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The microbial communities of sandy beaches are poorly described despite the biogeochemical importance and ubiquity of these ecosystems. Using metabarcoding of the 16S and 18S rRNA genes, we investigated the diversity, microhabitats (with or between sand grains) and intertidal distributions of microorganisms (including meiofauna) from pristine sandy beaches in British Columbia, Canada, and hypothesized that abiotic variations due to microhabitat or intertidal gradients influence the distribution of microorganisms on local scales. Bacterial, archaeal and protistan communities of the sand were clearly distinct from interstitial communities, and from planktonic communities of the overlying seawater, which correlated with differences in function and lifestyle (e.g., sulphur reduction and gliding motility). In contrast, meiofaunal communities could not be distinguished by sample type, suggesting that they are more frequently mobilized between these microhabitats. Across intertidal zones, high intertidal, mid intertidal and low intertidal/swash communities were distinct and correlated with moisture, organic carbon and phosphate content, implying that the distribution of microorganisms is influenced by intertidal abiotic gradients. However, few taxa at the genus or species level individually contributed to this zonation pattern; rather, a unique combination of multiple microbial taxa was probably responsible. Although significant differences in microbial community composition on sandy beaches can be attributed to microhabitat and intertidal gradients, further investigations are needed to assess community assembly processes, the consistency of these distributions, and the functions of the majority of the microorganisms observed in the sand and their effects on the biogeochemistry and ecology of sandy beaches.

Kleptoplasts (kP) are distinct among photosynthetic organelles in eukaryotes (i.e., plastids) because they are routinely sequestered from prey algal cells and function only temporarily in the new host cell. Therefore, the hosts of kleptoplasts benefit from photosynthesis without constitutive photoendosymbiosis. Here, we report that the euglenozoan Rapaza viridis has only kleptoplasts derived from a specific strain of green alga, Tetraselmis sp., but no canonical plastids like those found in its sister group, the Euglenophyceae. R. viridis showed a dynamic change in the accumulation of cytosolic polysaccharides in response to light–dark cycles, and 13 C isotopic labeling of ambient bicarbonate demonstrated that these polysaccharides originate in situ via photosynthesis; these data indicate that the kleptoplasts of R. viridis are functionally active. We also identified 276 sequences encoding putative plastid-targeting proteins and 35 sequences of presumed kleptoplast transporters in the transcriptome of R. viridis . These genes originated in a wide range of algae other than Tetraselmis sp., the source of the kleptoplasts, suggesting a long history of repeated horizontal gene transfer events from different algal prey cells. Many of the kleptoplast proteins, as well as the protein-targeting system, in R. viridis were shared with members of the Euglenophyceae, providing evidence that the early evolutionary stages in the green alga-derived secondary plastids of euglenophytes also involved kleptoplasty.

Dinoflagellates are a diverse protist group possessing many unique traits. These include (but are not limited to) expansive genomes packaged into permanently condensed chromosomes, photosynthetic or cryptic plastids acquired vertically or horizontally in serial endosymbioses, and a ruffle-like transverse flagellum attached along its length to the cell. When reconstructing character evolution, early branching lineages with unusual features that distinguish them from the rest of the group have proven useful for inferring ancestral states. The Noctilucales are one such lineage, possessing relaxed chromosomes in some life stages and a trailing, thread-like transverse flagellum. However, most of the cellular and molecular data for the entire group come from a single cultured species, Noctiluca scintillans, and because its phylogenetic position is unresolved it remains unclear if these traits are ancestral or derived. Here, we use single cell transcriptomics to characterize three diverse Noctilucales genera: Spatulodinium, Kofoidinium, and a new lineage, Fabadinium gen. nov. We also provide transcriptomes for undescribed species in Amphidinium and Abediniales, critical taxa for clarifying the phylogenetic position of Noctilucales. Phylogenomic analyses suggests that the Noctilucales are sister to Amphidinium rather than an independent branch outside the core dinoflagellates. This topology is consistent with observations of shared characteristics between some members of Noctilucales and Amphidinium and provides the most compelling evidence to date that the unusual traits within this group are derived rather than ancestral. We also confirm that Spatulodinium plastids are photosynthetic and of ancestral origin, and show that all non-photosynthetic Noctilucales retain plastid genes indicating a cryptic organelle.

This is a pre-print of an article published in Organisms Diversity and Evolution. The final authenticated version is available online at:https://link.springer.com/article/10.1007/s00435-021-00519-3 The post-print hasopen accessthrough the first author's web pagehttps://mariaherranzm.com/publications Open access to read-only published article: https://rdcu.be/ciOg9 (Provided by the Springer Nature SharedIt content-sharing initiative) This project received funding from the European Union’s Horizon 2020 research and innovation programme, under the Marie Sklodowska-Curie grant agreement No 797140 Abstract Aberrant kinorhynchs show several modifications deviating from the typical kinorhynch body plan, including a modified introvert with very elongated and flexible scalids, a weakly developed neck, and a slender trunk with less distinct segmentation. How these aberrant external features are reflected in the inner anatomy and how their aberrant body plan evolved are not understood. Here, we provide a comprehensive and comparative myoanatomical study of three putatively, distantly related worm-like species:Cateria styx,Franciscideres kalenesos andZelinkaderes yong. Despite the weak external segmentation of the trunk, the studied species show a distinct segmental arrangement of the musculature. However, this arrangement is shifted posteriorly with respect to the external segmentation, because the extremely thin and flexible cuticle is lacking the apodeme-like cuticular thickenings (pachycycli) where the longitudinal muscles usually attach. The muscular arrangement in the three species is overall similar, yet,C. styxshows most resemblance to the allomalorhagidF. kalenesos, whereas the cyclorhagidZ. yongdiffers in several ways. This suggests a closer relationship ofC. styxto Allomalorhagida. Whereas most kinorhynchs prefer muddy sediments, both the allomalorhagid and cyclorhagid worm-like kinorhynchs are mainly found in sandy environments, suggesting that a flexible, slender body evolved at least twice independently as an adaptation to the interstitial environment.

Species of rhabdocoels within the family Dalyelliidae (Platyhelminthes) generally inhabit fresh water and are rare in marine and brackish environments. Here we describe a new species of marine daly...

Marine gastrotrichs of the Pacific Ocean are poorly known. Here, we report on the finding of a marine chaetonotid gastrotrich of the genus Diuronotus from an intertidal beach within the Sea of Japan in Hokkaido (Japan). The Japanese individual shows a very close resemblance to Diuronotus aspetos. This new record is a consequential extension of its biogeographic range; previous records for representatives of this genus are confined to West Greenland, the North Sea and the east coast of North America. This rarely encountered, but seemingly widespread genus of marine gastrotrichs exemplifies our limited understanding of meiofaunal diversity and distribution patterns caused by sampling bias and insufficient knowledge on nominal species complexes.

Dinoflagellates possess many cellular characteristics with unresolved evolutionary histories. These include nuclei with greatly expanded genomes and chromatin packaged using histone-like proteins and dinoflagellate-viral nucleoproteins instead of histones, highly reduced mitochondrial genomes with extensive RNA editing, a mix of photosynthetic and cryptic secondary plastids, and tertiary plastids. Resolving the evolutionary origin of these traits requires understanding their ancestral states and early intermediates. Several early-branching dinoflagellate lineages are good candidates for such reconstruction, however these cells tend to be delicate and environmentally sparse, complicating such analyses. Here, we employ transcriptome sequencing from manually isolated and microscopically documented cells to resolve the placement of two cells of one such genus, Abedinium, collected by remotely operated vehicle in deep waters off the coast of Monterey Bay, CA. One cell corresponds to the only described species, Abedinium dasypus, whereas the second cell is distinct and formally described as Abedinium folium, sp. nov. Abedinium has classically been assigned to the early-branching dinoflagellate subgroup Noctilucales, which is weakly supported by phylogenetic analyses of small subunit ribosomal RNA, the single characterized gene from any member of the order. However, an analysis based on 221 proteins from the transcriptome places Abedinium as a distinct lineage, separate from and basal to Noctilucales and the rest of the core dinoflagellates. The transcriptome also contains evidence of a cryptic plastid functioning in the biosynthesis of isoprenoids, iron–sulfur clusters, and heme, a mitochondrial genome with all three expected protein-coding genes (cob, cox1, and cox3), and the presence of some but not all dinoflagellate-specific chromatin packaging proteins.

The vast majority of eukaryotic life is made up of single cells commonly referred to as protists. In this primer, Leander provides an introduction to predatory protists - cells that eat other cells. This lifestyle, in particular the use of phagocytosis, makes endosymbiosis possible and enabled the evolution of complex cells.

7 pages, 2 figures, supporting information https://doi.org/10.1111/1462-2920.14733, The application of metabarcoding to study animal‐associated microeukaryotes has been restricted because the universal barcode used to study microeukaryotic ecology and distribution in the environment, the Small Subunit of the Ribosomal RNA gene (18S rRNA), is also present in the host. As a result, when host‐associated microbial eukaryotes are analysed by metabarcoding, the reads tend to be dominated by host sequences. We have done an in silico validation against the SILVA 18S rRNA database of a non‐metazoan primer set (primers that are biased against the metazoan 18S rRNA) that recovers only 2.6% of all the metazoan sequences, while recovering most of the other eukaryotes (80.4%). Among metazoans, the non‐metazoan primers are predicted to amplify 74% of Porifera sequences, 4% of Ctenophora, and 15% of Cnidaria, while amplifying almost no sequences within Bilateria. In vivo, these non‐metazoan primers reduce significantly the animal signal from coral and human samples, and when compared against universal primers provide at worst a 2‐fold decrease in the number of metazoan reads and at best a 2800‐fold decrease. This easy, inexpensive, and near‐universal method for the study of animal‐associated microeukaryotes diversity will contribute to a better understanding of the microbiome, This work was funded by a grant from the Canadian Institutes for Health Research (MOP‐42517). JdC was supported by a grant from the Tula Foundation to the Centre for Microbial Biodiversity and Evolution and the Marie Curie International Outgoing Fellowship FP7‐PEOPLE‐2012‐IOF‐331450 CAARL

A comprehensive morphological and taxonomic account of the members of the genus Cheliplana de Beauchamp, 1927 is presented. Six new species are described: Cheliplana asinaraensis n. sp., C. cubana n. sp., C. curacaoensis n. sp., C. hawaiiensis n. sp., C. longissima n. sp. and C. mauii n. sp. The new species are mainly distinguished from each other and from other representatives of Cheliplana by the organisation of the reproductive system and the structure of the cirrus. Furthermore, C. triductibus Van Steenkiste, Volonterio, Schockaert & Artois, 2008 is considered a junior synonym of Cheliplana deverticula Ax, 2008. The two subspecies of Cheliplana asica Marcus, 1952, C. asica asica and C. asica terminalis Brunet, 1968, are considered separate species. The systematic position of the genus Dactyloplana Armonies, 2018 is discussed, and its synonymy with Cheliplana is retained. As such, this brings the total number of species of Cheliplana to 49. Finally, we provide an identification key to the members of the genus, based on characters that enable identification to species level in the field.

We describe new insights into the morphology and life history of the bizarre parasite Haplozoon axiothellae (Dinoflagellata) using light microscopy (LM), scanning electron microscopy (SEM), and confocal laser scanning microscopy (CLSM). Trophonts were isolated from the intestines of host maldanid polychaetes, Axiothella rubrocincta, collected from San Juan Island, Washington, USA. LM and SEM confirmed features previously observed, such as amphiesmal projections, mature and immature junctions between the nucleated compartments of the vermiform syncytium and visible polygonal alveoli. CLSM of adult trophonts fluorescently stained for DNA, tubulin, centrin, and plasma membrane demonstrated several new ultrastructural traits: (1) an extensive basket of parallel microtubules within the trophomere used for host attachment, (2) two physically separated MTOCs (i.e. putative pairs of basal bodies) beneath pores on the ventral side of each compartment, (3) robust mitotic and/or meiotic spindles associated with one to four nuclei in each compartment, (4) spindles with polar bodies that are disconnected from the MTOCs, (5) a centrin-stained fibril within the trophomeres that potentially functions to retract the motile stylet, and (6) cytokinesis in the posterior-most compartments. This study renames haplozoan compartments using the suffix "-mere" rather than "-cyte" (i.e. trophomere, gonomere, sporomere) to reflect their status within a single syncytium.

The taxon Koinocystididae is the third most species-rich family within Eukalyptorhynchia. However, its diversity and phylogeny have been largely neglected in former studies. We introduce three new genera and twelve new species of Koinocystididae including Simplexcystis asymmetrica gen. n. sp. n., Galapagetula cubensis sp. n., eight species of Reinhardorhynchus gen. n. and two species of Itaipusa. This raises the total number of species within Koinocystididae from 51 to 63. We also report on new distribution records for six known species: I. divae (Cuba, Panama and New Caledonia), I. karlingi (Sardinia and Lanzarote), Reinhardorhynchus riegeri comb. n. (Cuba), R. ruffinjonesi comb. n. (Cuba and Panama), Utelga heinckei (Cuba and Lanzarote), and U. pseudoheinckei (Sardinia). Simplexcystis asymmetrica gen. n. sp. n. is characterised by a male duct running eccentrically through the copulatory bulb, lack of any hard structures in the male system, lack of a bursa, and the fact that the epithelia of the female, the male, and part of the common atrium are covered by a brush border. Galapagetula cubensis sp. n. has a caudal gonopore, a divisa-type copulatory bulb with an unarmed penis papilla, and a female duct without a sphincter. The new species of Itaipusa and Reinhardorhynchus gen. n. differ from their congeners in the detailed structure of the copulatory bulb and especially the hard structures associated with it. In a molecular phylogenetic analysis based on all available 18S and 28S rDNA sequences of koinocystidids, we found support for the monophyly of the family and the genus Utelga Marcus, 1949. The genus Itaipusa is not monophyletic in that I. sinensis forms a clade with Rhinolasius dillonicus, while other species of Itaipusa that have a copulatory bulb armed with hooks form a clade together with Sekerana stolzi. As the type species of Itaipusa (I. divae) is in neither of these clades, we erected a new genus for I. sinensis (Koinogladius gen. n.) and one for species of Itaipusa having a hook-bearing copulatory bulb (Reinhardorhynchus gen. n.), respectively. Whether the remaining species of Itaipusa form a monophylum remains uncertain. The taxon Koinocystididae is the third most species-rich family within Eukalyptorhynchia. However, its diversity and phylogeny have been largely neglected in former studies. We introduce three new genera and twelve new species of Koinocystididae including Simplexcystis asymmetrica gen. n. sp. n., Galapagetula cubensis sp. n., eight species of Reinhardorhynchus gen. n. and two species of Itaipusa. This raises the total number of species within Koinocystididae from 51 to 63. We also report on new distribution records for six known species: I. divae (Cuba, Panama and New Caledonia), I. karlingi (Sardinia and Lanzarote), Reinhardorhynchus riegeri comb. n. (Cuba), R. ruffinjonesi comb. n. (Cuba and Panama), Utelga heinckei (Cuba and Lanzarote), and U. pseudoheinckei (Sardinia). Simplexcystis asymmetrica gen. n. sp. n. is characterised by a male duct running eccentrically through the copulatory bulb, lack of any hard structures in the male system, lack of a bursa, and the fact that the epithelia of the female, the male, and part of the common atrium are covered by a brush border. Galapagetula cubensis sp. n. has a caudal gonopore, a divisa-type copulatory bulb with an unarmed penis papilla, and a female duct without a sphincter. The new species of Itaipusa and Reinhardorhynchus gen. n. differ from their congeners in the detailed structure of the copulatory bulb and especially the hard structures associated with it. In a molecular phylogenetic analysis based on all available 18S and 28S rDNA sequences of koinocystidids, we found support for the monophyly of the family and the genus Utelga Marcus, 1949. The genus Itaipusa is not monophyletic in that I. sinensis forms a clade with Rhinolasius dillonicus, while other species of Itaipusa that have a copulatory bulb armed with hooks form a clade together with Sekerana stolzi. As the type species of Itaipusa (I. divae) is in neither of these clades, we erected a new genus for I. sinensis (Koinogladius gen. n.) and one for species of Itaipusa having a hook bearing copulatory bulb (Reinhardorhynchus gen. n.), respectively. Whether the remaining species of Itaipusa form a monophylum remains uncertain.

20 pages, 10 figures, 1 table, supporting information https://doi.org/10.1111/jpy.12984, Marine benthic dinoflagellates are interesting not only because some epiphytic genera can cause harmful algal blooms but also for understanding dinoflagellate evolution and diversification. Our understanding of their biodiversity is far from complete, and many thecate genera have unusual tabulation patterns that are difficult to relate to the diverse known phytoplankton taxa. A new sand-dwelling genus, Pachena gen. nov., is described based on morphological and DNA sequence data. Three species were discovered in distant locations and are circumscribed, namely, P. leibnizii sp. nov. from Canada, P. abriliae sp. nov. from Spain, and P. meriddae sp. nov. from Italy. All species are tiny (about 9–23 μm long) and heterotrophic. Species are characterized by their tabulation (APC 4′ 3a 6′′ 5c 5s 5′′′ 2′′′′), an apical hook covering the apical pore, an ascending cingulum, and a sulcus with central list. The first anterior intercalary plate is uniquely “sandwiched” between two plates. The species share these features and differ in the relative sizes and arrangements of their plates, especially on the epitheca. The ornamentation of thecal plates is species-specific. The new molecular phylogenies based on SSU and LSU rDNA sequences contribute to understanding the evolution of the planktonic relatives of Pachena, the Thoracosphaeraceae, This work was supported by a postdoctoral research salary to MH and AY from the Assembling the Tree of Life grant (NSF #EF‐0629624) and operating funds to BSL from the National Science and Engineering Research Council of Canada (NSERC 2019‐03986); AR thanks R. Gallisai (ICM‐CSIC) and T. Slámová (Univ. Prague, Czech Republic) for their help during samplings and samples processing. AR was funded by a MECD grant “Estancia de Movilidad en el extranjero José Castillejo” (CAS17/00237), a “Senckenberg Taxonomy Grant 2017″, and a DAAD “Research Stays for University Academics and Scientists 2018” Grant (91644317). CTS thanks Prof. Antonella Lugliè for the continuous and important economic and scientific support. CTS was funded by a DAAD Grant within the “Research Stays for University Academics and Scientists 2014” Grant (A/14/01530), With the funding support of the ‘Severo Ochoa Centre of Excellence’ accreditation (CEX2019-000928-S), of the Spanish Research Agency (AEI)

Kalyptorhynchs are abundant members of meiofaunal communities worldwide, but knowledge on their overall species diversity and distribution is poor. Here we report twenty species of eukalyptorhynchs...

We isolated Plagiodinium belizeanum into clonal culture from the Pacific island of Okinawa (Japan) and characterized it using a combination of light microscopy, scanning electron microscopy, transmission electron microscopy and 18S/28S ribosomal (r) gene sequences. Although molecular phylogenetic analyses of 18S rDNA and 28S rDNA sequences were unable to resolve the phylogenetic position of P. belizeanum within dinoflagellates, the ultrastructural data provided some new traits for the species. For instance, double-membrane-bound vesicles, distinct from the mitochondria, were interpreted as autolysosomes containing electron-dense virus particles. The thecal plate pattern was Po 1′ 0a 5′′ 5(6)C 4S 5′′′ 0p 1′′′′, which is slightly different from the original description in having an additional epithecal plate and four sulcal plates. The laterally flattened cells were 22–34 μm long, 11–13 μm deep, and 15–18 μm wide and contained a peridinin-type plastid with lobes radiating from a central pyrenoid t...

Euglenids are a well-known group of single-celled eukaryotes, with phototrophic, osmotrophic and phagotrophic members. Phagotrophs represent most of the phylogenetic diversity of euglenids, and gave rise to the phototrophs and osmotrophs, but their evolutionary relationships are poorly understood. Symbiontids, in contrast, are anaerobes that are alternatively inferred to be derived euglenids, or a separate euglenozoan group. Most phylogenetic studies of euglenids have examined the SSU rDNA only, which is often highly divergent. Also, many phagotrophic euglenids (and symbiontids) are uncultured, restricting collection of other molecular data. We generated transcriptome data for 28 taxa, mostly using a single-cell approach, and conducted the first multigene phylogenetic analyses of euglenids to include phagotrophs and symbiontids. Euglenids are recovered as monophyletic, with symbiontids forming an independent branch within Euglenozoa. Spirocuta, the clade of flexible euglenids that contains both the phototrophs (Euglenophyceae) and osmotrophs (Aphagea), is robustly resolved, with the ploeotid Olkasia as its sister group, forming the new taxon Olkaspira. Ploeotids are paraphyletic, although Ploeotiidae (represented by Ploeotia spp.), Lentomonas, and Keelungia form a robust clade (new taxon Alistosa). Petalomonadida branches robustly as sister to other euglenids in outgroup-rooted analyses. Within Spirocuta, Euglenophyceae is a robust clade that includes Rapaza, and Anisonemia is a well-supported monophyletic group containing Anisonemidae (Anisonema and Dinema spp.), ‘Heteronema II’ (represented by H. vittatum), and a clade of Neometanema plus Aphagea. Among ‘peranemid’ phagotrophs, Chasmostoma branches with included Urceolus, and Peranema with the undescribed ‘Jenningsia II’, while other relationships are weakly supported and consequently the closest sister group to Euglenophyceae remains unresolved. Our results are inconsistent with recent inferences that Entosiphon is the evolutionarily pivotal sister either to other euglenids, or to Spirocuta. At least three transitions between posterior and anterior flagellar gliding occurred in euglenids, with the phylogenetic positions and directions of those transitions remaining ambiguous.

Gregarine apicomplexans are unicellular parasites commonly found in the intestines and coeloms of invertebrate hosts. Traits associated with the conspicuous feeding stage of gregarines, known as the trophozoite, have been used in combination with molecular phylogenetic data for species delimitation and the reconstruction of evolutionary history. Trophozoite morphology alone is often inadequate for inferring phylogenetic relationships and delimiting species due to frequent cases of high intraspecific variation combined with relatively low interspecific variation. The current study combined morphological data with small subunit (SSU) rDNA sequences to describe and establish two novel marine gregarine species isolated from the intestine of a polychaete host Lumbrineris inflata collected in British Columbia (Canada): Paralecudina anankea n. sp. and Lecudina caspera n. sp. The sister species to the host is Lumbrineris japonica, which can be found on the opposite side of the Pacific Ocean (Japan) and contains two different species of gregarine parasites: Paralecudina polymorpha and Lecudina longissima. Molecular phylogenetic analyses placed P. anankea n. sp. as the sister species to P. polymorpha and L. caspera n. sp. as the sister species to L. longissima. This phylogenetic pattern demonstrates a co-evolutionary history whereby speciation of the host (Lumbrineris) corresponds with simultaneous speciation of the two different lineages of intestinal gregarines (Paralecudina and Lecudina). This article is protected by copyright. All rights reserved.

Surveys of kinorhynch diversity in the northeastern Pacific Ocean are rare, and only eight species have been described from the region so far. We explored the diversity of kinorhynchs from a pristine coastal habitat of British Columbia (Calvert Island) and discovered a new subtidal species, Echinoderes hakaiensis sp. nov. We also redescribed one of the oldest described echinoderid species, E. pennaki. Both species were characterized with high-resolution light microscopy (LM), scanning electron microscopy (SEM), and DNA sequences from the mitochondrial cytochrome c oxidase subunit 1 (COI) gene. Echinoderes hakaiensis sp. nov. can be differentiated from other species by a unique pattern of spines and tubes (i.e., four pairs of tubes on segment 2 combined with three middorsal spines). Echinoderes pennaki shows a configuration of traits on segment 2 that questions the reliability of conventional morphological characters used to distinguish different genera within the Echinoderidae. Reevaluation of these characters will require a resolved molecular phylogenetic context including a more comprehensive sampling of species within the group.

Two monospecific genera of marine benthic dinoflagellates, Adenoides and Pseudadenoides, have unusual thecal tabulation patterns (lack of cingular plates in the former; and no precingular plates and a complete posterior intercalary plate series in the latter) and are thus difficult to place within a phylogenetic framework. Although both genera share morphological similarities, they have not formed sister taxa in previous molecular phylogenetic analyses. We discovered and characterized a new species of Pseudadenoides, P. polypyrenoides sp. nov., at both the ultrastructural and molecular phylogenetic levels. Molecular phylogenetic analyses of SSU and LSU rDNA sequences demonstrated a close relationship between P. polypyrenoides sp. nov. and Pseudadenoides kofoidii, and Adenoides and Pseudadenoides formed sister taxa in phylogenetic trees inferred from LSU rDNA sequences. Comparisons of morphological traits, such as the apical pore complex (APC), demonstrated similarities between Adenoides, Pseudaden...

Denis,It’s ironic that when facing a problem like this – being asked to write a piece about the way you affected us in the labs we share – our first instinct would be to come and talk to you, to ge...

Since its discovery in the mid-1950'ies, Cateria has been an enigmatic kinorhynch genus due to its aberrant worm-like shape and extremely thin cuticle. However, the rare occurrence of the species, only found in sandy intertidal habitats, and the poor preservation of the type material have hampered detailed studies of the genus over time. Now, sixty years after the original description of Cateria styx, we present an extensive morphological and functional study based on new material collected from its type locality in Macae, Brazil. We combine live observations with detailed scanning electron microscopy data, new light microscopy material, confocal laser scanning microscopy and three-dimensional rendering. These observations show that C. styx displays a complex array of cuticular structures (spines, spinoscalids and extraordinarily complex cuticular ornamentation) that we interpret to be adaptations for mechanical adhesion, through friction and interlocking, in an interstitial habitat; the enigmatic dorsal organ, is a hydrostatic structure which function is inferred to be adhesive. Additional morphological traits in C. styx include: extremely elongated primary spinoscalids that cannot be completely retracted in the trunk; a reduced number of spinoscalids; sixteen elongated hairy patches in the introvert; fifteen trichoscalids that vary in length; the absence of a neck; dorsal spines being alternatingly displaced to either left or right side paradorsal positions; and high intraspecific variation in the number and position of glandular openings in the trunk.

Background The Scalidophora (Kinorhyncha, Loricifera and Priapulida) have an important phylogenetic position as early branching ecdysozoans, yet the architecture of their nervous organ systems is notably underinvestigated. Without such information, and in the absence of a stable phylogenetic context, we are inhibited from producing adequate hypotheses about the evolution and diversification of ecdysozoan nervous systems. Here, we utilize confocal laser scanning microscopy to characterize serotonergic, tubulinergic and FMRFamidergic immunoreactivity patterns in a comparative neuroanatomical study with three species of Echinoderes, the most speciose, abundant and diverse genus within Kinorhyncha. Results Neuroanatomy in Echinoderes as revealed by acetylated α-tubulin immunoreactivity includes a circumpharyngeal brain and ten neurite bundles in the head region that converge into five longitudinal nerves within the trunk. The ventral nerve cord is ganglionated, emerging from the brain with two connectives that converge in trunk segments 2–3, and diverge again within segment 8. The longitudinal nerves and ventral nerve cord are connected by two transverse neurites in segments 2–9. Differences among species correlate with the number, position and innervation of cuticular structures along the body. Patterns of serotoninergic and FMRFamidergic immunoreactivity correlate with the position of the brain neuropil and the ventral nerve cord. Distinct serotonergic and FMRFamidergic somata are associated with the brain neuropil and specific trunk segments along the ventral nerve cord. Conclusions Neural architecture is highly conserved across all three species, suggesting that our results reveal a pattern that is common to more than 40% of the species within Kinorhyncha. The nervous system of Echinoderes is segmented along most of the trunk; however, posterior trunk segments exhibit modifications that are likely associated with sensorial, motor or reproductive functions. Although all kinorhynchs show some evidence of an externally segmented trunk, it is unclear whether external segmentation matches internal segmentation of nervous and muscular organ systems across Kinorhyncha, as we observed in Echinoderes. The neuroanatomical data provided in this study not only expand the limited knowledge on kinorhynch nervous systems but also establish a comparative morphological framework within Scalidophora that will support broader inferences about the evolution of neural architecture among the deepest branching lineages of the Ecdysozoa. Electronic supplementary material The online version of this article (10.1186/s12862-019-1405-4) contains supplementary material, which is available to authorized users.

Although the dispersal ability of kinorhynchs is known to be limited, the distribution of certain kinorhynch species appears to extend over vast geographical areas. Combining molecular phylogenetic data with biogeographical investigations can test this paradox by discerning cryptic species with restricted distributions from species with potentially large geographical distributions. In this paper, we (1) redescribe two species of kinorhynchs ( Echinoderes ohtsukai and E. kozloffi ) found in the northeastern Pacific Ocean using molecular and morphological data and (2) provide the first evidence for a disjunct geographical distribution in kinorhynchs that is consistent with the introduction of an invasive species. Although we collected E. ohtsukai from the northeastern Pacific Ocean (British Columbia, Canada), this species was originally described from Japan. We demonstrated that specimens of E. ohtsukai collected from Japan and British Columbia have identical DNA sequences for the mitochondrial cytochrome c oxidase subunit 1 (COI) gene. These results are most consistent with a recent introduction of this species into one of the habitats on the opposite side of the Pacific Ocean through human-mediated dispersal.

The eugregarines are a group of apicomplexan parasites that mostly infect the intestines of invertebrates. The high level of morphological variation found within and among species of eugregarines makes it difficult to find consistent and reliable traits that unite even closely related lineages. Based mostly on traits observed with light microscopy, the majority of described eugregarines from marine invertebrates has been classified into a single group, the Lecudinidae. Our understanding of the overall diversity and phylogenetic relationships of lecudinids is very poor, mainly because only a modest amount of exploratory research has been done on the group and very few species of lecudinids have been characterized at the molecular phylogenetic level. In an attempt to understand the diversity of marine gregarines better, we surveyed lecudinids that infect the intestines of Pacific ascidians (i.e. sea squirts) using ultrastructural and molecular phylogenetic approaches; currently, these species fall within one genus,Lankesteria. We collected lecudinid gregarines from six ascidian host species, and our data demonstrated that each host was infected by a different species ofLankesteria: (i)Lankesteria hesperidiiformissp. nov., isolated fromDistaplia occidentalis, (ii)Lankesteria metandrocarpaesp. nov., isolated fromMetandrocarpa taylori, (iii)Lankesteria halocynthiaesp. nov., isolated fromHalocynthia aurantium, (iv)Lankesteria herdmaniaesp. nov., isolated fromHerdmania momus, (v)Lankesteriacf.ritterellae, isolated fromRitterella rubra, and (vi)Lankesteria didemnisp. nov., isolated fromDidemnum vexillum. Visualization of the trophozoites with scanning electron microscopy showed that four of these species were covered with epicytic folds, whereas two of the species were covered with a dense pattern of epicytic knobs. The molecular phylogenetic data suggested that species ofLankesteriawith surface knobs form a clade that is nested within a paraphyletic assemblage species ofLankesteriawith epicytic folds.

Multicellularity is often considered a prerequisite for morphological complexity, as seen in the camera-type eyes found in several groups of animals. A notable exception exists in single-celled eukaryotes called dinoflagellates, some of which have an eye-like ‘ocelloid’ consisting of subcellular analogues to a cornea, lens, iris, and retina1. These planktonic cells are uncultivated and rarely encountered in environmental samples, obscuring the function and evolutionary origin of the ocelloid. Here we show, using a combination of electron microscopy, tomography, isolated-organelle genomics, and single-cell genomics, that ocelloids are built from pre-existing organelles, including a cornea-like layer made of mitochondria and a retinal body made of anastomosing plastids. We find that the retinal body forms the central core of a network of peridinin-type plastids, which in dinoflagellates and their relatives originated through an ancient endosymbiosis with a red alga2. As such, the ocelloid is a chimaeric structure, incorporating organelles with different endosymbiotic histories. The anatomical complexity of single-celled organisms may be limited by the components available for differentiation, but the ocelloid shows that pre-existing organelles can be assembled into a structure so complex that it was initially mistaken for a multicellular eye3. Although mitochondria and plastids are acknowledged chiefly for their metabolic roles, they can also be building blocks for greater structural complexity.

Although environmental DNA surveys improve our understanding of biodiversity, interpretation of unidentified lineages is limited by the absence of associated morphological traits and living cultures. Unidentified lineages of marine stramenopiles are called "MAST clades". Twenty-five MAST clades have been recognized: MAST-1 through MAST-25; seven of these have been subsequently discarded because the sequences representing those clades were found to either (1) be chimeric or (2) affiliate within previously described taxonomic groups. Eighteen MAST clades remain without a cellular identity. Moreover, the discarded "MAST-13" has been used in different studies to refer to two different environmental sequence clades. After establishing four cultures representing two different species of heterotrophic stramenopiles and then characterizing their morphology and molecular phylogenetic positions, we determined that the two different species represented the two different MAST-13 clades: (1) a lorica-bearing Bicosoeca kenaiensis and (2) a microaerophilic flagellate previously named "Cafeteria marsupialis". Both species were previously described with only light microscopy; no cultures, ultrastructural data or DNA sequences were available from these species prior to this study. The molecular phylogenetic position of three different "C. marsupialis" isolates was not closely related to the type species of Cafeteria; therefore, we established a new genus for these isolates, Cantina gen. nov.

Increasing evidence suggests that many widespread species of meiofauna are in fact regional complexes of (pseudo-)cryptic species. This knowledge has challenged the ‘Everything is Everywhere’ hypothesis and also partly explains the meiofauna paradox of widespread nominal species with limited dispersal abilities. Here, we investigated species diversity within the marine microturbellarian Astrotorhynchus bifidus sensu lato in the Northeast Pacific Ocean. We used a multiple-evidence approach combining multi-gene (18S, 28S, COI) phylogenetic analyses, several single-gene and multi-gene species delimitation methods, haplotype networks and conventional taxonomy to designate Primary Species Hypotheses (PSHs). This included the development of rhabdocoel-specific COI barcode primers, which also have the potential to aid in species identification and delimitation in other rhabdocoels. Secondary Species Hypotheses (SSHs) corresponding to morphospecies and pseudo-cryptic species were then proposed based on the minimum consensus of different PSHs. Our results showed that (a) there are at least five species in the A. bifidus complex in the Northeast Pacific Ocean, four of which can be diagnosed based on stylet morphology, (b) the A. bifidus complex is a mixture of sympatric and allopatric species with regional and/or subglobal distributions, (c) sympatry occurs on local (sample sites), regional (Northeastern Pacific) and subglobal (Northern Atlantic, Arctic, Northeastern Pacific) scales. Mechanisms for this co-occurrence are still poorly understood, but we hypothesize they could include habitat differentiation (spatial and/or seasonal) and life history characteristics such as sexual selection and dispersal abilities. Our results also suggest the need for improved sampling and exploration of molecular markers to accurately map gene flow and broaden our understanding of species diversity and distribution of microturbellarians in particular and meiofauna in general.

Beyond the potent stings of jellyfish, nematocysts have evolved independently in single-celled phytoplankton., We examine the origin of harpoon-like secretory organelles (nematocysts) in dinoflagellate protists. These ballistic organelles have been hypothesized to be homologous to similarly complex structures in animals (cnidarians); but we show, using structural, functional, and phylogenomic data, that nematocysts evolved independently in both lineages. We also recorded the first high-resolution videos of nematocyst discharge in dinoflagellates. Unexpectedly, our data suggest that different types of dinoflagellate nematocysts use two fundamentally different types of ballistic mechanisms: one type relies on a single pressurized capsule for propulsion, whereas the other type launches 11 to 15 projectiles from an arrangement similar to a Gatling gun. Despite their radical structural differences, these nematocysts share a single origin within dinoflagellates and both potentially use a contraction-based mechanism to generate ballistic force. The diversity of traits in dinoflagellate nematocysts demonstrates a stepwise route by which simple secretory structures diversified to yield elaborate subcellular weaponry.

Summary In order to obtain insights into the evolution of colorless (apochlorotic) diatoms, we investigated newly established apochlorotic strains of Nitzschia spp. using light and electron microscopy and molecular phylogenetic analyses. Fluorescence microscopic observations demonstrated that the apochlorotic diatoms lack chlorophylls. Transmission electron microscopy of two apochlorotic strains also demonstrated that their plastids lacked thylakoids; instead, having four-membrane-bound organelles without thylakoids, similar to nonphotosynthetic plastid remnants. From the apochlorotic strains, we also found plastid small subunit rRNA genes that were unusually long branched in phylogenetic analyses, as observed in other nonphotosynthetic plastids. Molecular phylogenetic analysis of the nucleus-encoded large subunit rRNA genes showed eight distinct lineages for apochlorotic diatoms. The eight apochlorotic lineages were not monophyletic, suggesting that the loss of photosynthesis took place multiple times independently within Nitzschia. Several diatoms, including Nitzschia spp., are mixotrophic, which is an expected mode of nutrition that would help explain the evolutionary switch from a photosynthetic lifestyle to a heterotrophic lifestyle.

Dicyemids are enigmatic parasites found within the excretory systems of benthic cephalopods. The phylogenetic position and overall diversity of dicyemids remains poorly understood, in part because current species delimitation criteria are based solely on morphological traits. Understanding the diversity of parasite species is particularly problematic because they tend to be devoid of consistent (informative) morphological traits while simultaneously rich in morphological variation associated with developmental stages and environmental conditions. In this study, we tested the boundaries of currently described morphospecies of dicyemids using molecular phylogenetic data. Variation within sequences of the small subunit (18S) rRNA gene was explored because this marker (1) is known to be fast-evolving in parasitic eukaryotes, (2) is one of the few molecular markers to have been previously sequenced in some dicyemids, and (3) has been used successfully as a DNA barcode in other groups of parasites. Thre...

Gregarines are a diverse group of apicomplexan parasites with a conspicuous extracellular feeding stage, called a "trophozoite", that infects the intestines and other body cavities of invertebrate hosts. Although the morphology of trophozoites is very diverse in gregarines as a whole, high degrees of intraspecific variation combined with relatively low degrees of interspecific variation make the delimitation of different species based on trophozoite morphology observed with light microscopy difficult. The coupling of molecular phylogenetic data with comparative morphology has shed considerable light onto the boundaries and interrelationships of different gregarine species. In this study, we isolated a novel marine gregarine from the hepatic region of a Pacific representative of the hemichordate Glossobalanus minutus, and report the first ultrastructural and molecular data from any gregarine infecting this distinctive group of hosts. Molecular phylogenetic analyses of an SSU rDNA sequence derived from two single-cell isolates of this marine gregarine demonstrated a strong and unexpected affiliation with a clade of terrestrial gregarines (e.g. Gregarina). This molecular phylogenetic data combined with a comparison of the morphological features in previous reports of gregarines collected from Atlantic representatives of G. minutus justified the establishment of a new binomial for the new isolate, namely Caliculium glossobalani n. gen. et sp. The molecular phylogenetic analyses demonstrated a clade of terrestrial gregarines associated with a sequence acquired from a marine species, which suggest that different groups of terrestrial/freshwater gregarines evolved independently from marine ancestors.

The discovery and characterization of protist communities from diverse environments are crucial for understanding the overall evolutionary history of life on earth. However, major questions about the diversity, ecology, and evolutionary history of protists remain unanswered, notably because data obtained from natural protist communities, especially of heterotrophic species, remain limited. In this review, we discuss the challenges associated with "field protistology", defined here as the exploration, characterization, and interpretation of microbial eukaryotic diversity within the context of natural environments or field experiments, and provide suggestions to help fill this important gap in knowledge. We also argue that increased efforts in field studies that combine molecular and microscopical methods offer the most promising path toward (1) the discovery of new lineages that expand the tree of eukaryotes; (2) the recognition of novel evolutionary patterns and processes; (3) the untangling of ecological interactions and functions, and their roles in larger ecosystem processes; and (4) the evaluation of protist adaptations to a changing climate.

The spaces between sand grains are home to a myriad of microscopic marine eukaryotes, including kalyptorhynch rhabdocoels equipped with an eversible proboscis that enables them to capture microscopic prey living in these environments. The structure of the kalyptorhynch proboscis separates the two major subclades within the group: the Schizorhynchia (bifurcated proboscis) and the Eukalyptorhynchia (unbranched proboscis). A survey of meiofaunal metazoans in the Pacific north-west led to the discovery of three new schizorhynch species (Undicola tofinoensisgen. nov., sp. nov.,Schizorhinos vancouverensisgen. nov., sp. nov. andLinguabana tulaigen. nov., sp. nov.) and two new eukalyptorhynch species (Thinodactylaina tlaoquiahtensisgen. nov., sp. nov. andRostracilla nuuchahnulthensisgen. nov., sp. nov.). This survey also recovered the putative cosmopolitan eukalyptorhynch (Polycystididae)Gyratrix hermaphroditusEhrenberg, 1831. We performed molecular phylogenetic analyses on 18S rDNA sequences from all five novel isolates and from all available kalyptorhynch species in GenBank. The molecular data supported the monophyly of the Eukalyptorhynchia and Schizorhynchia and helped demonstrate the boundaries between different species within the Kalyptorhynchia.

Dinoflagellates are key species in marine environments, but they remain poorly understood in part because of their large, complex genomes, unique molecular biology, and unresolved in-group relationships. We created a taxonomically representative dataset of dinoflagellate transcriptomes and used this to infer a strongly supported phylogeny to map major morphological and molecular transitions in dinoflagellate evolution. Our results show an early-branching position of Noctiluca, monophyly of thecate (plate-bearing) dinoflagellates, and paraphyly of athecate ones. This represents unambiguous phylogenetic evidence for a single origin of the group's cellulosic theca, which we show coincided with a radiation of cellulases implicated in cell division. By integrating dinoflagellate molecular, fossil, and biogeochemical evidence, we propose a revised model for the evolution of thecal tabulations and suggest that the late acquisition of dinosterol in the group is inconsistent with dinoflagellates being the source of this biomarker in pre-Mesozoic strata. Three distantly related, fundamentally nonphotosynthetic dinoflagellates, Noctiluca, Oxyrrhis, and Dinophysis, contain cryptic plastidial metabolisms and lack alternative cytosolic pathways, suggesting that all free-living dinoflagellates are metabolically dependent on plastids. This finding led us to propose general mechanisms of dependency on plastid organelles in eukaryotes that have lost photosynthesis; it also suggests that the evolutionary origin of bioluminescence in nonphotosynthetic dinoflagellates may be linked to plastidic tetrapyrrole biosynthesis. Finally, we use our phylogenetic framework to show that dinoflagellate nuclei have recruited DNA-binding proteins in three distinct evolutionary waves, which included two independent acquisitions of bacterial histone-like proteins.

Summary The apicomplexans are a group of obligate animal pathogens that include Plasmodium (malaria), Toxoplasma (toxoplasmosis), and Cryptosporidium (cryptosporidiosis) [ 1 ]. They are an extremely diverse and specious group but are nevertheless united by a distinctive suite of cytoskeletal and secretory structures related to infection, called the apical complex, which is used to recognize and gain entry into animal host cells. The apicomplexans are also known to have evolved from free-living photosynthetic ancestors and retain a relict plastid (the apicoplast), which is non-photosynthetic but houses a number of other essential metabolic pathways [ 2 ]. Their closest relatives include a mix of both photosynthetic algae (chromerids) and non-photosynthetic microbial predators (colpodellids) [ 3 ]. Genomic analyses of these free-living relatives have revealed a great deal about how the alga-parasite transition may have taken place, as well as origins of parasitism more generally [ 4 ]. Here, we show that, despite the surprisingly complex origin of apicomplexans from algae, this transition actually occurred at least three times independently. Using single-cell genomics and transcriptomics from diverse uncultivated parasites, we find that two genera previously classified within the Apicomplexa, Piridium and Platyproteum, form separately branching lineages in phylogenomic analyses. Both retain cryptic plastids with genomic and metabolic features convergent with apicomplexans. These findings suggest a predilection in this lineage for both the convergent loss of photosynthesis and transition to parasitism, resulting in multiple lineages of superficially similar animal parasites.

Dinoflagellates are some of the most common eukaryotic cells in the ocean, but have very unusual nuclei. Many exhibit a form of closed mitosis (dinomitosis) wherein the nuclear envelope (NE) invaginates to form one or more trans-nuclear tunnels. Rather than contact spindles directly, the chromatids then bind to membrane-based kinetochores on the NE. To better understand these unique mitotic features, we reconstructed the nuclear architecture of Polykrikos kofoidii in 3D using focused ion beam scanning electron microscopy (FIB-SEM) in conjunction with high-pressure freezing, freeze-substitution, TEM, and confocal microscopy. We found that P. kofoidii possessed six nuclear tunnels, which were continuous with a reticulating network of membranes that has thus far gone unnoticed. These membranous extensions interconnect the six tunnels while ramifying throughout the nucleus to form a “nuclear net.” To our knowledge, the nuclear net is the most elaborate endomembrane structure described within a nucleus. Our findings demonstrate the utility of tomographic approaches for detecting 3D membrane networks and show that nuclear complexity has been underestimated in Polykrikos kofoidii and, potentially, in other dinoflagellates.

A novel parasitic ciliate Fusiforma themisticola n. gen., n. sp. was discovered infecting 4.4% of the hyperiid amphipod Themisto libellula. Ciliates were isolated from a formaldehyde-fixed whole amphipod and the DNA was extracted for amplification of the small subunit (SSU) rRNA gene. Sequence and phylogenetic analyses showed unambiguously that this ciliate is an apostome and about 2% divergent from the krill-infesting apostome species assigned to the genus Pseudocollinia. Protargol silver impregnation showed a highly unusual infraciliature for an apostome. There are typically 8 (6-9) bipolar somatic kineties covering the banana-shaped body. The anterior end of the oral cavity begins about 1/3 of the body length from the anterior end and is composed of an inpocketing that is lined on its anterior and left wall with an oral field of densely packed ciliated kinetosomes. Stomatogenesis begins with some dedifferentiation of the parental oral field and elongation of its paroral and oral kineties. A new oral field develops midventrally and the paroral and oral kineties break to form the oral apparatus of the opisthe, which completes development by additional kinetosomal proliferation and migration of the paroral. This morphology is novel among apostomes and justifies the establishment of a new genus and species.