Echiniscoides Sigismundi Research Articles

New insights into osmobiosis and chemobiosis in tardigrades

Tardigrades are renowned for their ability to enter the extremotolerant state of latent life known as cryptobiosis. While it is widely accepted that cryptobiosis can be induced by freezing (cryobiosis) and by desiccation (anhydrobiosis), the latter involving formation of a so-called tun, the exact mechanisms underlying the state—as well as the significance of other cryptobiosis inducing factors—remain ambiguous. Here, we focus on osmotic and chemical stress tolerance in the marine tidal tardigrade Echiniscoides sigismundi. We show that E. sigismundi enters the tun state following exposure to saturated seawater and upon exposure to locality seawater containing the mitochondrial uncoupler DNP. The latter experiments provide evidence of osmobiosis and chemobiosis, i.e., cryptobiosis induced by high levels of osmolytes and toxicants, respectively. A small decrease in survival was observed following simultaneous exposure to DNP and saturated seawater indicating that the tardigrades may not be entirely ametabolic while in the osmobiotic tun. The tardigrades easily handle exposure to ultrapure water, but hypo-osmotic shock impairs tun formation and when exposed to ultrapure water the tardigrades do not tolerate DNP, indicating that tolerance towards dilute solutions involves energy-consuming processes. We discuss our data in relation to earlier and more contemporary studies on cryptobiosis and we argue that osmobiosis should be defined as a state of cryptobiosis induced by high external osmotic pressure. Our investigation supports the hypothesis that the mechanisms underlying osmobiosis and anhydrobiosis are overlapping and that osmobiosis likely represents the evolutionary forerunner of cryptobiosis forms that involve body water deprivation.

New marine heterotardigrade lineages (Echiniscoididae) from the tropics

Echiniscoididae are the only family of the clade Echiniscoidea residing in marine habitats. Their characteristic feature is the multiplication of claws on legs, commonly regarded as an adaptation to unstable tidal environment. Although DNA barcoding data indicated the presence of numerous unnamed candidate species within Echiniscoides about a decade ago, only recently have new species been formally described. In this paper, we present new genetic, morphological and morphometric data and establish five Echiniscoides species found in samples acquired from various tropical regions: Echiniscoides basalticus sp. nov. (Mauritius, Indian Ocean), Echiniscoides bufocephalus sp. nov. (Qatar, Indian Ocean), Echiniscoides lichenophilus sp. nov. (Dominican Republic, Atlantic Ocean), Echiniscoides musa sp. nov. (Brazil, Atlantic Ocean), and Echiniscoides trichosus sp. nov. (Rapa Nui, Pacific Ocean). We discuss the taxonomic importance of cuticular sculpturing in Echiniscoididae. Finally, we elevate all subspecies of Echiniscoides sigismundi to species level. The key to all echiniscoidids is provided. http://www.zoobank.org/urn:lsid:zoobank.org:pub:63154710-C3BA-402A-A11B-C211DAE59CAB

Diversity in Expression Biases of Lineage-Specific Genes During Development and Anhydrobiosis Among Tardigrade Species.

Lineage-specific genes can contribute to the emergence and evolution of novel traits and adaptations. Tardigrades are animals that have adapted to tolerate extreme conditions by undergoing a form of cryptobiosis called anhydrobiosis, a physical transformation to an inactive desiccated state. While studies to understand the genetics underlying the interspecies diversity in anhydrobiotic transitions have identified tardigrade-specific genes and family expansions involved in this process, the contributions of species-specific genes to the variation in tardigrade development and cryptobiosis are less clear. We used previously published transcriptomes throughout development and anhydrobiosis (5 embryonic stages, 7 juvenile stages, active adults, and tun adults) to assess the transcriptional biases of different classes of genes between 2 tardigrade species, Hypsibius exemplaris and Ramazzottius varieornatus. We also used the transcriptomes of 2 other tardigrades, Echiniscoides sigismundi and Richtersius coronifer, and data from 3 non-tardigrade species (Adenita vaga, Drosophila melanogaster, and Caenorhabditis elegans) to help identify lineage-specific genes. We found that lineage-specific genes have generally low and narrow expression but are enriched among biased genes in different stages of development depending on the species. Biased genes tend to be specific to early and late development, but there is little overlap in functional enrichment of biased genes between species. Gene expansions in the 2 tardigrades also involve families with different functions despite homologous genes being expressed during anhydrobiosis in both species. Our results demonstrate the interspecific variation in transcriptional contributions and biases of lineage-specific genes during development and anhydrobiosis in 2 tardigrades.

Comparative myoanatomy of Tardigrada: new insights from the heterotardigrades Actinarctus doryphorus (Tanarctidae) and Echiniscoides sigismundi (Echiniscoididae)

BackgroundTardigrada is a group of microscopic invertebrates distributed worldwide in permanent and temporal aquatic habitats. Famous for their extreme stress tolerance, tardigrades are also of interest due to their close relationship with Arthropoda and Cycloneuralia. Despite recent efforts in analyzing the musculature of a number of tardigrade species, data on the class Heterotardigrada remain scarce. Aiming to expand the current morphological framework, and to promote the use of muscular body plans in elucidating tardigrade phylogeny, the myoanatomy of two heterotardigrades, Actinarctus doryphorus and Echiniscoides sigismundi, was analyzed by cytochemistry, scanning electron and confocal laser scanning microscopy and 3D imaging. We discuss our findings with reference to other tardigrades and internal phylogenetic relationships of the phylum.ResultsWe focus our analyses on the somatic musculature, which in tardigrades includes muscle groups spanning dorsal, ventral, and lateral body regions, with the legs being musculated by fibers belonging to all three groups. A pronounced reduction of the trunk musculature is seen in the dorsoventrally compressed A. doryphorus, a species that generally has fewer cuticle attachment sites as compared to E. sigismundi and members of the class Eutardigrada. Interestingly, F-actin positive signals were found in the head appendages of A. doryphorus. Our analyses further indicate that cross-striation is a feature common to the somatic muscles of heterotardigrades and that E. sigismundi—as previously proposed for other echiniscoidean heterotardigrades—has relatively thick somatic muscle fibers.ConclusionsWe provide new insights into the myoanatomical differences that characterize distinct evolutionary lineages within Tardigrada, highlighting characters that potentially can be informative in future phylogenetic analyses. We focus our current analyses on the ventral trunk musculature. Our observations suggest that seven paired ventromedian attachment sites anchoring a large number of muscles can be regarded as part of the ground pattern of Tardigrada and that fusion and reduction of cuticular attachment sites is a derived condition. Specifically, the pattern of these sites differs in particular details between tardigrade taxa. In the future, a deeper understanding of the tardigrade myoanatomical ground pattern will require more investigations in order to include all major tardigrade lineages.

Modelling extreme desiccation tolerance in a marine tardigrade

It has recently been argued that the enigmatic tardigrades (water bears) will endure until the sun dies, surviving any astrophysical calamities in Earth’s oceans. Yet, our knowledge of stress tolerance among marine tardigrade species is very limited and most investigations revolve around species living in moist habitats on land. Here, we investigate desiccation tolerance in the cosmopolitan marine tidal tardigrade, Echiniscoides sigismundi, providing the first thorough analysis on recovery upon desiccation from seawater. We test the influence on survival of desiccation surface, time spent desiccated (up to 1 year) and initial water volume. We propose analysis methods for survival estimates, which can be used as a future platform for evaluating and analysing recovery rates in organisms subjected to extreme stress. Our data reveal that marine tidal tardigrades tolerate extremely rapid and extended periods of desiccation from seawater supporting the argument that these animals are among the toughest organisms on Earth.

Comparative Investigation of Copper Tolerance and Identification of Putative Tolerance Related Genes in Tardigrades.

Tardigrades are microscopic aquatic animals renowned for their tolerance toward extreme environmental conditions. The current study is the first to investigate their tolerance toward heavy metals and we present a novel tardigrade toxicant tolerance assay based on activity assessments as a measure of survival. Specifically, we compare tolerance toward copper in four species representing different evolutionary lineages, habitats and adaptation strategies, i.e., a marine heterotardigrade, Echiniscoides sigismundi, a limno-terrestrial heterotardigrade, Echiniscus testudo, a limno-terrestrial eutardigrade, Ramazzottius oberhaeuseri, and a marine eutardigrade, Halobiotus crispae. The latter was sampled at a time of year, when the population is predominantly represented by aberrant P1 cysts, while the other species were in normal active states prior to exposure. Based on volume measurements and a general relation between body mass and copper tolerance, expected tardigrade EC50 values were estimated at 0.5–2 μg l−1. Following 24 h of exposure, tolerance was high with no apparent link to lineage or habitat. EC50s (95% CI), 24 h after exposure, were estimated at 178 (168–186) and 310 (295–328) μg l−1, respectively, for E. sigismundi and R. oberhaeuseri, whereas E. testudo and H. crispae were less affected. Highest tolerance was observed in H. crispae with a mean ± s.e.m. activity of 77 ± 2% (n = 3) 24 h after removal from ~3 mg l−1 copper, suggesting that tardigrade cysts have increased tolerance toward toxicants. In order to identify putative tolerance related genes, an E. sigismundi transcriptome was searched for key enzymes involved in osmoregulation, antioxidant defense and copper metabolism. We found high expression of Na/K ATPase and carbonic anhydrase, known targets for copper. Our transcriptome, furthermore, revealed high expression of antioxidant enzymes, copper transporters, ATOX1, and a Cu-ATPase. In summary, our results indicate that tardigrades express well-known key osmoregulatory enzymes, supporting the hypothesis that copper inhibits sodium turnover as demonstrated for other aquatic organisms. Tardigrades, nevertheless, have high tolerance toward the toxicant, which is likely linked to high expression of antioxidant enzymes and an ability to enter dormant states. Tardigrades, furthermore, seem to have a well-developed battery of cuproproteins involved in copper homeostasis, providing basis for active copper sequestering and excretion.

Tolerance to Gamma Radiation in the Marine Heterotardigrade, Echiniscoides sigismundi.

Tardigrades belong to the most radiation tolerant animals on Earth, as documented by a number of studies using both low-LET and high-LET ionizing radiation. Previous studies have focused on semi-terrestrial species, which are also very tolerant to desiccation. The predominant view on the reason for the high radiation tolerance among these semi-terrestrial species is that it relies on molecular mechanisms that evolved as adaptations for surviving dehydration. In this study we report the first study on radiation tolerance in a marine tardigrade, Echiniscoides sigismundi. Adult specimens in the hydrated active state were exposed to doses of gamma radiation from 100 to 5000 Gy. The results showed little effect of radiation at 100 and 500 Gy but a clear decline in activity at 1000 Gy and higher. The highest dose survived was 4000 Gy, at which ca. 8% of the tardigrades were active 7 days after irradiation. LD50 in the first 7 days after irradiation was in the range of 1100–1600 Gy. Compared to previous studies on radiation tolerance in semi-terrestrial and limnic tardigrades, Echiniscoides sigismundi seems to have a lower tolerance. However, the species still fits into the category of tardigrades that have high tolerance to both desiccation and radiation, supporting the hypothesis that radiation tolerance is a by-product of adaptive mechanisms to survive desiccation. More studies on radiation tolerance in tardigrade species adapted to permanently wet conditions, both marine and freshwater, are needed to obtain a more comprehensive picture of the patterns of radiation tolerance.

Tun formation is not a prerequisite for desiccation tolerance in the marine tidal tardigradeEchiniscoides sigismundi

The so-called ‘tun’ state is best known from limno-terrestrial tardigrades and rotifers that rely on this compact body shape for anhydrobiotic survival. Little is known of tun formation in marine species and the evolutionary origin of the state is presently unknown. Here, we investigate desiccation tolerance and tun formation in the marine tidal echiniscoidean tardigrade, Echiniscoides sigismundi (M. Schultze, 1865). Groups of approximately 20 E. sigismundi sampled from Lynaes (Denmark) were dehydrated on filter paper from seawater as well as ultrapurified water and kept for 48 h at 5 °C, after which they were rehydrated in seawater. The activity and behaviour of the tardigrades was examined under a light microscope, whereas scanning electron microscopy was used for high-resolution three-dimensional imaging. When dehydrated from seawater, E. sigismundi enters a tun, however, when exposed to ultrapurified water, the tardigrade swells and becomes incapable of movement, and thus incapable of tun formation. Nonetheless, E. sigismundi tolerates being dehydrated from ultrapurified water, revealing an exceptional and unparalleled resilience towards losing structural integrity. Our results confirm previous investigations, which suggest that tun formation relies on a functional musculature. They further suggest that tun formation may have evolved as a response to elevated external pressure rather than desiccation per se.

First record of cysts in the tidal tardigrade Echiniscoides sigismundi

Tardigrades are microscopic metazoans that withstand environmental extremes by entering dormant states, such as cryptobiosis (latent life). In addition, they may also form cysts. Here, we present the first report of cyst formation in a marine heterotardigrade, i.e., Echiniscoides sigismundi, which constitutes a cryptic species complex present worldwide in tidal zones. The cysts were initially discovered during experimental series constructed to investigate osmotic stress tolerance. The animals, which eventually formed cysts, showed signs of an imminent molt at the beginning of experimentation. We use the term “cyst” for stages, where a total of three or more cuticles have been synthesized. Our observations show that encystment in E. sigismundi involves synthesizing of at least two new cuticle layers. Legs with discharged claws are present in connection with the first outer cuticle, as well as the second cuticular layer. In the most developed cyst, a third cuticle lacking claws seems to surround the animal, which is delineated by a fourth cuticle. Many features are shared with the well-studied cysts of eutardigrades. The cysts of E. sigismundi, however, lack pigmentation and have an extra set of claws, and the animal inside retains buccopharyngeal sclerified parts, until discharging the third cuticle. The finding of cysts in a marine heterotardigrade is novel and confirms that encystment also occurs within this major evolutionary lineage.

Phylogeography of North Atlantic intertidal tardigrades: refugia, cryptic speciation and the history of the Mid-Atlantic Islands

Aim To analyse the phylogeographical history of intertidal tardigrades in the North Atlantic in order to improve our understanding of geographical differentiation in microscopic organisms, and to understand the potential importance of the Mid-Atlantic Islands as stepping stones between the American and European coasts of the Atlantic Ocean. Location Twenty-four localities from the Mid-Atlantic Islands (Greenland, Iceland and the Faroe Islands) and both sides of the North Atlantic Ocean. Methods A mitochondrial marker (cytochrome c oxidase subunit I) was sequenced from individual tardigrades belonging to the genus Echiniscoides. The existence of cryptic species was detected using generalized mixed Yule coalescence analysis; lineage ages were estimated with relaxed clock methods; and the degree of geographical differentiation was analysed with samova analyses, haplotype networks and Mantel tests. Results Echiniscoides hoepneri, previously known only from Greenland, was recovered throughout the Mid-Atlantic Islands. The Faroe Islands population was isolated from Greenland and Iceland, but overall genetic variation was low. The morphospecies Echiniscoides sigismundi had high genetic variation and consisted of at least two cryptic species. A northern and a southern species were both recovered on both sides of the Atlantic, but only the northern species was found on the Mid-Atlantic Islands. The northern species showed signs of long-term isolation between the Western and Eastern Atlantic, despite the potential of the Mid-Atlantic islands to act as stepping-stones. There was no sign of long-term isolation in the southern species. The Mid-Atlantic individuals of the northern species were of Eastern Atlantic origin, but Greenland and Iceland showed signs of long-term isolation. The genetic pattern found in the southern species is not clearly geographical, and can probably be best explained by secondary contact between former isolated populations. Main conclusions North Atlantic intertidal tardigrades from the genus Echiniscoides showed strong geographical differentiation, and the Mid-Atlantic Islands seemed unimportant as stepping stones across the Atlantic. The geographical variation of the northern species of E. sigismundi suggests post-glacial recolonization from several refugia.

The spermatozoon of the Echiniscidae (Tardigrada, Heterotardigrada) and its phylogenetic significance

The spermatozoon ultrastructure of four species of moss-dwelling Heterotardigrada belonging to four genera of Echiniscidae, namely Pseudechiniscus juanitae, Echiniscus duboisi, Novechiniscus armadilloides and Antechiniscus parvisentus, was investigated. In all species, the testicular male gamete is similar in morphology and in length. The spermatozoon is made up of a long head, consisting of a cylindrical acrosome and an oval or rod-shaped nuclear region which contains a nucleus with osmiophilic and electron-dense chromatin, and a tapering tail, with a "9+2" axoneme. An elongated sack-like structure originates from the posterior part of the head, extending beyond the main axis of the cell and running parallel to the tail. It consists of two parallel tubular regions which sometimes form a strict double helix and contain two voluminous, "free" mitochondria with unmodified cristae. In addition, a voluminous vesicle is present laterally to the centriole or between the end of the nucleus and the beginning of the mitochondria, limited by two cytomembranes and filled with electron-lucent and granular material. The male gametes representative of these moss-dwelling Echiniscidae are very similar to the spermatozoa of the marine Echiniscoididae Echiniscoides sigismundi. This close similarity emphasises that habitat changes have had little influence on the organisation of the sperm cell representative of Echiniscoidea. Spermatozoon characters which could be useful for phylogenetic studies on Tardigrada are discussed.

A New Method for Making SEM Preparations of the Tardigrade Buccopharyngeal Apparatus

The buccopharyngeal apparatus of several species of tardigrades, including both heterotardigrades and eutardigrades, has been prepared for scanning electron microscopy (SEM) by using a new method developed and refined for this purpose. The buccalpharyngeal apparatus from three well-known species are described in detail and preliminary attempts to interpret the function of some of the structures are presented. The chosen species are used as examples and the new findings are compared to studies from the literature. The species used are: Echiniscoides sigismundi, Halobiotus crispae and Milnesium tardigradum. A redescription of the buccopharyngeal apparatus, in particular the stylets, the stylet supports, the stylet sheaths and their interrelationships, is presented for each species. For H. crispae the cuticular elements of the pharynx are also redescribed from this material. The potential for new information available with this method is discussed, together with the possible artifacts and difficulties the method may cause.

New records of marine Tardigrada in the Mediterranean Sea

Abstract Meiofauna from Chafarinas archipelago (western Mediterranean, N. Africa) were studied, and marine Tardigrada were identified to species level. A total of 15 species, belonging to families Stygarctidae, Halechiniscidae (Subfamilies Halechiniscinae, Florarctinae, Styraconyxinae and Tanarctinae), Batillipedidae, and Echiniscoididae, were found for the first time from the Alboran Sea and southwestern Mediterranean area. Four species and one subspecies are new records for the Mediterranean Sea: Floractus acer , F. asper , F. stellatus , Actinarctus doryphorus and Echiniscoides sigismundi hispaniensis . The male of Floractus stellatus , previously unknown to science, is described. Many species have been recorded for the first time in different sedimentary environments, such as midlittoral pools, very shallow gravels, pools in a mixed rocky environment or between Posidonia oceanica meadows, ‘Amphioxus’ gravels, and coastal detritic.

Marine Interstitial Heterotardigrada from the Pacific Coast of the United States, including a Description of Batillipes tridentatus n. sp.

Marine Interstitial Heterotardigrada from the Pacific Coast of the United States, including a Description of Batillipes tridentatus n. sp.

Structure and adaptation of the fauna in the black zone (littoral fringe) along rocky shores in northern Europe

The black zone within the littoral fringe of world-wide rocky shores is overgrown with lichens (Lichina pygmaea, Verrucaria maura), and blue-green algae. The zone is inhabited by a few coenophil animals: Hyadesia fusca (Acari), Telmatogeton japonicus (Chironomidae), Mniobia symbiotica (Rotatoria), Echiniscoides sigismundi (Tardigrada), and several aquatic or terrestrial intruders (Nematoda, Littorina, Copepoda-Harpacticoidea, Lrgia, Collembola, terrestrial mites and insects). The main adaptation of foundation species lies in their ability to reduce metabolism by anhydrobiosis or anaerobosis and to survive unfavourable circumstances in quiescence. Fauna1 structure is influenced by unpredictably changing abiotic factors such as moisture, salinity, temperature and eutrophication, which influence quantitative rather than qualitative aspects.

On Some British Marine Tardigrada, Including two New Species of Batillipes

INTRODUCTIONUntil recently only two marine members of the phylum Tardigrada were known to occur in Britain. Echiniscoides sigismundi (Schultze), was found associated with mussels (Green, 1950) and intertidal barnacles (Crisp & Hobart, 1954). The interstitial species, Batillipes mirus Richters, was located at beaches in North Wales (Boaden, 1963) and Strangford Lough in Northern Ireland (Boaden, 1966).Interstitial tardigrades have been reported from several localities around the North Sea. The fauna of the German coast is particularly well studied; Schultze, 1867; Richters, 1908, 1909; Schulz, 1935, 1951, 1953; Grell, 1937; Remane, 1940; Freidrich, 1963; Riemann, 1966a, b; Schmidt, 1969. This German fauna includes three species of Batillipes and single representatives of five other genera. In addition, E. sigismundi and B. mirus have been found in Norway (Tambs-Lyche, 1939–40). Along the French coast of the English Channel, eight species from six genera are reported. While most studies originated at RoscofT, including Cuénot, 1892; Cantacuzène, 1951; Guèrin, 1960; Swedmark, 1951, 1955, 1956a; Swedmark & Teissier, 1967; Thulin, 1942, two species of Batillipes have been located elsewhere on the northern coast of France (Renaud-Debyser & Salvat, 1963).In recent surveys of beaches in Britain, additional tardigrades have been located. Gray & Rieger (1971) reported their occurrence at several beaches on the north-east coast of Yorkshire, especially at Stoupe Beck, Robin Hood's Bay and Filey Beach near Scar-borough. Also, A. D. Mclntyre has found them in his collections at Firemore Bay in Loch Ewe on the west coast of Scotland. This paper describes specimens generously provided from these two studies.

Habits of the Marine Tardigrade, Echiniscoides sigismundi

IT would appear that, although tardigrades are often common, in many situations in fresh waters, they are comparatively rare, or at least have escaped frequent observation, in the sea. This is particularly true of British waters, for which only two records can, be traced. Both of them refer to the same species, Echiniscoides sigismundi (Schultze). The first record is that of Murray1 from the west coast of Ireland, and the second is that of Cole2 in 1934 from North Wales. In both instances the creature was found among seaweeds. Yet it is quite clear both from Marcus3 and Cuenot4 that E. sigismundi is widely distributed in the Mediterranean, North Sea and, Caribbean Sea, and even off the coasts of China. It is difficult to account for its apparent absence from most British coasts except on the supposition that it has not so far attracted attention.