ULTRASTRUCTURE DURING DEVELOPMENT OF THE NUCLEUS OF BATOPHORA OERSTEDII (CHLOROPHYTA; DASYCLADACEAE)1

The primary nucleus of Batophora oerstedii J. Agardh like that of the related Acetabularia, undergoes a great increase in size throughout vegetative development. Formation of small secondary nuclei represents an irreversible stage in the development of reproductive structures in the gametangia. Changes observed in the course of the life cycle include: i) an increase from 3 to 200 μm diam, of the nucleus; ii) increase in number of nucleoli; iii) development of the perinuclear region; iv) increased pore density in the nuclear membrane; v) development of chromosomes in the nucleoplasm; vi) formation of secondary nuclei; and, vii) division of secondary nuclei.     

Life History,Reproductive Morphology and Development of the Antarctic Brown Alga Desmarestia menziesii J. Agardh*

The life history, reproduction and development of Desmarestia menziesii J. Agardh from Antarctica is described. Unilocular sporangia occur singly or in small groups in the outermost cortical layer of the sporophyte. They are formed by periclinal division of cortex cells into a stalk cell and the sporangium initial. Meiospores germinate into dioecious microscopic filamentous gametophytes. As in other perennial Antarctic species of the Desmarestiales, gametangia are formed in culture under short-day conditions or in darkness. In nature, juvenile sporophytes should therefore be formed in winter. They develop only attached to the oogonium. At first they are uniseriate and elongate by means of an intercalary meristem located in their middle part. Laterals are formed predominantly in this region, and they subsequently give rise to secondary laterals. The branching pattern is opposite to alternate in both young and adult plants. Cortication of the main axis is initiated by filaments growing out from the lowermost cells of the primary laterals. In sporophytes of this developmental stage the meristem of the main axis is confined to a small region where cortication starts and above. Lateral branches elongate and become corticated in the same way as the main axis. In mature plants, cells of the inner cortex can become meristematic again and form a meristoderm which contributes to axis thickness by periclinal and anticlinal divisions. The observations are discussed in relation to possible evolutionary relationships in the genus Desmarestia and in the order Desmarestiales.     

Behavior of nuclei during zoosporogenesis in Bryopsis plumosa (Bryopsidales, Chlorophyta)

The behavior of nuclei during zoosporogenesis in Bryopsis plumosa (Bryopsidales, Chlorophyta) was examined by fluorescence and electron microscopy. Each mature filamentous sporophyte had a single lenticular nucleus, which was about 25 m in diameter and embedded in a thick cytoplasmic layer. At the commencement of multinucleation, giant nuclei with large vacuolated nucleoli, giant nuclei containing chromosomes, and dumbbell-shaped nuclei were observed. Sometimes, two small nuclei also appeared in the thick cytoplasm where the giant nucleus had presumably been present. Electron microscopy revealed the existence of ribbon-like structures resembling synaptonemal complexes within the nucleus having a large vacuolated nucleolus. Nuclei extended their distribution by repetitive divisions. A pair of centrioles was adjacent to the interphase nucleus. When the nuclei were distributed throughout the cell, they became localized nearly equidistantly from one another, each being surrounded by several chloroplasts. At this stage, many centrioles lay along the nuclear surface. The bulk of cytoplasm was then divided into many masses of protoplasm, each of which developed into a uninucleate, stephanokontic zoospore with a whorl of flagella.     

Chromosome behavior in the primary nucleus ofAcetabularia calyculus as revealed by epifluorescent microscopy

Summary Chromosome behavior preceding secondary nuclei formation within a giant primary nucleus (50–100 m in diameter) inAcetabularia calyculus was observed by the fluorescence emitted from 4-6-diamidino-2-phenylindole (DAPI)-stained DNA.Throughout the period when the large nucleolus was present in the primary nucleus, thin chromonemata were observed twining around the nucleolus. Nuclear division was initiated by degeneration of the sausage-shaped nucleolus into a number of spherical subunits soon after the initiation of cap formation. On the fourth day of cap development, the chromonemata became thicker and chromomeres appeared. They accumulated adjacent to the single spherical nucleolus. The lump of chromosomes became loosened and thick chromosomes were scattered in the nucleus. The peculiar shapes of chromosomes which suggest the existence of chiasmata were frequently observed until the chromosome segregation started. This sequence of chromosome behavior seems to be the prophase of meiotic division. Chromosome segregation, the first meiotic division, occurred on the seventh day of cap development, probably being accompanied by the second meiotic division. Immediately after nuclear division of the primary nucleus, secondary nuclei were formed and cyst formation started 24 hours after repeated mitoses of the secondary nuclei.     

LIGHT AND ELECTRON MICROSCOPIC OBSERVATIONS OF PREFERENTIAL DESTRUCTION OF CHLOROPLAST AND MITOCHONDRIAL DNA AT EARLY MALE GAMETOGENESIS OF THE ANISOGAMOUS GREEN ALGA DERBESIA TENUISSIMA (CHLOROPHYTA)1

Gametogenesis in male and female gametophytes was studied by light microscopy and EM in the dioecious multinucleate green alga Derbesia tenuissima (Moris & De Notaris) P. Crouan & H. Crouan, where male and female gametes differ in size. Gametogenesis was divided into five stages: 32 h (stage 1), 24 h (stage 2), 16 h (stage 3), 8 h (stage 4), and 0.5 h (stage 5) before gamete release. At stage 1, the first sign of gametogenesis observed was the aggregation of gametophyte protoplasm to form putative gametangia. At stage 2, gametangia were separated from the vegetative protoplasm of gametophytes. Morphological changes of nuclei and organelles occurred at this early stage of male gametogenesis, and organelle DNA degenerated. At stage 3, male organelle DNA had completely degenerated, whereas in female gametangia, organelle DNA continued to exist in both chloroplasts and mitochondria. Gametogenesis was almost completed at stage 4 and fully at stage 5. Small male gametes had a DNA‐containing nucleus and a large mitochondrion and one or several degenerated chloroplasts. The mitochondria and plastids were devoid of DNA. The large female gametes had a nucleus and multiple organelles, all of which contained their own DNA. Thus, degeneration of chloroplast DNA along with morphological changes of organelles occurred at male gametogenesis in anisogamous green algae (Bryopsis and D. tenuissima), in contrast with previous studies in isogamous green algae (Chlamydomonas, Acetabularia caliculus, and Dictyosphaeria cavernosa) in which degeneration of chloroplast DNA occurred after zygote formation.     

ANATOMY OF SEEDLING ROOTS OF PSEUDOTSUGA MENZIESII

The seedling root system of Pseudotsuga menziesii (Mirb.) Franco consists of the primary root, active long laterals, long laterals that become mycorrhizal, and short roots that may or may not become mycorrhizal. Numerous adventitious roots arise from the pericycle in young roots and from the vascular cambium and pericycle in older roots following pruning. All actively growing apices have a single plate of initials, a complex zonation of mother cells, and a similar pattern of primary tissue differentiation. Short roots and mycorrhizal short roots have 2 plates of initials, one producing the stele and the other the root cap and cortex, and differentiation occurs close to the apex. Primary and adventitious roots are usually triarch, while long laterals are usually diarch as are all short roots. The latter lack secondary xylem, but mycorrhizal short roots may produce a small amount of secondary phloem.     

The fluorescence localization of mouse satellite DNA in interphase nuclei

Nuclei from various mouse tissues exhibit a pattern of fluorescence characteristic of the cell type when stained with the fluorescent compound Hoechst 33258. When such preparations are hybridized in situ with ³H-RNA complementary to the A-T rich satellite of mouse, it is clearly seen that only the fluorescent regions of the nuclei contain the satellite DNA. Thus Hoechst 33258 allows the precise localization of satellite DNA at all stages of the mouse cell cycle.     

Cytophotometric and autoradiographic evidence for functional apomixis in a gynogenetic fish,Poecilia formosa and its related,triploid unisexuals

Summary Amounts of DNA in individual Feulgen-stained nuclei from squash preparations of ovaries and testes from wild-caught and laboratory-reared stocks of Poecilia spp. were determined with an integrating microdensitometer. The DNA content of primary spermatocytes (4C) at zygotene, pachytene, or at metaphase I (3.3–3.4 pg) was approximately twice that found in secondary spermatocytes (2C) and four times that found for young spermatids (1C). Rarely, mature sperm were found with 2C DNA amounts. Nuclei from follicular epithelium and oogonia from both bisexual and diploid unisexual fish contained about 1.6–1.7 pg DNA; whereas, the DNA content of primary oocyte nuclei was about 3.5–3.7 pg DNA, indicating that just one cycle of chromosomal replication had occurred in these cells during the period of DNA synthesis before the visible onset of meiotic prophase. Similar results were obtained for triploid unisexuals whose 6C primary oocyte nuclei contained 5.0–5.1 pg DNA, which was twice the DNA content of 3C oogonia and follicular epithelial cells (2.4–2.5 pg DNA). Autoradiographic studies, designed to monitor the incorporation of ³H-thymidine by oogonia and primary oocytes in vivo and in vitro, also showed that there is no additional synthesis of DNA during the course of meiotic prophase in these unisexual fish. Therefore, we conclude that apomixis, not endoreduplication, is the cytological basis of reproduction in Poecilia formosa and its related, triploid biotypes.     

Synaptic behaviour of the tetraploid wheat Triticum timopheevii

Triticum timopheevii (2n=4x=A^tA^tGG) is an allotetraploid wheat which shows a diploid-like behaviour at metaphase-I. The synaptic process was analyzed in fully traced spread nuclei at mid-zygotene, late-zygotene and pachytene. The length and type of synaptonemal complexes, as well as the number of bivalent and multivalent associations, were determined in each nucleus. A high number of bivalents per nucleus was detected at all three stages. Nuclei at pachytene showed a lower frequency of multivalents than did zygotene nuclei, which suggests the existence of a pairing correction mechanism. At metaphase-I only homologous bivalents and, rarely, one pair of univalents were observed. Similarities between the diploidization mechanism of T. timopheevii and that of allohexaploid wheat, controlled by chromosome 5B, are discussed.     

The lid forming apparatus in cysts of the green algaAcetabularia mediterranea

Summary Cross sections and cross tangential sections of 1 to 3-day-old cysts (gametangia) ofAcetabularia mediterranea were examined by electron microscopy. In a defined zone of the peripheral cytoplasm of the cysts, where the lid is to be formed, a characteristic circular band-like structure, the putative lid forming apparatus, can be identified. In 1 -day-old cysts this structure is characterized by two electron dense amorphous layers close and parallel to the plasma membrane. In 3-day-old cysts the lower layer consists of rod-like structures. The position of the circular band-like lid forming apparatus is correlated to the position of the cyst organizing secondary nucleus which occupies a non central position. Usually the center of the lid forming apparatus lies on the shortest line between the secondary nucleus and the cyst wall. This suggests that the cyst organizing secondary nucleus plays an important role in the formation of the cyst lid.     

MORPHOLOGY AND SYSTEMATICS OF SCIUROTHAMNION STEGENGAE GEN. ET SP. NOV. (CERAMIACEAE,RHODOPHYTA) FROM THE INDO‐PACIFIC1

A new ceramiaceous alga, Sciurothamnion stegengae De Clerck et Kraft, gen. et sp. nov., is described from the western Indian Ocean and the Philippines. Sciurothamnion appears related to the tribe Callithamnieae on the basis of the position and composition of its procarps and by the majority of post‐fertilization events. It differs, however, from all current members of the tribe by the presence of two periaxial cells bearing determinate laterals per axial cell. Additionally, unlike any present representative of the subfamily Callithamnioideae, no intercalary foot cell is formed after diploidization of the paired auxiliary cells. The genus is characterized by a terminal foot cell (“disposal cell”), which segregates the haploid nuclei of the diploidized auxiliary cell from the diploid zygote nucleus. The nature of three types of foot cells reported in the Ceramiaceae (intercalary foot cells containing only haploid nuclei, intercalary foot cells containing haploid nuclei and a diploid nucleus, and terminal foot cells containing only haploid nuclei) is discussed.     

Accumulation of peroxidase in the cap rays of Acetabularia during the development of gametangia.

Accumulation of peroxidase was demonstrated by light and electron microscopy to occur in Acetabularia in certain regions of the cap rays in relation to the development of the gametangia (cysts). Peroxidase was found to be incorporated into special, cell wall-like obstructions that separate the cap rays from the stalk when the secondary nuclei have settled in the cap rays. It is assumed that peroxidase acts as an anti-microbial protectant of the gametangia.     

The role of microtubules in establishing nuclear spatial patterns in multinucleate green algae

Summary In uninucleate cells, cytokinesis follows karyokinesis, thereby reestablishing a specific nucleus-to-cytoplasm ratio. In multinucleate cells, cytokinesis is absent or infrequent; no plasmalemma boundary defines the cytoplasmic territory of an individual nucleus. Several genera of large multinucleate green algae were examined with epifluorescence light microscopy to determine whether the patterns of cytoplasmic organization establish nuclear cytoplasmic domains. Randomly spaced nuclei, singular mitotic events and cytoplasmic streaming characterize the organization of two genera,Derbesia andBryopsis (Caulerpales). The cells ofValonia, Valoniopsis, Boergesenia, Ventricaria (Siphonocladales), andHydrodictyon (Chlorococcales) display regularly spaced nuclei which undergo synchronous divisions in a stationary cytoplasm. In the cytoplasm of genera with regularly spaced nuclei, microtubules radiate from all nuclei in late telophase-early interphase. These internuclear microtubule arrays are not found in algal genera with randomly spaced nuclei. It is hypothesized that these microtubule arrays play a role in establishing the cytoplasmic domain of each nucleus in genera with regularly spaced nuclei. Loss of microtubule arrays during the events of mitosis correlated positively with the increasing randomization of nuclear patterns in algae grown in microtubule inhibitors. Cytoplasmic domains were maintained when cells were grown in the same media in the dark. This suggests that, after a round of division, regular nuclear spacing in certain multinucleate algae is reestablished by internuclear microtubule arrays, which are not, however, required to maintain spacing during interphase.Dedicated to the memory of Professor Oswald Kiermayer     

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.     

Ion currents associated with root tips, emerging laterals and induced wound sites in Nicotians tabacum: spatial relationship proposed between resulting electrical fields and phytophthoran zoospore infection

Abstract Growing roots of Nicotiana tabacum var. Havana generate transcellular ion currents which traverse developing and wounded tissues. Positive current of around 10 mA m^?2 enters meristematic and elongating cells at the tip of primary roots. The growing tips of first order laterals are also traversed by a similar positive current with a density of around 2.0 mA m^?2, as are immature laterals emerging at the primary root surface. These self-generated ion currents flow basipetally through developing tissues and leave from mature non-elongating tissue. A large positive current of around 70 mA m^?2 also enters induced wound sites on the primary root surface. Motile zoospores of the fungal pathogen Phytophthora parasitica var. nicotianae have been reported to associate preferentially with these regions of the root. This might suggest that electrotaxis may be part of the mechanism by which zoospores locate root regions susceptable to fungal infection.     

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.     

18S rDNA AND EVOLUTION IN THE DASYCLADALES (CHLOROPHYTA): MODERN LIVING FOSSILS1

Phylogenetic relationships were inferred from parsimony and distance analyses of nuclear small-subunit ribosomal DNA sequences taken from 14 species representing 8 of the 11 extant genera in the Dasycladales. Of 1733 aligned positions, 412 (23.8%) were variable and 251 (61%) of those were phylogenetically informative within the Dasycladales. Secondary structure was analyzed and taken into account during all phases of data analysis. Robustness of the trees was assessed using bootstrap analysis and g₁ statistics of tree-length decay. Strongly supported branches were robust to all methods of analysis regardless of weighting schemes used. The secondary structure of the 18S within the Dasycladales agrees with that of other green algae with the exception of a shared deletion in stemloop E10-1 (ca. 13 nucleotides long), which provides additional support for the uniqueness of this monophyletic group. A molecular clock was calibrated from the dasyclad fossil record and suggests a radiation of the Acetabulariaceae at 120 ± 30 million years (Ma) ago and the Dasycladaceae 215 ± 40 Ma ago. The split of the two lineages from a shared ancestor is estimated at 265 ± 50 Ma ago. Within the Dasycladaceae, Neomeris and Cymopolia are sister taxa, as are Batophora and Chlorocladus. Bornetella groups with the Neomeris and Cymopolia clade in 78% of the bootstrap replicates. Relationships among the Acetabulariaceae show that Acetabularia and Polyphysa do not form monophyletic groups as presently circumscribed. No evidence indicates that Acicularia is the oldest genus. Halicoryne, Chalmasia, and Dasycladus were not included in the analysis. Molecular data provide afresh background perspective from which to discuss the evolution of one of the most ancient lineages of green plants.     

Spermatogenic ultrastructure in the anomalodesmatan bivalve Myochama anomioides (Mollusca: Myochamidae) – does the nucleus help position the ‘temporary’ acrosome?

Spermatogenic ultrastructure in the marine bivalve mollusc Myochama anomioides (Myochamidae) is described and contrasted with other bivalves, especially other euheterodonts. Small (0.1 μm diameter), primary proacrosomal vesicles produced in spermatocytes give rise to much larger (0.4 μm diameter) secondary proacrosomal vesicles in early spermatids, which in turn form the dished‐shaped, definitive acrosomal vesicle (diameter 1.0 μm) of later spermatids. The acrosomal vesicle acquires a deposit of subacrosomal material and comes to lie close to or in contact with the plasma membrane. The acrosomal complex (acrosomal vesicle + subacrosomal material) initially positions itself at the apex of the condensing, fibrous nucleus (the so‐called temporary acrosome position), but subsequently begins to move posteriorly. The condensing nucleus becomes markedly folded so that its apex is posteriorly orientated towards the migrating acrosomal complex and the midpiece (mitochondria and centrioles). The close spatial relationship of nuclear apex to acrosomal complex during this folding strongly suggests that acrosomal migration in M. anomioides is assisted, at least in part, by movement of the late spermatid nucleus. Similar nuclear folding has previously been demonstrated in an early stage of fertilization in another anomalodesmatan (Laternula limicola) raising the possibility that one event might be a reversal of the other.