Ice-inhabiting species of Bdelloidea Rotifera reveal a pre-Quaternary ancestry in the Arctic cryosphere.

Historical climate data indicate that the Earth has passed through multiple geological periods with much warmer-than-present climates, including epochs of the Miocene (23-5.3 mya BP) with temperatures 3-4°C above present, and more recent interglacial stages of the Quaternary, for example, Marine Isotope Stage 11c (approx. 425-395 ka BP) and Middle Holocene thermal maximum (7.5-4.2 ka BP), during which continental glaciers may have melted entirely. Such warm periods would have severe consequences for ice-obligate fauna in terms of their distribution, biodiversity and population structure. To determine the impacts of these climatic events in the Nordic cryosphere, we surveyed ice habitats throughout mainland Norway and Svalbard ranging from maritime glaciers to continental ice patches (i.e. non-flowing, inland ice subjected to deep freezing overwinter), finding particularly widespread populations of ice-inhabiting bdelloid rotifers. Combined mitochondrial and nuclear DNA sequencing identified approx. 16 undescribed, species-level rotifer lineages that revealed an ancestry predating the Quaternary (> 2.58 mya). These rotifers also displayed robust freeze/thaw tolerance in laboratory experiments. Collectively, these data suggest that extensive ice refugia, comparable with stable ice patches across the contemporary Norwegian landscape, persisted in the cryosphere over geological time, and may have facilitated the long-term survival of ice-obligate Metazoa before and throughout the Quaternary.

Genome assembly of Pseudomonas sp. strain SED1T, a psychrotolerant bacterium isolated from Deception Glacier (Washington, USA).

Strain SED1T was isolated from glacial samples collected on Mount Deception, Washington, USA. Genome sequencing and assembly identified a DNA G + C content of 60.4 mol% with 6,125 predicted proteins. Analysis by the Type Strain Genome Server is consistent with the isolate representing a previously undescribed species in the genus Pseudomonas.

Transcriptomic Profiling at the Maternal-to-Zygotic Transition in Leech, Helobdella austinensis.

The glossiphoniid leech, Helobdella austinensis, is an experimentally tractable member of the superphylum, Lophotrochozoa. Its large embryonic cells, stereotyped asymmetric cell divisions and ex vivo development capabilities makes it a favorable model for studying the molecular and cellular events of a representative spiralian. In this study, we focused on a narrow developmental time window of ~6-8 h, comprising stages just prior to and immediately following zygote deposition. Employing RNA-Seq methodology, we identified differentially expressed transcripts at this fundamental ontogenic boundary, known as the maternal-to-zygotic transition (MZT). Gene expression changes were characterized by the massive degradation of maternal RNAs (~45%) coupled with the rapid transcription of ~5000 zygotic genes (~20% of the genome) in the first mitotic cell cycle. The latter transcripts encoded a mixture of cell maintenance and regulatory proteins that predictably influence downstream developmental events.

Biocontrol of snail-borne parasites with the glossiphoniid leech, Helobdella austinensis.

Parasites in which freshwater snails are intermediate hosts pose a serious threat to human health worldwide. We show here that freshwater snails can potentially be controlled by leech predation; in principle, this approach could significantly reduce snail-borne parasitic diseases (SBPDs). Specifically, glossiphoniid leeches, Helobdella austinensis and congener species, consume freshwater snails indiscriminately, while other common leeches do not. A single adult H. austenensis, for example, can consume up to its weight in snails, e.g. Physella acuta, per day. Our predator-prey models suggest that snail populations could be eliminated in relatively short time periods (approximately six months) using a leech biocontrol approach. This could have considerable impact on global SBPDs by breaking the intermediate host life cycle.

Two new tardigrade genera from New Zealand's Southern Alp glaciers display morphological stasis and parallel evolution.

Tardigrada is an invertebrate phylum that often constitutes a dominant micrometazoan group on glaciers worldwide. We investigated tardigrades residing in surface ice above the equilibrium line altitude (ELA) on three temperate glaciers of New Zealand's Southern Alps. Morphological, morphometric and multilocus DNA analyses (CO1, 18S rRNA, 28S rRNA, ITS-2) revealed two new genera comprising four species, of which two are formally described here: Kopakaius gen. nov. nicolae sp. nov. and Kararehius gen. nov. gregorii sp. nov. The former is represented by three genetically distinct phyletic lineages akin to species. According to CO1, Kopakaius gen. nov. nicolae sp. nov. inhabits Whataroa Glacier only while the remaining two Kopakaius species occur on Fox and Franz Joseph Glaciers, suggesting low dispersal capabilities. Although morphological characteristics of the new genera could indicate affinity with the subfamily Itaquasconinae, phylogenetic analysis placed them confidently in the subfamily Diphasconinae. Kopakaius gen. nov. lack placoids in the pharynx similar with some Itaquasconinae, whereas dark pigmentation and claw shape aligns them with the glacier-obligate genus, Cryobiotus (subfamily Hypsibiinae), which is an example of parallel evolution. The second genus, Kararehius gen nov. could be classified as Adropion-like (subfamily Itaquasconinae), but differs greatly by genetics (placed in the subfamily Diphasconinae) as well as morphology (e.g., lack of septulum), exemplify deep stasis in Hypsibiidae. Our results suggest that glacier fragmentation during the Pleistocene triggered tardigrade speciation, making it a suitable model for studies on allopatric divergence in glacier meiofauna.

The Glacier Ice Worm, Mesenchytraeus solifugus, Elevates Mitochondrial Inorganic Polyphosphate (PolyP) Levels in Response to Stress.

The inorganic polymer, polyphosphate (polyP), is present in all organisms examined to date with putative functions ranging from the maintenance of bioenergetics to stress resilience and protein homeostasis. Bioenergetics in the glacier-obligate, segmented worm, Mesenchytraeus solifugus, is characterized by a paradoxical increase in intracellular ATP levels as temperatures decline. We show here that steady-state, mitochondrial polyP levels vary among species of Annelida, but were elevated only in M. solifugus in response to thermal stress. In contrast, polyP levels decreased with temperature in the mesophilic worm, Enchytraeus crypticus. These results identify fundamentally different bioenergetic strategies between closely related annelid worms, and suggest that I worm mitochondria maintain ATP and polyP in a dynamic equilibrium.

A living bdelloid rotifer from 24,000-year-old Arctic permafrost.

In natural, permanently frozen habitats, some organisms may be preserved for hundreds to tens of thousands of years. For example, stems of Antarctic moss were successfully regrown from an over millennium-old sample covered by ice for about 400 years1. Likewise, whole campion plants were regenerated from seed tissue preserved in relict 32,000-year-old permafrost2, and nematodes were revived from the permafrost of two localities in northeastern Siberia, with source sediments dated over 30,000 years BP3. Bdelloid rotifers, microscopic multicellular animals, are known for their ability to survive extremely low temperatures4. Previous reports suggest survival after six to ten years when frozen between -20° to 0°C4-6. Here, we report the survival of an obligate parthenogenetic bdelloid rotifer, recovered from northeastern Siberian permafrost radiocarbon-dated to ∼24,000 years BP. This constitutes the longest reported case of rotifer survival in a frozen state. We confirmed the finding by identifying rotifer actin gene sequences in a metagenome obtained from the same sample. By morphological and molecular markers, the discovered rotifer belongs to the genus Adineta, and aligns with a contemporary Adineta vaga isolate collected in Belgium. Experiments demonstrated that the ancient rotifer withstands slow cooling and freezing (∼1°C min-1) for at least seven days. We also show that a clonal culture can continuously reproduce in the laboratory by parthenogenesis.

Five animal phyla in glacier ice reveal unprecedented biodiversity in New Zealand's Southern Alps.

Glacier ice is an extreme environment in which most animals cannot survive. Here we report the colonization of high elevation, climate-threatened glaciers along New Zealand's southwestern coast by species of Arthropoda, Nematoda, Platyhelminthes, Rotifera and Tardigrada. Based on DNA barcoding and haplotype-inferred evidence for deep genetic variability, at least 12 undescribed species are reported, some of which have persisted in this niche habitat throughout the Pleistocene. These findings identify not only an atypical biodiversity hotspot but also highlight the adaptive plasticity of microinvertebrate Animalia.

Biophysical mechanism for zinc as an anticancer agent.

The experimental observation that an increase in calcium above micromolar concentrations results in a slowing or stopping of anaphase-A motion is evidence for an electrostatic mechanism for poleward mitotic chromosome motions. Specifically, higher concentrations of doubly-charged calcium ions screen negative charges at microtubule free "plus" ends at kinetochores and at centrosomes. These structures normally interact with positive charges at kinetochores and positively charged microtubule free ends vicinal to centrosomes to generate poleward force. As with calcium ions, doubly-charged zinc cations can also shield these negative charges, thereby interfering with force generation for anaphase-A chromosome motion, aborting mitosis. Experimental evidence reveals that dysregulation of free cytosolic zinc homeostasis contributes to cancerous transformation. Treatment of cancers by increasing zinc concentration has unknowingly been accomplished by utilizing zinc ionophores to facilitate zinc transport across the plasma membrane, revealing an inverse relationship between malaria incidence - and malaria treatment with zinc ionophores - and cancer mortality. Here we hypothesize a biophysical mechanism for cancer therapy employing zinc supplementation enhanced by zinc ionophores.

Key rules of life and the fading cryosphere: Impacts in alpine lakes and streams.

Alpine regions are changing rapidly due to loss of snow and ice in response to ongoing climate change. While studies have documented ecological responses in alpine lakes and streams to these changes, our ability to predict such outcomes is limited. We propose that the application of fundamental rules of life can help develop necessary predictive frameworks. We focus on four key rules of life and their interactions: the temperature dependence of biotic processes from enzymes to evolution; the wavelength dependence of the effects of solar radiation on biological and ecological processes; the ramifications of the non-arbitrary elemental stoichiometry of life; and maximization of limiting resource use efficiency across scales. As the cryosphere melts and thaws, alpine lakes and streams will experience major changes in temperature regimes, absolute and relative inputs of solar radiation in ultraviolet and photosynthetically active radiation, and relative supplies of resources (e.g., carbon, nitrogen, and phosphorus), leading to nonlinear and interactive effects on particular biota, as well as on community and ecosystem properties. We propose that applying these key rules of life to cryosphere-influenced ecosystems will reduce uncertainties about the impacts of global change and help develop an integrated global view of rapidly changing alpine environments. However, doing so will require intensive interdisciplinary collaboration and international cooperation. More broadly, the alpine cryosphere is an example of a system where improving our understanding of mechanistic underpinnings of living systems might transform our ability to predict and mitigate the impacts of ongoing global change across the daunting scope of diversity in Earth's biota and environments.

Detailed ultrastructure of the Hirudo (Annelida: Hirudinea) salivary gland.

Medical leeches have been widely used in medical applications and treatments for millennia. Studies on the salivary glands of blood-sucking leeches have focused on their bioactive secretions and mechanisms of action, with little attention to ultrastructure. In this study, we examined dissected embryonic and adult Hirudo verbana salivary glands by scanning electron microscopy (SEM). Gland cells of embryos were physically separated while adults displayed highly developed cell bunches in which each cell was connected to others by fine channels. Channels from each bunch combined to form a larger canal that opened to the jaw. Secreted material from these glands prevent blood from clotting and allow the adult to feed while sucking blood.

Structural Evolution of the Glacier Ice Worm Fo ATP Synthase Complex.

The segmented annelid worm, Mesenchytraeus solifugus, is a permanent resident of temperate, maritime glaciers in the Pacific northwestern region of North America, displaying atypically high intracellular ATP levels which have been linked to its unusual ability to thrive in hydrated glacier ice. We have shown previously that ice worms contain a highly basic, carboxy terminal extension on their ATP6 regulatory subunit, likely acquired by horizontal gene transfer from a microbial dietary source. Here we examine the full complement of F1F0 ATP synthase structural subunits with attention to non-conservative, ice worm-specific structural modifications. Our genomics analyses and molecular models identify putative proton shuttling domains on either side of the F0 hemichannel, which predictably function to enhance proton flow across the mitochondrial membrane. Other components of the ice worm ATP synthase complex have remained largely unchanged in the context of Metazoan evolution.

Correct spelling of a recently described species of Helobdella (Annelida: Clitellata: Hirudinea).

In a recently published article (Saglam et al. 2018) the name of a new species of Hirudinea (Annelida, Clitellata) was simultaneously published in three different ways, as Helobdella serendipitious (Saglam et al. 2018: 61, 70, 71, 73), Helobdella serendipitous (ibid.: 63, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74), and as Helobdella serendipidous (ibid.: 75). Acting as first reviser according to Article 24 of the International Code of Zoological Nomenclature (ICZN 1999) here we determine serendipitous as the correct spelling of the species epitheton.

Disappearing Kilimanjaro snow-Are we the last generation to explore equatorial glacier biodiversity?

Glaciation accompanied our human ancestors in Africa throughout the Pleistocene. Regrettably, equatorial glaciers and snow are disappearing rapidly, and we are likely the last generation who will get to know these peculiar places. Despite the permanently harsh conditions of glacier/snow habitats, they support a remarkable diversity of life ranging from bacteria to animals. Numerous papers have been devoted to microbial communities and unique animals on polar glaciers and high mountains, but only two reports relate to glacial biodiversity in equatorial regions, which are destined to melt completely within the next few decades. Equatorial glaciers constitute "cold islands" in tropics, and discovering their diversity might shed light on the biogeography, dispersal, and history of psychrophiles. Thus, an opportunity to protect biota of equatorial glaciers hinges on ex situ conservation. It is timely and crucial that we should investigate the glacial biodiversity of the few remaining equatorial glaciers.

Long-distance dispersal, ice sheet dynamics and mountaintop isolation underlie the genetic structure of glacier ice worms.

Disentangling the contemporary and historical factors underlying the spatial distributions of species is a central goal of biogeography. For species with broad distributions but little capacity to actively disperse, disconnected geographical distributions highlight the potential influence of passive, long-distance dispersal (LDD) on their evolutionary histories. However, dispersal alone cannot completely account for the biogeography of any species, and other factors-e.g. habitat suitability, life history-must also be considered. North American ice worms ( Mesenchytraeus solifugus) are ice-obligate annelids that inhabit coastal glaciers from Oregon to Alaska. Previous studies identified a complex biogeographic history for ice worms, with evidence for genetic isolation, unexpectedly close relationships among geographically disjunct lineages, and contemporary migration across large (e.g. greater than 1500 km) areas of unsuitable habitat. In this study, we analysed genome-scale sequence data for individuals from most of the known ice worm range. We found clear support for divergence between populations along the Pacific Coast and the inland flanks of the Coast Mountains (mean FST = 0.60), likely precipitated by episodic ice sheet expansion and contraction during the Pleistocene. We also found support for LDD of ice worms from Alaska to Vancouver Island, perhaps mediated by migrating birds. Our results highlight the power of genomic data for disentangling complex biogeographic patterns, including the presence of LDD.

Atypical Evolution of the F1Fo Adenosine Triphosphate Synthase Regulatory ATP6 subunit in Glacier Ice Worms (Annelida: Clitellata: Mesenchytraeus).

The glacier ice worm, Mesenchytraeus solifugus, is among a few animals that reside permanently in glacier ice. Their adaptation to cold temperature has been linked to relatively high intracellular adenosine triphosphate (ATP) levels, which compensate for reductions in molecular motion at low physiological temperatures. Here, we show that ATP6-the critical regulatory subunit of the F1Fo-ATP synthase and primary target of mitochondrial disease-acquired an unprecedented histidine-rich, 18-amino acid carboxy-terminal extension, which counters the strong evolutionary trend of mitochondrial genome compaction. Furthermore, sequence analysis suggests that this insertion is not of metazoan origin, but rather is a product of horizontal gene transfer from a microbial dietary source, and may act as a proton shuttle to accelerate the rate of ATP synthesis.

Phylogenetic and morphological resolution of the Helobdella stagnalis species-complex (Annelida: Clitellata: Hirudinea).

The glossiphoniid freshwater leech, Helobdella stagnalis, was described by Linnaeus 1758 based on common European specimens. The presence of a brown, chitinous scute on the dorsal-anterior surface, as observed on leeches elsewhere in the world, has generally led to the classification of all scute-bearing members of the genus as H. stagnalis. Here we describe the morphology and behavior of the type species from Europe, and analyze H. stagnalis-like specimens collected worldwide. We present evidence for at least four distinct scute-bearing Helobdella species that can be morphologically resolved. Maximum Parsimony (MP) and Bayesian Inference (BI) analyses at the mitochondrial cytochrome c oxidase subunit I (COI) locus further supported this notion, with divergence values suggesting a mid-Miocene ancestor of this successful group of cosmopolitan hirudineans. New species, Helobdella echoensis nov. sp., Helobdella eriensis nov. sp., and Helobdella serendipitious nov. sp., are described, based on morphological, anatomical and molecular data. Current distribution patterns of Helobdella spp. suggest a robust, global dispersal mechanism, as well as local pockets of endemism.

Phylogeny and cocoon production in the parasitic leech Myzobdella lugubris Leidy, 1851 (Hirudinidae, Piscicolidae).

Myzobdella lugubris is a commensal leech on crustaceans and a parasite to fishes, surviving predominantly in brackish waters throughout North America. Specimens in this study were collected within the tidal zone of the Delaware River basin (New Jersey and Pennsylvania). To compare regional M. lugubris specimens, defined characters were scored after dorsal and ventral dissections, and phylogenetic relationships were resolved using cytochrome c oxidase subunit 1 (CO1), 12S ribosomal RNA (rDNA) and 18S rDNA gene fragments. Variance between regional populations was low, suggesting recent dispersal events and/or strong evolutionary constraints. The reproductive biology of M. lugubris was explored by quantitative analysis of secreted cocoons. Specimens produced 32.67 ± 4.50 cocoons with fertilization ratios of 88.1% and hatching times of 48 ± 7 days at 17°C under laboratory conditions. At 22°C, 46 ± 28 cocoons were produced with fertilization ratios of 70.27% and hatching times of 28 ± 5 days. Surprisingly, each cocoon supported only one embryo, which is unusual among oligochaetes.

Elaborate ultrastructure of the Hirudo (Annelida: Hirudinae) cocoon surface.

Species of medicinal leeches (Hirudo medicinalis, H. verbana, and H. sulukii) secrete hard-shelled cocoons. When initially deposited, a cocoon is surrounded by a foam. Over a short time, the foam is transformed into a three-dimensional structure. We show here that this peripheral structure likely forms by the solidification and dehydration of a moderately viscous, proteinaceous substance that surrounds bubbles of various sizes. The resulting matrix-like structure comprises a network of curved branches juxtaposed at ∼120° and taper in width as a function of distance from the outer cocoon wall. The material is proteinaceous, and traps environmental material in its composition, especially silicon. The geometry of compartments and abundance of silicon on branch surfaces suggest a mechanism for trapping water to prevent desiccation in a terrestrial environment.

Punctuated invasion of water, ice, snow and terrestrial ecozones by segmented worms (Oligochaeta: Enchytraeidae: Mesenchytraeus).

Segmented worms (Annelida) are among the most successful animal inhabitants of extreme environments worldwide. An unusual group of enchytraeid oligochaetes of genus Mesenchytraeus are abundant in the Pacific northwestern region of North America and occupy geographically proximal ecozones ranging from low elevation rainforests and waterways to high altitude glaciers. Along this altitudinal transect, Mesenchytraeus representatives from disparate habitat types were collected and subjected to deep mitochondrial and nuclear phylogenetic analyses. Our data identify significant topological discordance among gene trees, and near equivalent interspecific divergence levels indicative of a rapid radiation event. Collectively, our results identify a Mesenchytraeus 'explosion' coincident with mountain building in the Pacific northwestern region that gave rise to closely related aquatic, ice, snow and terrestrial worms.