Embryogenesis and acquisition of algal symbionts by planulae of Xenia umbellata (Octocorallia: Alcyonacea)

Early embryogenesis of the internally brooding soft coral Xenia umbellata and acquisition of algal symbionts in the course of its planular ontogenesis have been examined by scanning and transmission electron microscopy and by light microscopy. The endoderm of adult X. umbellata harbours symbionts mainly in the tentacles and in the peripheral solenia system. The colonies are gonochoric brooders. Algal symbionts were never found in the sperm sacs, and were only rarely found in the follicular tissue enclosing the oocytes. Fertilized eggs pass into endodermal brood pouches where embryogenesis occurs. Cleavage is holoblastic and leads to formation of a solid blastula. Algal symbionts are conspicuously embedded in the parental mesoglea that coats the young embryo, most probably transmitted by surface adherence. At a further stage, this integument disappears and the algae reside extracellularly among the cells of the newly-formed blastula. After subsequent cell proliferation developing planulae possess an inner mass of yolk-laden cells that contain numerous symbiotic algae. Gradually the yolk disintegrates, leaving a cavity enclosed by ectoderm, a thin mesoglea and an inner endoderm with intracellular symbionts. The mature planulae have already been provided with numerous intracellular symbionts by the time they are expelled from the brood pouches. The markedly early symbiont acquisition by the embryos of X. umbellata may help support their developmental requirements in the course of planular ontogenesis.     

Reproduction in Anthelia glauca (Octocorallia: Xeniidae). II. Transmission of algal symbionts during planular brooding

The soft coral Anthelia glauca Lamarck, 1816, of the family Xeniidae, is found on the reefs of KwaZulu-Natal, South Africa. Its gastrodermal cells contain numerous endosymbiotic unicellular algae (zooxanthellae). A. glauca is a gonochoric species that simultaneously broods its planulae within the pharyngeal cavity of the polyps. Symbiotic algae appear with zygote formation within the pharynx, embedded in amorphous material. The algal cells adhere to the ciliated ectodermal surface of immature planulae and are most probably endocytosed by them. Zooxanthellae are translocated towards the basal part of the ectoderm. Gaps are subsequently opened in the mesoglea into which symbionts surrounded by ectodermally derived material, including plasma membrane, pass. The basal membrane of endodermal cells disintegrates, and the algae bulge into spaces formed in the underlying endoderm. Throughout the process, each zooxanthella resides within a vacuolar membrane in the detached ectodermal cytoplasm. The acquisition process is essentially one in which zooxanthellae are translocated from the pharyngeal cavity into the ectoderm and then through the mesoglea into the endoderm, culminating in the final symbiotic state. The direct transmission of symbiotic algae to the eggs or larvae probably provides the most efficient means whereby zooxanthellae are acquired by the host progeny. Received: 15 July 1997 / Accepted: 25 February 1998     

Bud formation and metamorphosis inCassiopea andromeda (Cnidaria: Scyphozoa): A developmental and ultrastructural study

Asexual reproduction by formation of swimming buds which metamorphose directly into polyps plays a most important role in the propagation ofCassiopea andromeda (Cnidaria: Scyphozoa). (C. andromeda polyps, originally supplied by the Löbbecke Museum and Aquarium Düsseldorf, FRG, were cultured in our laboratories.) We have defined five budding stages and investigated epithelial recruitment and dynamics during bud formation using intracellular vital stains. The region of cell recruitment was found to encircle the budding site asymmetrically. The aboral side contributing considerably less to the developing bud than the oral and lateral sides. Furthermore, it was found that the epithelial flow involved in bud formation is part of a permanent apico-basal displacement of ectodermal cells. Light and electronmicroscopic investigations revealed that no drastic changes occur at the cellular level, neither in the ectoderm nor in the endoderm which both participate in bud formation. Scanning and transmissionelectron microscopic investigations of the swimming bud revealed that the ectoderm is composed of three, and the endoderm of two, cell types. Nerve elements have been detected near the mesoglea between both ecto- and endodermal cells. Amoebocytes are regularly found either at the basis of epidermal cells or within the mesoglea, whereas symbionts are located in the endoderm. Buds induced to metamorphose by a bacterial-inducing factor were used to investigate morphological and ultrastructural changes occurring during metamorphosis and scyphistoma morphogenesis. Metamorphosis starts with the settling of a bud, followed by the formation of the pedal disk in which desmocytes, as typical cnidarian adhesive structures, are differentiated. Metamorphosis is completed with the formation of the mouth and tentacles. Whereas metamorphosis of cnidarian planulae implies considerable changes at the cellular level, tissue remodeling processes prevail in bud metamorphosis ofC. andromeda.     

The functional morphology and development of the alimentary tract of larval and juvenile barnacles (Cirripedia: Thoracica)

The alimentary tract of the nauplius larva of Balanus spp. consists of cuticle-lined foregut and hindgut, with intervening endodermal midgut constricted into anterior and posterior regions. The anterior midgut cells in the region of the constriction (constriction cells) secrete proteins (probably digestive enzymes). The remaining anterior midgut cells, often containing lipid droplets, form the absorptive region of the tract. Glycoprotein globules and lipid droplets within anterior midgut cells are the remants of the yolk in a pre-hatched larva, this yolk additionally supporting the larva through the non-feeding first nauplius stage. Nauplius Stages II to VI are actively feeding planktonic stages which increase in size and build up lipid reserves. These accumulated reserves support the non-feeding cyprid, first through its planktonic life and then through settlement and subsequent metamorphosis to the juvenile barnacle. Juvenile barnacles start to feed between 2 and 5 days after metamorphosis.     

Reproduction and development in the pycnogonid Pycnogonum littorale

Aspects of the reproductive biology of Pycnogonum littorale (Ström) are examined. Vitellogenesis occurs between November and August, the process resembling that in Nymphon gracile, annelids, and Limulus polyphemus, in which the majority of the yolk is synthesized within the oöcyte, with only a small contribution from outside, thus contrasting with the method in insects. The mode of formation of the mature yolk platelets shows similarities with the process described in Cambarus spp. Growth occurs in the post larva by a series of moults, and sometimes in the adult by a process not involving moults. The male usually has 9 moults and the female 10 or 11.     

Slow developing demersal embryos and larvae of the antarctic sea star Odontaster validus

The early development of Odontaster validus at McMurdo Sound, Antarctica, is indirect and includes equal cleavage, a convoluted blastula, a free-swimming coeloblastula, a gastrula, and a feeding bipinnaria larva. Development differs from that of other asteroids in two respects: (1) The developmental rate is extremely slow; blastulae form nearly 2 days after fertilization, gastrulation begins after 7 days, and the bipinnaria develops in about 40 to 55 days. The slow developmental rate appears to be only partly related to the low environmental temperature (-1.5°C). (2) The embryos and larvae are largely demersal. Such behavior may be an adaptation to keep the larvae out of antarctic surface waters, as does brooding in many other polar echinoderms.     

Development,settling behaviour,metamorphosis and pentacrinoid feeding and growth of the feather star Florometra serratissima

Embryonic development of the northeastern Pacific feather star Florometra serratissima takes place within a ridged fertilization membrane. Cleavage is radial, resulting in a coeloblastula, and gastrulations is by invagination. Cilia are swollen terminally during ciliogenesis whereas fully grown cilia possess several swellings along the length of their shafts. Young doliolaria larvae begin to hatch from the fertilization membranes 35 h after fertilization (9.5° to 11.5°C); by 4 d the doliolaria has acquired ciliated bands, a vestibular invagination and an antero-ventral adhesive pit. The surface of the larva is covered with a delicate glycocalyx supported by microvilli. Larvae swim along a vertical sinusoidal path just below the water surface; they begin to explore the substratum at 4.5 d and settlement begins as early as 4.6 d, but can be delayed for up to 9 more days. Larvae settle gregariously in culture and it is suggested that gregarious settlement plays a role in the formation and maintenance of adult aggregations of F. serratissima. Metamorphosis into a stalked cystidean following settlement is rapid. Major changes at this period include: loss of cilia; withdrawal of ectoderm from the glycocalyx; covering over of the vestibular invagination; and a 90 degree rotation of the vestibule to the former posterior end of the doliolaria. Transformation from cystidean to pentacrinoid includes the opening of the 5 oral plates, the extension of the 15 papillate tube feet and further elongation of the stalk. The pentacrinoid is able to feed on small food particles. Rudiments of all 10 adult arms are present by 4 months; at 6 months the pentacrinoid has an arm span of 6.5 mm but cirri and pinnules are not yet present.     

Unusual lecithotrophic development of the caribbean brittle star Ophiothrix oerstedi

The ophiuroid Ophiothrix oerstedi Lütken spawned in the laboratory at Barbados, West Indies, from August, 1975 until early December. The embryo passes through a wrinkled blastula stage, and the larva is a reduced lecithotrophic ophiopluteus with a shortened pelagic existence. A larva of this type is unusual for brittle stars in general and unique for ophiothricids for which development has been described. Metamorphosis is completed 4.3 days (24° to 27°C) after fertilization with a single pair of ciliated larval arms, the postero-lateral arms, being retained as a swimming device for the late larva. Settlement, with subsequent separation of the postero-lateral arms from the young brittle-star, begins as early as 4.5 days, but can be delayed for at least one week, at the end of which time midwater separation can occur resulting in the pelagic dispersal of post-larvae. A comparison of gametic and larval characteristics of O. oerstedi with the literature suggests that the larva of this species is most closely allied to the abbreviated developers. The adaptive significance of this larval form is discussed.     

X-ray diffraction study of calcification processes in embryos and larvae of the brooding oyster Ostrea edulis

X-ray powder diffraction was used to study shell calcifications of the oyster Ostrea edulis, sampled in the Limski Kanal, Istria (Adriatic Sea), in May 1992. All the developmental stages were followed, from the embryonic stage through the transition between the trochophore and veliger larva (prodissoconch I and II) and later, after swarming, the pelagic free-swimming larval stages, up to their settlement and attachment (from the D-shaped to the fully formed pediveliger larva), and finally during metamorphosis and juvenile stages (dissoconch). In the first gastrula stage, only an amorphous tissue is present (a periostracum and organic matrix). The beginning of shell formation (at the end of gastrulation) in early trochophores is manifested by the appearance of calcite (up to 1–7% of total volume) and then aragonite (about 1%). In the later stage of the veliger larva the fraction of calcite decreases as well as the amorphous fraction, while the fraction of aragonite rapidly increases. In the prodissoconch II stage and during the whole pelagic period aragonite is dominant, accompanied by a very small amorphous fraction and traces of calcite. The shell mineral composition does not change until metamorphosis, whereupon the fraction of calcite rapidly increases and the fraction of aragonite decreases. The postmetamorphic valves of the juvenile and adult oyster consist mainly of calcite, except the resilium and myostracum which remain aragonitic, possibly as a continuation of the inner layer of the larval shell. Received: 28 May 1997 / Accepted: 1 July 1997     

Effects of temperature on yolk utilization,initial growth,and behaviour of unfed marine fish-larvae

The effect of temperature on yolk utilization, initial growth and behaviour of larvae of four species of marine fishes, i.e.,Acanthopagrus schlegeli, Engraulis japonica, Pagrus major andParalichthys olivaceus, was investigated under laboratory conditions at Hiroshima in 1989. The yolk sac was absorbed earlier with increasing temperature for all species. Morphological characters such as pectoral fin appearance, mouth opening and eye pigmentation differed from species to species over the range of experimental temperatures, and the sequence of development of these characters varied with temperature even within the same species. Temperature did not have a large influence on the maximum growth of unfed larvae, but had a clear effect on the time to starvation, occurring earlier at higher temperatures. Larval behaviour as indicated by time spent moving was also influenced by temperature; behavioural activity was greater at higher temperatures for all species examined. The effect of temperature on the early stage of larval life is discussed in terms of behavioural and morphological developments, as well as on yolk utilization, and its influence on larval survival in nature and under rearing conditions is evaluated.     

Development of the echinothurid sea urchin Asthenosoma ijimai

Development of an echinothurid sea urchin is described for the first time; eggs of Asthenosoma ijimai (Yoshiwara) have been fertilized in the laboratory, and development has been followed light-microscopically up to the early juvenile stage. The eggs, which are orange and float, are 1.2 mm in diameter, by far the largest echinoid eggs on record. The embryos, larvae and early juveniles are lecithotrophic, and no exogenous food is needed for development. The embryonic stages include a remarkable biscuit-shaped late blastula, which has never been described for any echinoid before. The larva, with its para-arms unsupported by skeletal ossicles, is unlike the echinoid prism or pluteus and more resembles the early bipinnaria or early auricularia of asteroids and holothurians, respectively. All stages through late larvae float just beneath the surface until settlement occurs during the third and fourth weeks at 20°C. Metamorphosis from late larva to juvenile is gradual and no part of the larval body appears to be cast off.This paper is dedicated to Professor Juro Ishida on the occasion of his seventieth birthday.     

Localization of bioluminescence in the siphonophore Nanomia cara

The base of the tentacle of the developing physonect larva (Nanomia cara) has a bioluminescent region. The ability to produce light in the larva is transitory; this ability first appears at about two days of development and is disappearing by eight days, as the larva begins to feed. Subsequently paired bilaterally symmetrical bioluminescent organs are found on the nectophores and the bracts of the adult colony. In both the larva and the adult, bioluminescence is mediated by a calcium specific photoprotein. In all cases the photocytes lack a green fluorescent protein.     

Route of egg yolk protein uptake in the oocytes of kuruma prawn,Penaeus japonicus

The route of egg yolk protein uptake into the oocytes of kuruma prawn, Penaeus japonicus, was studied using immunohistochemical and electron microscopical methods. Although a significant immunofluorescence with anti-vitellin-immunoglobulin was observed in the enlarged follicle cells surrounding oil globule stage oocytes of the early vitellogenic ovary, no fluorescence was detected in shrunken follicle cells surrounding oocytes in the yolk granule stage. Electron microscopically, yolk granule stage oocytes have an irregular surface with numerous well-developed microvilli. In contrast, the surface of follicle cells is relatively smooth. The irregular surface of yolk granule stage oocytes was covered with a layer of electron dense material. Similar dense material was found in the spaces between the neighboring follicle cells on the yolk granule stage oocytes. The outer surface of the follicle cells on yolk granule stage oocytes was covered by dense materials which were similar to those found on the irregular surface of oocytes. Micropinocytotic vesicles containing dense material were found in the ooplasm near the irregular surface with numerous well-developed microvilli. Dense material was concentrated in the peripheral part of the small forming yolk bodies of yolk granule stage oocytes. This suggests that the electron dense material, probably egg yolk protein, transferred to the surface of yolk granule stage oocytes from the spaces between the neighboring follicle cells may be incorporated into the ooplasm by pinocytosis through the microvilli and subsequently aggregate to form yolk bodies.     

Ovarian morphology and oogenesis in Aurelia aurita (Scyphozoa: Semaeostomae): ultrastructural evidence of heterosynthetic yolk formation in a primitive metazoan

In Aurelia aurita, the ovaries arise as horseshoe-shaped evaginations of the gastrodermis in the floor of four interradial gastric pouches. Germ-cell islands arise within endodermally-derived gastrodermal cells. Oocytes grow and gradually bulge into the mesoglea while maintaining physical contact throughout vitellogenesis with specialized cells called trophocytes. Ultrastructural changes suggest that these cells transport yolk precursors from the coelenteron to the oocytes in a manner similar to that reported for the trophonema cells of anthozoan ovaries. Vitellogenesis involves both the autosynthetic activity of the oocyte organelles (Golgi complex and rough endoplasmic reticulum) and the heterosynthetic incorporation of precursors through endocytotic processes involving both coated pits and vesicles and smooth-surfaced tubules. Ultrastructural data suggest that different types or classes of yolk precursors enter the oocyte through the trophocytes and via the surrounding mesoglea. To our knowledge, this is the most primitive animal in which this type of yolk synthesis has been described. The trophocyte-oocyte relationship in oocytes of A. aurita is reminiscent of the trophonema-oocyte relationship in anthozoans and supports the belief that the Anthozoa and Scyphozoa share a close phylogenetic relationship.     

Determination of copper in embryos and very young specimens of Sepia officinalis

The total amount of copper in embryos and newly hatched young individuals of Sepia officinalis L. has been determined by microtechnique, using bathocuproine-sulfonate as complexing reagent. During embryonic life, the total amount of copper does not change; it remains at a level close to 3.8 g. The copper is found in the yolk sac of very early embryos; it is subsequently transferred into the embryo proper. After hatching, the copper content diminishes quickly in starved individuals. Fed S. officinalis also usually lose copper. The reason for this may be that the inner yolk sac of newly hatched individuals contains a great deal of the total copper, which is excreted with the yolk after the latter has become superfluous. Later on, copper must be taken up from the food. The mobilization of protein and copper from the yolk into the blood may account for the early appearance of embryonic hemocyanin in the blood.     

Oocyte development in the kuruma prawn Penaeus japonicus

Female kuruma prawns (Penaeus japonicus Bate) with undeveloped, early developing, developing, nearly ripe and ripe ovaries, were collected from Ise Bay, Japan, in 1984. Oocyte development of the kuruma prawn was classified into ten stages according to morphological characters, namely: (1) synapsis stage, (2) chromatin nucleolus stage, (3) early perinucleolus stage, (4) late perinucleolus stage, (5) oil globule Stage I, (6) oil globule Stage II, (7) yolkless stage, (8) yolk granule stage, (9) prematuration stage, and (10) maturation stage. The synapsis stage is a multiplication stage. The chromatin nucleolus stage, early and late perinucleolus stages are previtellogenesis and primary growth stages. Oil globule Stage I is an initial stage of primary vitellogenesis and secondary growth. Follicle cells on the oil globule Stage I oocytes expand rapidly and reach maximum size during oogenesis. Yolk granule stage oocytes are in the initial stages of secondary vitellogenesis. Strongly acidophilic yolk granules accumulate within basophilic vesicles of the cytoplasm. The yolk granules are first concentrated in the inner part of the cytoplasm, then gradually spread to the periphery. Cortical crypts, which are separated from the oocyte cytoplasm by the cytoplasmic membrane, are situated outside of oocyte cytoplasm. Germinal vesicle breakdown (GVBD) is initiated in the late phase of prematuration and continues until the late phase of maturation immediately prior to spawning. At the beginning of the maturation stage, the oocytes are ovulated, after which the nuclei further shrink and migrate out-wards. After ovulation, meiotic division of the ovarian oocyte progressed up to the metaphase of primary maturation division. Finally, the meiotic metaphase is visible just beneath the cytoplasmic membrane in the mature oocyte. Though ovulation is synchronous within the same ovary, GVBD is not completely synchronous. Ovulated mature oocytes have many club-shaped cortical crypts in the peripheral part of the cytoplasm and contain extensive accumulations of yolk granules dispersed throughout the cytoplasm. The apical end of the club-shaped cortical crypts and cytoplasmic membrane are coated by the vitellin envelope in the mature oocyte.     

Preliminary observations on the circulatory system in eggs and early larvae of some teleostean fishes

Observations on the pattern of blood circulation in eggs and early larvae of Ambassis, Mugil, Dorosoma and Thrissocles species have been made. In the initial stages, the tubular heart lies on the left side, near the cephalic region. The anterior end of the heart lies near the snout. The posterior end is attached to a point mid-way between the orbits and auditory vesicles; the position of the anterior end is greatly influenced by the size of yolk. Initially, the tubular heart and dorsal aorta lie in a line. As a result of yolk absorption, the anterior end of the tubular heart gradually sinks to a position posterior to the attached end. Liquified yolk is circulated through the heart in early stages; blood corpuscles become apparent only in larvae. The heart-beat, which is initially irregular, gradually becomes regular as development progresses. In the initial stages the blood flows to the caudal and not to the cephalic region. The blood vessels are narrow, and the oval-shaped blood corpuscles must therefore pass through individually, with their long axes parallel to the passage. A definite capillary system is not evident. The blood vessels can dilate and contract when required.     

Permeability of the oral epithelial layers in cnidarians

In order to characterize the permeability of the oral epithelial layers in cnidarians, we investigated the kinetics of transport of labelled ions (⁴⁵Ca,²²Na,³⁶Cl) and organic molecules (¹⁴C-inulin-carboxyl,¹⁴C-ala) through the oral tissue of two cnidarian species,Anemonia viridis (Forsskål, 1775) andHeliofungia actiniformis (Quoy and Gaimard, 1833) using the Ussing chamber method. In both species, unidirectional Ca, Na and Cl fluxes were the same in both directions (ectoderm towards endoderm and vice versa), the net flux being equal to zero. The insensitivity of these unidirectional transepithelial fluxes to metabolic inhibitor (1 mM sodium cyanide) and calcium channel inhibitor (100 M verapamil) and their linear dependence on calcium concentration suggest that these fluxes are simple driven by diffusion via a paracellular pathway. The epithelial layers were not permeable to inulin. Low-molecular weight amino acids such as alanine did not cross the epithelia but were absorbed by the ectoderm. The permeability coefficients indicate that the oral epithelial layers are leaky. It is suggested that the coelenteric cavity represents a compartment in which the ionic pool can be entirely renewed by simple diffusion. This process seems efficient enough to meet all calcium requirements in scleractinian corals.     

The use of the embryo‐culture techniques in the research on the Teratogenic and Mutagenic properties of metals†

Actually, embryos can be cultured from the one‐cell stage up to the blastocyst stage, and their development can be easily monitored at any time: severe effects caused by toxic compounds are traduced by rapid embryonic death, less pronounced effects can be expressed by a lowered cleavage activity or by an arrest of the development from a particular stage. The system can be improved by transferring the embryos at the blastocyst stage in another more complete medium where they can “implant”; and form an inner cell mass with differentiated ectoderm and endoderm. Since last years, it has also become possible to culture postimplantation rodent embryos for short periods involving a number of particularly critical stages of organogenesis, such as the formation and closure of the anterior neuropore. Embryo‐culture also represents a useful system to study cytogenetic effects of chemicals which are often linked to lethal or teratogenic effects. These different possibilities are illustrated by examples of studies already performed with metals, and dealing with their teratogenic and/or cytogenetic effects on pre‐ and postimplantation rodents.