EFFECT OF DNP ON ACCELERATION OF THE CLEAVAGE FOLLOWING INSUFFICIENT PARTHENOGENETIC ACTIVATION IN THE SEA URCHIN EGG*

Unfertilized eggs of sea urchins, Hemicentrotus pulcherrimus and Pseudocentrotus depressus, were treated with 4–5% butyric acid-sea water for 40–60 sec so that they were activated partheno-genetically without visible cortical changes. When these insufficiently activated eggs were inseminated 90–120 min after butyric acid-treatment, they divided much earlier than the control eggs in the first cleavage cycle. In the present paper, it becomes clear that if eggs are put into m /2,000-m /16,000 DNP-sea water at 60 min after insufficient activation and 30 min later, returned to normal sea water and then inseminated, they still show acceleration of the first cleavage in the same degree as the eggs which are not treated with DNP, while if eggs are exposed to DNP for 30 min prior to the insufficient activation or within 60 min after the activation, they do not show any acceleration of the cleavage. From these results, it may be concluded that some preparations for cleavage acceleration which are arrested by DNP become ready in the eggs at an early period in the first cleavage cycle and these preparations cannot be cancelled by DNP-treatment once they have been completed.     

ON THE ROLE OF CALCIUM IONS IN OOCYTE MATURATION IN THE POLYCHAETE CHAETOPTERUS PERGAMENTACEUS*

The germinal vesicle of mechanically released Chaetopterus oocytes disintegrates in natural sea water (NSW), but not in artificial sea water of normal composition (ASW), calcium-free sea water (CaFSW), magnesium-free sea water (MgFSW) or calcium and magnesium-free sea water (CaMgFSW). Several methods of inducing oocyte maturation using chemically well-defined medium have been established. (1) Germinal vesicle breakdown was induced by the treatment of immature oocytes with KCl (60 mM) in ASW or MgFSW. The presence of Ca²⁺ is necessary for inducing oocyte maturation with high potassium concentration. “Differentiation without cleavage” was observed after this treatment. (2) Trypsin (0.3%) induced oocyte maturation in ASW, but not in CaFSW. Oocytes matured in this manner developed to trochophores upon insemination. (3) Immature oocytes, treated with isotonic CaCl₂ for less than 1 min and then transferred to ASW, underwent germinal vesicle breakdown. The oocytes were arrested at the first meiotic metaphase and upon insemination developed to trochophore larvae. (4) Tetracaine (0.4 mM) induced oocyte maturation in the absence of Ca²⁺ in the medium. In ASW, CaFSW or CaMgFSW containing the drug, oocytes were arrested at the first meiotic metaphase, while in MgFSW with tetracaine they developed parthenogenetically up to the 4- and 8-cell stages. The role of calcium in oocyte maturation was established and its importance was discussed based on the results obtained with the different ways of inducing oocyte maturation.     

A correlative study of experimentally changed first cleavage and Janus development in the trunk of Platynereis dumerilii (Annelida,Polychaeta)

Summary Among zygotes of Platynereis dumerilii treated with cytochalasin B (CCB) prior to first cleavage, a wide variety of developmental effects were observed. One effect is a delay in the first cleavage. Treated embryos may skip the first or even more than one cleavage cycle and become multinucleated. Once these eggs start cleaving their cleavage plane takes the same position as in synchronously fertilized controls. Accordingly, the first cleavage in embryos having skipped the first normal cleavage cycle is meridional and equal, but their second cleavage is equatorial as in the third cleavage in controls. None of the embryos that were observed to skip early cleavages showed normal organogenesis, but developed into vesicle-shaped embryos with little cytological differentiation. Another effect of CCB treatment is altered blastomere size in those embryos which begin cleaving in synchrony with controls. While the majority of treated embryos followed a normal cleavage pattern, i.e. they cleaved at the right time and inequally, some of them cleaved equally or almost equally (adequally). Most of these embryos showed cleavage defects in subsequent cleavage cycles and became abnormal vesicle-shaped embryos. However, some of these embryos cleaving on schedule and equally or adequally developed into juvenile worms showing complete duplication of urites and parapodial rows (0.3% of all treated eggs) and are described as Janus duplicitates. This means that the occurrence of duplicitates and geometrically altered first cleavage patterns are correlated phenomena. The character and origin of the duplications and the consequences for dorsoventral polarity are discussed.     

Alginate films for delivery of root-knot nematode inoculum and evaluation of microbial interactions

A method was developed for utilizing alginate films to deliver inoculum into soil and evaluate microbial antagonistic activity against nematode eggs. Eggs of Meloidogyne incognita were harvested from galled tomato roots (Lycopersicon esculentum), surface disinfested, suspended in 2% (w/v) aqueous sodium alginate, and applied to 2.5 × 5.0 cm polyvinyl chloride coated fiberglass screens (1.5 mm² mesh size) at a uniform thickness of 0.5 mm. The alginate solution was gelled by dipping in 0.25 M CaCl₂. Films containing eggs were observed in vitro and egg development was evaluated. The number of immature eggs and eggs with first stage juveniles declined linearly over time while the number of empty eggs shells, and hatched juveniles increased over time, indicating that the alginate gel did not inhibit development and motility of M. incognita juveniles. In a greenhouse experiment using cucumber (Cucumis sativus) the number of galls g^-1 root was correlated with the number of eggs in alginate films placed in each pot at planting. Films containing M. incognita eggs were buried in field soil containing organic amendments, incubated, removed from soil, rinsed with water, and observed. The number of immature eggs in grids from soil amended with chitin or flax seed meal were lower than in untreated soil; percent parasitized eggs was also greater in films from amended soil than from untreated soil.     

Mitochondrial distribution, ATP-GSH contents, calcium [Ca2+] oscillation during in vitro maturation of dromedary camel oocytes

Dromedary camel oocytes have the ability to spontaneous parthenogenetic activation and development in vivo and in vitro. The present study was conducted to investigate changes in mitochondrial distribution, adenosine triphosphate (ATP), and glutathione (GSH) contents and [Ca²⁺] oscillation during in vitro maturation and spontaneous parthenogentic activation of dromedary camel oocytes. Dromedary camel cumulus-oocyte complexes (COCs) were matured in TCM199 medium supplemented with 10% FCS + 10 μg/mL FSH + 10 IU hCG + 10 IU eCG + 10 ng/mL EGF and 50 μg/mL gentamycine. Maturation was performed at 38.5 °C under 5% CO₂ in humidified air for 40 h. After maturation and removal of cumulus cells, oocytes were classified into: immature cultured (Group 1); metaphase II (M II, Group 2); and spontaneously parthenogenetically activated (with 2 polar bodies, Group 3); cleaved embryos (Group 4); and immature oocytes served as a control (Group 5). Cytoplasmic mitochondrial distribution, ATP-GSH contents, calcium [Ca²⁺] oscillation were determined. Results indicated that M II and spontaneously parthenogenetically activated oocytes represent 37.53% and 32.67% of the cultured oocytes, respectively, and 3.3% cleaved and developed to 2-16-cell stage embryos. Mitochondrial distribution, ATP-GSH contents and [Ca²⁺] oscillation were significantly (P < 0.01) differ between immature and matured dromedary camel oocytes. Mitochondrial distribution showed clustering form in matured oocytes without polar body. High polarized mitochondrial distribution (HPM) was detected in M II and spontaneously parthenogenetically activated oocytes, and the intensity of MitoTracker Red was higher in spontaneously parthenogenetically activated than M II. ATP-GSH contents and the duration of [Ca²⁺] oscillation were significantly (P < 0.01) higher in spontaneously parthenogenetically activated than M II oocytes or that matured without polar body. In conclusion, the higher incidence of spontaneously parthenogenetically activated in vitro matured dromedary camel oocytes could be attributed to the high polarized mitochondrial distribution associated with significantly higher ATP-GSH contents and duration of [Ca²⁺] oscillation.     

RELATIONSHIPS BETWEEN DISTRIBUTION OF TUBULIN-PARACRYSTALS,GRANULES STAINING WITH NEUTRAL RED AND CLEAVAGE ACTIVITY IN SEA URCHIN EGG-FRAGMENTS1,2

Unfertilized eggs of Japanese sea urchins (Temnopleurus toreumaticus and Hemicentrotus pulcherrimus)were separated by centrifugation into two fractions (nucleated light and enucleated heavy fragments) or three fractions (nucleated light, enucleated middle, and enucleated heavy fragments). These fragments were stained with neutral red and then fertilized. Cleavage took place only in fragments containing cytoplasmic granules staining with neutral red: no cleavage occurred in fragment without these granules. When fragments of unfertilized eggs were incubated in a solution in sea water of 10^?4M vinblastine, a mitotic poison that specifically binds to tubulin, tubulin-paracrystals were found in all kinds of fragments, irrespective of whether they had stained granules and cleavage activity. These results suggest that lack of cleavage activity in the fragments is not due to the absence of polymerizable tubulin molecules in the cytoplasm, but rather to other factors, such as the absence of granules staining with neutral red. In other words, there is no relation between the distribution of these granules and polymerizable tubulin, but a close relation between the number of stainable granules and cleavage activity. Quantitative analysis of tubulin molecules in the egg fragments is necessary for confirmation of this idea.     

Morphological features,development and reproduction of Melittobia acasta on Bombus terrestris

Morphological features, development and reproduction behavior of the parasite Melittobia acasta (Walker) were studied when reared on the pupae of the bumblebee Bombus terrestris L. in the laboratory under 23°C, 50% relative humidity and 12 h light : 12 h dark conditions. The parasites laid transparent white and elongated eggs. Newly hatched larval size and shape were very similar to eggs but they were identified by their body segments. Larvae increased their body size through moulting and transformed into a vermiform shape. Male pupae were shiny brown with dots. The female pupae were distinguished by their black shiny color, shorter size and the presence of compound eyes. Adult male pupae were dark brown and dwarf‐winged, whereas female pupae were macropterous and brachypterous. Reproduction took place by fertilization and also parthenogenetically. Mean fecundity within 5 days by mated (47.9 ± 30.5 female^?1) and virgin (7.4 ± 6.8 female^?1) females were statistically different. Mated females laid fertilized eggs that produced adult males or females, whereas virgin females laid unfertilized eggs that produced males. Development durations of the virgin female originated eggs, larvae, pupae and adults were statistically identical with those of mated females. The parasites were female‐biased and foundress number did not affect offspring sex ratio. This study shows that both mated and virgin females of M. acasta can produce many offspring on B. terrestris pupae within a short period, indicating that they are dangerous parasites of the bumblebee in a mass rearing system.     

Quantitative and Qualitative Aspects of Dissolved Organic Carbon Leached from Senescent Plants in an Oligotrophic Wetland

We conducted a series of experiments whereby dissolved organic matter (DOM) was leached from various wetland and estuarine plants, namely sawgrass (Cladium jamaicense), spikerush (Eleocharis cellulosa), red mangrove (Rhizophora mangle), cattail (Typha domingensis), periphyton (dry and wet mat), and a seagrass (turtle grass; Thalassia testudinum). All are abundant in the Florida Coastal Everglades (FCE) except for cattail, but this species has a potential to proliferate in this environment. Senescent plant samples were immersed into ultrapure water with and without addition of 0.1% NaN₃ (w/ and w/o NaN₃, respectively) for 36 days. We replaced the water every 3 days. The amount of dissolved organic carbon (DOC), sugars, and phenols in the leachates were analyzed. The contribution of plant leachates to the ultrafiltered high molecular weight fraction of DOM (>1 kDa; UDOM) in natural waters in the FCE was also investigated. UDOM in plant leachates was obtained by tangential flow ultrafiltration and its carbon and phenolic compound compositions were analyzed using solid state ¹³C cross-polarization magic angle spinning nuclear magnetic resonance (¹³C CPMAS NMR) spectroscopy and thermochemolysis in the presence of tetramethylammonium hydroxide (TMAH thermochemolysis), respectively. The maximum yield of DOC leached from plants over the 36-day incubations ranged from 13.0 to 55.2 g C kg⁻¹ dry weight. This amount was lower in w/o NaN₃ treatments (more DOC was consumed by microbes than produced) except for periphyton. During the first 2 weeks of the 5 week incubation period, 60–85% of the total amount of DOC was leached, and exponential decay models fit the leaching rates except for periphyton w/o NaN₃. Leached DOC (w/ NaN₃) contained different concentrations of sugars and phenols depending on the plant types (1.09–7.22 and 0.38–12.4 g C kg⁻¹ dry weight, respectively), and those biomolecules comprised 8–34% and 4–28% of the total DOC, respectively. This result shows that polyphenols that readily leach from senescent plants can be an important source of chromophoric DOM (CDOM) in wetland environments. The O-alkyl C was found to be the major C form (55±9%) of UDOM in plant leachates as determined by ¹³C CPMAS NMR. The relative abundance of alkyl C and carbonyl C was consistently lower in plant-leached UDOM than that in natural water UDOM in the FCE, which suggests that these constituents increase in relative abundance during diagenetic processing. TMAH thermochemolysis analysis revealed that the phenolic composition was different among the UDOM leached from different plants, and was expected to serve as a source indicator of UDOM in natural water. Polyphenols are, however, very reactive and photosensitive in aquatic environments, and thus may loose their plant-specific molecular characteristics shortly. Our study suggests that variations in vegetative cover across a wetland landscape will affect the quantity and quality of DOM leached into the water, and such differences in DOM characteristics may affect other biogeochemical processes.     

Cytogenetic study of silver-staining NOR in 8-cell-stage mouse blastomeres fused to 1-cell-stage embryos

Isolated blastomeres from 8- to 16-cell-stage embryos were fused by standard micromanipulatory means with either unfertilized eggs or fertilized or haploid parthenogenetically activated pronuclear-stage embryos. The hybrid eggs/embryos were incubated overnight in the presence of Colcemid until they had entered the first cleavage division. Air-dried chromosome preparations were then stained with silver nitrate in order to detect active nucleolar organizing regions (NOR). While control unfertilized eggs and 1-cell-stage fertilized and parthenogenetically activated embryos showed no evidence of silver-staining NOR-positive regions, the metaphase plates from 8- to 16-cell embryos showed characteristic NOR-positive regions, while their interphase nuclei also showed a characteristic reticular staining appearance. When hybrids between blastomere nuclei and unfertilized eggs were examined, none of the blastomere nuclei entered mitosis. However, when hybrids between blastomere nuclei and fertilized embryos were examined, in two thirds of the embryos, a single blastomere-derived diploid metaphase plate was present in association with two pronuclear-derived haploid metaphase plates. In most instances, the blastomere-derived chromosomes did not display silver-nitrate-staining NOR. Similar findings were observed when the blastomere-derived chromosomes in hybrids between blastomere nuclei and haploid parthenogenetic embryos were analysed. In the majority of cases, when blastomere nuclei remained in interphase, the characteristic silver-nitrate-staining fine reticular material either was not seen, or the nuclear contents were dispersed into clumps of chromatin-like material. Occasionally, the diploid chromosomes in the hybrids displayed morphological abnormalities. Our findings suggest that the cytoplasm of activated (but not nonactivated) 1-cell embryos is capable of influencing the nucleolar activity of the introduced 8- to 16-cell nuclei, effectively erasing from their chromosomes the memory of at least three previous rounds of rRNA synthesis.     

Plasma membrane block to sperm entry occurs in mouse eggs upon parthenogenetic activation

The ability of parthenogenetically activated mouse eggs to establish a plasma membrane (PM) block to sperm penetration was studied. Zona-free eggs preloaded with Hoechst 33342 were activated by exposure to ethanol or OAG (1-oleoyl-2-acetyl-sn-glycerol) and inseminated after different periods. Eggs challenged with sperm at 30- or 60-min postactivation displayed a fertilization frequency significantly lower than that of control eggs. Conversely, when insemination was carried out at 120-min postactivation, the proportion of fertilized eggs was equivalent to that observed in the control group. Moreover, we report that when the eggs were induced to resume meiosis without any notable loss of CGs (egg exposure to OAG at 100 μM external Ca²⁺ or to heat shock), a normal ability to be penetrated was recorded at 30-min postactivation. Similar behaviour was exhibited by eggs that underwent a CG exocytosis close to that triggered by sperm in absence of nuclear activation (microinjection of inositol 1,4,5-trisphosphate into the egg at 1 μM cytosolic concentration). Present data support the conclusion that parthenogenetically activated mouse eggs are capable of a transitory PM block response that requires both CG exocytosis and meiosis resumption to occur. © 1994 Wiley-Liss, Inc.     

The effect of incubation media on the water exchange of snapping turtle (Chelydra serpentina) eggs and hatchlings

Summary The effect of two different incubation media, sand and vermiculite, on the water exchange of eggs and the mass of hatchlings of snapping turtles (Chelydra serpentina) was assessed. The eggs were incubated fully buried in either sand or vermiculite at 30 °C and egg mass was measured periodically throughout incubation. The wet and dry masses of each hatchling and its residual yolk were measured at the end of incubation. The media had similar water potentials () but their thermal conductivities differed 2.8-fold. The eggs experienced a net water gain during incubation. The rates of water uptake between treatments were not statistically different throught the first 36 days of incubation but were statistically different thereafter, with eggs incubating in sand taking up water at about twice the rate of eggs incubating in vermiculite. Hatchling masses were similar to both media but hatchling water contents were significantly different. Hatchlings incubated in sand had lower water contents than hatchlings incubated in vermiculite even though the eggs in sand took up more water. Hatchling mass was correlated with egg water exchange for eggs incubated in vermiculite but not for eggs incubated in sand. The difference in egg water exchange in the two media appears to be attributable to differences in the thermal conductivity of the media. The presence of such a thermal effect supports the hypothesis that the eggs exchanged water with the media as water vapor. Egg water exchange was limited by the shell and shell membranes and not by the media. The shell and shell membranes appear to present an effective barrier to water uptake.Abbreviations M _{H 2 O} water flux (cm³·day^-1) - L _p hydraulic conductivity (cm·day^-1·kPa^-1) - A shell area (cm²) - A _p pore area (cm²) - l shell thickness (cm) - r pore radius (cm) - viscosity (kPa·day) - P _{EH 2 O} egg water potential (kPa) - P _{AH 2 O} medium water potential (kPa) - G _{H 2 O} water vapor conductance (cm³·day^-1·kPa^-1) - D _{H 2 O} diffusion coefficient (cm²·day^-1) - R gas constant (cm³·kPa·K^-1·cm^-3) - T temperature (K) - P _{EH 2 O} egg water vapor pressure (kPa) - P _{AH 2 O} medium water vapor pressure (kPa) - d egg diameter - K soil hydraulic conductivity (cm²·day^-1·kPa^-1) - DHM hatchling dry mass - WHM hatchiling wet mass - WU water uptake - IM initial egg mass     

Cytological changes and DNA and protein synthesis in parthenogenetically activated sea urchin eggs

Summary The present study deals with cytological observations, DNA and protein synthesis in artificially activated sea urchin eggs. The eggs were activated by means of Loeb's double treatment with butyric acid and hypertonic sea water. Most of the eggs ofHemicentrotus pulcherrimus divided when the chromosomes duplicated after formation of the first monaster and other eggs divided at a later cell cycle. In the eggs ofTemnopleurus toreumaticus, however, haploid division at the first cell cycle was observed predominantly.Activated eggs that were treated for 25 min with hypertonic sea water showed a marked uptake of³H-thymidine during the two periods of 30–40 min and 90–100 min after the double treatment. These periodic changes in the³H-thymidine uptake paralleled morphological changes within the nucleus. However, these periods of increased uptake were not observed in the eggs treated with hypertonic sea water for 60 min. During exposure to hypertonic sea water, the³H-thymidine-uptake by eggs activated with butyric acid decreased gradually. When the uptake of¹⁴C-valine by eggs was measured, a very low level was seen in unfertilized eggs. The level of uptake increased strikingly when the eggs were activated with butyric acid but was suppressed by the hypertonic treatment. However, removal of the eggs to sea water allowed the uptake to return to the former high level. This pattern suggests that the hypertonic treatment has an inhibitory effect on the synthesis of protein (or enzymes) which obstruct cleavage induction.     

Properties of adenylate cyclase activity during early sea urchin development

The total adenylate cyclase activity in homogenates of eggs of the sea urchins Strongylocentrotus purpuratus and Lytechinus pictus was assayed in vitro and found to remain constant in eggs before and at intervals after fertilization. In S. purpuratus egg homogenates virtually all of the enzyme activity was sedimented by centrifugation at 20 000 g. The enzyme specific activity in the 20 000 g pellet remained unchanged at each point through first cleavage, though it was several-fold higher than in the whole homogenate. The adenylate cyclase from both fertilized and unfertilized eggs was maximally active in vitro when assayed with 10 mM MgSO₄ and 10 mM NaF at pH 8 using 0.2 mM AMP-PNP (an ATP analog) as the substrate. Sucrose density gradient centrifugation of egg homogenates showed that adenylate cyclase activity was present in fractions which sedimented at a variety of densities. The adenylate cyclase specific activity in cortices isolated by the method of Sakai [10] from eggs at first cleavage was 4- to 6-fold higher than in unfertilized egg cortices. The increased enzyme activity in egg cortices at first cleavage suggests that adenylate cyclase-containing membranes may become localized within the egg cortex after fertilization.     

Synthesis of variously coupled conjugates of d-glucose, 1,3,4-oxadiazole, and 1,2,3-triazole for inhibition of glycogen phosphorylase

5-(O-Perbenzoylated-β-d-glucopyranosyl)tetrazole was obtained from O-perbenzoylated-β-d-glucopyranosyl cyanide by Bu₃SnN₃ or Me₃SiN₃–Bu₂SnO. This tetrazole was transformed into 5-ethynyl- as well as 5-chloromethyl-2-(O-perbenzoylated-β-d-glucopyranosyl)-1,3,4-oxadiazoles by acylation with propiolic acid–DCC or chloroacetyl chloride, respectively. The chloromethyl oxadiazole gave the corresponding azidomethyl derivative on treatment with NaN₃. These compounds were reacted with several alkynes and azides under Cu(I) catalysed cycloaddition conditions to give, after removal of the protecting groups by the Zemplén protocol, β-d-glucopyranosyl-1,3,4-oxadiazolyl-1,2,3-triazole, β-d-glucopyranosyl-1,2,3-triazolyl-1,3,4-oxadiazole, and β-d-glucopyranosyl-1,3,4-oxadiazolylmethyl-1,2,3-triazole type compounds. 5-Phenyltetrazole was also transformed under the above conditions into a series of aryl-1,3,4-oxadiazolyl-1,2,3-triazoles, aryl-1,2,3-triazolyl-1,3,4-oxadiazoles, and aryl-1,3,4-oxadiazolylmethyl-1,2,3-triazoles. The new compounds were assayed against rabbit muscle glycogen phosphorylase b and the best inhibitors had inhibition constants in the upper micromolar range (2-phenyl-5-[1-(β-d-glucopyranosyl)-1,2,3-triazol-4-yl]-1,3,4-oxadiazole 36: K_i = 854 μM, 2-(β-d-glucopyranosyl)-5-[1-(naphthalen-2-yl)-1,2,3-triazol-4-yl]-1,3,4-oxadiazole 47: K_i = 745 μM).     

Cleavage and blastoderm formation in normal and experimentally deformed naked eggs of a dipteran insect

Summary The eggs of the gall midgeHeteropeza pygmaea develop parthenogenetically inside of the mother larva. They lack a chorion and remain enveloped by the follicular epithelium. After experimental elimination of the follicular epithelium naked eggs are formed, which reach the blastoderm stage but remain spherical instead of assuming an elongated shape. To analyze this peculiar egg development and the roles of egg shape and envelope during development, the ultrastructure of cleaving normal and naked eggs was investigated. It was shown that the number of elements of Golgi apparatus and endoplasmic reticulum strongly increases during early cleavage. Their association with cleavage furrows and nuclei suggests that these organelles play a dominant role in membrane production. Egg yolk consists of lipids and glycogen, wheareas no proteins are found. Cleaving eggs contain numerous vesicles with lysosomal characteristics, indicating intense autophagic processes. Cleavage furrow formation occurs independently from the positioning of cleavage nuclei. The numerous microtubules, which are associated with cleavage furrows and nuclei and located in the egg periphery, the intercellular bridges, and in the central part of the egg, suggest that the cytoskeleton has an important role in cleavage furrow formation, blastoderm layer establishment, and yolk localization. Since these processes are accurately accomplished in naked spherical eggs, they can be considered as independent of normal egg shape and the follicular epithelium.     

EFFECTS OF CHEMICAL REAGENTS ON THE CYCLIC CHANGES OF CORTEX AND CYTOPLASM IN THE ACTIVATED ENUCLEATED EGG-FRAGMENTS OF THE SEA URCHIN*

Unfertilized eggs of sea urchins. Anthocidaris crassispina and Hemicentrotus pulcherrimus, were separated by centrifugation into two fractions (nucleated light and enucleated heavy fragments). The enucleated egg-fragments were activated by treatment with 1 M urea and then put into sea water solutions of the following three reagents; colcemid, cytochalasin B and Monogen at a concentration by which cleavage was suppressed. It was then examined whether the egg-fragments can exhibit cyclic changes of cytoplasm and cortex in correlation with the cleavage cycle in normally fertilized eggs without any influence of nuclear activity. The results obtained clearly showed that colcemid can suppress the cyclic appearance of cytoplasmic changes, but not that of cortical changes; on the contrary, in cytochalasin B- and Monogen-treated fragments, the periodicity in cortical activities is suppressed, while the periodic changes in the cytoplasm appear according to a timeschedule of the cleavage cycle. Therefore, it may be said that: 1) cyclic changes can occur in both the cytoplasm and the cortex independently, without the direct influence of nuclear activity; 2) if either of them is arrested, the cleavage does not take place; 3) the normal cleavage requires the simultaneous occurrence of periodic activities both in the cortex and in the cytoplasm after fertilization.     

Taxonomic status of the grasshopper Podisma tyatiensis and reproductive isolation between P. tyatiensis and Podisma sapporensis

The grasshopper Podisma tyatiensis, which is distributed only at the summit of Mount Tyatya on Kunashiri Island, the Kuril Islands, is closely related to Podisma sapporensis, which has a broad distribution range on the islands of northern Japan and the Russian Far East (Hokkaido, Sakhalin and Kunashiri). The present study examined the taxonomic status of P. tyatiensis by crossing P. tyatiensis males with P. sapporensis females from Sakhalin. More than 90% of eggs from intrapopulation crosses developed to at least the last embryonic stage, whereas only 64% of eggs from the interpopulation crosses developed into that embryonic stage. Cytogenetic observations of prediapause embryos showed that the interpopulation crosses always led to the production of unfertilized eggs, and that all of the developing embryos had the maternal genome only. A mixture of haploid and diploid cells of maternal origin was found in most of those embryos. This result shows that unfertilized eggs produced by P. sapporensis females from Sakhalin developed parthenogenetically to at least the embryonic stage before hatching. The present crossing experiments revealed a high level of incompatibility between the genomes of the Sakhalin population and the Tyatya population, and confirmed the full species status of P. tyatiensis.     

Parthenogenetic reproduction in neotenics of the subterranean termite Reticulitermes speratus (Isoptera: Rhinotermitidae)

Experiments with artificial colonies consisting of only female workers and those of female nymphs and workers of a subterranean termite, Reticulitermes speratus, showed that neotenics (ergatoids and nymphoids) can reproduce parthenogenetically. The larvae hatched from the eggs produced partheno‐genetically by the neotenics were all female and did not have any aberrant external morphology. They had 42 chromosomes and were diploid. The analysis of genotypes at microsatellite loci revealed that the eggs produced parthenogenetically had only one of the two alleles that the mother neotenics possessed, indicating that the eggs were produced by automixis.     

Chemistry of Benzeneglycols

Diacetyl-trans-aminodeoxybenzeneglycol (trans-1-acetamido-2-acetoxy-cyclohexadiene-3, 5) was synthesized by starting from α-B. T. C.-cis-diol²⁾ through its dehydrochlorination, NaN₃ treatment, reduction, and then dehalogenation with zinc. During NaN₃ treatment, the formation of some diazido compounds was also observed.     

The effect of temperature on the development times of eggs,naupliar and copepodite stages of five species of cyclopoid copepods

The duration times of eggs, combined naupliar instars and of the different copepodite stages of five species of cyclopoid copepods — Acanthocyclops robustus, Cyclops vicinus, Diacyclops bicuspidatus, Mesocyclops leuckarti, and Thermocyclops crassus — were investigated at five different temperatures. The five species can be divided in two groups: two species, C. vicinus and D. bicuspidatus, adapted to cold water conditions and three species, A. robustus, M. leuckarti and T. crassus adapted to warm water conditions. The cold water species showed a faster egg development than M. leuckarti and T. crassus at 5–15 °C. The eggs and instars of the warm water species M. leuckarti tend to develop faster than those of the former two species at higher temperatures. A. robustus showed the shortest egg and instar development at 10–25 °C. The warm water species T. crassus produced no eggs at 10 °C and temperatures below. At higher temperatures (20, 25 °C) the egg and instar duration times were similar or longer than those of the other species. When cultured in total darkness a great part of the C_IV respectively C_V copepodites of the summer forms entered arrest and the percentage of copepodites that showed an arrest of development was highest at lowest temperatures. The present results are compared with data from literature and differences are discussed.