The Stilbonematinae live in symbiosis with ectosymbiotic bacteria covering their cuticle, which evidently constitute their food. In different Stilbonematinae genera, two pharynx types are found, depending on the arrangement of the bacterial coat. Species descriptions show that most Stilbonematinae species with a thick multilayer of symbionts have a two-part pharynx with a predominantly muscular posterior bulb. In contrast, in cases of a thin monolayer of bacteria, the nematodes predominantly show a three-part pharynx with a distinctly swollen muscular corpus at their anterior end. This indicates a shift of the main pumping structure from the terminal bulb to the anterior corpus. Consequently, the amount of contractile filaments in the terminal bulb should decrease. Using phalloidin staining in combination with confocal laser scanning microscopy, light microscopy, and transmission electron microscopy, we measured and compared the filamentous actin (F-actin) volume in the posterior bulb in several Stilbonematinae species representing both pharynx types. Two-part pharynges had a larger relative F-actin volume in the terminal bulb than three-part pharynges. In the latter, prominent gland tissue occupied most of the space between the reduced muscles. This supports our hypothesis of two distinct feeding modes: ingestion of large amounts of food in species with a two-part pharynx ("gourmands") requiring a muscular terminal bulb vs discriminant grazing on a thin bacterial coat in species with a three-part pharynx ("gourmets").

Recent molecular phylogenies of Nematoda have suggested a sister-group relationship between Dorylaimia and Chromadoria, but supporting morphological evidence has so far been lacking. Using immunohistochemistry in combination with confocal laser scanning microscopy, we identified pharynx-associated musculature in Stilbonematinae, consisting of buccal dilators, somato-pharyngeal muscles, and a longitudinal spiral muscle encircling nearly the entire pharynx. To trace the evolutionary origin of these structures, we extended our investigation to closely related marine outgroups within Desmodorida but also the basally branching Chromadorida and Enoplia. Somato-pharyngeal muscles, which arise from the body wall musculature and attach to the pharynx surface, function as retractors in Chromadoria and Dorylaimia. In some Enoplia, however, they extend towards the posterior pharynx and act as protractors. These muscles are homologous and thus represent part of the ancestral body plan of Nematoda. Homologous buccal dilators and spiral muscles occur in both Dorylaimia and Chromadoria, whereas in Oncholaimina (Enoplia) buccal dilators—acting as protractors—have evolved independently. Taken together, these morphological findings strongly support a sister-group relationship between Dorylaimia and Chromadoria. A statistical analysis of the Stilbonematinae pharynx and its spiral musculature showed that the number of spiral coils is strongly correlated with pharynx slenderness (length-to-width ratio), in line with Roggen’s pharynx model. In both Dorylaimia and Chromadoria, the spiral musculature surrounding the posterior pharynx likely generates injection pressure, aiding the posterior transport of ingested food into the intestine as well as the anterior movement of secretions from pharyngeal glands.

Nematodes are one of the most widespread and abundant animal taxa across aquatic and terrestrial environments. In marine shallow-water, porous sediments, members of the subfamily Stilbonematinae may be found in high numbers. Stilbonematinae are characterized by their coat of symbiotic bacteria, which give the nematodes a white appearance, while the nematodes themselves are usually colourless. We identified several species of the genus Robbea (Desmodoridae: Stilbonematinae) in which live specimens had a conspicuous dark purple coloration of the glandular sense organs (GSOs), highly specialized epidermal glands that are prominent in the Stilbonematinae. The dark inclusions in the GSOs of Robbea contained high concentrations of iron and bromide, regardless of their habitat. Morphological and phylogenetic analyses show that the genus Robbea is paraphyletic and we define the new genus Cyathorobbea gen. nov. for species with sucker-shaped post-pharyngeal supplements in males. Furthermore, we describe five new Robbea species from around the world. Some Robbea species are morphologically very similar and required the use of principal component analysis of morphometric data to untangle the diversity in this taxon. Our work emphasizes the value of live animal observations that enable the identification of important characters and highlights the overlooked diversity within the Stilbonematinae.

Stilbonematinae are nematodes commonly found in shallow marine sands. They are overgrown by a genus- and species-specific coat of chemoautotrophic sulphur-oxidizing ectosymbiotic bacteria which profit from the vertical migration of their hosts through the chemocline by alternately gaining access to oxidizing and reducing chemical species, while in return, the host feeds on its symbionts. The subfamily exhibits a large morphological variability; e.g. the anterior pharynx is cylindrical in genera possessing a voluminous coat, but species with a bacterial monolayer possess a distinctly swollen corpus and therefore a tripartite pharynx. Since 18S-based phylogenetic analyses do not show close relationships between corpus-bearing species, we investigated the pharynx morphology using phalloidin staining in combination with confocal laser scanning microscopy, transmission electron microscopy and light microscopy in order to assess an independent evolution. The class-wide stable position of the subventral pharynx ampullae was used as a morphological marker. Ampullae are positioned at the anterior-most end of the isthmus in Cyathorobbea, further posterior in Catanema and Robbea and inside the corpus in Laxus oneistus. We therefore conclude an independent evolution of corpus enlargements within Stilbonematinae. This further suggests that pharynx morphology is driven by the volume of the symbiotic bacterial coat rather than phylogeny. Based on an existing mathematical model, an enlarged corpus should enable its bearer to ingest food in smaller quantities, in gourmet style, whereas a cylindrical pharynx would restrict its bearer to ancestral gourmand feeding. A review of pharynx types of Nematoda showed that the Stilbonematinae pharynx is substantially different compared to other tripartite pharynges. The lack of pharyngeal tubes and valves, the undivided corpus and evenly distributed nuclei in the isthmus warrant the definition of the “stilbonematoid” three-part pharynx.

A new genus of the marine Stilbonematinae (Nematoda, Desmodoridae) is described from the Pacific coast of the United States of America. The worms inhabit the sulfidic sediment among the roots of the surfgrass Phyllospadix sp. in the rocky intertidal. The ectosymbiotic coat is of a new type for Stilbonematinae. It consists of rod-shaped bacteria pointed at both poles densely attached with one pole to the host cuticle. This is the first report of this symbiotic nematode subfamily from the US West Coast.

Evolutionary theory predicts potential shifts between cooperative and uncooperative behaviour under fluctuating environmental conditions. This leads to unstable benefits to the partners and restricts the evolution of dependence. High dependence is usually found in those hosts in which vertically transmitted symbionts provide nutrients reliably. Here we study host dependence in the marine, giant colonial ciliate Zoothamnium niveum and its vertically transmitted, nutritional, thiotrophic symbiont from an unstable environment of degrading wood. Previously, we have shown that sulphidic conditions lead to high host fitness and oxic conditions to low fitness, but the fate of the symbiont has not been studied. We combine several experimental approaches to provide evidence for a sulphide-tolerant host with striking polyphenism involving two discrete morphs, a symbiotic and an aposymbiotic one. The two differ significantly in colony growth form and fitness. This polyphenism is triggered by chemical conditions and elicited by the symbiont's presence on the dispersing swarmer. We provide evidence of a single aposymbiotic morph found in nature. We propose that despite a high fitness loss when aposymbiotic, the ciliate has retained a facultative life style and may use the option to live without its symbiont to overcome spatial and temporal shortage of sulphide in nature.

The level of integration between associated partners can range from ectosymbioses to extracellular and intracellular endosymbioses, and this range has been assumed to reflect a continuum from less intimate to evolutionarily highly stable associations. In this study, we examined the specificity and evolutionary history of marine symbioses in a group of closely related sulphur-oxidizing bacteria, called Candidatus Thiosymbion, that have established ecto-and endosymbioses with two distantly related animal phyla, Nematoda and Annelida. Intriguingly, in the ectosymbiotic associations of stilbonematine nematodes, we observed a high degree of congruence between symbiont and host phylogenies, based on their ribosomal RNA (rRNA) genes. In contrast, for the endosymbioses of gutless phallodriline annelids (oligochaetes), we found only a weak congruence between symbiont and host phylogenies, based on analyses of symbiont 16S rRNA genes and six host genetic markers. The much higher degree of congruence between nematodes and their ectosymbionts compared to those of annelids and their endosymbionts was confirmed by cophylogenetic analyses. These revealed 15 significant codivergence events between stilbonematine nematodes and their ectosymbionts, but only one event between gutless phallodrilines and their endosymbionts. Phylogenetic analyses of 16S rRNA gene sequences from 50 Cand. Thiosymbion species revealed seven well-supported clades that contained both stilbonematine ectosymbionts and phallodriline endosymbionts. This closely coupled evolutionary history of marine ecto-and endosymbionts suggests that switches between symbiotic lifestyles and between the two host phyla occurred multiple times during the evolution of the Cand. Thiosymbion clade, and highlights the remarkable flexibility of these symbiotic bacteria.

Marine EcologyVolume 37, Issue 1 p. 234-234 Book Review A Guide to Southern Temperate Seagrasses M. Waycott, K. McMahon, P. Lavery Collingwood, Vic., Australia: CSIRO Publishing, 2014. 108 pp. ISBN: 9781486300150, Paperback, AU$29.95; 9781486300167 (epdf); 9781486300174 (epub). Jörg Ott, Jörg Ott University of Vienna, Vienna, AustriaSearch for more papers by this author Jörg Ott, Jörg Ott University of Vienna, Vienna, AustriaSearch for more papers by this author First published: 24 October 2014 https://doi.org/10.1111/maec.12221Read the full textAboutPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Share a linkShare onFacebookTwitterLinkedInRedditWechat No abstract is available for this article. Volume37, Issue1February 2016Pages 234-234 RelatedInformation

Eubostrichus fertilis sp. n. is described from fine subtidal sands in the Belize Barrier Reef system using LM and SEM illustrations and the sequence of the 18S rRNA gene. The new species is one of the smallest (mature specimens ranging from 1.88 to 3.03 mm) and the stoutest (a = 36-80) of all previously described Eubostrichus species. The closest relatives are E. parasitiferus and E. hopperi . It differs from the former in the more posterior position of the vulva and the postanal porids, and from the latter in the smaller size of the amphids, the shorter cephalic setae and the shape of the tail. Furthermore, it is remarkable for the prominent extent of the female genital system. Females have up to 18 eggs of similar size in their uteri. The body of the worm is covered by large (up to 45 μm long) crescent-shaped bacteria attached with both poles to the cuticle of the worm in a spiral pattern. The genus Eubostrichus is phylogenetically well supported on the basis of the 18S rRNA gene sequence. Eubostrichus gerlachi nom. nov. (= E. parasitiferus apud Gerlach, 1963 nec Chitwood, 1936) is proposed.

Two long-standing paradigms in biology are that cells belonging to the same population exhibit little deviation from their average size and that symmetric cell division is size limited. Here, ultrastructural, morphometric and immunocytochemical analyses reveal that two Gammaproteobacteria attached to the cuticle of the marine nematodes Eubostrichus fertilis and E. dianeae reproduce by constricting a single FtsZ ring at midcell despite being 45 μm and 120 μm long, respectively. In the crescent-shaped bacteria coating E. fertilis , symmetric FtsZ-based fission occurs in cells with lengths spanning one order of magnitude. In the E. dianeae symbiont, formation of a single functional FtsZ ring makes this the longest unicellular organism in which symmetric division has ever been observed. In conclusion, the reproduction modes of two extraordinarily long bacterial cells indicate that size is not the primary trigger of division and that yet unknown mechanisms time the localization of both DNA and the septum.

The Stilbonematinae are a monophyletic group of marine nematodes that are characterized by a coat of thiotrophic bacterial symbionts. Among the ten known genera of the Stilbonematinae, the genus Robbea Gerlach 1956 had a problematic taxonomic history of synonymizations and indications of polyphyletic origin. Here we describe three new species of the genus, R. hypermnestra sp. nov., R. ruetzleri sp. nov. and R. agricola sp. nov., using conventional light microscopy, interference contrast microscopy and SEM. We provide 18S rRNA gene sequences of all three species, together with new sequences for the genera Catanema and Leptonemella. Both our morphological analyses as well as our phylogenetic reconstructions corroborate the genus Robbea. In our phylogenetic analysis the three species of the genus Robbea form a distinct clade in the Stilbonematinae radiation and are clearly separated from the clade of the genus Catanema, which has previously been synonymized with Robbea. Surprisingly, in R. hypermnestra sp. nov. all females are intersexes exhibiting male sexual characters. Our extended dataset of Stilbonematinae 18S rRNA genes for the first time allows the identification of the different genera, e.g. in a barcoding approach. http://zoobank.org/urn:lsid:zoobank.org:pub:D37C3F5A-CF2B-40E6-8B09-3C72EEED60B0.

The remarkable totipotent stem-cell-based regeneration capacities of the Platyhelminthes have brought them into the focus of stem cell and regeneration research. Although selected platyhelminth groups are among the best-studied invertebrates, our data provide new insights into regenerative processes in the most basally branching group of the Platyhelminthes, the Catenulida. The mouth- and gutless free-living catenulid flatworm Paracatenula galateia harbors intracellular bacterial symbionts in its posterior body region, the trophosome region, accounting for up to 50% of the volume. Following decapitation of this flatworm, we have analyzed the behavior of the amputated fragments and any anterior and posterior regeneration. Using an EdU-pulse-chase/BrdU-pulse thymidine analog double-labeling approach combined with immunohistochemistry, we show that neoblasts are the main drivers of the regeneration processes. During anterior (rostrum) regeneration, EdU-pulse-chase-labeled cells aggregate inside the regenerating rostrum, whereas BrdU pulse-labeling before fixation indicates clusters of S-phase neoblasts at the same position. In parallel, serotonergic nerves reorganize and the brain regenerates. In completely regenerated animals, the original condition with S-phase neoblasts being restricted to the body region posterior to the brain is restored. In contrast, no posterior regeneration or growth of the trophosome region in anterior fragments cut a short distance posterior to the brain has been observed. Our data thus reveal interesting aspects of the cellular processes underlying the regeneration of the emerging catenulid-bacteria symbiosis model P. galateia and show that a neoblast stem cell system is indeed a plesiomorphic feature of basal platyhelminths.

Paracatenula galateia sp. nov. is a mouthless marine catenulid platyhelminth with bacterial intracellular endosymbionts. The worms live in shallow back-reef sands in the Belize Barrier Reef system and are distinguished from the four previously described members of the genus by their large size combined with a ribbon-shaped body and characteristic bipartite inclusions in cells, which are interpreted as sperm. The bacteria are presumed to be sulphur-oxidizing chemoautotrophs. They are found in bacteriocytes which fill the body region ('trophosome region') posterior to the brain, whereas the anterior part of the worm (rostrum) is bacteria-free. Phalloidin staining reveals a delicate system of subepitheliar circular and longitudinal muscles and dorsoventral fibres. The serotonergic nervous system consists of a brain at the base of the rostrum and longitudinal fibres extending both anteriorly and posteriorly, the latter being concentrated in a structure called the 'dorsal cord'.

Harnessing chemosynthetic symbionts is a recurring evolutionary strategy. Eukaryotes from six phyla as well as one archaeon have acquired chemoautotrophic sulfur-oxidizing bacteria. In contrast to this broad host diversity, known bacterial partners apparently belong to two classes of bacteria—the Gamma - and Epsilonproteobacteria . Here, we characterize the intracellular endosymbionts of the mouthless catenulid flatworm genus Paracatenula as chemoautotrophic sulfur-oxidizing Alphaproteobacteria . The symbionts of Paracatenula galateia are provisionally classified as “ Candidatus Riegeria galateiae” based on 16S ribosomal RNA sequencing confirmed by fluorescence in situ hybridization together with functional gene and sulfur metabolite evidence. 16S rRNA gene phylogenetic analysis shows that all 16 Paracatenula species examined harbor host species-specific intracellular Candidatus Riegeria bacteria that form a monophyletic group within the order Rhodospirillales . Comparing host and symbiont phylogenies reveals strict cocladogenesis and points to vertical transmission of the symbionts. Between 33% and 50% of the body volume of the various worm species is composed of bacterial symbionts, by far the highest proportion among all known endosymbiotic associations between bacteria and metazoans. This symbiosis, which likely originated more than 500 Mya during the early evolution of flatworms, is the oldest known animal–chemoautotrophic bacteria association. The distant phylogenetic position of the symbionts compared with other mutualistic or parasitic Alphaproteobacteria promises to illuminate the common genetic predispositions that have allowed several members of this class to successfully colonize eukaryote cells.

Marine oligochaete and nematode thiotrophic symbionts (MONTS) form a phylogenetic cluster within the Gammaproteobacteria. For the symbionts that live on the nematode surface, environmental transmission is likely. However, until now, no free-living relatives have been found. In this study, we detected MONTS cluster members in offshore surface seawater of both the Caribbean and the Mediterranean Sea by PCR amplification of their 16S rRNA genes. This is the first evidence of members of this cluster in the pelagic environment. These may either be free-living forms of the symbionts or closely related, nonsymbiotic strains. In either case, their existence sheds light on the evolution of beneficial symbioses between shallow water invertebrates and sulfur-oxidizing bacteria.

Marine catenulid platyhelminths of the genus Paracatenula lack mouth, pharynx and gut. They live in a symbiosis with intracellular bacteria which are restricted to the body region posterior to the brain. The symbiont-housing cells (bacteriocytes) collectively form the trophosome tissue, which functionally replaces the digestive tract. It constitutes the largest part of the body and is the most important synapomorphy of this group. While some other features of the Paracatenula anatomy have already been analyzed, an in-depth analysis of the trophosome region was missing. Here, we identify and characterize the composition of the trophosome and its surrounding tissue by analyzing series of ultra-thin cross-sections of the species Paracatenula cf. polyhymnia. For the first time, a protonephridium is detected in a Paracatenula species, but it is morphologically reduced and most likely not functional. Cells containing needle-like inclusions in the reference species Paracatenula polyhymnia Sterrer and Rieger, 1974 were thought to be sperm, and the inclusions interpreted as the sperm nucleus. Our analysis of similar cells and their inclusions by EDX and Raman microspectroscopy documents an inorganic spicule consisting of a unique magnesium–phosphate compound. Furthermore, we identify the neoblast stem cells located underneath the epidermis. Except for the modifications due to the symbiotic lifestyle and the enigmatic spicule cells, the organization of Paracatenula cf. polyhymnia conforms to that of the Catenulida in all studied aspects. Therefore, this species represents an excellent model system for further studies of host adaptation to an obligate symbiotic lifestyle.

Selection of a specific microbial partner by the host is an all-important process. It guarantees the persistence of highly specific symbioses throughout host generations. The cuticle of the marine nematode Laxus oneistus is covered by a single phylotype of sulfur-oxidizing bacteria. They are embedded in a layer of host-secreted mucus containing the mannose-binding protein Mermaid. This Ca(2+)-dependent lectin mediates symbiont aggregation and attachment to the nematode. Here, we show that Stilbonema majum-a symbiotic nematode co-occurring with L. oneistus in shallow water sediment-is covered by bacteria phylogenetically distinct to those covering L. oneistus. Mermaid cDNA analysis revealed extensive protein sequence variability in both the nematode species. We expressed three recombinant Mermaid isoforms, which based on the structural predictions display the most different carbohydrate recognition domains (CRDs). We show that the three CRDs (DNT, DDA and GDA types) possess different affinities for L. oneistus and S. majum symbionts. In particular, the GDA type, exclusively expressed by S. majum, displays highest agglutination activity towards its symbionts and lowest towards its L. oneistus symbionts. Moreover, incubation of L. oneistus in the GDA type does not result in complete symbiont detachment, whereas incubation in the other types does. This indicates that the presence of particular Mermaid isoforms on the nematode surface has a role in the attachment of specific symbionts. This is the first report of the functional role of sequence variability in a microbe-associated molecular patterns receptor in a beneficial association.