Zygote Germination Research Articles

INFLUENCE OF CRUDE OIL ON THE DEVELOPMENT OF ZYGOTES AND SEEDLINGS OF THE GREEN ALGA <i>ULVA LACTUCA</i> (ULVACEAE) OF THE BARENTS SEA

The ability of the zygotes and seedlings of the green alga Ulva lactuca to develop under the influence of crude oil in various concentrations was determined. The oil at a content of 1 mg/l in water does not have a damaging effect on the germination of zygotes and their further development into multicellular seedlings. Juvenile seedlings have a wider tolerance range than zygotes and can develop under oil content in marine water up to 3 mg/l. The real threat of the death of zygotes, which rovide replenishment of ulva algae thickets in the littoral with young thalli, is observed under short-term oil pollution of more than 25 mg/l, or with constant oil pollution of more than 5 mg/l. The oil content in water of 25 mg/l and above stops the development of most seedlings by irreversibe destroying the structure of their cells.

Features of the population structure of Fucus vesiculosus L. (Phaeophyceae) in different areas of the Murmansk coast of the Barents Sea, 2021

The age, size-mass and reproductive structure of the Fucus vesiculosus population in the areas of the Murmansk coast has been studied on boulder soils of the middle horizon of the littoral with weakened surf. In the studied areas characterized by different temperature conditions and the degree of anthropogenic impact the biomass of F. vesiculosus does not differ statistically significantly. The minimum population size has been found in the middle knee of the Kola Bay, the maximum – at the eastern stations. In the warmer Pechenga and Retinskaya Bays, receptacles form on thalli with more branching than in the Zelenetskaya and Yarnyshnaya Bays; thalli in western Murman have a lower age of appearance of receptacles. The linear dimensions of thalli and their mass do not differ in the studied areas. The beginning of zygote germination judging by the size of seedlings is the same on the entire coast. Maximum fertility is observed at the age of 2.5–3 years. Differences between the coenopopulations of F. vesiculosus in Western and Eastern Murman indicate the important role of climate in shaping the appearance of littoral vegetation. The current level of pollution in the Kola Bay has a negative impact on the survival of fucus seedlings, however, it can be concluded that the life span of thalli is high and the population is stable.

Cryopreservation of vegetative cells and zygotes of the multicellular volvocine green alga Gonium pectorale

BackgroundColonial and multicellular volvocine green algae have been extensively studied recently in various fields of the biological sciences. However, only one species (Pandorina morum) has been cryopreserved in public culture collections.ResultsHere, we investigated conditions for cryopreservation of the multicellular volvocine alga Gonium pectorale using vegetative colonies or cells and zygotes. Rates of vegetative cell survival in a G. pectorale strain after two-step cooling and freezing in liquid nitrogen were compared between different concentrations (3% and 6%) of the cryoprotectant N,N-dimethylformamide (DMF) and two types of tubes (0.2-mL polymerase chain reaction tubes and 2-mL cryotubes) used for cryopreservation. Among the four conditions investigated, the highest rate of survival [2.7 ± 3.6% (0.54–10%) by the most probable number (MPN) method] was obtained when 2.0-mL cryotubes containing 1.0 mL of culture samples with 6% DMF were subjected to cryogenic treatment. Using these optimized cryopreservation conditions, survival rates after freezing in liquid nitrogen were examined for twelve other strains of G. pectorale and twelve strains of five other Gonium species. We obtained ≥ 0.1% MPN survival in nine of the twelve G. pectorale strains tested. However, < 0.1% MPN survival was detected in eleven of twelve strains of five other Gonium species. In total, ten cryopreserved strains of G. pectorale were newly established in the Microbial Culture Collection at the National Institute for Environmental Studies. Although the cryopreservation of zygotes of volvocine algae has not been previously reported, high rates (approximately 60%) of G. pectorale zygote germination were observed after thawing zygotes that had been cryopreserved with 5% or 10% methanol as the cryoprotectant during two-step cooling and freezing in liquid nitrogen.ConclusionsThe present study demonstrated that cryopreservation of G. pectorale is possible with 6% DMF as a cryoprotectant and 1.0-mL culture samples in 2.0-mL cryotubes subjected to two-step cooling in a programmable freezer.

MENGENAL MAKROALGA Turbinaria DAN PEMANFAATANNYA

KNOWING MACROALGAE Turbinaria AND THEIR USAGES. Turbinaria is a member of brown macroalgae that can be found in almost all Indonesian waters. Turbinaria is often regarded as Sargassum because they have a similar morphology and both belong to the Family Sargassaceae. There are 35 species of Turbinaria in the world and 11 of which can be found in Indonesia. Turbinaria conoides, T. decurrens and T. ornata are the most common ones in Indonesian waters. Turbinaria grows on rocky intertidal coastlines, tidepools and reef flats. The life cycle of Turbinaria occurs through alternations between sexual and asexual. Their biphasic life cycle contains gametophytes and sprorophytes. In their sexual reproduction, sporophytes resulted from the germination of zygotes formed by fusion (fertilization) of male and female gametes. Asexual spores develop from both sporophytes and gametophytes. Turbinaria was commonly used as a source of alginate, fucoidan and other bioactive polysaccharides (antioxidant, anticancer, antibacterial, anticoagulant, antiploriferative and antivirus activities).

Hybridization between Japanese and North American Chlamydomonas reinhardtii (Volvocales, Chlorophyceae)

SummaryThe microalga Chlamydomonas reinhardtii is a model organism whose whole genome has been sequenced. Although considered a cosmopolitan species, only eastern North American isolates of C. reinhardtii were available before 2010, when new Japanese isolates were reported. In the study describing the new Japanese isolates, zygote formation between Japanese and North American strains was shown, but germination was not demonstrated. In this study, the germination of intercontinental hybrid zygotes was examined using wild‐type Japanese strains and mutant American strains that cannot utilize nitrate. Several clonal progeny strains were established, and the progeny strains were screened based on mating type and nitrate utilization to confirm their hybrid nature. The establishment of four intercontinental hybrid strains with different phenotypic combinations was confirmed by sequencing mating type‐specific and nitrate reductase‐related genes. The potential for hybrid formation between Japanese and North American strains suggests the existence of a worldwide mating population of C. reinhardtii.

Effects of dopamine, a compound released by the green-tide macroalga Ulvaria obscura (Chlorophyta), on marine algae and invertebrate larvae and juveniles

:Dopamine is produced by Ulvaria obscura, a bloom-forming green alga that occurs from the mid intertidal to the shallow subtidal zones of North Pacific and North Atlantic shores. Its concentrations in the alga are approximately 0.5–1% of the alga's fresh mass. When experimentally desiccated and rehydrated at ecologically realistic densities, Ulvaria releases dopamine, resulting in seawater dopamine concentrations that can exceed 500 μM. Thus, dopamine could be responsible for previous reports of bioactivity by Ulvaria exudates. We tested this hypothesis by measuring the effects of dopamine in seawater on co-occurring macroalgae and crab larvae and juveniles. We ran assays that examined the effects of a range of dopamine concentrations on the growth of the green alga Ulva lactuca, on the germination of zygotes of the brown alga Fucus distichus, and on the survival, time to metamorphosis and time to first molt of crab (Metacarcinus magister and Cancer oregonensis) larvae and juveniles. Dopamine began to inhibit Fucus germination at concentrations above 5 μM, Ulva growth at concentrations above 50 μM, and the survival of Metacarcinus zoeae at concentrations above 168 μM. It did not affect the survival of Cancer megalopae or juveniles or the time to metamorphosis of megalopae. It had no effect on the time to first molt of Cancer juveniles, except at the highest concentration tested (738 μM), where it delayed molting by an average of a day and a half. These toxic effects could have been due to the dopamine or to its oxidation products. We concluded that the large-scale release of dopamine by U. obscura following stressful environmental conditions could significantly affect co-occurring species in intertidal pools as well as intertidal and shallow subtidal marine communities where the alga can form large blooms.

Life history ofChaetomorpha valida(Cladophoraceae, Chlorophyta) in culture

AbstractThe marine green algaChaetomorpha valida(Cladophoraceae, Cladophorales) has been found for several years fouling aquaculture ponds along coasts of the cities Dalian and Rongcheng, P.R. China. Using unialgal culture and microscopic observation, we completed the life history ofC. validaand discovered fragment regeneration for the first time. The life history comprised an isomorphic alternation between gametophytes and sporophytes typical of most other members of the Cladophorales. Male and female gametes fused into zygotes then developed into sporophytes. Quadriflagellate zoospores produced by mature sprophytes gave rise to new gametophytes that yielded biflagellate gametes. Germination of zygotes and zoospores was of the bipolar erect type, as observed in some other species ofChaetomorpha. Fragment regeneration occurred in both gametophytes and sporophytes via production of polar algal fragments, which directly developed into entire filaments to give a self-replicating gene­ration. The results improved understanding of the seasonal abundances ofC. validain spring and summer.

Uniparental Inheritance of Chloroplast DNA Is Strict in the Isogamous Volvocalean Gonium

BackgroundA problem has remained unresolved regarding the exceptions to the unilateral inheritance of chloroplast DNA (cpDNA) from MT+/female in Chlamydomonas and other volvocaleans demonstrated by the previous genetic analyses. For identification of the parental types of cpDNA, these studies used parents that have differences in restriction fragment length polymorphisms and exhibit partial sexual incompatibility.Methodology/Principal FindingsIn the present study, we used sexually compatible parents of the isogamous colonial volvocalean Gonium maiaprilis that seemed an ideal species to identify the pattern of cpDNA inheritance based on the length difference in the putative group I intron interrupted in the Rubisco large subunit gene and objective identification of mating types by the presence or absence of the minus-dominance (MID) gene. We examined patterns of inheritance of cpDNA and presence/absence of a MID ortholog (GmMID) in 107 F1 progeny of G. maiaprilis that were obtained by inducing germination of separated single zygotes. The results demonstrated no exception of the uniparental inheritance of cpDNA from the MT+ parent (lacking GmMID) in sexually compatible or genetically less divergent strains of G. maiaprilis.Conclusions/SignificanceThe present data suggest that the uniparental inheritance of cpDNA is likely more strict in crossings of less diverged strains or sexually compatible parental volvocaleans, and some genetic inconsistency between the parents may cause exceptional uniparental inheritance of cpDNA.

Mating and Tetrad Separation of &lt;em&gt;Chlamydomonas reinhardtii&lt;/em&gt; for Genetic Analysis

The unicellular green alga Chlamydomonas reinhardtii (Chlamydomonas) has become a popular organism for research in diverse areas of cell biology and genetics because of its simple life cycle, ease of growth and manipulation for genetic analysis, genomic resources, and transformability of the nucleus and both organelles. Mating strains is a common practice when genetic approaches are used in Chlamydomonass, to create vegetative diploids for analysis of dominance, or following tetrad dissection to ascertain nuclear vs. organellar inheritance, to test allelism, to analyze epistasy, or to generate populations for the purpose of map-based cloning. Additionally, genetic crosses are routinely used to combine organellar genotypes with particular nuclear genotypes. Here we demonstrate standard methods for gametogenesis, mating, zygote germination and tetrad separation. This protocol consists of an easy-to-follow series of steps that will make genetic approaches amenable to scientists who are less familiar with Chlamydomonas. Key parameters and trouble spots are explained. Finally, resources for further information and alternative methods are provided.

Zygote germination in Pleodorina starrii (Volvocaceae, Chlorophyta)

Zygote germination of the anisogamous/oogamous colonial green flagellate Pleodorina starrii was observed. After the zygotes were transferred to the usual, illuminated conditions from the dark treatment on the agar plate, they began to germinate. The germinating zygotes gave rise to one or two viable biflagellate gone cells. This type of zygote germination is rare in the colonial Volvocales and may characterize a certain lineage within the anisogamous/oogamous members of the colonial Volvocales.

개다시마 Kiellmaniella crassifolia Miyabe (Phaeophyta)의 암.수배우체 분리 및 보존

We examined the conditions for separating and preserving the male and female gametophytes of Kjellmaniella crassifolia. The highest percentage of zygote germination (85%) was on semi-solid medium composed of 1.0% transfer gel agar at <TEX>$15\;^{\circ}C$</TEX> and <TEX>$20\;{\mu}mol/m^2/s$</TEX> after a 4-week culture. Zygote germination in PESI liquid medium was 93.5% at <TEX>$20\;^{\circ}C$</TEX> and <TEX>$20\;{\mu}mol/m^2/s$</TEX>. The maximum zygote growth was <TEX>$252{\pm}19.7\;{\mu}m$</TEX> on 1.0% transfer gel agar at <TEX>$15\;^{\circ}C$</TEX> and <TEX>$40\;{\mu}mol/m^2/s$</TEX> after 5-week culture, and was <TEX>$76.7{\pm}2.8\;{\mu}m$</TEX> in PESI liquid medium at <TEX>$20\;^{\circ}C$</TEX> and <TEX>$40\;{\mu}mol/m^2/s$</TEX>. The respective numbers of separated male and female gametophytes from germinated zygotes were 157 and 93 on 1.0% transfer gel agar and 14 and 28 in PESI liquid medium. The maximum growth of separated male and female gametophytes was <TEX>$575{\pm}28.3\;{\mu}m$</TEX> at <TEX>$5\;^{\circ}C$</TEX> and <TEX>$60\;{\mu}mol/m^2/s$</TEX> and <TEX>$686{\pm}35.4\;{\mu}m$</TEX> at <TEX>$20\;^{\circ}C$</TEX> and <TEX>$20\;{\mu}mol/m^2/s$</TEX> in PESI liquid medium after 3 weeks, respectively. The highest percentage fertilized was <TEX>$93.3{\pm}5.8%$</TEX> at <TEX>$15\;^{\circ}C$</TEX> and <TEX>$20\;{\mu}mol/m^2/s$</TEX> in PESI liquid medium. These results show that the best conditions for the separation and preservation of gametophytes (male and female) consisted of culturing on 1.0% transfer gel agar at <TEX>$15\;^{\circ}C$</TEX> and <TEX>$20\;{\mu}mol/m^2/s$</TEX>.

The toxicity of Irgarol 1051 and Sea-Nine 211 to the non-target macroalga Fucus serratus Linnaeus, with the aid of an image capture and analysis system

A programme of research was initiated to provide information on the efficacy of the antifouling agents Irgarol 1051™ and Sea-Nine 211™ to zygotes of the fucoid alga Fucus serratus Linnaeus. The development of methodologies that incorporated the use of image capture and analysis, which minimised the problems of subjectivity and bias that can be typical, are also described. The generation of macro-instructions allowed rapid generation and interpretation of accurate quantitative data. Emphasis was placed on the use of methodologies that did not rely on the subjective determination of spore germination. Consequently, affects due to the biocides were determined that, otherwise, would not have been realised through normal analytical routines. Zygotes were exposed, in static non-renewal systems, for 3 days, to a range of concentrations of biocide dissolved in seawater. Data for seven zygote parameters were recorded daily and values of percentage germination were calculated. From these data, NOEC values of 8 μg l −1 were calculated for both Sea-Nine 211 and Irgarol 1051. An EC 50 of 19.4 μg l −1 was also determined for Sea-Nine 211. It is, therefore, suggested that F. serratus zygote germination and early development is unaffected by published environmental concentrations of these biocides. Additionally, it has been demonstrated that the image capture and analysis methodologies developed provide an invaluable tool for use in future bioassays.

Electron microscopy of sexual reproduction in Nephroselmis olivacea (Prasinophyceae, Chlorophyta)

SUMMARY The electron microscopy of zygote formation and the early stages of zygote germination in Nephroselmis olivacea Stein are presented. Although the gametes differ behaviorally during the early stages of gamete fusion, the alga is isogamous. The minus gamete settled on the substrate, and attached with its left side. The plus gamete swam to the minus gamete, attached ventral to the right side of the minus gamete, while slightly on its left side, and plasmogamy started. No specialized organelle for gamete fusion was seen using either scanning or transmission electron microscopy. Gametic fusion was uniform; the right side of the minus gamete always fused with the ventral, slightly left side of the plus gamete, which suggests the participation of the d-rootlets of the flagellar apparatus of the two gametes. Body scales were retained throughout the entire sexual process. Before karyogamy, a network of endoplasmic reticulum developed between the nuclei. This position corresponded to the contractile vacuole of the plus gamete. Fusion proceeded as the minus gamete was drawn to the plus gamete and resulted in a hemispherical zygote. Fibrous material appeared on the cell surface, embedding the body scales to form a layer that thickened and contributed to the strong adhesion of the zygote to the substrate. During this stage, karyogamy was completed. A thick zygotic wall composed of two layers, an electron-dense outer layer and a straticulate electron-lucent inner layer developed beneath the layer of fibrous material and scales. Zygote germination was induced. After the first meiotic division, the layer of fibrous material and scales ruptured and the inner layer of the zygotic wall thinned, allowing the emergence of two germ cells. They had newly formed scales and two starch grains, but no typical pyrenoid.

Phototropin is the blue-light receptor that controls multiple steps in the sexual life cycle of the green alga Chlamydomonas reinhardtii.

Blue light as an environmental cue plays a pivotal role in controlling the progression of the sexual life cycle in the green alga Chlamydomonas reinhardtii. Phototropin was considered a prime candidate for the blue-light receptor involved. By using the RNA interference method, knockdown strains with reduced phototropin levels were isolated. Those with severely reduced levels of this photoreceptor were partially impaired in three steps of the life cycle: in gametogenesis, the maintenance of mating ability, and the germination of zygotes. These observations suggest that phototropin is the principal sensory molecule used by this alga for the control of its life cycle by light.

Comparison of Eudorina/Pleodorina ITS sequences of isolates from nature with those from experimental hybrids

Internal transcribed spacer (ITS) regions of nuclear ribosomal repeats were compared among 50 Eudorina and Pleodorina isolates and two Volvox species known to clade with Eudorina species. Of the six major subclades found, four containing Eudorina and Pleodorina illinoisensis isolates, one containing Eudorina and Pleodorina indica, and one containing Volvox gigas and V. powersii, the basal branching order remains uncertain, but the positioning of isolates known to mate was always as nearest neighbors on the terminal branches of the tree. Four hybrid clones from a cross of E. elegans with P. illinoisensis, known from chromosome counts to be products of the failure of meiosis at zygote germination, contain both parental ITS repeat regions, as expected. However, they have in addition both crossover and other variant ITS cistrons among their many repeats of ITS. Such variation is limited to terminal regions of helices, as recognized from knowledge of RNA transcript secondary structure. Proper alignment then utilizes all of the nucleotide positions; the hybrid variants appear in positions intermediate between their parents in the tree. In fact, such variants seem to be hallmarks of recent hybridization events, since they were not found in any of the other 50 isolates.

A New Idea of Sinian Doushantuo‘ an Metaphyte Fossils from Xiuning, Anhui Province

AbstractThis paper gives a summary of the fossil data from the Sinian Doushantuo' an Lantian Formation in the Xiuning area, Anhui Province, China, and reports the genital butt structure comparable to that of some modern advanced algae and the fossil structure comparable to the dehiscence of sporangia as well as the ontogenetic phenomenon occurring possibly from the formation of embryos by zygote germination to the production of sessile benthonic algal adults. This phenomenon furnishes a useful clue to the further study of the life history of alternate generations in advanced algae and provides megafossil evidence for the sex differentiation of metaphytic algae.

Morphology and evolution of sexual reproduction in the Volvocaceae (Chlorophyta)

Morphological features of sexual reproduction in the Volvocaceae are reviewed, focusing particularly on gametic union and zygote gemination. Both of the two conjugating gametes of the isogamous generaPandorina, Volvulina andYamagishiella bear a tubular mating structure (mating papilla), and plasmogamy is initiated by union of the papillae tips. On zygote germination, a single biflagellate gone cell is released from the zygote wall. Although all the anisogamous and oogamous genera of the Volvocaceae produce “sperm packets” during gametogenesis and a single gone cell at zygote germination, some difference can be recognized in the male gametes. The male gametes ofEudorina bear a tubular cytoplasmic protuberance (putative mating papllla) near the base of the flagella, whereas such a structure recognized at the light microscopic level is not evident inPleodorina andVolvox. Evolution of the sexual reproduction characteristics of volvocacean algae is discussed on the basis of recent cladistic analysis of morphological data as well as of the ribosomal (r) RNA phylogeny and large subunit of the ribulose-1, 5-biphosphate carboxylase/oxygenase(rbcL) gene trees.

Effect of sewage effluents on germination of three marine brown algal macrophytes

Inhibition of germination of zygotes of the fucoid macroalgae Hormosira banksii and Phyllospora comosa and zoospores of the laminarian Macrocystis angustifolia was used as an end-point to assess the toxicity of three sewage effluents of differing quality. For each species, between-assay variation was low and results of tests with the reference toxicant 2,4-dichlorophenoxyacetic acid suggested that results are reproducible, especially in R. comosa. Each species showed a greater sensitivity to primary-treated effluent than to secondary-treated effluent, and higher variability in response to the primary effluent. High variation in response for each species when exposed to the primary effluent (compared with that for the secondary effluent) is presumably indicative of variation in quality of the primary effluent. The capacity to reproduce these assays, the sensitivity of species employed, and the ecological relevance of germination as a toxicological end-point suggest that germination tests of this nature may be useful in biological testing of effluent quality at discharge sites in south-eastern Australia.

Genes involved in light control of sexual differentiation in Chlamydomonas reinhardtii.

Gamete formation requires the sequential action of two extrinsic cues, nitrogen deprivation and blue light. The mutants described here are specifically altered in the light-dependent step. Mutations lrg1, lrg3, and lrg4 overcome this light dependence while mutation lrg2 results in a delayed execution of the light-mediated step. The four mutations are linked. The recessive nature of the lrg1, lrg3, and lrg4 mutations implies that they encode elements of negative control in this light response pathway. Analyses of diploids suggest an interaction between the gene products of the mutated loci with a central role for lrg4. The lrg4 mutation is unique also because it overcomes the light dependence of Chlamydomonas zygote germination when present in homozygous form. These data indicate that there are common components in the signal chains that control gametogenesis and zygote germination.

MORPHOLOGY AND PAEDOGAMOUS SEXUAL REPRODUCTION IN CHLOROGONIUM CAPILLATUM SP. NOV. (VOLVOCALES, CHLOROPHYTA)1

ABSTRACT Morphology and sexual reproduction in Chlorogonium capillatum Nozaki, Watanabe &amp; Aizawa sp. nov. (Volvocales, Chlorophyta) originating from Miyatoko Mire, Japan, were studied under controlled laboratory conditions. Vegetative cells of this new species were fusiform with blunt anterior and posterior ends, and they had a massive parietal chloroplast and numerous contractile vacuoles distributed throughout the protoplast. Several to many pyrenoids were randomly distributed in the chloroplast, but they disappeared under the light microscope when grown photoheterotrophically. During asexual reproduction, the first division took place transversely without a preceding rotation of the parental protoplast. In sexual reproduction, the parental protoplast divided successively to form 32 or 64 small, biflagellate isogametes. After gametogenesis, the gametes did not escape from the parental cell (gametangial) wall, within which pairs of the adjoining gametes fused to form quadriflagellate zygotes. Such zygotes were then released from the parental cell wall and developed into hypnozygotes, which at maturity developed numerous thin spines or hairs on the zygote wall. On zygote germination, four biflagellate germ cells were released from the zygote wall separately. This type of gametic union, “paedogamy,” has not previously been described in the green algae except for Chlorococcum echinozygotum Starr. Chlorogonium capillatum can be clearly distinguished from other described species of Chlorogonium by its numerous contractile vacuoles and blunt anterior and posterior ends in vegetative cells as well as by its unique sexual reproduction, in which paedogamous conjugation occurs, and numerous thin spines or hairs that develop on the hypnozygote walls.