National Park (GSMNP). Two Milnesium species have been evaluated, and one of them, Milnesium bohleberi sp. nov., is new to science. The new species is most similar to M. eurystomum but differs by shorter claws and a shorter, narrower, and more cylindrical buccal tube. The other Milnesium species, very rare in our collection, is morphologically indistinguishable from Milnesium granulatum Ramazzotti 1962, which was previously known only from Chile, Italy and Romania. Based on the recently revised description of M. tardigradum sensu stricto, this nominal species for the genus has not been found in the GSMNP samples.

New tardigrade records for the Baltic states with a description of Minibiotus formosus sp. n. (Eutardigrada, Macrobiotidae).

In sixteen moss, lichen and mixed (moss/lichen) samples, collected from Estonia, Latvia and Lithuania, 291 specimens, 48 simplexes, including one exuvium with 6 eggs, and 8 free-laid eggs of eutardigrades were found. In total, 17 species, together with one new to science, were identified (all are new records for the Baltic states): Astatumen bartosi, Diphascon (Adropion) prorsirostre, D. (Diphascon) bullatum, D. (D.) pingue pingue, D. (D.) recamieri, D. (D.) rugosum, Hypsibius convergens, H. dujardini, H. cf. scabropygus, Isohypsibius ronsisvallei, I. sattleri, Macrobiotus harmsworthi harmsworthi, M. hufelandi hufelandi, Milnesium asiaticum, Milnesium tardigradum tardigradum, Minibiotus formosus sp. n. and Paramacrobiotus richtersi. The new species is most similar to Minibiotus gumersindoi, but differs from it mainly by the presence of two types of cuticular pores, the absence of a triangular or pentagonal arrangement of pores above a single large pore on legs, the presence of granulation on all legs and a different macroplacoid length sequence. In this paper we also provide photographs and morphometrics of H. cf. scabropygus.

Terrestrial and freshwater Tardigrada of the Americas.

This paper provides a comprehensive list of the freshwater and terrestrial tardigrade fauna reported from the Americas (North America, South America, Central America and the West Indies), their distribution in the Americas, and the substrates from which they have been reported. Data were obtained from 316 published references. Authors' identifications were accepted at face value unless subsequently amended. Taxa were assigned to sub-national units (states, provinces, etc.). Many areas, in particular large portions of Central America and the West Indies, have no reported tardigrade fauna. The presence of 54 genera and 380 species has been reported for the Americas; 245 species have been collected in the Nearctic ecozone and 251 in the Neotropical ecozone. Among the tardigrade species found in the Americas, 52 are currently considered cosmopolitan, while 153 species have known distributions restricted to the Americas. Based on recent taxonomic revision of the genus Milnesium, the vast majority of records of M. tardigradum in the Americas should now be reassigned to Milnesium tardigradum sensu lato, either because the provided description differs from M. tardigradum sensu stricto or because insufficient description is provided to make a determination; the remainder should be considered Milnesium cf. tardigradum. Most terrestrial tardigrade sampling in the Americas has focused on cryptogams (mosses, lichens and liverworts); 90% of the species have been collected in such substrates. The proportion of species collected in other habitats is lower: 14% in leaf litter, 20% in soil, and 24% in aquatic samples (in other terrestrial substrates the proportion never exceeds 5%). Most freshwater tardigrades have been collected from aquatic vegetation and sediment. For nine species in the Americas no substrates have been reported.

Phylogenetic position of the enigmatic clawless eutardigrade genus Apodibius Dastych, 1983 (Tardigrada), based on 18S and 28S rRNA sequence data from its type species A. confusus

The systematics of Eutardigrada, the largest lineage among the three classes of the phylum Tardigrada, is based mainly on the morphology of the leg claws and of the buccal apparatus. However, three members of the rarely recorded and poorly known limno-terrestrial eutardigrade genus Apodibius have no claws on their strongly reduced legs, a unique character among all tardigrades. This absence of all claws makes the systematic position of Apodibius one of the most enigmatic among the whole class. Until now all known associates of the genus Apodibius have been located in the incertae sedis species group or, quite recently, included into the Necopinatidae family. In the present study, phylogenetic analyses of 18S and 28S rRNA sequence data from 31 tardigrade species representing four parachelan superfamilies (Isohypsibioidea, Hypsibioidea, Macrobiotoidea, Eohypsibioidea), the apochelan Milnesium tardigradum, and the type species of the genus Apodibius, A. confusus, indicated close relationship of the Apodibius with tardigrade species recently included in the superfamily Isohypsibioidea. This result was well-supported and consistent across all markers (separate 18S rRNA, 28S rRNA, and combined 18S rRNA+28S rRNA datasets) and methods (MP, ML) applied.

The Aquaporin Channel Repertoire of the TardigradeMilnesium tardigradum

Limno-terrestrial tardigrades are small invertebrates that are subjected to periodic drought of their micro-environment. They have evolved to cope with these unfavorable conditions by anhydrobiosis, an ametabolic state of low cellular water. During drying and rehydration, tardigrades go through drastic changes in cellular water content. By our transcriptome sequencing effort of the limno-terrestrial tardigrade Milnesium tardigradum and by a combination of cloning and targeted sequence assembly, we identified transcripts encoding eleven putative aquaporins. Analysis of these sequences proposed 2 classical aquaporins, 8 aquaglyceroporins and a single potentially intracellular unorthodox aquaporin. Using quantitative real-time PCR we analyzed aquaporin transcript expression in the anhydrobiotic context. We have identified additional unorthodox aquaporins in various insect genomes and have identified a novel common conserved structural feature in these proteins. Analysis of the genomic organization of insect aquaporin genes revealed several conserved gene clusters.

Comparative proteome analysis of Milnesium tardigradum in early embryonic state versus adults in active and anhydrobiotic state

Tardigrades have fascinated researchers for more than 300 years because of their extraordinary capability to undergo cryptobiosis and survive extreme environmental conditions. However, the survival mechanisms of tardigrades are still poorly understood mainly due to the absence of detailed knowledge about the proteome and genome of these organisms. Our study was intended to provide a basis for the functional characterization of expressed proteins in different states of tardigrades. High-throughput, high-accuracy proteomics in combination with a newly developed tardigrade specific protein database resulted in the identification of more than 3000 proteins in three different states: early embryonic state and adult animals in active and anhydrobiotic state. This comprehensive proteome resource includes protein families such as chaperones, antioxidants, ribosomal proteins, cytoskeletal proteins, transporters, protein channels, nutrient reservoirs, and developmental proteins. A comparative analysis of protein families in the different states was performed by calculating the exponentially modified protein abundance index which classifies proteins in major and minor components. This is the first step to analyzing the proteins involved in early embryonic development, and furthermore proteins which might play an important role in the transition into the anhydrobiotic state.

Effect of very high pressure on life of plants and animals

We studied the tolerance of living organisms, such as a small animal (Milnesium tardigradum), a small crustacean (Artemia), non-vascular plants or moss (Ptichomitrium and Venturiella), and a vascular plant (Trifolium) to the extremely high hydrostatic pressure of 7.5 GPa. It turned out that most of the high pressure exposed seeds of white clover were alive. Those exposed to 7.5 GPa for up to 1 day and seeded on agar germinated roots. Those exposed for up to 1 hour and seeded on soil germinated stems and leaves. Considering the fact that proteins begins to unfold around 0.3 GPa, it seems difficult to understand that all the living samples which have been investigated can survive after exposure to 7.5 GPa.

Integrated pathway modules using time-course metabolic profiles and EST data from Milnesium tardigradum

BackgroundTardigrades are multicellular organisms, resistant to extreme environmental changes such as heat, drought, radiation and freezing. They outlast these conditions in an inactive form (tun) to escape damage to cellular structures and cell death. Tardigrades are apparently able to prevent or repair such damage and are therefore a crucial model organism for stress tolerance. Cultures of the tardigrade Milnesium tardigradum were dehydrated by removing the surrounding water to induce tun formation. During this process and the subsequent rehydration, metabolites were measured in a time series by GC-MS. Additionally expressed sequence tags are available, especially libraries generated from the active and inactive state. The aim of this integrated analysis is to trace changes in tardigrade metabolism and identify pathways responsible for their extreme resistance against physical stress.ResultsIn this study we propose a novel integrative approach for the analysis of metabolic networks to identify modules of joint shifts on the transcriptomic and metabolic levels. We derive a tardigrade-specific metabolic network represented as an undirected graph with 3,658 nodes (metabolites) and 4,378 edges (reactions). Time course metabolite profiles are used to score the network nodes showing a significant change over time. The edges are scored according to information on enzymes from the EST data. Using this combined information, we identify a key subnetwork (functional module) of concerted changes in metabolic pathways, specific for de- and rehydration. The module is enriched in reactions showing significant changes in metabolite levels and enzyme abundance during the transition. It resembles the cessation of a measurable metabolism (e.g. glycolysis and amino acid anabolism) during the tun formation, the production of storage metabolites and bioprotectants, such as DNA stabilizers, and the generation of amino acids and cellular components from monosaccharides as carbon and energy source during rehydration.ConclusionsThe functional module identifies relationships among changed metabolites (e.g. spermidine) and reactions and provides first insights into important altered metabolic pathways. With sparse and diverse data available, the presented integrated metabolite network approach is suitable to integrate all existing data and analyse it in a combined manner.

Form and function of the feeding apparatus in Eutardigrada (Tardigrada)

Tardigrade feeding apparatus is a complex structure with considerable taxonomic significance that can be schematically divided into four parts: buccal ring, buccal tube, stylet system, and pharynx. We analyzed the fine morphology and the tridimensional organization of the tardigrade buccal–pharyngeal apparatus in order to clarify the relationships between form and function and to identify new characters for systematic and phylogenetic studies. We conducted a comparative analysis of the cuticular structures of the buccal–pharyngeal apparatuses of twelve eutardigrade species, integrating data obtained by SEM and LM observations. Morphological diversity was observed and new cuticular structures such as the stylet coat of the stylet system were identified. The synthesis of the buccal–pharyngeal apparatus during molting was also analyzed obtaining a clear developmental sequence of its resynthesis. These findings lead us to redefine the previous interpretations of the functioning mechanisms of the buccal–pharyngeal apparatus and provide a more specific relationship between tardigrade diet and the anatomy of their feeding apparatuses. In addition, the detection by energy-dispersive X-ray spectroscopy of calcium in the stylets, buccal tube, and placoids of eutardigrade species (i.e., Milnesium tardigradum, Paramacrobiotus richtersi) indicates that CaCO3 incrustations are not an exclusive feature of heterotardigrades and lead to suppose that this trait was present in the ancestors of both classes.

Redescriptions of three Milnesium Doyère, 1840 taxa (Tardigrada: Eutardigrada: Milnesiidae), including the nominal species for the genus

In this paper we redescribe two species (Milnesium eurystomum Maucci, 1991 and Milnesium tardigradum Doyère, 1840) and one subspecies (Milnesium tardigradum granulatum Ramazzotti, 1962), which we elevate to the species level. We establish a new type series for M. tardigradum sensu stricto along with the sequences of the cytochrome c oxidase subunit I (COI) and the internal transcribed spacer 2 region (ITS2). Moreover, we define two species groups within the genus based on the appearance of the dorso-lateral cuticle (the tardigradum and the granulatum group, with smooth and reticulated cuticle, respectively) and propose a system for denotation of the number of points on secondary branches of claws in the genus. We also provide a diagnostic key to all described Milnesium species and discuss the world distribution of the genus.