Patterns of host-parasite associations between marine meiofaunal flatworms (Platyhelminthes) and rhytidocystids (Apicomplexa).

Microturbellarians are abundant and ubiquitous members of marine meiofaunal communities around the world. Because of their small body size, these microscopic animals are rarely considered as hosts for parasitic organisms. Indeed, many protists, both free-living and parasitic ones, equal or surpass meiofaunal animals in size. Despite several anecdotal records of "gregarines", "sporozoans", and "apicomplexans" parasitizing microturbellarians in the literature-some of them dating back to the nineteenth century-these single-celled parasites have never been identified and characterized. More recently, the sequencing of eukaryotic microbiomes in microscopic invertebrates have revealed a hidden diversity of protist parasites infecting microturbellarians and other meiofaunal animals. Here we show that apicomplexans isolated from twelve taxonomically diverse rhabdocoel taxa and one species of proseriate collected in four geographically distinct areas around the Pacific Ocean (Okinawa, Hokkaido, and British Columbia) and the Caribbean Sea (Curaçao) all belong to the apicomplexan genus Rhytidocystis. Based on comprehensive molecular phylogenies of Rhabdocoela and Proseriata inferred from both 18S and 28S rDNA sequences, as well as a molecular phylogeny of Marosporida inferred from 18S rDNA sequences, we determine the phylogenetic positions of the microturbellarian hosts and their parasites. Multiple lines of evidence, including morphological and molecular data, show that at least nine new species of Rhytidocystis infect the microturbellarian hosts collected in this study, more than doubling the number of previously recognized species of Rhytidocystis, all of which infect polychaete hosts. A cophylogenetic analysis examining patterns of phylosymbiosis between hosts and parasites suggests a complex picture of overall incongruence between host and parasite phylogenies, and varying degrees of geographic signals and taxon specificity.

Microscopic marine invertebrates are reservoirs for cryptic and diverse protists and fungi.

BACKGROUND: Microbial symbioses in marine invertebrates are commonplace. However, characterizations of invertebrate microbiomes are vastly outnumbered by those of vertebrates. Protists and fungi run the gamut of symbiosis, yet eukaryotic microbiome sequencing is rarely undertaken, with much of the focus on bacteria. To explore the importance of microscopic marine invertebrates as potential symbiont reservoirs, we used a phylogenetic-focused approach to analyze the host-associated eukaryotic microbiomes of 220 animal specimens spanning nine different animal phyla. RESULTS: Our data expanded the traditional host range of several microbial taxa and identified numerous undescribed lineages. A lack of comparable reference sequences resulted in several cryptic clades within the Apicomplexa and Ciliophora and emphasized the potential for microbial invertebrates to harbor novel protistan and fungal diversity. CONCLUSIONS: Microscopic marine invertebrates, spanning a wide range of animal phyla, host various protist and fungal sequences and may therefore serve as a useful resource in the detection and characterization of undescribed symbioses. Video Abstract.

Taxonomy and phylogeny of Koinocystididae (Platyhelminthes, Kalyptorhynchia), with the description of three new genera and twelve new species.

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 curious and neglected soft-bodied meiofauna: Rouphozoa (Gastrotricha and Platyhelminthes).

Gastrotricha and Platyhelminthes form a clade called Rouphozoa. Representatives of both taxa are main components of meiofaunal communities, but their role in the trophic ecology of marine and freshwater communities is not sufficiently studied. Traditional collection methods for meiofauna are optimized for Ecdysozoa, and include the use of fixatives or flotation techniques that are unsuitable for the preservation and identification of soft-bodied meiofauna. As a result, rouphozoans are usually underestimated in conventional biodiversity surveys and ecological studies. Here, we give an updated outline of their diversity and taxonomy, with some phylogenetic considerations. We describe successfully tested techniques for their recovery and study, and emphasize current knowledge on the ecology, distribution and dispersal of freshwater gastrotrichs and microturbellarians. We also discuss the opportunities and pitfalls of (meta)barcoding studies as a means of overcoming the taxonomic impediment. Finally, we discuss the importance of rouphozoans in aquatic ecosystems and provide future research directions to fill in crucial gaps in the biology of these organisms needed for understanding their basic role in the ecology of benthos and their place in the trophic networks linking micro-, meio- and macrofauna of freshwater ecosystems.

A new case of kleptoplasty in animals: Marine flatworms steal functional plastids from diatoms.

To date, sea slugs have been considered the only animals known to sequester functional algal plastids into their own cells, via a process called "kleptoplasty." We report here, however, that endosymbionts in the marine flatworms Baicalellia solaris and Pogaina paranygulgus are isolated plastids stolen from diatoms. Ultrastructural data show that kleptoplasts are located within flatworm cells, while algal nuclei and other organelles are absent. Transcriptomic analysis and rbcL amplicons confirm the absence of algal nuclear mRNA and reveal that the plastids originate from different species of diatoms. Laboratory experiments demonstrated photosynthetic activity and short-term retention of kleptoplasts in starved worms. This lineage of flatworms represents the first known case of functional kleptoplasty involving diatoms and only the second known case of kleptoplasty across the entire tree of animals.

Four new species of the genus Paraustrorhynchus (Rhabdocoela: Kalyptorhynchia: Polycystididae).

Four new species of Paraustrorhynchus (Rhabdocoela: Kalyptorhynchia: Polycystididae) are described: two from Southern China (P. shenda n. sp., P. sinensis n. sp.), one from New Caledonia (P. isoldeae n. sp.), and one from India (P. smeetsae n. sp.). All four species show a typical dark blue dorsal pigmentation pattern, which is unique to the genus. This pigment forms a broad longitudinal line in P. shenda n. sp. and P. sinensis n. sp., shows an additional transverse band at the level of the pharynx in P. smeetsae n. sp. and forms a Y-shaped pattern in P. isoldeae n. sp. The four species can also be distinguished from each other and their congeners by the detailed morphology of the hard parts of the male system, which in all cases consists of an L-shaped (P. shenda n. sp., P. isoldeae n. sp) or slightly-curved (P. smeetsae n. sp., P. sinensis n. sp.) prostate stylet type II and a prostate stylet type III, which consists of three plates: a basal plate from which two longer plates depart, each with a distal serrated rim. A phylogenetic analysis based on 18S and 28S-rDNA data indicates that Paraustrorhynchus is a monophyletic taxon within Polycystidinae, with Alcha evelinae as the sister group.

Species diversity in the marine microturbellarian Astrotorhynchus bifidus sensu lato (Platyhelminthes: Rhabdocoela) from the Northeast Pacific Ocean.

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.