Phascolosoma Agassizii Research Articles

Novel cytoskeletal traits in the intestinal parasites (Squirmida, Platyproteum vivax) of Pacific peanut worms (Sipuncula, Phascolosoma agassizii).

The cytoskeletal organization of a squirmid, namely Platyproteum vivax, was investigated with confocal laser scanning microscopy (CLSM) to refine inferences about convergent evolution among intestinal parasites of marine invertebrates. Platyproteum inhabits Pacific peanut worms (Phascolosoma agassizii) and has traits that are similar to other lineages of myzozoan parasites, namely gregarine apicomplexans within Selenidium, such as conspicuous feeding stages, called "trophozoites," capable of dynamic undulations. SEM and CLSM of P. vivax revealed an inconspicuous flagellar apparatus and a uniform array of longitudinal microtubules organized in bundles (LMBs). Extreme flattening of the trophozoites and a consistently oblique morphology of the anterior end provided a reliable way to distinguish dorsal and ventral surfaces. CLSM revealed a novel system of microtubules oriented in the flattened dorsoventral plane. Most of these dorsoventral microtubule bundles (DVMBs) had a punctate distribution and were evenly spaced along a curved line spanning the longitudinal axis of the trophozoites. This configuration of microtubules is inferred to function in maintaining the flattened shape of the trophozoites and facilitate dynamic undulations. The novel traits in Platyproteum are consistent with phylogenomic data showing that this lineage is only distantly related to Selenidium and other marine gregarine apicomplexans with dynamic intestinal trophozoites.

Genetic structure in two Phascolosoma species in the Pacific Ocean

ABSTRACTPhascolosoma agassizii, a commonly reported species of sipunculan worm, occurs at various depths throughout the north Pacific. However, previous studies have shown that this nominal species is actually comprised of two genetically divergent lineages. Phascolosoma agassizii is isolated to the eastern Pacific, whereas a second species, here referred to Phascolosoma sp., is located in the western Pacific and represents a unique, non-sister taxon to P. agassizii. Both species exhibit the same developmental mode with a long-lived, planktotrophic, pelagosphera larval stage, which may facilitate long-distance dispersal, although developmental timing differs. Using an inter-simple sequence repeat polymerase chain reaction (ISSR-PCR) genetic fingerprinting approach, this study examines non-coding polymorphic regions of the nuclear genomes of P. agassizii from four eastern Pacific populations and of Phascolosoma sp. from six western Pacific locations. Small-scale spatial genetic variation was analysed separately for each coastline using an Analysis of Molecular Variance. The data pointed to the potential presence of fine-scale genetic structure within P. agassizii, and recovered significant genetic structure within Phascolosoma sp. from the western Pacific. This study shows that ISSR-PCR is a relatively fast and cost-effective way to study fine-scale genetic diversity in marine invertebrates, although possible analyses and interpretations of the data are subject to the same limitations as other size polymorphism-based approaches.

Cryptic species in Pacific sipunculans (Sipuncula: Phascolosomatidae): east‐west divergence between non‐sister taxa

Cryptic diversity, i.e. diversity observable in genetic but not in morphological traits, is prevalent in marine invertebrates and presents one of the greatest obstacles to obtaining accurate estimates of species richness. Sipunculans, commonly called peanut worms, are marine annelids in which high levels of cryptic diversity have previously been documented. In this study, we use genetic identification techniques to examine divergence of two lineages of Pacific sipunculans, both known under the name Phascolosoma agassizii. One lineage is isolated to the eastern Pacific coast while the other one inhabits the western Pacific coast. These clades are reciprocally monophyletic and are not recovered as sister taxa, suggesting relatively early divergence within Phascolosoma. Furthermore, we did not find support for a genetic distinction between the western Pacific Phascolosoma agassizii agassizii and Phascolosoma agassizii kurilense, a subspecies reported from the Kuril Islands. Considering that the type locality for P. agassizii is in the Eastern Pacific, we suggest that the western clade, including the samples from the Kuril islands, represent a new, undescribed species.

Peanut worms of the phylum Sipuncula from the Sea of Japan with a key to species

Sipunculan worms from the Russian waters of the Sea of Japan are still poorly investigated while they are much better known from the Japanese coast. The aim of this paper is to describe sipunculans from the Russian coast and from the deepest part of the Sea of Japan near the Primorye Province collected by SoJaBio expedition, and to provide keys for identification of sipunculan species from the Sea of Japan. At the Russian coast of the Sea of Japan only 8 valid species of sipunculans were found and identified: Golfingia margaritacea, G. vulgaris, Nephasoma capilleforme, N. wodjanizkii, Phascolion strombus, Thysanocardia nigra, Themiste hexadactyla (=T. pyroides), Phascolosoma agassizii. Taking into account 4 other valid species noted for this area, Nephasoma eremite, Thysanocardia catharinae, Themiste blanda and Phascolosoma scolops, which were not found, the sipunculan fauna of the Russian waters of the Sea of Japan now comprises 12 valid species. Nephasoma capilleforme and Nephasoma wodjanizkii are the first records for the North-West Pacific and the Sea of Japan. Species accounts include the most important taxonomic characters and specific biotope data. Accordingly, a key up to species level is provided. Totally, the fauna of the Sea of Japan is now estimated as having 31 valid species of sipunculans.

Embryonic and larval development of the peanut worm Phascolosoma agassizii (Keferstein 1867) from the Sea of Japan (Sipuncula: Phascolosomatidea)

All stages of the embryonic and larval development of Phascolosoma agassizii from Peter the Great Bay (Sea of Japan) were studied and illustrated using light and electron microscopy. The eggs of P. agassizii have the form of an ellipsoid (long and short axes about 100 and 70 µm, respectively). Egg cleavage is typical, spiral, and unequal. Gastrulation occurs by epiboly. This species possesses two pelagic larval stages, a lecithotrophic trochophore and a planktotrophic pelagosphera. The transformation of trochophore into pelagosphera occurs 80–90 h after fertilization. After 120–180 h, the larva has developed all systems of organs characteristic of the pelagosphera and is capable of feeding. At day 10, pelagospheras can settle, for some time, on the aquarium bottom and move on a ciliated lip, collecting food with the aid of a buccal organ. In addition, the larvae periodically attach themselves to the aquarium bottom or to the surface film of the water by means of a terminal organ. The trunk of the larva elongates by enlargement of the region behind the dorsal anal opening, which is located almost in the middle of the trunk region in the 15-day old larva. In the laboratory, 1-month old larvae spend the greater part of time in the attached state. Being attached by a glandular terminal organ to the aquarium bottom, they characteristically bend the body, actively feeding on microalgae from the substratum surface. The differences in the development of P. agassizii in the isolated West-Pacific and East-Pacific populations are shown and discussed.

Cellular and muscular growth patterns during sipunculan development.

Sipuncula is a lophotrochozoan taxon with annelid affinities, albeit lacking segmentation of the adult body. Here, we present data on cell proliferation and myogenesis during development of three sipunculan species, Phascolosoma agassizii, Thysanocardia nigra, and Themiste pyroides. The first anlagen of the circular body wall muscles appear simultaneously and not subsequently as in the annelids. At the same time, the rudiments of four longitudinal retractor muscles appear. This supports the notion that four introvert retractors were part of the ancestral sipunculan bodyplan. The longitudinal muscle fibers form a pattern of densely arranged fibers around the retractor muscles, indicating that the latter evolved from modified longitudinal body wall muscles. For a short time interval, the distribution of S-phase mitotic cells shows a metameric pattern in the developing ventral nerve cord during the pelagosphera stage. This pattern disappears close to metamorphic competence. Our findings are congruent with data on sipunculan neurogenesis, as well as with recent molecular analyses that place Sipuncula within Annelida, and thus strongly support a segmental ancestry of Sipuncula.

Reproduction and development of common species of peanut worms (Sipuncula) from the Sea of Japan

This study deals with the reproduction and development of the most common species of peanut worms from the Sea of Japan: Thysanocardia nigra, Themiste pyroides, and Phascolosoma agassizii. Data on the time of reproduction and larval settlement and the distribution of these species in Peter the Great Bay are provided. The peculiarities of gametogenesis, spawning, and embryonic, larval, and postlarval development are described. The reproductive biology of representatives of these species from the western and eastern Pacific is examined in a comparative aspect.

Molecular Phylogeny and Surface Morphology of Marine Archigregarines (Apicomplexa), Selenidium spp., Filipodium phascolosomae n. sp., and Platyproteum n. g. and comb. from North‐Eastern Pacific Peanut Worms (Sipuncula)

The trophozoites of two novel archigregarines, Selenidium pisinnus n. sp. and Filipodium phascolosomae n. sp., were described from the sipunculid Phascolosoma agassizii. The trophozoites of S. pisinnus n. sp. were relatively small (64-100 microm long and 9-25 microm wide), had rounded ends, and had about 21 epicytic folds per side. The trophozoites of F. phascolosomae n. sp. were highly irregular in shape and possessed hair-like surface projections. The trophozoites of this species were 85-142 microm long and 40-72 microm wide and possessed a distinct longitudinal ridge that extended from the mucron to the posterior end of the cell. In addition to the small subunit (SSU) rDNA sequences of these two species, we also characterized the surface morphology and SSU rDNA sequence of Selenidium orientale, isolated from the sipunculid Themiste pyroides. Molecular phylogenetic analyses demonstrated that S. pisinnus n. sp. and S. orientale formed a strongly supported clade within other Selenidium and archigregarine-like environmental sequences. Filipodium phascolosomae n. sp. formed the nearest sister lineage to the dynamic, tape-like gregarine Selenidium vivax. Overall, these data enabled us to reassess the molecular systematics of archigregarines within sipunculid hosts and make the following revisions: (1) Filipodium was transferred from the Lecudinidae (eugregarines) to the Selenidiidae (archigregarines), and (2) Platyproteum n. g. was established for Platyproteum vivax n. comb. (ex. S. vivax) in order to account for the highly divergent morphological features and better resolved phylogenetic position of this lineage.

Segmental Mode of Neural Patterning in Sipuncula

Recent molecular phylogenetic analyses suggest a close relationship between two worm-shaped phyla, the nonsegmented Sipuncula (peanut worms) and the segmented Annelida (e.g., earthworms and polychaetes) [1-5]. The striking differences in their bodyplans are exemplified by the annelids' paired, ladder-like ventral nervous system, which contains segmentally arranged ganglia, and the sipunculans' single ventral nerve cord (VNC), which is devoid of any segmental structures [6, 7]. Investigating central nervous system (CNS) formation with serotonin and FMRFamide labeling in a representative sipunculan, Phascolosoma agassizii, we found that neurogenesis initially follows a segmental pattern similar to that of annelids. Starting out with paired FMRFamidergic and serotonergic axons, four pairs of associated serotonergic perikarya and interconnecting commissures form one after another in an anterior-posterior progression. In late-stage larvae, the two serotonergic axons of the VNCs fuse, the commissures disappear, and one additional pair of perikarya is formed. These cells (ten in total) migrate toward one another, eventually forming two clusters of five cells each. These neural-remodeling processes result in the single nonmetameric CNS of the adult sipunculan. Our data confirm the segmental ancestry of Sipuncula and render Phascolosoma a textbook example for the Haeckelian hypothesis of ontogenetic recapitulation of the evolutionary history of a species [8].

Multiple Isolations of a Culturable, Motile Ichthyosporean (Mesomycetozoa, Opisthokonta), Creolimax fragrantissima n. gen., n. sp., from Marine Invertebrate Digestive Tracts

A fragrant, spherical, osmotrophic eukaryote was isolated 27 times from the digestive tracts of marine invertebrates collected from the Northeast Pacific. The isolates were cultured from 7 animal collections over a 2-year period, most from the peanut worm, Phascolosoma agassizii. A small subunit ribosomal DNA phylogeny placed the spherical organism within the ichthyosporea, closest to Sphaeroforma arctica and Pseudoperkinsus tapetis. Supporting the close relationship of isolates, the sequences of ribosomal gene internal transcribed spacers determined for 26 isolates were identical, as were the elongation factor 1-alpha-like gene fragments from 7 isolates. Dispersal via amoeboid cells distinguished this species from its closest relatives and led to the erection of a new genus and species, "Creolimax fragrantissima." Vegetative cells reproduced asexually in vitro after they reached 30-60 microm in diameter by producing amoebae or endospores, which escaped through openings in the parent cell wall. Ultrathin sections of vegetative cells prepared by high-pressure-freeze substitution provided some of the first images of ichthyosporean spindle pole bodies and document, for the first time, tubular extensions of the plasma membrane into an electron-translucent inner layer of the cell wall. Ichthyosporeans are parasites and commensals of animals and culturable species are few. Because "C. fragrantissima" can be isolated regularly and repeatedly from nature and then grown easily through cycles of asexual reproduction, it has the potential to serve as a model organism for further research into marine ichthyosporeans.

Fine structure of the pharyngeal apparatus of the pelagosphera larva in Phascolosoma agassizii (Sipuncula) and its phylogenetic significance

Sipuncula is a small taxon of worm-like marine organisms of still uncertain phylogenetic position. Si- punculans are characterized by an unsegmented body composed of a trunk into which the anterior part, the introvert, can be withdrawn. The group has been placed at various positions within Metazoa; currently, it is ei- ther seen as sister group of a clade comprising Mollusca and Annelida or as sister to each of these. An in-group position in either Mollusca or Annelida has usually been precluded till now due to the lack of so-called annelid or molluscan ''key-characters'' such as segmentation and chaetae or the radula. In the development of certain taxa the trochophore stage is followed by a planktonic larva, the pelagosphera, which might exhibit phylogenetically important structures. Among these is the buccal organ, which has been considered homologous either to the ventral pharyngeal organ present in many sedentary polychaetes or to the radular apparatus of molluscs. In the present paper, the ventral pharynx of the pelagosp- hera larva of Phascolosoma agassizii is investigated by transmission electron microscopy. The pharynx com- prises dorsolateral ciliary folds, a muscle bulb formed by transverse muscle fibres with large intercellular spaces, and an investing muscle. A tongue-like organ is lacking. These results show great structural correspondences to the ventral pharynx of polychaetes, especially to that of the flabelligerid Diplocirrus longisetosus. In contrast, there are no signs of structural similarities to the cor- responding structures of molluscs. Thus evidence in- creases that Sipuncula are closely related to annelids; moreover, an in-group position of Sipuncula within Annelida, as suggested by recent molecular studies, is not precluded by the present data. Instead these studies find additional support. Hence the lack of segmentation and chitinous chaetae in Sipuncula would be a second- ary rather than a primary situation, as has recently been shown for Echiura and Pogonophora.

Ultrastructure of the archigregarine Selenidium vivax (Apicomplexa) – A dynamic parasite of sipunculid worms (host: Phascolosoma agassizii)

Selenidium vivax is a large and unusual unicellular parasite that inhabits the intestinal lumen of the dotted peanut worm, Phascolosoma agassizii. Molecular phylogenies suggest that this archigregarine lineage diverges near the nexus of the apicomplexan radiation and could shed light on to the early evolution of parasitism within the group. The behaviour and ultrastructure of the trophozoites were described using digital videography and scanning and transmission electron microscopy. The trophozoites were extremely flat and capable of dynamic cellular deformations. An intimate association between a superficial layer of mitochondria and longitudinal clusters of subpellicular microtubules formed a distinct functional configuration that helped explain the mechanism behind the cellular motility. Although inconclusive, the presence of small mitochondria-like profiles and narrow connections between larger mitochondrial profiles suggested that an expansive mitochondrial reticulum might surround the trophozoites. The nucleus was highly convoluted and gave rise to blebs of different sizes. The nuclear blebs were connected to the nucleus proper and surrounded by one cisterna of endoplasmic reticulum, giving the impression of four membrane-bound organelles that were misleadingly reminiscent of apicoplasts. The novel attachment apparatus consisted of a transverse ridge, a linear arrangement of pores that contained thread-like structures and a network of dense bodies and endoplasmic reticulum.

Selenidium vivax n. sp. (Protozoa, Apicomplexa) from the Sipunculid Phascolosoma agassizii Keferstein, 1867

Selenidium vivax n. sp. (Protozoa, Apicomplexa, Eugregarinorida, Selenidiidae) is described from the intestine of the sipunculid Phascolosoma agassizii from the central Califomian coast. Gamonts were up to 550 Am long, flat, variable in shape, with an asymmetric rostrum and without permanent surface striations.

CELLULAR DEFENSE REACTIONS OFPHASCOLOSOMA AGASSIZII, A SIPUNCULAN WORM: PHAGOCYTOSIS BY GRANULOCYTES

Based on Wright's blood stain, uninucleate circulating granulocytes of the sipunculan Phascolosoma agassizii Keferstein are of two types: acidophils and basophils. The granulocytes are characterized by numerous granules scattered throughout the cytoplasm. Morphology of the granulocytes viewed with the light and electron microscope is illustrated and described. The granules of the granulocytes contain the enzymes acid phosphatase, alkaline phosphatase, lipase, and peroxidase. Fibrin-like strands entrap foreign particles—polystyrene latex beads and Staphylococcus aureus—introduced into the coelomic fluid in vivo or in vitro. Granulocytes accumulate in the vicinity of the trapped foreign particles. Both acidophilic and basophilic granulocytes phagocytize polystyrene latex beads and Staphylococcus aureus in vivo and in vitro. The cytoplasmic granules of the granulocytes degranulate into the phagocytic vacuoles and new granules form from the concave face of the Golgi complex.

Uptake of Hydrocarbons from Marine Sediments Contaminated with Prudhoe Bay Crude Oil: Influence of Feeding Type of Test Species and Availability of Polycyclic Aromatic Hydrocarbons

Selected benthic animals were exposed to marine sediments contaminated with Prudhoe Bay crude oil, and uptake of hydrocarbons was monitored under various experimental schemes. When uptake of aliphatic and diaromatic hydrocarbons by two deposit feeders, Macoma inquinata and Phascolosoma agassizii, was compared with that of a suspension feeder, Protothaca staminea, we found that the deposit feeders generally accumulated hydrocarbons to a greater extent than the suspension feeder. However, other factors, such as the intrinsic capabilities of species to accumulate hydrocarbons, also played an important role in the extent of contamination. Hydrocarbon concentrations in the bivalve species (M. inquinata and P. staminea) increased during the 60-d exposure period; whereas concentrations in the sipunculid Phascolosoma agassizii appeared to have reached an equilibrium relatively early in the exposure. Experiments on uptake of 14C-phenanthrene, -chrysene, -dimethylbenz[a]anthracene, and -benzo[a]pyrene by M. inquinata indicated that compounds directly associated with sediment were less available for uptake than those released from sediment to the surrounding seawater. Concentration factors for uptake from sediment were ≤ 0.2, while those for uptake from seawater were 10–1349. Additionally, the heavier molecular weight aromatic compounds tended to be more concentrated in tissue and retained for longer periods of time than the lighter compounds. Key words: petroleum, hydrocarbons, sediments, bivalve, sipunculid, feeding type

A light and electron microscopic study of the ciliated urn of Phasolosoma agassizii (Sipunculida).

The free swimming ciliated urn found in the coelomic fluid of Phascolosoma agassizii has been studied by electron microscopy. The urn is a multicellular structure composed of three cell types: (a) ciliated cells which possibly function in capturing cell debris and foreign particles; (b) cupola cells which are capable of phagocytozing latex particles; and (c) lobe cells which are capable of phagocytozing carbon particles. The lobes are separated from the ciliated cells by a semilunar area, with mucoprotein staining characteristics, containing fibrils which appear to be the structural support for the urn. Ciliated cells and lobes are attached to the semilunar area by hemidesmosomes.

Cell within a cell or a circulating cell pair.

I WISH to report that one of the coelomocytes of Phascolosoma agassizii Keferstein, 18661 (collected from Arroyo de Frijoles State Beach, California) which was previously considered to be one granulocyte is seen, under the electron microscope, to consist of two separate cells in a unique relationship.

The quaternary structure of Phascolosoma agassizii coelomic hemerythrin

The coelomic hemerythrin of Phascolosoma agassizii gave a molecular weight of 32,200 on Sephadex gel chromatography and an s20,ω of 3.65 S in sedimentation velocity experiments. A combination of the sedimentation coefficient and the Stoke's radius determined by gel chromatography permitted the calculation of a molecular weight of 40,600. A subunit molecular weight of 12,800 was obtained by SDS-polyacrylamide gel electrophoresis. Iron analyses indicated the hemerythrin contained 0.885% Fe. Assuming 2Fe/subunit, a minimum subunit molecular weight of 12,600 was calculated. Crosslinking experiments of native protein with dimethyl suberimidate yielded three bands on SDS-polyacrylamide gel electrophoresis. The molecular weights of these species were 13,000, 26,500, and 40,500. Nine bands which stainded for protein and iron were observed in disc gel electrophoresis of the native hemerythrin. Electrophoresis in gels containing 8 m urea showed three major and one minor band. Individual native hemerythrin bands, isolated by preparative disc electrophoresis, gave one, two, or three bands when reelectrophoresed on 8 m urea gels. Based on this evidence, it was concluded that P. agassizii coelomic hemerythrin exist as a trimeric protein. A model is proposed for the subunit composition of the native protein which accounts for the nine bands observed in disc gel electrophoresis.

Some osmotic responses of the sipunculid worm Themiste dyscritum

1. 1. Themiste dyscritum, apparently stenohaline in nature, tolerates a wide range of salinities in the laboratory (168–639 mM Cl −, or 30–114% sea water). Long-term survival probably does not occur below about 50% sea water. 2. 2. T. dyscritum is a passive osmoconformer over this entire salinity range, but has an apparent ion deficit in the coelomic fluid, as indicated by low internal chloride concentrations relative to external levels. This apparent ion deficit is most marked in worms in low salinities, and may be made up by small molecular weight organic molecules. 3. 3. Water content is strongly dependent upon salinity, and the worms absorb large amounts of water osmotically after a transfer to a lower salinity. Volume regulation is restricted to worms adapted to high salinities and to transfers which are not excessive. 4. 4. Nephridial contents are little different in concentration from the coelomic fluid. 5. 5. Limited data on Phascolosoma agassizii in 84% sea water indicate the coelomic fluid is slightly hyperosmotic to the medium, but iso-ionic with respect to chloride.

A comparative study of the development ofPhascolosoma agassizii,Golfingia pugettensis, andThemiste pyroideswith a discussion of developmental patterns in the Sipuncula

Abstract Breeding seasons, eggs, cleavage and development are described for three species of sipunculids, Phascolosoma agassizii, Golfingia pugettensis, and Themiste pyroides, from the San Juan Archipelago on the Northwestern Pacific Coast of the United States. Phascolosoma agassizii breeds from the middle of June to early September, Golfingia pugettensis from late October through January, and Themiste pyroides from March to August. The egg of P. agassizii is the smallest (140 × 110 microns) with a low yolk content, whereas that of G. pugettensis is intermediate in size (160 microns in diameter) and yolk content, and the egg of T. pyroides is the largest of the three (190 microns in diameter) with a dense concentration of yolk. Cleavage in all three species is of the typical spiral pattern except that in the 8-cell stage of T. pyroides the micromeres of the A, B, and C quadrants arc larger than the macromeres. Both P. agassizii and G. pugettensis undergo an indirect development with a pelagic trochophore ...