Auxosporulation of Licmophora communis (Bacillariophyta) and a review of mating systems and sexual reproduction in araphid pennate diatoms

The auxosporulation of Licmophora communis is allogamous and dioecious. Pairing between sessile, shortstalked cells of compatible clones is followed by meiosis and gametogenesis, to form two gametes in each gametangium. The behavior of the gametes differs between the gametangia. In the male gametangium, the gametes detach from the frustule, round up, and migrate out of the gametangium after its dehiscence at the broader, unattached pole. In the female gametangium, both gametes remain attached to the adjacent theca over almost their whole length and do not move. Plasmogamy therefore occurs within the female gametangium and this is where the zygotes are formed and remain. After fertilization, the zygotes detach from the thecae of the female gametangia, contract, and become ellipsoidal, before expanding parallel to the apical axis of the gametangium. We review the types of auxosporulation in other pennate diatoms and the systems used for classifying these. Dioecy and cis‐type anisogamy (in which one gametangium produces active gametes and the other produces passive gametes), as in L. communis, are probably primitive within the pennate group (although there is no information on the AsterionellopsisRhaphoneis clade). However, size can also be restored in various araphid pennates by allogamous sexual reproduction involving the formation of only one gamete per gametangium, or in rare cases by automixis or (apparently) vegetative enlargement.     

Sexual auxosporulation of the marine diatom Navicula directa var. directa

Sexual auxosporulation was observed in a mixed culture of two strains of Navicula directa var. directa. Two gametes did not re‐arrange in their gametangium and each adhered to the inner surfaces of the gametangial theca. Each of the two gametes of one gametangium fused with a gamete of the other gametangium iso‐gamously. As a result, two zygotes and hence two auxo‐spores were produced per paired gametangia. As the gametangial thecae kept close to the gametes during fusion, the zygote became associated with two different thecae. The presence of type IB2a of Geitler's (1973) system was confirmed by the present observations.     

Patterns of sexual reproduction in diatoms

Sexual reproduction takes many forms within the diatoms. The variation has been classified by several authors, but in most cases the distinctions between their main categories have depended on the number of gametes produced per gametangium (and thus on how many zygotes per pair of copulating cells), and upon whether fusion is oogamous, anisogamous or isogamous. These classifications are not themselves an adequate basis for taxonomic comparison, which should be based on individual characteristics of the sexual process. Diatoms seem to be primitively oogamous. In araphid pennate diatoms and some raphid diatoms the gametes and gametangia are morphologically alike but physiologically distinct; one gametangium produces active gametes and the other passive ones. This may be the primitive condition in pennate diatoms, providing a link to the oogamy of centrics via the morphological anisogamy of Rhabdonema Kütz.     

SEXUAL REPRODUCTION IN NAVICULA CRYPTOCEPHALA (BACILLARIOPHYCEAE)1

Homothallic sexual reproduction and auxosporulation were studied in monoclonal cultures and seminatural populations of the freshwater epipelic diatom Navicula cryptocephala Kütz. Gametangia paired via the girdle, one gamete was formed per gametangium (and hence one zygote per pair of gametangia), and gamete fusion took place without the formation of any copulation envelope or copulation canal. Superfluous nuclei from meiosis survived unusually long, so that gametes and young zygotes were probably functionally polyploid; later, all but two haploid nuclei degenerated. Expanded auxospores had a swollen center, but during formation of the initial valves, the auxospore contracted away from the perizonium to produce linear‐lanceolate valves. The pattern of reproductive behavior found in N. cryptocephala can be classified as type IIA2a auxosporulation in Geitler's system. The same type of zygote and auxospore formation seen in clonal cultures was observed in seminatural material from four lakes in Scotland and the Czech Republic. Variation in nuclear structure and auxosporulation in the N. cryptocephala species complex is discussed, as is the evolution of type II auxosporulation (one zygote per pair of gametangia) from type I auxosporulation (two zygotes per pair). The penalty of smaller numbers of zygote produced in type II may be outweighed by formation of larger auxospores (prolonging the vegetative phase) or more vigorous auxospores. The variation present among members of the N. cryptocephala complex indicates that biogeographical analyses based on use of the name N. cryptocephala, as performed recently to support the ubiquity hypothesis of protist distributions, are almost meaningless.     

THE ULTRASTRUCTURE OF DERBESIA TENUISSIMA (DE NOTARIS) CROUAN. I. ORGANIZATION OF THE GAMETOPHYTE PROTOPLAST,GAMETANGIUM, AND GAMETANGIAL PORE1,2,3

The fine structure of vegetative and reproductive gametophytes of Derbesia tenuissima is described. Development of the gametangium and release of the gametes progress as follows: (1) In initial stages of gametangium formation, prior to 24 hr before gamete release, there is an accumulation and proliferation of nuclei, chloroplasts, and other organelles. (2) This is followed by separation of the gametangium from the rest of the plant by a gametangial membrane; segregation of organelles into gametes has begun by 12 hr before release and the process is completed by 2.5 hr before release. (3) Enzymatic wall dissolution of the pore area occurs between 2.5 and, 12 hr before normal lights-on time. (4) The release mechanism appears to be an instantaneous light-induced increase in lurgor pressure rupturing the weakened pore area, of the wall and causing a forcible expulsion of the gametes. (5) Following release, the pore is sealed by organellar debris and the gametangial membrane. Additional wall layers are presumed to be laid down internal to the plugged pore by the vegetative protoplasm which migrates into the area.     

GAMETOGENESIS AND GAMETE RELEASE OF ULVA MUTABILIS AND ULVA LACTUCA (CHLOROPHYTA): REGULATORY EFFECTS AND CHEMICAL CHARACTERIZATION OF THE “SWARMING INHIBITOR”1

Gametophytes of Ulva mutabilis Føyn and Ulva lactuca L. were artificially induced to form gametangia by removal of sporulation inhibitors. After this treatment, U. mutabilis gametes were ready for swarming on the third morning after induction, while U. lactuca gametangia needed 1–2 d longer for maturation. Release of gametes of U. lactuca was dependent solely upon exposure to the first light in the morning. Gametangia of U. mutabilis, however, also required sufficient dilution of the swarming inhibitor (SWI). SWI was excreted transiently by both Ulva species early during gametogenesis. While the SWI concentration in U. mutabilis medium remained above the inhibitory concentration until the gametangia were mature, the concentration of U. lactuca‐SWI dropped rapidly below this level. In the presence of sufficient SWI, mature gametes of U. mutabilis remained motionless within the gametangia despite light and open exit pores. However, using SEM, an additional seal was detected within these pores, which probably prevented premature swarming until dilution of SWI and exposure to light. Observations by time lapse microscopy and experiments with the myosin kinase inhibitor BDM suggest that the gametes may be either extruded by the gametangium or leave the exit pore by active gliding motion, driven by a myosin‐like motor protein. The SWIs were purified from both Ulva species, and mass spectral analysis showed their molecular masses (292 Da) were identical.     

Gamete discharge by Bryopsis plumosa (Codiales, Chlorophyta) induced by blue and UV-A light

Gamete discharge by the coenocytic green alga Bryopsis plumosa (Hudson) C. Agardh is induced by light. The mature male gametangia consist of a mass of quiescent male gametes and a large central vacuole. Within a few minutes after the onset of irradiation, breakdown of the tonoplast of the central vacuole and initiation of gamete motility occur simultaneously. This is followed by a forced discharge of moving gametes through a hole ruptured at the subapical region of the gametangium. The action spectrum for the light-induced gamete discharge was determined from a series of fluence-response curves. This action spectrum, having two major maxima at 370 and 450 nm, indicates the involvement of a blue light/UV-A-absorbing photoreceptor previously described as ‘cryptochrome’.     

Vegetative growth and reproduction of Fossombronia brasiliensis Steph.: the influence of photoperiod,temperature and inorganic nitrogen source

Abstract

Monoclonal cultures of Fossombronia brasiliensis were grown with different photoperiods, temperatures and inorganic nitrogen sources. The subsequent vegetative growth and production of gametangia is described. A quantitative analysis of the numbers of antheridia and archegonia and their relative proportions is given. At 18°C, F. brasiliensis was found to be a short-day plant, with a critical night length of between six and twelve hours, while at 10°C it exhibited quantitative SD plant properties. Plants produced more male gametangia at 18°C and more female gametangia at 10°C. Nitrogen as nitrate consistently produced more gametangia than the ammonium source.     

Improved mating efficiency inAcetabularia acetabulum: recovery of 48–100% of the expected zygotes

Summary We have improved zygote recovery 11–1,000 fold by optimizing the physiology of gamete release and mating inAcetabularia acetabulum. Gamete release was affected by agar purity, concentration, and volume/gametangial pair. Cold pre-treatment of gametangia (14–30 d at 10°C in the dark) synchronized subsequent gamete release at 21°C in the light. Cold pre-treatment was nearly twice as effective in synchronizing subsequent gamete release when intact, gametangia-bearing caps rather than isolated gametangia were pretreated. Synchronizing gamete release doubled mating efficiency. In a wild-type laboratory strain ofA. acetabulum, there were 1,561±207 gametes/gametangium which had half-lives of 14.5 d in 0.1% seawater-agar. We recovered 48–93% of the expected numbers of zygotes from a mass mating of 8 to 1,226 gametangia and 11–128% of the expected numbers of zygotes from mating single gametangial pairs: the large range in the calculated mating efficiency may be attributable to the variation in the numbers of gametes made per gametangium. Zygote recovery from single gametangial pairs was highly dependent on the volume of mating matrix. In addition, most zygotes recovered were unattached to any other zygotes in the subsequent generation (> 95% single cells from matings of 1–500 gametangial pairs). Our improvements in mating conditions and zygote recovery (1) have facilitated cell manipulation and culture ofA. acetabulum in the laboratory; and (2) have made controlled crosses for selection and genetic analysis of mutants feasible. These advances have removed a major barrier to genetic analysis of development inAcetabularia.Abbreviations LB Luria-Bertani bacteriological broth - SE standard error of the mean - T_g agar gelling temperatures - DAPI 4,6-diamidino-2-phenylindole     

Le corps paranucléaire des gamètes géants d'Allomyces javanicus traité à l'acide borique

Summary The gametes ofAllomyces javanicus exhibit a nuclear cap essentially made of ribonucleoprotein with sulphydryl groups.When dipped in phosphate buffered solutions of boric acid M/20–M/400, the male and female gametangia ofAllomyces liberate giant male and female gametes. Such giant gametes exhibit either a single regular nuclear cap (with 2 or 3 nuclei) or a massive and irregular basophilic formation (with 4–10 nuclei).Boric acid may interfere with ribonucleic acid, essential constituent of the nuclear cap, during the cleavage of the gamete-units in the gametangia.     

Production of anisogametes and gamete motility dimorphism in Monostroma angicava

 The reproductive strategy of a marine alga with a heteromorphic biphasic life cycle was studied by analyzing various sexual reproductive characters in light of the evolution of anisogamy. Gametophytes of Monostroma angicava were dioecious and their gametes were slightly anisogamous. Volume of gametangium, density of gametangia and area of mature gametangial parts on each gametophyte did not differ from male to female. Therefore, the reproductive biomass investment for gamete production was considered to be the same for each sex. Anisogamy in this alga appeared to be derived from the difference in the number of cell divisions during gametogenesis, because the majority of male gametangia each produced 64 (2⁶) gametes and the female produced 32 (2⁵) gametes. This corresponded with measurements of cell size in male and female gametes. Further, the sex ratio was 1:1 for sexually mature plants sampled at Charatsunai. Therefore, it was suggested that in the field twice as many male gametes are released as female gametes. Liberated gametes of both sexes showed positive phototaxis. The swimming velocity of freshly liberated male gametes was a little higher than that of female gametes. Male gametes had the potential to swim for ca. 72 h and female gametes for ca. 84 h. The difference in gamete motility between the two sexes seemed to be related to cell size. Planozygotes were negatively phototactic and swam more rapidly than gametes of either sex. Received: 5 March 1997 / Revision accepted: 18 July 1997     

CULTURE STUDIES ON THE MARINE GREEN ALGA HALICYSTIS PARVULA—DERBESIA TENUISSIMA. II. SYNCHRONY AND PERIODICITY IN GAMETE FORMATION AND RELEASE

Unialgal cultures of the macroscopic, vesicular, coenocytic gametophyte (Halicystis parvula Schmitz) of Derbesia tenuissima (DeNotaris) Crouan fr. were grown under various environmental regimes to elucidate the cytology of gamete formation and the factors controlling synchronous gamete formation and release. No synchrony of nuclear division was observed in vegetative plants or during the early stages of gamete formation. In the later stages of gamete formation in plants in a light-dark cycle, nuclear divisions within any gametangium were synchronous, and the stages of gamete formation were synchronous for the population. This synchrony was not as great for plants in continuous light. Gametes of plants in a light-dark cycle were released explosively immediately following the dark-to-light transition. Release was random and much less forceful for plants in continuous light. After a certain stage of gamete formation, gamete release was timed to occur after a particular interval of darkness, but release could be triggered by light during the last portion of this interval. The length of the dark interval was shorter for male plants than for females, but the period of light sensitivity was longer for females. Formation of gametangia by series of isolated plants was also synchronous and sometimes periodic under certain conditions. Intervals between gametangia on the same plant varied from 2 to 14 days but were usually 4 or 5 days (unlike plants in nature, which show a bi- or tri-weekly periodicity). Male and female plants did not differ in synchrony or periodicity. Different media affected the number of gametangia formed over a period of time but not the synchrony of formation. Under some conditions changing the medium had a stimulating or synchronizing effect. Non-repeated temperature changes also synchronized gamete formation. Optimum temperature for continued gamete formation was about 21 C. Regular daily light and temperature variation together maintained synchronous and periodic gamete formation in populations of isolated plants. Reproduction diminished and became less synchronous at constant temperature either in continuous light or under a light-dark schedule, although in the light-dark regime steps in the formation of any given gametangium remained synchronous with the light-dark cycle. Length of times between gametangial formation on individual plants showed a tendency to occur in multiples of the usual period lengths; e.g., plants sometimes tend to “skip” intervals, thus maintaining the synchrony of the population. These results suggest that interaction between daily environmental cycles and an endogenous physiological cycle may maintain periodic reproduction.     

SEXUAL REPRODUCTIO IN CODIUM FRAGILE SSP. TOMENTOSOIDES (CHLOROPHYCEAE) FROM THE NORTHEAST COAST OF NORTH AMERICA1

Male and female gametes were formed within the same gametangiurn on reproductive plants of Codium fragile (Sur.) Hariot ssp. tomentosoides (Van Goor) Silva growing off Appledore Island in the Gulf of Maine, USA. The female gametes were approximately twice the diameter of male gametes and outnumbered male gametes in the same gametangium by approximately six times. Fusion appears to require gametes from different gametangia if not different plants.     

The Life History of Acetabularia mobii Solms-Laubach

Acetabularia mobii Solms-Laubach is a perennial and littoralalga along the coast of Karachi. Plants bear 15–18 gametangiawhich are strongly calcined. The contents of gametangia aretransformed into 64–85 thick-walled multinucleate sphericalcysts which produce anisogamous biflagellate gametes. Micro-and macro-gametes are produced in two different cysts of thesame gametangium. The germinating zygotes produce directly erectcylindrical tube-like structures which ultimately form adultplants.     

CULTURE STUDY OF REPRODUCTIVE CYCLES AND SYSTEMATICA IN MOUGEOTIA TRANSEAUI (CHLOROPHYTA)1,2

Asexual and sexual reproductive cyries are described and illustrated for a cultured homothallic strain of Mougeotia transeaui Collins. Reproduction is by aplanospores and zygospores. Aplanospore formation precedes zygospore formation and continues longer with regreening of older cultures occurring regularly from precocious germination of aplanospores. Aplanospores typically form when most of the protoplast moves into the central swollen region of a cell and two new cross walls form to delimit an aplanosporangium. Conjugation is scalariform without the movement and fusion of well-organized gametes. During zygospore maturation, three new cross walls form in the receptive gametangium and conjugation tube to produce a zygosporangium.     

Morphology and life history of Halopteris filicina (Sphacelariales, Phaeophyceae) from Korea

The vegetative morphology and life history of Halopteris filicina (Grateloup) Kutzing, collected from Korea, were examined in laboratory culture. Field plants attaining 3–5 cm in height were epilithic, tufted, yellowish-brown, and produced numerous erect axes with alternately distichous branches from compact basal discs. They were cultured under a 12:12 h LD photoperiod at 10°-C, 15°C and 20°C to observe the influence of temperature on reproduction. At 10°C plants grew only vegetatively, whereas at 15°C and 20°C they produced unilocular sporangia. Unispores released from sporangia developed into monoecious, anisogamous gametophytes that formed plurilocular female and male gametangia on the same lateral branches. The zygotes, by fusion of female macrogametes and male microgametes, developed into sporophytes bearing unilocular sporangia, whereas the unfused female gametes germinated parthenogenetically. This species was confirmed to have an isomorphic life history, basically similar to the other species of Sphacelariales.     

Induction of abnormal male gametes and androgenesis in the aquatic PhycomyceteAllomyces

Summary Induction of cleavage of the cytoplasm in the gametangium of a predominantly male strain of the aquatic PhycomyceteAllomyces under conditions of oxygen starvation, in the presence of dilute lactic acid, or dilute phosphate buffer, pH 7.0, resulted in multinucleate-multi-flagellate gametes. Under certain conditions the frequency for such abnormal gametes approached 70%. Phosphate buffer pH 7.0, at a concentration of 5 × 10^–3-1×10^–2 M and an incubation time of 90 minutes was found to be most effective in inducing multi-flagellate-multinucleate gametes. Nuclear fusion and multiple nuclear fusions were observed in these abnormal gametes as they developed into sporophytic thalli on dilute nutrient agar (YpSS/10). Multinuclear gamete development and nuclear fusion was analysed by electron microscopy. The reported observations reveal the occurrence of androgenesis from multinuclear male gametes inAllomyces.     

Female gametogenesis and female gamete germination in the anisogamous green alga Codium fragile subsp. novae‐zelandiae (Bryopsidophyceae,Chlorophyta)

Female gametogenesis was studied in the dioecious siphonous green alga Codium fragile subsp. novae‐zelandiae (J. Agardh) P. C. Silva using light and electron microscopy. Early during gametogenesis the protoplasm was uniform; then it separated in portions, while fusiform chloroplasts and nuclei increased in numbers. Some features of the nuclear divisions were similar to those of other Bryopsidophyceae. They were acentric and semi‐open. Pairs of parallel electron‐dense lines resembling synaptonemic complexes were observed in several prophase nuclei indicating meioses. In metaphase the nuclear envelope showed polar fenestrae from which the spindle emerged. No spindle microtubule nucleating material was visible and chromosome kinetochores were evident. Mature female gametes were pyriform with a hyaline anterior end from which the two flagella emerged. Mature gametes had a spherical nucleus surrounded by a mitochondrion and numerous discoid chloroplasts. Female gametes germinated parthenogenetically in culture and also inside gametangia, involving loss of flagella, rounding and lengthening of cells, multiplication of chloroplasts with well developed thylakoid systems, vacuolization and synthesis of a fibrillar cell wall.     

GROWTH AND PROBABLE GAMETE FORMATION IN THE MARINE DINOFLAGELLATE CERATIUM SCHRANKII1

Growth and formation of probable gametes in clonal cultures of Ceratium schrankii Kofoid isolated from the Gulf of Naples were monitored at 15, 20 and 25°C (12:12h LD period). Sustained division rates (k) of 0.15 div d ^?1 at 15°C and elevated ones at 20°C (0.27 div d^?1) and 25°C (0.25 div d^?1) were obtained. “Gamete” formation occurred at each temperature with few “gametes” observed at 15°C, and the greatest numbers at 20° and 25°C. This process in each parent cell involved two successive divisions with the first division forming tow “pregametes” which, within a few hours, divided again to form four motile unicells. These “gametes” remained motile for at least 24 h and normally survived for no longer tahn 48 h. Depending on the initial population at the onset of “gamete” formation, the impact on Ceratium populations was to cause either a momentary pause in log growth or a reversible decline in the populaiton.     

Preferential digestion of chloroplast DNA in male gametangia during the late stage of gametogenesis in the anisogamous algaBryopsis maxima

Summary The fate of chloroplast nuclei (cp-nuclei) and mitochondrial nuclei (mt-nuclei) was followed during gametogenesis in male and female coenocytic thalli in the anisogamous algaBryopsis maxima by epifluorescence microscopy, after staining with 4,6-diamidino-2-phenylindole (DAPI), by quantification of chloroplast DNA (cp-DNA) by fluorimetry using a video-intensified, photon-counting system (VIMPICS), and by CsCl density gradient centrifugation. The male and female coenocytic thalli, 48 h before the release of gametes, contain a large number of chloroplasts, each of which is larger in size than the cell nucleus and the mitochondria and contains about 150 cp-nuclei. The size of each chloroplast in the female and male gametangia decreases markedly during gametogenesis as a result of continuous divisions till about 10 h before the release of gametes and, eventually, the numbers of cp-nuclei per chloroplast in the male and female gametangia fall to about 20 and 5, respectively. Two hours later, as the preferential digestion of cp-DNA in the male gametangium occurs, the number of cp-nuclei in the chloroplast of each male gamete falls to zero while the number of cp-nuclei in female gamete does not change, even after release of female gametes. Several mt-nuclei are observed in all of the female gametes. By contrast, the mt-nuclei in the bulk of the male gametes disappear but those in a few gametes remain. The profiles after CsCl density gradient centrifugation of DNAs extracted from male and female plants and gametes support the cytological data. The results suggest that the preferential digestion of cp-DNA in male plants occurs about 8 h before the release of gametes and that there is differential digestion of cp-DNA and mitochondrial DNA (mt-DNA).