Myzostomida: a link between trochozoans and flatworms?

Myzostomids are obligate symbiotic invertebrates associated with echinoderms with a fossil record that extends to the Ordovician period. Due to their long history as host-specific symbionts, myzostomids have acquired a unique anatomy that obscures their phylogenetic affinities to other metazoans: they are incompletely segmented, parenchymous, acoelomate organisms with chaetae and a trochophore larva. Today, they are most often classified within annelids either as an aberrant family of polychaetes or as a separate class. We inferred the phylogenetic position of the Myzostomida by analysing the DNA sequences of two slowly evolving nuclear genes: the small subunit ribosomal RNA and elongation factor-1alpha. All our analyses congruently indicated that myzostomids are not annelids but suggested instead that they are more closely related to flatworms than to any trochozoan taxon. These results, together with recent analyses of the myzostomidan ultrastructure, have significant implications for understanding the evolution of metazoan body plans, as major characters (segmentation, coeloms, chaetae and trochophore larvae) might have been independently lost or gained in different animal phyla.

Evidence from polychromatism and bioluminescence that the cosmopolitan ophiuroid Amphipholis squamata might not represent a unique taxon.

Individuals of the cosmopolitan ophiuroid Amphipholis squamata were collected from eight stations. Eleven colour varieties were described and their distribution was non-random among stations. This suggests that the varieties differ in ecophysiologic tolerance and that their geographical distribution is modulated by environmental conditions. Varieties also differed in bioluminescence. Contrary to kinetics, intensity of light production varied among co-occurring varieties, meaning that they have similar bioluminescent reactions but a different amount of bioluminescent reagent. Light intensity differed in absolute value among stations but the rank position of each variety relative to others remained constant from one station to another. The 'colour-bioluminescence' link appeared clearly fixed (the same level of bioluminescence for the same variety) and is suggested to be of genetic origin. The species 'A. squamata' may then be a mosaic of genetically different entities (the varieties) rather than a unique cosmopolitan taxonomic entity.

Evidence of seasonal variation in bioluminescence of Amphipholis squamata (Ophiuroidea, Echinodermata): effects of environmental factors.

The bioluminescence of Amphipholis squamata was assessed from freshly collected individuals for 16 successive months, and from individuals maintained in the laboratory under various experimental conditions of salinity, temperature and photoperiodic regime. Field investigations showed that bioluminescence intensity and kinetics varied seasonally, with the light produced being brighter and faster in winter and summer. The seasonal variation was not correlated with changes of ambient salinity. However, it was correlated with changes in temperature, the luminescence being brighter and faster in coldest and warmest seasons, and with the changes of photoperiod, the luminescence being brighter and faster in seasons with shortest and longest day length. Laboratory investigations also demonstrated that luminescence was not affected by salinity conditions. Conversely, luminescence was affected by temperature, the light production being brighter and faster in warmer conditions (in agreement with field observations) and dimmer and slower in colder conditions (in disagreement with field observations). Light production was also affected by photoperiod since experimental changes of natural light:dark regime caused the bioluminescence to decrease. Considering that photoperiod guides the biology of A. squamata and that reproduction takes place during coldest months in the species, an endogenous factor of neurophysiological nature linked to the ophiuroid reproductive cycle is proposed to induce the luminescence to peak in winter. This was confirmed by the fact that seasonal variation of luminescence was different between adult and juveniles, the latter showing no winter peak of luminescence. It is suggested that the luminescence normally associated with defense could also be part of an intraspecific visual signal related to individuals aggregating for reproduction during winter.

Maintaining the line of defense: regeneration of Cuvierian tubules in the sea cucumber Holothuria forskali (Echinodermata, Holothuroidea).

When irritated, individuals of the sea cucumber Holothuria forskali expel a few Cuvierian tubules which lengthen, instantly become sticky, and rapidly immobilize most organisms with which they come into contact. After expulsion, the lost tubules are readily regenerated. When only a few tubules have been expelled, there is often a latent period before the regeneration starts. In contrast, when many tubules have been expelled, the regenerative process starts immediately but proceeds in successive waves of 10 to 30 tubules that begin to regenerate at 10-day intervals. However, in all cases, the complete regeneration of a given tubule takes about 5 weeks and may be divided into three successive phases: an initial repair phase including the overall 48-h post-autotomy period, a true regenerative phase taking about 4 weeks to complete, and a growth phase of about one more week. Initial regeneration events occur by epimorphosis, cell proliferation being essential to the regenerative process, whereas late events occur mainly by morphallaxis, with migration of the newly differentiated cells. The mesothelium is the tissue layer in which cell proliferation is the most precocious and the most important, involving both peritoneocytes and undifferentiated cells (which seem to be dedifferentiated peritoneocytes). As regeneration proceeds, the percentage of undifferentiated cells regularly decreases in parallel with the differentiation of granular (adhesive-secreting) cells and myocytes. The myocytes then separate off from the mesothelium and migrate within the connective tissue layer. Three types of pseudopodial cells follow one another in the tubule connective tissue during regeneration. Type 1 cells have all the characteristics of echinoderm phagocytes and may have a fibroblastic function, cleaning the connective tissue compartment before new collagen synthesis starts. Type 2 cells are rather undifferentiated and divide actively. The presence of type 3 cells is closely associated with the appearance of collagen fibers, and it is suggested that they have a fibroblastic function. In the inner epithelium, cells also divide actively, but only those in which spherules have not yet differentiated in the basal intraconnective processes. It appears, therefore, that in the three tissue layers of the tubules, regeneration proceeds by cell dedifferentiation, then proliferation, and finally by differentiation. Cuvierian tubules thus constitute a very efficient defensive mechanism: their large number, sparing use, and particular regeneration dynamics make them an almost inexhaustible line of defense maintained at limited energy cost.

Alteration of bioluminescence in Amphipholis squamata (Ophiuroidea: Echinodermata) by heavy metals contamination: a field study.

The ophiuroid Amphipholis squamata (Echinodermata) is a bioluminescent species whose light production varies with physico-chemical parameters of the medium. Individuals collected in the bay of Portman along a gradient of heavy metal contamination show different patterns of light production: the ones from the highest contaminated area showing a bioluminescence weaker and slower than those from the lowest contaminated area. Individuals that were transferred for 3 days from the lowest to the highest contaminated area displayed a light production that became weaker and slower. It is suggested that the decrease of the bioluminescent capability due to heavy metal pollution could indirectly affect the ophiuroid ecological success (bioluminescence is associated with defense functions in ophiuroids.

Cytological changes during bioluminescence production in dissociated photocytes from the ophiuroid Amphipholis squamata (Echinodermata).

Bioluminescence in the ophiuroid Amphipholis squamata is produced by photocytes located within the spinal ganglia of arm spines. Ganglionic cells were dissociated (pronase digestion) and photocytes separated from other cell types by using a continuous density Percoll gradient. Aliquots from a stock suspension of photocytes in artificial sea water were stimulated to produce light by using KCl or acetylcholine and fixed for ultrastructural observation at different times of the luminous process. Preluminescent, luminescent, and postluminescent photocytes contained various intracytoplasmic structures, such as Golgi, flat and distended rough endoplasmic reticulum cisternae, bundles of fibrils, and up to six types of membrane-bounded vesicles. These structures either co-occurred or succeeded one another during the process of light production, indicating that they were most probably participating in the luminescence reaction. Two types of vesicles, sharing some ultrastructural features, probably represented the microsources of the photocytes. One type occurred almost exclusively in luminescent photocytes, and the other almost exclusively in postluminescent photocytes, suggesting that one may be transformed into the other. The latter type of vesicle contained densely packed fibro-tubular units, giving a characteristic paracrystalline appearance to postluminescent photocytes.

Symbioses in Amphipholis squamata (Echinodermata, Ophiuroidea, Amphiuridae): geographical variation of infestation and effect of symbionts on the host's light production.

Populations of the polychromatic and bioluminescent species Amphipholis squamata from eight locations were examined for internal and external symbionts. At three locations (two in the United Kingdom and one in Papua New Guinea), no symbionts were present, while four species were recovered from the remaining locations: Cancerilla tubulata and Parachordeumium amphiurae (copepods), Rhopalura ophiocomae (orthonectid) and an undescribed species of rhabdocoel turbellarian. No ophiuroid individual hosted more than one symbiont species, despite the presence of two or more within a population. Symbiont presence and prevalence varied with location, and with colour variety, but with no apparent pattern or trends. Light-production characteristics of the host were affected by the presence of all symbionts except C. tubulata. These effects, however, did not vary between colour varieties or between geographical locations, but were specific to the symbiont species: the presence of P. amphiurae resulted in enhanced intensity of light production, while that of R. ophiocomae and the turbellarian species resulted in reduced intensity. The kinetics of light production (time until maximum output) were altered only by the presence of the turbellarian. Changes in the light-production characteristics are discussed in relation to morphological, energetical and physiological effects of the symbioses.

A study of the temporary adhesion of the podia in the sea star asterias rubens (Echinodermata, asteroidea) through their footprints

Sea stars are able to make firm but temporary attachments to various substrata owing to secretions released by their podia. A duo-glandular model has been proposed in which an adhesive material is released by two types of non-ciliated secretory (NCS1 and NCS2) cells and a de-adhesive material is released by ciliated secretory (CS) cells. The chemical composition of these materials and the way in which they function have been investigated by studying the adhesive footprints left by the asteroids each time they adhere to a substratum. The footprints of Asterias rubens consist of a sponge-like material deposited as a thin layer on the substratum. Inorganic residues apart, this material is made up mainly of proteins and carbohydrates. The protein moiety contains significant amounts of both charged (especially acidic) and uncharged polar residues as well as half-cystine. The carbohydrate moiety is also acidic, comprising both uronic acids and sulphate groups. Polyclonal antibodies have been raised against footprint material and were used to locate the origin of footprint constituents in the podia. Extensive immunoreactivity was detected in the secretory granules of both NCS1 and NCS2 cells, suggesting that their secretions together make up the bulk of the adhesive material. No immunoreactivity was detected in the secretory granules of CS cells, and the only other structure strongly labelled was the outermost layer of the cuticle, the fuzzy coat. This pattern of immunoreactivity suggests that the secretions of CS cells are not incorporated into the footprints, but instead might function to jettison the fuzzy coat, thereby allowing the podium to detach.

Fine structure of the dorsal papillae in the holothurioid Holothuria forskali (Echinodermata).

The dorsal surface of the holothurioid Holothuria forskali bears several longitudinal rows of modified podia called papillae. Each papilla consists of a conical stem topped by an hemispherical bud. Their gross tissue stratification is the same all along the papilla being made up of four tissue layers, viz. an inner mesothelium, a connective tissue layer, a nerve plexus and an outer epidermis. The latter is differently organized according to whether it belongs to the stem or to the bud. The epidermis of the bud is built up by ciliated cells that intimately contact the nerve plexus and have the classical structure of echinoderm sensory cells. The papillae are thus sensory organs involved in mechanoreception and possibly chemoreception.

The Role of Podial Secretions in Adhesion in Two Species of Sea Stars (Echinodermata).

Individuals of Asterias rubens and Marthasterias glacialis use their podia in locomotion, anchorage, and feeding. Each podium consists of a stem with a disk at its tip. The stem allows the podium to lengthen, flex, and retract, and the disk allows the podium to adhere to the substratum. Adhesion of sea star podia seems to rely on the epidermal secretions of the disk and not on a mechanical sucker-like operation. The disk epidermis is made up of five cell types: nonciliated secretory cells (NCS cells) of two different types (NCS1 and NCS2), both containing granules that are at least partly mucopolysaccharidic in composition; ciliated secretory cells (CS cells) containing small granules of unknown content; nonsecretory ciliated cells (NCS cells); and support cells. The epidermal cells of the podial disk are presumably functioning as a duogland adhesive system that is involved in an adhesive/deadhesive process. The following model is presented. Adhesive secretions are produced by NCS1 and NCS2 cells (both of them have extruded some of their secretory granules in attached podia). These secretions constitute a layer of adhesive material between the podium and the substratum, this layer being the footprint left by the podium after it has become detached from the substratum. Deadhesion, on the other hand, would be due to CS cell secretions. All these secretions would be controlled by stimuli perceived by the two types of ciliated cells (receptor cells), which presumably interact with the secretory cells via the nerve plexus.