Ovarian Fluid pH Research Articles

Effect of temperature and spawning agents on wild female asp (Leuciscus aspius) reproductive efficiency under controlled conditions

Artificial reproduction is a bottleneck to produce stocking material for many species of freshwater fish. One of these species is the asp, Leuciscus aspius. Research in the field of artificial reproduction of this species is very scarce and often incomplete. There are no breeding protocols specifying optimal environmental conditions and hormonal stimulation for many species of rheophilic cyprinids, including asp. Since the number of natural asp populations is constantly decreasing, it is important to support natural stocks by restocking with high quality stocking material. For this reason, optimized protocols are needed to breed this species under controlled conditions to produce stocking material with high biodiversity and good health. Such an approach will make it possible to maintain the population of natural asp at a constant level. The aim of this study was to develop the protocol of asp artificial reproduction using optimized thermal conditions and appropriate hormonal stimulation. In experiment I, the influence of constant temperature (10.0, 12.0 and 14.0 °C) on the effectiveness of artificial reproduction of asp. In experiment II, the effectiveness of asp reproduction was checked after the application of spawning agents: Ovopel, Ovaprim or a combination of these two agents The obtained results indicate that for the final maturation of oocytes (FOM) and artificial reproduction of asp in controlled conditions, water temperatures of 10–12 °C are the most useful. Under these thermal conditions, the highest percentages of female’s ovulation and embryo survival, as well as the percentage of hatching, were obtained. Hormone injections are necessary to perform final oocyte maturation (FOM) in female asp in captivity. All spawning agents used were especially useful for artificial reproduction of asp, however, the best values of the studied indices, such as ovulation rate and embryo survival, were obtained after the application of Ovaprim or the combination of Ovopel and Ovaprim in water temperature at a range of 10–12 °C. It was found that the pH of ovarian fluid may be a preliminary indicator of the biological quality of eggs in asps. The optimal pH value is 8.0–8.4. At pH below 7.4, no viable embryos were observed.

Artificial reproduction of vimba bream (Vimba vimba) cultured generation F2 and eggs management under controlled conditions

The experiments were conducted to investigate the hormonal stimulation of oocyte maturation in cultured (F2 generation) vimba bream (Vimba vimba) under controlled conditions and some aspects of egg management. In artificial reproduction, four hormonal agents (CPH, Ovopel, Ovaprim, and hCG) were tested. Fish were intraperitoneally (IP) injected with two doses of the hormonal agents at the base of the ventral fin. The highest percentage of ovulating vimba bream females (100%) was obtained in the Ovaprim group. Fertilization and hatchability percentages were nearly 95% and 93% in this group. In the control group, which was not injected with hormones, only one female from seven ovulated. Regarding the egg management aspects, the effects of 1) time adding sperm to activated eggs, 2) portional adding sperm to eggs, and 3) pH of the ovarian fluid on fertilization success of vimba bream were also assessed. The results indicate that vimba bream eggs retain fertilization capacity as long as 150–180 s after their activation. When fertilization was carried out within 30 s from the activation of the eggs, the hatchability rate of the larvae was over 80%. The analysis of the results of the present study indicates that there is a statistically significant correlation between vimba bream ovarian fluid pH and the hatching rate of larvae (R2 = 0.954). Moreover, a hatching rate (%) of >50% was noted in the females, of which the ovarian fluid pH was above 7.6. Thus, the results of the present study indicate that the pH of the ovarian fluid is an important factor affecting the quality of vimba bream gametes. The obtained results can be successfully used to produce vimba bream stocking material in a fish farm with the use of cultured spawners.

Extended in vitro storage of eggs and milt increases maternal but not paternal variation in embryo viability of landlocked Atlantic salmon (Salmo salar m. sebago)

Appropriate fertilization practices play a crucial role in the maintenance of genetic diversity within captive fish broodstocks. We assessed the influence of in vitro storage of gametes (0-, 7- and 14-days post-stripping) at 1 °C on ovarian fluid pH, sperm motility, and embryo viability in the critically endangered Finnish landlocked salmon (Salmo salar m. sebago) population. A repeated full-factorial design used in both sperm-ovarian fluid activation analyses and fertilizations allowed us to separate the effects of female, male and their interaction on gamete quality and embryo survival across storage times. The ovarian fluid pH decreased most significantly during the first week of storage and showed considerable variation among females. The decrease of sperm motility varied across males, females and male–female (sperm-ovarian fluid) combinations. The mean proportion of viable embryos was initially 96.4%, but dropped to 45.5% after 7 days, and to 13.1% after 14 days of gamete storage. Embryo viability responses to gamete storage time predominantly increased among- and within-female variation but also showed major differences between female × male combinations (non-additive genetic effects). Interestingly, embryo viability showed virtually complete re-ranking (change of order) among full-sib families between the first two fertilization times (rs = − 0.03) but consistency between the first and last times (rs = 0.59). Overall, our results indicate that even when the primary aim is to maximize the genetic diversity of endangered fish populations, it may be advisable to prioritize minimal egg storage time rather than maximizing the number of parental combinations in fertilizations.

Optimization of barbel (Barbus Barbus L.) fertilization and effects of ovarian fluid when there are controlled conditions for gamete activations

Fertilization is one of the most important procedures in artificial reproduction and it directly affects the reproduction outcome. When there is optimization of fertilization, there can be a positive effect on subsequent reproductive processes and economic aspects of aquaculture. This study was conducted to determine time for which oocytes and sperm of barbel Barbus barbus retain fertilization capacity following placement in freshwater. Furthermore, the amount of ovarian fluid, excreted by fish during spawning with eggs (OFI; %) was determined, along with the chemical composition and effects on fertilization were determined. Gametes, ovarian fluid, and seminal plasma from barbel spawning specimens of the F4 generation were used to conduct the study. Ovarian fluid accounted for 14%–68% of contents of the mass released at spawning and post-spawning composition differed depending on whether hormonal treatments were utilized for control of reproduction. There was an association (R2 = 0.982; P = 0.000) between the pH of ovarian fluid and the barbel embryo survival rate. There was the greatest survival rate (>60 %) when the pH range of 7.9–8.4 and there was a lesser embryo viability when pH values were lesser or greater than values within this range (P < 0.05). The results from the study indicate that barbel eggs retain fertilization capacity longer (as long as 210 s) after activation by placement in fresh water than spermatozoa (about 30 s).

Biochemical composition of the ovarian fluid and its effects on the fertilization capacity of turbot Scophthalmus maximus during the spawning season.

This study investigated the biochemical composition of ovarian fluid and its effect on the fertilization capacity of turbot Scophthalmus maximus during the spawning season. The fertilization rate and pH of ovarian fluid varied throughout the spawning season, with the highest values recorded at the mid-season. Positive correlations were found between the fertilization rate and the ovarian fluid pH. The composition of major inorganic ions (Na(+) , K(+) , Ca(2+) and Cl(-) ) showed no significant changes during the spawning season. Alkaline phosphatase (AKP) activity was significantly higher during mid-season than other seasons. The lowest levels of protein, acid phosphatase (ACP) and aspartate aminotransferase (AAT) were in the ovarian fluid released at the mid-season. Moreover, significant relationships were observed between the fertilization rate and the levels of protein, ACP, AKP and AAT. These observations suggest that the biochemical profile of ovarian fluid affects the insemination microenvironment as well as the fertilization capacity of S. maximus eggs. Determination of such profiles may prove to be a useful strategy to improve S. maximus breeding techniques.

Quantitative characteristics of Atlantic halibut (Hippoglossus hippoglossus L.) egg quality throughout the reproductive season

Assessment of egg quality is an important aspect in finfish hatchery management, but guidelines for such assessment are scarce, especially for marine fish production. In the present work, potential indicators of egg and larval quality were measured in 39 batches of eggs of Atlantic halibut (Hippoglossus hippoglossus L.) throughout the reproductive season. The paternal influence on offspring was minimized by using the same cryopreserved semen for all fertilizations; consequently, maternal effects were emphasized. The progression of the spawning season and ovarian fluid parameters, including pH, electrical conductivity (EC), osmolality, and the amount of fluid were registered. The behavior of the broodfish at egg collection was registered and the eggs were analyzed for cortisol content. Fertilization and hatching rates as well as larvae survival were calculated, and larval standard length and myotome height were measured. Furthermore, the occurrence of major types of larval deformities was registered. The myotome height was significantly (P < 0.05) affected by the spawning season progress. Cortisol content in the eggs was decreasing with the progressing spawning season and correlated positively with the occurrence of yolk-sac edema. The ovarian fluid pH and EC were significantly related to fertilization and hatching rates. High fertilization and hatching success was associated with pH greater than 7.9 and EC less than 2.5 mS/cm. Low fertilization rates (<50%) resulted in further low hatchability from such egg batches. Ovarian fluid EC was significantly and positively related to increased occurrence of yolk-sac edema. High quantity of ovarian fluid in egg batches was associated with reduced egg quality in terms of fertilization and hatching rates and occurrence of yolk-sac edema. A cumulative effect of ovarian fluid pH, EC, osmolality, and quantity explained up to 62% of the total variation in fertilization rates. The findings from the present study indicate that parameters measurable at the initial phase of production, in particular ovarian fluid pH and EC, might have a potential for future use as egg quality indicators in hatchery management.

Synchronization of ovulation in cultured northern whitefish (Coregonus peled, Gmelin 1788) using [D-Arg6Pro9Net]-sGnRH analogue and its effect on egg quality

Female northern whitefish were treated with salmon [D-Arg6Pro9Net]-sGnRHa emulsified in Freund′s incomplete adjuvant as a sustained release treatment (sGnRHa-FIA) or with [D-Arg6Pro9Net]-sGnRHa dissolved in physiological saline as acute release treatment (sGnRHa-PS). Females in four experimental groups (A, B, C, D) and one control group (E) were injected intraperitoneally as follows: A and B – sGnRHa-FIA at doses of 50 and 25 μg kg−1, respectively; C – double injection (DI) of sGnRHa-PS at 25 μg kg−1 administered 3 days apart; D – single injection (SI) of sGnRHa-PS at 25 μg kg−1; and E – control group receiving a 1:1 mixture of FIA and physiological saline. All treatments induced and synchronized ovulation. Mean time to ovulation was significantly reduced (P < 0.05) in GnRHa-treated groups compared with control. Hormone treatments did not affect the relative fecundity. Slight differences were found in ovarian fluid pH between group A and D (P < 0.05). Except for group C, egg size was significantly reduced (P < 0.01) in hormonally synchronized groups compared with the control. Survival to the eyed stage of eggs from females in groups A, B and C was significantly lower (P < 0.01) than in groups D and E. Per cent hatched alevins was lower (P < 0.01) in groups A, B and C than in groups D and E. We conclude that synchronization of ovulation in northern whitefish can be induced by a single acute injection of [D-Arg6Pro9NEt]-sGnRHa at 25 μg kg−1 BW if given near the natural spawning time determined by water temperature below 2°C.

Synchronization of ovulation in brook char (Salvelinus fontinalis, Mitchill 1814) using emulsified d-Arg6Pro9NEt sGnRHa

Effects of physiological saline (PS)-dissolved or Freund′s incomplete adjuvant (FIA)-emulsified sGnRHa on the induction and advancement of ovulation in brook char were assessed. Two-year-old females were randomly divided into 5 groups. Groups A and B received intraperitoneal injection of FIA-emulsified sGnRHa (sGnRHa-FIA) at dosages of 50 and 25 μg kg−1 body weight (BW), respectively. Females in group C were treated with a double injection (DI) of PS-dissolved sGnRHa (sGnRHa-PS) at 25 μg kg−1 BW spaced 3 days apart. Fish in group D received a 25 μg kg−1 BW single injection (SI) of sGnRHa-PS. Group E was established as a control group. After stripping, ovarian plasma pH level was measured, and an egg sample was taken from each female to record egg weight and diameter and survival to the eyed stage. Females in the GnRHa-treated groups ovulated significantly earlier than did females in the control group (P < 0.01). No significant differences were found among GnRHa-treated groups in ovulation dynamics and mean time to ovulation. Ovarian fluid pH was significantly higher in groups A, B, and C compared to control group E (P < 0.05). Significantly lower egg weight was found only in group B (P < 0.01), although all advanced groups tended to have lower egg weight than the control group. Egg diameters paralleled egg weight. Survival to the eyed stage was significantly higher in GnRHa-advanced groups compared to the control. A negative relationship was found between egg weight end eyed eggs percent (R2 = 0.26). No pre-spawning or post-spawning mortality was observed during a 6-month period. Neither sGnRHa-PS nor sGnRHa-FIA are associated with negative influences on the health of females. The sGnRHa-FIA injections proved to exhibit the same efficacy as the DI protocol with sGnRHa-PS. Although no statistical differences were found in ovulation dynamics, we do not recommend the use of SI with sGnRHa-PS in brook char.

The Influence of Ovarian Fluid pH of Stripped Unfertilized Channel Catfish, Ictalurus punctatus, Eggs on the Hatching Success of Channel Catfish ♀ x Blue Catfish, Ictalurus furcatus♂, Hybrid Catfish Eggs

This study was designed to determine the effect of ovarian fluid pH of stripped unfertilized channel catfish, Ictalurus punctatus, eggs on fertilization and hatch rate of channel catfish ♀ x blue catfish, Ictalurus furcatus♂, hybrid catfish eggs. A significant correlation was established between ovarian fluid pH of stripped channel catfish eggs and hybrid embryo hatch rate (R2 = 0.75, P = 0.01), suggesting ovarian fluid pH of stripped catfish eggs prior to fertilization can be predictive of the hatching success of hybrid catfish embryos. These data were used to categorize pH of stripped eggs: pH &lt;7.0 as “low pH eggs,” pH 7.0–7.4 as “medium pH eggs,” or pH &gt;7.4 as “high pH eggs” quality eggs. The range in percent hatch rate for these pH categories was &lt;15%, 15–30%, and &gt;30%, respectively. Higher calcium concentrations during incubation do not appear to improve hatching success of “low pH eggs.” The predictive model presented herein describes a quick method for evaluating stripped catfish eggs for hybrid fry production in catfish hatcheries.

Broken Eggs Influence on Fertilization Capacity and Viability of Eggs, Turbidity and pH of Ovarian Fluid and Fertilization Water in the Endangered Caspian Brown Trout, Salmo Trutta Caspius

To identify a simple tool for quick evaluation of quality of endangered Caspian brown trout, Salmo trutta caspiuseggs, the changes of pH (both in ovarian fluid and fertilization water), turbidity (both in ovarian fluid andfertilization water), fertilization and eyeing rates were investigated in ovarian fluid samples containing perfecteggs as well as different concentrations of broken eggs. The pH of ovarian fluid and water as well as fertilizationand eyeing rates decreased significantly (P<0.05) with increasing of broken eggs in ovarian fluid. In contrast, theturbidity of ovarian fluid and water as well as mortality rate of eggs increased significantly (P<0.05) withincreasing of broken eggs in ovarian fluid. Also, significant correlations (P<0.01) were found between measuredparameters as follow: pH of ovarian fluid vs. pH of water and fertilization and eyeing rates; turbidity of ovarianfluid vs. turbidity of water; turbidity of ovarian fluid and water vs. fertilization and eyeing rates. Our resultsconclude that pH and turbidity of ovarian fluid and water effectively influence on efficiency of artificialpropagation. Therefore, these could be used as two simple tools for quick evaluation of quality of Caspian browntrout eggs during artificial reproduction in the hatchery.

Broken eggs decrease pH of rainbow trout (Oncorhynchus mykiss) ovarian fluid

Low pH of ovarian fluid is attributed to its inability to activate sperm motility. In the present study we sought if broken eggs are reason for the decrease of ovarian fluid pH. We examined if 1) there is relationship between turbidity of ovarian fluid (indicator of presence of broken eggs) and pH, and 2) there is a decrease in ovarian fluid pH after addition of broken eggs. We found a negative relationship between the turbidity and ovarian fluid pH. Addition of the content of broken eggs to ovarian fluid caused a decrease of pH and a decrease in ovarian fluid ability to activate sperm motility. Therefore it can be suggested that content of broken eggs decreases or negates the ability of ovarian fluid to activate sperm motility due to a decrease of pH.

Ovarian fluid pH enhances motility parameters of rainbow trout ( Oncorhynchus mykiss) spermatozoa

All evidence to date suggest that sperm motility is the primary determinant of fertilization success in externally fertilizing fish species. Ovarian fluid, which comprises 10–30% of the total egg volume in salmonids, enhances sperm motility with respect to swimming speed, trajectory and the duration of movement. It was recently demonstrated that there is individual variability in sperm motility enhancing potential of ovarian fluid of particular females. In the present study we examined the effect of particular ovarian fluids collected from 31 females on the sperm motility parameters of one male of rainbow trout ( Oncorhynchus mykiss) using computer-assisted sperm analysis (CASA). During our experiment we also monitored the pH of ovarian fluid. We found that particular fluids differed in the ability to activate spermatozoa; sperm remained immotile in four fluids and exhibited 50–100% motility in 27 samples. The percentage of motile sperm, velocity and duration of movement positively correlated with ovarian fluid pH ( r 2 = 0.34–0.62). These data strongly suggest that the pH of the ovarian fluid is the primary determinant of sperm motility in rainbow trout under natural conditions of fertilization.

Effects of external pH on hormonally regulated ovarian follicle maturation and ovulation in Atlantic croaker

In vitro studies of ovarian follicle maturation and ovulation in teleost fishes typically are conducted within a narrow range (7.5–7.8) of constant external (medium) pH, although there is evidence that pH can influence ovulation. Therefore, this study with Atlantic croaker investigated the effects of external pH on hormonally regulated in vitro maturation and ovulation as well as changes in the pH of ovarian fluid during in vivo maturation and ovulation. For the in vitro experiments, follicles were first incubated with human chorionic gonadotropin (hCG) to induce maturational and ovulatory competencies, and then with maturation-inducing hormone (MIH) to induce completion of maturation and ovulation. At a constant external pH within the range of 7.0–8.2, the lower pH levels (7.0–7.3) generally inhibited or slowed down hormonally induced maturation and ovulation whereas higher pH (7.6–8.2) facilitated these processes. When ovarian follicles were incubated at a constant pH of 7.6 during the priming incubation with hCG, changing the external pH during the incubation with MIH had relatively little effect on oocyte maturation or ovulation. Thus, the inhibitory effect of constant low levels of external pH (7.0–7.3) on maturation and ovulation may be primarily due to disruptions in the gonadotropin-dependent acquisition of maturational and ovulatory competencies. The pH of ovarian fluid remained constant at 8.5 during in vivo ovarian follicle maturation and ovulation. Subsequent in vitro tests showed that external pH of 8.5 enhances hormonally induced maturation and ovulation relative to pH of 7.6. These observations suggest that attention should be paid to the pH of incubation media used in basic research and in biotechnological applications relying on in vitro maturation and ovulation in teleosts. Further, an understanding of the physiological significance of the enhancing effect of alkaline pH on maturation and ovulation will require determination of the intrafollicular pH around the oocyte during the periovulatory period.

Effects of post-ovulatory oocyte ageing and temperature on egg quality and on the occurrence of triploid fry in rainbow trout, Oncorhynchus mykiss

The present experiment was designed to evaluate the effect of spawning season rearing temperature and post-ovulatory oocyte ageing on egg developmental success in rainbow trout, Oncorhynchus mykiss, assessed as percent survival at the eyeing, hatch and swim-up stages, and as percent occurrence of malformations and ploidy anomalies at yolk sac resorption. Moreover, female weight, egg wet weight, oocyte ATP concentration and ovarian fluid pH, osmolality and protein concentration were measured to investigate possible links with post-ovulatory oocyte ageing and egg developmental success. Thirty three mature females were held at 12 °C and 10 at 17 °C. Eggs and ovarian fluid were sampled at the time of detected ovulation and 1, 2 and 3 weeks later. Eggs were inseminated with fresh semen pooled from five males and embryo development was monitored until yolk sac resorption. Generally, survival rate decreased and malformation rate increased with an increase in post-ovulatory oocyte ageing. Egg developmental success was positively correlated to female weight and to ovarian fluid pH and osmolality. Ovarian fluid protein concentration significantly increased during post-ovulatory ageing, whereas oocyte ATP concentration remained constant. A significant increase in morphological anomalies was observed as soon as 7 days post-ovulation (dpo), though this effect was not correlated with decreased survival. Also, an increase in spontaneous triploidy was associated with increased post-ovulatory oocyte ageing and high temperature, sometimes in dramatically high proportions (50%).

Bio-markers for egg quality determination in cyprinid fish

The present study investigated potential bio-markers (ovarian fluid parameters, egg weight and weight increase during water hardening, biochemical egg composition, egg enzyme activities) for egg quality determination in the common carp ( Cyprinus carpio), the silver carp ( Hypophthalmichthys molitrix), the grass carp ( Ctenopharyngodon idella) and the bleak ( Chalcalburnus chalcoides). For all investigated species, the study revealed highly significant correlations between the egg fertilization rate and the weight of water-hardened eggs, the percent weight increase during water hardening and the ovarian fluid pH, protein concentration, and aspartate aminotransferase activity. The fertilization rate was further correlated with the activities of malate dehydrogenase and pyruvate kinase of the eggs, and in C. carpio and Cha. chalcoides with the respiration activity, too. Investigated biochemical parameters of the eggs (protein, peptides, fructose, galactose, glucose, non-esterified fatty acids, esterified fatty acids, total DNA and RNA) were not correlated with the fertilization rate. The possible use of the analysed parameters for prediction of egg quality during short-term storage was also investigated in C. carpio and Cte. idella: during short-term storage for 4 h at 4°C, the fertilization rate significantly decreased. Also, the weight of the eggs after water hardening, the percent weight increase due to water hardening and the ovarian fluid pH and ovarian fluid aspartate aminotransferase changed in comparison to fresh samples and were highly significantly correlated with the fertilization rate. In contrast, the biochemical composition of the eggs (protein, peptides, fructose, galactose, glucose, non-esterified fatty acids, esterified fatty acids, total DNA and RNA) and egg enzyme activities (phosphofructokinase, pyruvate kinase, protease, lipase, NAD-dependent malate dehydrogenase, respiration rate, NADP-dependent isocitrate dehydrogenase, aspartate aminotransferase) remained constant.

Changes in Eggs of Brown Trout, Rainbow Trout, and Grayling during Short-Term Storage

Short-term storage of the eggs of rainbow trout Oncorhynchus mykiss, brown trout Salmo trutta, and grayling Thymallus thymallus was investigated to examine factors that limited egg viability and to compare storage with and without ovarian fluid. Egg viability was similar after 2 h, but after 4 h, egg fertilization rate was lower by 14–26% for eggs stored in ovarian fluid and lower by 23–51% in eggs stored without ovarian fluid. Substitution of a buffered physiological saline solution for ovarian fluid was tested for brown trout and grayling but did not improve egg viability. The wet weight of eggs during storage without ovarian fluid decreased significantly. Water influx during hardening was higher for eggs stored without ovarian fluid than in controls and eggs stored with ovarian fluid. However, water loss due to storage could not be completely compensated, so the wet weight of hardened eggs that had been stored without ovarian fluid remained lower. Ovarian fluid pH and activities of protease during egg storage with ovarian fluid significantly increased, and peptide levels decreased.

Physiological and biochemical parameters for egg quality determination in lake trout, Salmo trutta lacustris

The present study investigated the relationships between egg viability and ovarian fluid composition, egg physiology and egg metabolism in lake trout, Salmo trutta lacustris, to obtain biomarkers for egg quality determination. The ovarian fluid pH, protein levels and activities of aspartate aminotransferase and β-d-glucuronidase were significantly correlated with egg viability expressed as the number of eyed stage embryos. Regression models demonstrated that an ovarian fluid pH between 8.44 and 8.57, protein levels below 235.56 mg 100 ml−1ovarian fluid, aspartate aminotransferase activity below 31.65 μm min−1 l−1ovarian fluid and β-d-glucuronidase activity below 8.62 μm min−1 l−1 ovarian fluid characterized egg batches with high viability (≥80%).

Morphology, composition and fertilization of carp eggs: a review

This paper is a review of the literature on eggs of carp ( Cyprinus carpio L.) and some related species and includes data taken from the former USSR and eastern European countries. Reported data relate to fecundity, egg morphology and composition, and fertilization. Information is also available on the ovarian fluid composition with characterization of proteins and lipids. The fecundity of common carp is very high, ranging from 100 000 to 300 000 eggs · kg −1 body weight per oogenetic cycle, with one cycle/year in the case of females reared in an outside natural pond environment. The diameter and weight of the eggs are in the range of 1.24–1.42 mm and 0.86–1.41 mg, respectively. The ooplasm includes large amounts of yolk and various organelles (endoplasmic reticulum, Golgi apparatus, mitochondria), cortical granules and alveolae. The main characteristics of cyprinid ovarian fluid are an osmotic pressure of about 300 mOsm, a Mg 2+ concentration of 2.58 mM and a high ovarian fluid pH of 9. Energy in the form of ATP which is necessary for egg metabolism originates from glycolytic and oxidative reactions. The egg has a relatively thick vitelline envelope (VE) or zona radiata which is reorganized into the fertilization envelope (FE) after fertilization. The micropyle located at the animal pole is a funnel-shaped structure leading spermatozoa to the ooplasmic surface on which a fertilization cone develops after fertilization. A site of sperm attachment is identified on the plasma membrane at the level of the internal aperture of the micropylar canal. Major differences are observed between the two outermost VE and FE layers, as revealed by electron microscopy, enzyme or carbohydrate cytochemistry, and imunohistochemistry. FE extracts have strong bactericidal and fungicidal effects.

Reliable assessment of overripening in turbot ( Scophthalmus maximus) by a simple pH measurement

The pH of ovarian fluid was around 8.1 in turbot spawn collected by stripping shortly after ovulation. It decreased down to 7.1 when overripening developed. A highly significant correlation ( r=0.92, P<0.001) was found between the ovarian fluid pH and the fertilization rate. The ovarian fluid was found to have a strong buffer capacity affecting the insemination microenvironment, and hence affecting the fertilization process. Spawn pH may be used to assess egg quality.