Physiological and biochemical study of over-ripened oocyte in the cultivated Caspian brown trout (Salmo trutta caspius) and determination of egg quality markers

To determine the best time for egg stripping after ovulation and over-ripened oocyte in the Caspian brown trout (Salmo trutta caspius), the eggs were retained in the parental abdominal cavity for 40 days post-ovulation (DPO) at 7±0.6°C. Eggs were stripped every 10-day interval in 4 treatment and were fertilized with a pool of semen obtained from 8 males. Also, the physiology and biochemistry of the eggs and ovarian fluids were studied. Results showed that the level of eyed eggs and hatched alevins declined with over-ripening time: that is, the expected amounts (90.65 ± 6.28% for eyeing and 86.33 ± 6.82% for hatching) in newly ovulated eggs (0–10 DPO) decreased to 0.67 ± 1.34% and 0.49 ± 0.98%, respectively, in over-ripened eggs (30–40 DPO). However, larval abnormalities remained constant for 30-days after ovulation. During the course of oocyte over-ripening, the pH of the ovarian fluid significantly decreased and the concentration of glucose, protein, calcium, iron, and aspartate aminotransferase activity significantly increased. Moreover, the concentration of protein, triglycerides, and aspartate aminotransferase activity in the eggs also changed. In the newly ovulated egg, the yolk consisted of homogenous tissue and its perivitelline space diameter had no considerable differences. With over-ripening, the yolk became heterogeneous, while chorion diameter and micropyle did not change. The perivitelline space diameter varied among different areas. The present study demonstrated that the best time to take Caspian brown trout eggs after ovulation at 7± 0.6°C was up to 10 DPO. Among the studied parameters of the egg and ovarian fluid, egg quality was related to both ovarian fluid parameters (e.g., pH, protein, aspartate aminotransferase, glucose, cholesterol, triglycerides, calcium, iron) and egg parameters (e.g., cholesterol, triglycerides, iron, aspartate aminotransferase). Thus, these parameters can be used as a egg quality markers in this species.

A comparison of the protective action of added egg yolks from five avian species to the cryopreservation of bull sperm

Cryopreservation of domestic animal sperm has been widely used for artificial insemination (AI), and egg yolk is one of the most commonly used cryoprotectants during the freezing-thawing process. The objectives of this study were to compare the effectiven

A C-type lectin associated and translocated with cortical granules during oocyte maturation and egg fertilization in fish

Oocyte maturation and egg fertilization in both vertebrates and invertebrates are marked by orchestrated cytoplasmic translocation of secretory vesicles known as cortical granules. It is thought that such redistribution of cellular content is critical for asymmetrical cell division during early development, but the mechanism and regulation of the process is poorly understood. Here we report the identification, purification and cDNA cloning of a C-type lectin from oocytes of a freshwater fish species gibel carp (Carassius auratus gibelio). The purified protein has been demonstrated to have lectin activity and to be a Ca2+-dependent C-type lectin by hemagglutination activity assay. Immunocytochemistry revealed that the lectin is associated with cortical granules, gradually translocated to the cell surface during oocyte maturation, and discharged to the egg envelope upon fertilization. Interestingly, the lectin becomes phosphorylated on threonine residues upon induction of exocytosis by fertilization and returns to its original state after morula stage of embryonic development, suggesting that this posttranslational modification may represent a critical molecular switch for early embryonic development. (C) 2003 Elsevier Inc. All rights reserved.

Comparative studies on in vitro sperm decondensation and pronucleus formation in egg extracts between gynogenetic and bisexual fish

A cell-free system based upon the egg extracts from gynogenetic gibel carp (Carassius auratus gibelio) or bisexual red common carp (Cyprinus carpio red variety) was developed to investigate developmental behaviors of the demembranated sperm nuclei. Both red common carp and gibel carp sperm nuclei could decondense fully and form pronuclei in the red common carp egg extracts. Gibel carp sperm nuclei could also decondense fully and form pronuclei in the gibel carp egg extracts, but red common carp sperm nuclei could not decondense sufficiently in the same extracts. The significant differences of morphological changes were further confirmed by ultrastructural. observation of transmission electron microscopy. The data further offer cytological evidence for gonochoristic reproduction in the gynogenetically reproducing gibel carp. In addition, the sperm nuclei in vitro decondensation is dependent on the pH in the extracts, and the decondensed efficiency is optimal at pH 7. However, no DNA replication was observed in the two kinds of egg extracts during the incubation period of the sperm nuclei. It is suggested that the egg extracts prepared from the gynogenetic gibel carp should be a valid in vitro system for studying molecular mechanism on gynogenesis and reproduction mode diversity in fish.

Differential screening and characterization analysis of the egg envelope glycoprotein ZP3 cDNAs between gynogenetic and gonochoristic crucian carp

Gynogenetic silver crucian carp, Carassius auratus gibelio, is an intriguing model. system. In the present work, a systemic study has been initiated by introducing suppression subtractive hybridization technique into this model system to identify the differentially expressed genes in oocytes between gynogenetic silver crucian carp and its closely related gonochoristic color crucian carp. Five differential cDNA fragments were identified from the preliminary screening, and two of them are ZP3 homologues. Moreover, the full length ZP3 cDNAs were cloned from their oocyte cDNA libraries. The length of ZP3 cDNAs were 1378 bp for gyno-carp and 1367 bp for gono-carp, and they can be translated into proteins with 435 amino acids. Obvious differences are not only in the composition of amino acids, but also in the number of potential O-linked oligosaccharide sites. In addition, gyno-carp ZP3 amino acid sequence has an unexpected higher identity value with common carp (83.5%) than that with the closely related gono-carp (74.7%). The unique homology may be originated from the ancient hybridization. Northern blot analysis confirmed that expression of the ZP3 gene occurred exclusively in the oocytes. Because O-linked oligosaccharides on ZP3 have been demonstrated to play very important roles in fertilization, it is suggested that the extra O-linked glycosylation sites may be related to the unique sperm-egg recognition mechanism in gynogenesis.

STUDY ON THE MODE OF PRIMARY CONTROL IN THE EGG OF GYNOGENETIC CRUCIAN CARP FOR INHIBITING THE DEVELOPMENT OF 2 TYPES OF SPERM NUCLEI

By comparing the different developmental characteristics of two types of sperm nuclei which were from gynogenetic fish (crucian carp) and amphimictic fishes (red carp, red goldfish and sex-reversal red carp) respectively in the eggs of gynogenetic crucian carp, it was preliminarily revealed that there existed selective inhibiting actions of the primary control in the eggs of crucian carp for inhibiting the development of the two types of sperm nuclei. To homologous sperms, the primary control showed weak effect, thus leading to the decondensation of homologous sperm nuclei at different degrees in the eggs of crucian carp. But to heterologous sperms, the primary control showed strong effects, resulting in the total inhibition of the development of heterologous sperm nuclei. Moreover, our experimental results also showed that the different developmental behavior of the two types of sperm nuclei might have a great relationship to the changes of the sex ratio in the population of gynogenetic crucian carp. The infiltration of "the genetic materials in sperm nuclei" into the female nucleus at random might play an important role in male emergence in the naturally gynogenetic population of crucian carp.

Direct electrochemistry of hemoglobin in egg-phosphatidylcholine films and its catalysis to H2O2

Direct electrochemistry of hemoglobin was observed in stable thin film composed of a natural lipid (egg-phosphatidylcholine) and hemoglobin on pyrolytic graphite (PG) electrode. Hemoglobin in lipid films shows thin layer electrochemistry behavior. The formal potential Edegrees' of hemoglobin in the lipid film was linearly varied with pH in the range from 3.5 to 7.0 with a slope of -46.4 mV pH(-1) Hemoglobin in the lipid film exhibited elegant catalytic activity for electrochemical reduction of H202, based which a unmediated biosensor for H2O2 was developed.

Preparation and mass spectrometric study of egg yolk antibody (IgY) against rabies virus

Rabies virus was used as the antigen to immunize laying chickens. Anti-rabies virus immunoglobulin Y(IgY) was isolated from yolks of the eggs laid by these chickens using a two-step salt precipitation and one-step gel filtration protocol. The purified IgY was reduced with dithiothreitol, and heavy chains (HC) and light chains (LC) were obtained. In addition, the purified IgY was digested with pepsin and the fragment with specific antigen binding properties (Fab) was produced. Using matrix-assisted laser desorption/ionization mass spectrometry (MALDI-TOFMS), the average molecular weights of IgY, HC, LC, and Fab were determined as 167 250, 65 105, 18 660, and 45,359 Da, respectively. IgY has two structural differences compared with mammalian IgGs. First, the molecular weight of the heavy chain of IgY is larger than that of its mammalian counterpart, while the molecular weight of the light chain of IgY is smaller. Second, upon pepsin digestion, anti-rabies virus IgY is degraded into Feb, in contrast to mammalian IgG, which has been reported to be degraded into F(ab')(2) under the same conditions. Copyright (C) 2001 John Wiley & Sons, Ltd.

Effect of light period on egg-discharge of gametophyte clones of Undaria pinnatifida (Phaeophyta)

A sporeling culture method using gametophyte clones developed in the early 1990s led to egg discharge occurring in the dark 5 min after the start of the dark period under growth under a 11:13 L-D photoperiod. The course of egg discharge could be disturbed by light, with irradiance as low as 5-6 mu mol photon m(-2) s(-1) causing 75-80% of the discharged eggs to detach from the oogonia and consequently to die within several hours. In order to enhance outgrowth rate of young sporophytes, a study was conducted to test the effect of controlling darkness in the period 2-3 h after dusk. When the slides were transferred from the standard 11:13 L-D regime to continuous light, eggs were discharged 5 min after the end of the light phase and peaked 5-l5 min later on first day after transfer, indicating that the female gametes "remember" the light-dark regime. This suggests the existence of an endogenous circadian rhythm. During the second and third days, very few eggs were discharged throughout the 11 h of the normal light phase of the L-D regime, indicating the inhibitory effect of continuous light and that the rhtyhm is easily damped by light.

Feeding and egg production of the planktonic copepod Calanus sinicus in spring and autumn in the Yellow Sea, China

Shipboard incubations were conducted in spring (April) and autumn (October/November) 2006 to measure the feeding and egg production rates (EPR) of Calanus sinicus in the Yellow Sea, China. The ingestion rate (2.08-11.46 and 0.26-3.70 mu g C female(-1) day(-1) in spring and autumn, respectively) was positively correlated with microplankton carbon concentrations. In the northern part of the Yellow Sea, feeding on microplankton easily covers the respiratory and production requirements, whereas in the southern part in spring and in the frontal zone in autumn, C. sinicus must ingest alternative food sources. Low ingestion rates, no egg production and the dominance of the fifth copepodite (CV) stage indicated that C. sinicus was in quiescence inside the Yellow Sea Cold Bottom Water (YSCBW) area in autumn. Calanus sinicus ingested ciliates preferentially over other components of the microplankton. The EPR (0.16-12.6 eggs female(-1) day(-1) in spring and 11.4 eggs female(-1) day(-1) at only one station in autumn) increased with ciliate standing stock. Gross growth efficiency (GGE) was 13.4% (3-39%) in spring, which was correlated with the proportion of ciliates in the diet. These results indicate that ciliates have higher nutrient quality than other food items, but the low GGE indicates that the diet of C. sinicus is nutritionally incomplete.

Inhibition of egg hatching success and larvae survival of the scallop, Chlamys farreri, associated with exposure to cells and cell fragments of the dinoflagellate Alexandrium tamarense

We report an apparently novel toxic effect of the dinoflagellate Alexandrium tamarense, manifested by inhibition of the egg hatching success of the scallop, Chlamys farreri. The hatching rate of C. farreri approached only 30% of controls when its fertilised eggs were exposed for 36 h to A. tamarense cells or cellular fragments at a concentration of 100 cells/ml, and the hatching rate was just 5% after exposure to A. tamarense of 500 cells/ml. Similar exposures of the fertilised scallop eggs to two other algal species, the diatom Phaeodactylum tricornutum and the raphidophyte Heterosigma carterae, resulted in no such toxicity or inhibitory effects.. Likewise, exposure of eggs to standard STX toxin. as well as to A. tamarense cell contents (supernant of re-suspended algal cells following ultrasonication and centrifugation), did not elicit this inhibitory response. However, exposure of the scallop eggs to cell cultures, intact algal cells, or cell fragments of A. tamarense produced marked toxicity. The alga also influenced larvae at early D-shape stage of scallop. The survival rates began to decrease significantly after exposed for 6 days at concentration of 3000 cells/ml and above: no larvae could survive after 14-day exposure to A. tamarense at 10,000 cells/ml or 20-day at 5000 cells/ml. The results indicated the production of novel substances from A. tamarense which can cause adverse effects on egg hatching and survival of the scallop larvae, The experiment also found that the developmental stages before blastula was the developmental period most sensitive to the A. tamarense toxin(s) and the alga at early exponential stage had the strongest effect on egg hatching comparing with other growth phases. The adverse effect of A. tamarense on early development of scallops may cause decline of shellfish population and may have further impact on marine ecosystem. (C) 2001 Elsevier Science Ltd. All rights reserved.

Seasonal and spatial variation in abundance and egg production of Paracalanus parvus (Copepoda: Calanoida) in/out Jiaozhou Bay, China

The spatial distribution of stage-specific abundance and reproduction of the copepod Paracalanus parvus were studied from October 2005 to September 2006 in the Jiaozhou Bay. This copepod occurred continuously in this bay throughout the year. The species reached the lowest abundance in April and peaked in June. From October to December, distribution center mainly occurred in offshore water and at the mouth of the bay. In winter, early copepodites and adults gradually decreased and till February, most of the population was only comprised of CIV-CV stages. Overwintering copepodites matured in March and males tended to mature before female. From May to September, each stage occurred in the population and gradually reached high abundance. Temperature and chlorophyll a (Chl-a) concentration in the three stations can't clearly explain the seasonal variation in stage-specific abundance, so we surmised the important effect of the Yellow Sea. Egg production rate (EPR) reached its lowest in winter and peaked in June at 60.8 eggs female(-1) day(-1) in nearshore water. In the warming period, EPR in nearshore water was statistically higher and EPR > 10 eggs female(-1) day(-1) lasted longer than that in offshore water, showing the importance of nearshore water for recruitment of R parvus. Our study showed that EPR was positively related to temperature and total chlorophyll a in offshore water and mouth of the bay. In nearshore water, the relationships between EPR and temperature and Chl-a in three size fractions were not the same as those in offshore water, suggesting complicated ecosystem in such a eutrophic area in warming period. (C) 2008 Elsevier Ltd. All rights reserved.