On the hibernation of Spongilla lacustris (L.)

Summary The osmotic pressure of the summer-sponge is about 25–30 mM NaCl. At and after gemmulation it increases to about 110 mM (in a single case 175 mM was found), due to a liberation of small organic molecules. Osmotic pressure remains constant for a time, but in January and February (i. e. before the germination) it is again reduced to the summer values. The shell allows a high hydrostatic pressure to develop and thereby prevents osmotic rupture of the gemmula. The hibernation may be divided into three periods: The prehibernation, the posthibernation and intercalated between them the hibernation proper. In the prehibernation and in the posthibernation definite changes take place in the tissue, but in the hibernation proper no changes are observed. In Nature the hibernation lasts about six months at, say, 4°, but at 22 degrees the whole hibernation will abbreviate to about 13 days.This is due to an abbreviation of all the three periods of hibernation, but whereas the pre- and the posthibernation will only be accelerated in a manner similiar to that of other physiological processes, such as processes of growth, development and metabolism, the hibernation proper will be accelerated much more, and in fact it will be almost abolished at 22°. This strange effect of temperature on hibernation proper is discussed. Since development often occurs at very low temperatures (2–5°), we may conclude that a mechanism must be present in the gemmulae which ensures development after a certain time, here 6 months. The different phases of hibernation may serve as an indicator of this mechanism. It is suggested, that the transformations taking place during prehibernation and posthibernation, and resulting in the germination of the gemmula are inhibited during hibernation proper by a substance formed in prehibernation.     

Microbial Diversity of the Freshwater Sponge Spongilla lacustris

To provide insight into the phylogenetic bacterial diversity of the freshwater sponge Spongilla lacustris, a 16S rRNA gene libraries were constructed from sponge tissues and from lake water. Restriction fragment length polymorphism (RFLP) analysis of >190 freshwater sponge-derived clones resulted in six major restriction patterns, from which 45 clones were chosen for sequencing. The resulting sequences were affiliated with the Alphaproteobacteria (n = 19), the Actinobacteria (n = 15), the Betaproteobacteria (n = 2), and the Chloroflexi (n = 2) lineages. About half of the sequences belonged to previously described actinobacterial (hgc-I) and betaproteobacterial (beta-II) sequence clusters of freshwater bacteria that were also present in the lake water 16S rRNA gene library. At least two novel, deeply rooting alphaproteobacterial lineages were recovered from S. lacustris that showed <89% sequence similarity to known phylogenetic groups. Electron microscopical observations revealed that digested bacterial remnants were contained within food vacuoles of sponge archaeocytes, whereas the extracellular matrix was virtually free of bacteria. This study is the first molecular diversity study of a freshwater sponge and adds to a growing database on the diversity and community composition of sponge-associated microbial consortia.     

Cold hardiness of the green gemmules of Spongilla lacustris L. (Porifera: Spongillidae)

Most green gemmules of Spongilla lacustris survived enclosure in ice at –20 °C for up to 30 days; however, their rate of germination at 20 °C was less rapid than that of control gemmules. The length of time spent at low temperature had little effect on gemmule survival. In contrast, repeated cooling to –20 °C and warming to 4 °C led to a progressive decline in gemmule viability. These results indicate that cold injury occurs primarily during transitions between high and low temperatures.     

Growth of Stylaria lacustris (L.) (Oligochaeta,Naididae)

The peculiarities of the growth in weight and length of Stylaria lacustris (L.) on the basis of observations in experimental vessels are considered. The growth of this species fits a parabolic curve. The equations relating weight to absolute growth rate as well as weight to duration of life are given.     

Field experiments on egg production in the fresh-water Sponge Spongilla lacustris

Short (0.3 cm) and long (1.5–2.5 cm) lengths of laboratory-stored branches of gemmulated S. lacustris were implanted in the pond of origin on three dates approximately one, two, and three months after the time of normal gemmule hatching. The sponges derived from these implants produced eggs in both old and new tissue for three to four weeks following gemmule hatching. The significance of these results with respect to the control of egg production in natural populations is discussed.     

Contractile vacuoles in cells of a fresh water sponge,Spongilla lacustris

Summary Forty or more independently functioning contractile vacuoles (CVs) occupy the central region of fresh water sponge pinacocytes. Each CV undergoes a cycle of enlargement by fusion, movement, shape change, rounding up, and emptying over the course of 5–30 min. Diameter at discharge varies between 1 and 13 m. CVs in all cell types are associated with submicroscopic coated vesicles. Filled CVs are bounded by an unmodified trilaminar membrane, but vacuoles with excess membrane frequently show coated evaginations. These evaginations are thought to pinch off as coated vesicles, providing an avenue for membrane recycling in the CV system.Supported by NIH grants AS-T01-GM-0723 and GM-23708-CBY     

Ultrastructural investigation of spermatogenesis in Spongilla lacustris and Ephydatia fluviatilis (Porifera,Spongillidae)

Summary Spermatogenesis of the spongillids investigated here is similar in Spongilla lacustris and Ephydatia fluviatilis and proceeds, on the whole, as in other Eumetazoa. Sponges however lack true sex organs, the germ cells developing from somatic cells. The male germ cells originate in spongillids from choanocytes and the female ones from archaeocytes. In Spongilla lacustris single choanocytes leave the flagellated chambers and transform into spermatogonia; in Ephydatia fluviatilis they result from differential cell division. The spermatogonia gather in distinct mesenchyme regions and are surrounded by cyst-building cells. Thus spermatocysts are built in which spermatogenesis proceeds. The spermatogonia in the spermatocysts differentiate into flagellated spermatocytes of I. order. In this process, the early appearance of the flagellum and its mode of formation are uncommon. The following meiotic divisions generate spermatocytes of II. order in the first step and spermatids in the second. In both developmental stages the cells remain connected by cytoplasmic bridges. In the subsequent spermiocytogenesis the cytoplasm of the spermatids is reduced. The reduced parts of the cytoplasm appear as cell fragments in the lumen of the spermatocysts and are eventually ingested by the cystwall cells. The mature spermatozoa arrange in the spermatocysts in a characteristic pattern. Later the spermatocysts open into the excurrent canal system and the spermatozoa leave the sponge with the egestive water stream.     

The development of Spongilla lacustris from the oocyte to the free larva (Porifera,Spongillidae)

Summary During June and July oocytes appear in well-developed specimens of Spongilla lacustris. These differentiate from archeocytes, and during the first growth phase they reach a diameter of ca. 50 m. At this time each oocyte is enclosed in a single-layered follicle epithelium, which is retained until emergence of the larva.In the second phase the oocytes grow to about 220 m by phagocytosis of trophocytes. When phagocytosis has come to an end, there is a distinct layering of the yolk material that has formed within the cytoplasm of the oocyte. Small yolk granules surround the centrally located nucleus, and peripheral to these is a layer of larger spheres of yolk.Cleavage is totally equal to unequal. Some blastomeres are binucleate. In the 15-cell staged micro- and macromeres appear.The embryo consists of uniform cells with high yolk content; at the periphery they are slightly flattened rather than spherical. In this stage of development the first scleroblasts appear.Further development to the young larva is marked by the appearance of a cavity (the larval cavity) lined with pinacocytes. The cavity expands to occupy about half the volume of the larva at emergence, becoming hemispheric in shape. The cells at the periphery of the larva form a columnar, single-layered, multiseriate ciliated epithelium with teardrop-shaped nuclei.The emerging larva breaks through its follicle and the wall of the excurrent canal system; occasionally larvae can be found in the canals. At this time the larva has developed a few flagellated chambers, which may already be integrated into the primordia of the excurrent canal system. The previously discernible scleroblasts have now formed isolated spicules, which may adhere to form spicule-spongin complexes.     

Cellular src gene product detected in the freshwater sponge Spongilla lacustris.

Serum from Rous sarcoma virus tumor-bearing rabbits immunoprecipitated from extracts of the freshwater sponge Spongilla lacustris a tyrosine-specific protein kinase with characteristics similar to the chicken pp60c-src kinase activity. An immune competition assay confirmed the relationship between the protein from sponges and viral pp60v-src.     

Microscopical aspects on symbiosis of Spongilla lacustris (Porifera,Spongillidae) and green algae

Summary When growing in the sunlight, some specimens of Spongilla lacustris are coloured green due to the presence of symbiotic unicellular chlorellae. The algae live inside most sponge cells. The chlorellae were extracted from green sponges, cultivated, added to algae-free sponges and fixed after different incubation times. In this way the uptake of the algae, their distribution and their final whereabouts in the mesenchymatic cells could be followed by in vivo microscopy, phase-contrast microscopy and electron microscopy. A few minutes after addition, the chlorellae can be found inside the choanocyte chambers. Here they are taken up by the cell bodies and collars of the choanocytes. Pinacocytes are also involved in the uptake. The distribution of algae results from a specific transmission from the donor cell to the receiver cell. The chlorellae are not released from their host vacuoles until they are extensively enclosed by the cell taking them up. Six hours after addition, all sponge cells contain algae except granulocytes, microscleroblasts, the pinacocytes of the peripheral rim region and those of the pinacoderm. The chlorellae are able to divide inside the sponge cells.Abbreviations StM Stereo-microscopical photograph - PhC Phase-contrast microscopical photograph - EM Electron microscopical photograph     

Life-cycles of lotic populations of Spongilla lacustris and Eunapius fragilis (Porifera: Spongillidae)

SUMMARY. The life-cycles of green and white morphs of the freshwater sponge Spongilla lacustris were examined in the light of past evidence that zoochlorellae may augment their sponge host's nutrition. Field collections from a lotic population of S. lacustris were supplemented by laboratory experiments on gemmule hatching and gemmule size. Both white and green S. lacustris produced sperm for a 6-week period in 1976 starting in the middle of May. Out of thirty white and thirty green sponges examined during this period, twenty white and ten green sponges contained sperm. Sperm production in both morphs was limited primarily to the basal 3.18mm of sponge tissue, and the density of sperm packets in the two morphs was the same. Out of 180 white and green sponges examined in 1976, only four eggs, no embryos, and no larvae were observed. White sponges gemmulated a week or two earlier, and produced smaller gemmules which were more uniform in size than those of green sponges. White and green gemmules hatched synchronously in the spring. In 1977 one female and numerous male specimens of S. lacustris , and numerous females but no males of another sponge, Eunapius fragilis , were found. The life-cycles are discussed in the light of other recent studies on freshwater sponges.     

The ultrastructure of cell walls in Enteromorpha intestinalis (L.) link

Using transmission electron microscopy, the process of cell wall development following cytokinesis is described. Three distinct components appear to be localized in the wall and these are discussed in relation to previous work. In older cells the walls are stratified and the origin of the layers is described. The outer layers of wall are continually being shed and it is suggested that this process prevents epiphytes persisting on this species of alga.     

Trophic ecology of a freshwater sponge (Spongilla lacustris) revealed by stable isotope analysis

The vital roles that sponges play in marine habitats are well-known. However, sponges inhabiting freshwaters have been largely ignored despite having widespread distributions and often high local abundances. We used natural abundance stable isotope signatures of carbon and nitrogen (δ ¹³C and δ ¹⁵N) to infer the primary food source of the cosmopolitan freshwater sponge Spongilla lacustris. Our results suggest that S. lacustris feed largely on pelagic resources and may therefore link pelagic and benthic food webs. A facultative association between S. lacustris and endosymbiotic green algae caused S. lacustris to have significantly depleted carbon and nitrogen signatures that may reflect carbon and nitrogen exchange between sponges and their symbiotic algae. Isotopic data from specialist sponge consumers demonstrated that sponges hosting zoochlorellae were the major component of the diet of the spongillafly Climacia areolaris and the sponge-eating caddisfly Ceraclea resurgens suggesting that the symbiosis between freshwater sponges and algae is important to sponge predator trophic ecology. Our results help define the role of sponges in freshwater ecosystems and shed new light on the evolution and ecological consequences of a complex tri-trophic symbiosis involving freshwater sponges, zoochlorellae, and spongivorous insects.     

Life cycle of Spongilla lacustris (Porifera,Spongillidae): a cue for environment-dependent phenotype

We studied the life cycle and growth of Spongilla lacustris in a stream with three distinct habitats. Sponge populations in the habitats exhibited different adaptive strategies. Growth forms of S. lacustris ranged from encrusting to digitate and branched. Environmental factors controlled the appearance of each growth form. In the most hospitable habitat, a variety of colonization strategies and different growth forms were present. In the less hospitable habitat growth was restricted to small and encrusting specimens. In the optimal habitat, the largest and most luxuriant specimens developed. Gemmulation and hatching were dephased among specimens in the three habitats; hence gemmules were present for long periods of time. S. lacustris was found capable of displaying two life strategies: r in the short run, K in the long run.     

The role of intragemmular osmotic pressure in cell division and hatching of gemmules of the fresh-water sponge Spongilla lacustris (Porifera)

Summary The gemmule coat of Spongilla lacustris is histologically single-layered in the gemmules studied in this work. This single layer is comparable to the classically described internal chitinous membrane of Leveaux (1939). It has been found to contain collagen with an axial period in electron micrographs of about 120 Å and is bounded internally by a thin dense layer which is separate from the internal gemmular cells, and which may be chitinous.Gemmules of this sponge studied during March to June of 1973 respond to 230 mOsmolar solutions of small molecules by: 1. undergoing no change, in which case the substances are freely permeable to the gemmule coat and cells; 2. displaying shrinkage of the cell mass, in which case the substances are permeable to the coat but relatively impermeable to the cells; 3. displaying folding of the coat and cell mass shrinkage because the substances are relatively impermeable to both the coat and the cells; and 4. displaying complete collapse of the gemmule due to impermeability to the coat. The lipid solubility of a substance is directly related to its ability to penetrate the coat. Further, molecular size and charge are also of apparent importance.Substances which penetrate the coat and remain osmotically active (are not metabolized) inhibit hatching. Low concentrations of sodium chloride (23 mOsmolar) have been demonstrated to reversibly inhibit hatching. Higher concentrations cause irreversible damage at 20° C but have little effect at 4° C, indicating that damage is related to the metabolic level of the cells. Once hatching is stimulated by increased temperature the cells become progressively less sensitive to an increase in osmotically active substances.Inhibition of gemmule hatching can theoretically occur by: 1. an addition of solutes to the gemmular fluid, or 2. through an increase in concentration of intragemmular solutes by water withdrawal.Our results raise the question of whether the inhibition of hatching by gemmulostasine, reported by Rasmont (1965) and Rozenfeld (1970, 1971), is due to an osmotic effect rather than to a specific physiological one.Based upon the results reported here and on the work of Zeuthen (1939) and Schmidt (1970) we propose a tight coupling between the intragemmular osmotic pressure and the triggering of hatching (cell division). Any substance which increases intragemmular osmotic pressure to a large enough extent will inhibit hatching. Furthermore, it can be hypothesized that hatching is normally triggered by a decrease in osmotic pressure due to water movement into the gemmule, the movement of solutes out of the gemmule, or to a combination of these.This work was supported by a grant from the National Science Foundation (GB-37775) to T. L. S.     

Cytoskeletal organization and cell organelle transport in basal epithelial cells of the freshwater sponge Spongilla lacustris

Summary Different antibodies against actin, tubulin and cytokeratin were utilized to demonstrate the spatial organization of the cytoskeleton in basal epithelial cells of the freshwater sponge Spongilla lacustris. Accordingly, actin is localized in a cortical layer beneath the plasma membrane and in distinct fibers within the cytoplasmic matrix. Microtubules exhibit a different distributional pattern by radiating from a perinuclear sheath and terminating at, the cell periphery; in contrast, intermediate filaments are lacking. Cytoplasmic streaming activity was studied by in-vivo staining of mitochondria and endoplasmic reticulum by means of fluorescent dyes. Single-frame analysis of such specimens revealed a regular shuttle movement of mitochondria and other small particles between the cell nucleus and the plasma membrane, which can be stopped in a reversible manner with the use of colcemid or colchicine but not with cytochalasin D. The results point to the microtubular system as a candidate for cell organelle transport, whereas the actomyosin system rather serves for changes in cellular shape and motility.     

Ingestion,digestion, and egestion in Spongilla lacustris (Porifera,Spongillidae) after pulse feeding with Chlamydomonas reinhardtii (Volvocales)

Summary The route followed by food particles in Spongilla lacustris was clarified by light and electron microscopic examination of sponges fed with Chlamydomonas reinhardtii. The algal cells are phagocytosed by prosendopinacocytes and choanocytes. After some time they are transferred to archaeocytes, amoebocytes, and lophocytes. Changes in algal structure during digestion were observed and the egestion of algal remnants was documented in life for the first time. In light micrographs, digestion of the algal cells is manifest first in shrinkage of the cells, then in disintegration to form several spherical green fragments 2–3 m in diameter, and finally, after 12–18 h, in a reddish brown discoloration of the fragments. Signs of the digestive process in electron micrographs include disappearance of the cell-wall layers, the flagella, and the pyrenoid and its starch sheath, as well as a progressive increase in the density of the cytoplasm and karyoplasm.     

Correlation of cyclic GMP and cyclic AMP immunofluorescence with cytochemical patterns during dormancy release and development from gemmules in Spongilla lacustris L. (Porifera: Spongillidae)

The spongillid freshwater sponges asexually produce an encapsulated dormant stage, the gemmule. With release from dormancy, internal, yolk-laden, binucleate thesocytes differentiate into histoblasts or archeocytes. The histoblasts emerging first from the gemmule form the initial pinacoderm of the hatching sponge. Immunohistochemistry was employed to examine the distribution of cyclic GMP (cGMP) and cyclic AMP (cAMP) following dormancy release and during gemmule germination and hatching in the freshwater sponge, Spongilla lacustris L. Cyclic nucleotide fluorescence patterns were analyzed in relation to the distribution of cytochemically demonstrable macromolecular constituents and intracellular organelles. Twenty-four hours following temperature-activated release from dormancy, cGMP fluorescence levels are elevated in thesocytes at the gemmule periphery prior to histoblast formation. The cAMP fluorescence in the gemmule also occurs first in those thesocytes differentiating into histoblasts. Cytochemical patterns in germinating gemmules are comparable with those described by Ruthmann ('65) and Tessenow ('69). However, cytochemically demonstrable events of cytodifferentiation follow the earlier appearance of cGMP and cAMP in the histoblast precursors by approximately 12 hours. In addition, cGMP appears to be associated with the membranes of cytoplasmic organelles, possibly lysosomes or lipid inclusions, in the region of vitelline platelets and with symbiotic algae. cAMP is located primarily on the membranes of the vitelline platelets and on membranes of vacuoles involved in forming the spicular skeleton These observations suggest that cGMP and cAMP are involved in the mobilization of nutrient reserves and in ion transport during dormancy release and development from gemmules in freshwater sponges.