• Development and Reproduction
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• v.17 no.4
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• pp.345-351
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• 2013

This study examines the effects on fertilization rate (FR), hatching rate (HR), and normal individual rate after artificial fertilization using frozen thawed sperm according to the cryoprotectant (DMSO) concentration and the period of cryopreserved sperm of longtooth grouper, Epinephelus bruneus. Performing artificial fertilization using frozen-thawed sperm, after freezing the sperm at different DMSO concentration of 5.0%, 7.5%, 10.0% respectively, FR were (DMSO 5.0%: $99.5{\pm}0.8%$, DMSO 7.5%: $99.5{\pm}0.7%$, and DMSO 10.0%: $99.6{\pm}0.6%$). The results are not significantly different from the control fresh sperm (100%). HR also (DMSO 5.0%: $96.2{\pm}2.3%$, DMSO 7.5%: $95.3{\pm}3.6%$, 10.0%: $96.6{\pm}1.8%$) were not significantly different in each group. The normal individual rate after hatching using with control fresh sperm ($98.4%{\pm}0.5$) and DMSO concentration level of 5.0% ($97.8{\pm}0.1%$) were not significantly different. However, with 7.5% ($97.2{\pm}0.6%$) and 10.0% DMSO concentrations ($95.9{\pm}0.2%$) are lower than the normal individual rate after hatching observed in the control and 5.0% DMSO. Performing artificial fertilization using frozen-thawed sperm at different frozen period (2 days, 2 years, and 3 years), 10% DMSO FR and HR of 3 years (FR; $66.8{\pm}1.8%$, HR: $82.0{\pm}12.9%$) and 2 years (FR; $78.5{\pm}14.8%$, HR: $79.3{\pm}0.6%$) cryopreserved sperm were lower than control (FR; 100%, HR: $91.1{\pm}3.6%$) and 2 days cryopreserved sperm (FR; $99.6{\pm}0.6%$, HR: $96.6{\pm}1.8%$). These results suggest suitable DMSO concentration ranges of cryopreservation sperm for E. bruneus is 5 to 10% and with 2 to 3 years cryopreservation period, cryopreservation sperm can be useful for seed production.

• Journal of Aquaculture
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• v.17 no.3
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• pp.180-186
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• 2004

This study was carried out to obtain the fundamental data for the mass seedling production of grunt, Hapalogenys nitens in terms of the natural spawning and some characteristics of the eggs spawned. The wild grunt were reared at indoor tanks for three years. The adults spawners were 34.0∼44.0 cm (38.6$\pm$4.0 cm, n=7) in total length, 1.00∼2.23 kg (1.62$\pm$0.50 kg, n=7) in body weight. Spawning were observed 9 times from September 22 to October 1, 2000 and 37 times from August 22 to October 3, 2001, with a water temperature range of 19.8$\pm$28.5$^{\circ}C$. The total number of eggs collected was 2.29${\times}$10$^{7}$ (1.7${\times}$10$^{3}$/ml). The relative proportion of floating eggs to total eggs was 41.7%. The fertilization rate of floating eggs was ranged between 85.0 and 99.9% and the hatching rate was ranged between 2.9 and 93.0%. Fertilized eggs were buoyant and spherical in shape, and were 0.85∼0.98 mm in diameter. Each egg contained 1-5 oil globules which were, 0.18∼0.25 mm in diameter. The incubation time from fertilization to blastodisc formation was 10 minutes, to blastula was 3 hours, and to the hatched larvae at 26$^{\circ}C$ was 20 hours 30 minutes. The newly hatched larvae attained total length of 1.81$\pm$0.18 mm. The time required from fertilization to hatching was 31∼34 hours at 23$^{\circ}C$ and 17∼20 hours at 29$^{\circ}C$.

• Korean Journal of Veterinary Research
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• v.38 no.2
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• pp.394-400
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• 1998

Cryopreservation of embryos by vitrification is a simple method to preserve bovine embryos for subsequent embryo transfer, but embryonic viability after vitrification has been inconsistent and low compared with conventional slow freezing. The aim of the present study is to examine the effect of serum or serum albumin in a vitrification solution and epidermal growth factor(EGF) or fibroblast growth factor(FGF) on in vitro viability of bovine blastocysts frozen by vitrification. Bovine blastocysts were produced by in vitro maturation, fertilization of follicular oocytes and culture of embryos in a synthetic oviduct fluid medium(SOFM) containing BSA and 19 essential and nonessential amino acids. Blastocysts with excellent or good morphology were selected at 7 or 8 days after culture and utilized for vitrification. In experiment 1, blastocysts were vitrified in a solution containing semi-fetal calf serum(SFCS) or BSA(5 or 10mg/ml) and then their subsequent viabilities were examined by culturing thawed embryos in a SOFM containing BSA and 19 amino acids. Effect of EGF or FGF added to a SOFM containing polyvinyl alcohol(PVA) on the viability of vitrified-thawed blastocysts was investigated in experiment 2. BSA added at 5 or 10mg/ml to a vitrification solution showed significantly higher(p < 0.05) developmental rate to expanded and hatching blastocysts than SFCS, but there was no significant difference in the developmental rate to hatched blastocysts after thawing. Supplementation of a culture medium with EGF and/or FGF significantly increased(p < 0.05) embryo development to expanded blastocysts compared with control but showed no beneficial effect on the development to hatching or hatched blastocysts. Coculture of thawed embryos with granulosa cells in a TCM 199 containing 10% fetal calf serum(FCS) showed the highest developmental rate to expanded, hatching and hatched blastocysts among the groups tested. In conclusion, supplementation of a vitrification solution with BSA at 5mg/ml and culture of thawed blastocysts in a medium containing EGF and/or FGF can improve in vitro viability of bovine blastocysts frozen by vitrification.

• Korean Journal of Fisheries and Aquatic Sciences
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• v.37 no.2
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• pp.98-104
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• 2004

Early development and metamorphosis of the ascidian (Halocynthia hilgendorfi ritteri) were investigated from fertilized egg. The samples were collected in the coastal waters of Yongdam, northwest of Jeju Island in November 2002. H. hilgendorfi ritteri was solitary ascidian and produced spheral eggs with egg size ranging from $0.33\pm0.01\;mm.$ On the outer surface of the vitelline coat are attached many follicle cells. At $21.0\pm0.5^{\circ}C$ of water temperature, first cleavage took place in about 1.5 hrs after fertilization, and gastrulation followed in about 12.5 hrs. The formation of tailbud embryos and free swimming larvae were observed 13.3 hrs and 20.5 hrs after fertilization, respectively. The size of newly hatched tadpole larva was 1.30-1.45 mm, the larva swam for 2 hrs to 14 hrs. At 4 hrs after hatching, the palpi were lost and tail absorption began with an abrupt rupture of the anterior end of the notochord. At 17-18 hrs after hatching, tail completely absorption and remained trunk. The coniform adhesive papilla began protrusion at 30 hrs after hatching. The oral and atrial siphon formed at 6-7 days after settlement. At 17-18 days after settlement, metamorphosed the larvae developed into protoascidian of which the external morphology was similar to their adult.

• Clinical and Experimental Reproductive Medicine
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• v.20 no.1
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• pp.9-17
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• 1993

This study was carried out investigate the effect of water quality and the kind of media on the in vitro development of 1-cell and stage mouse embryos. $F_1$ hybrid mice were superovulated and timely mated. 1-cell stage and 2-cell stage mouse embryos were recruited and taken into Ham's F-10 or m-KRB media which was made of two of two kinds of water having different quality, highly purified water and tap water. 2-cell stage embryos grew up well in vitro to blastocyst or hatching blastocyst regardless of the composition of culture media, but 1-cell stage mouse embryo didn't develop well to blastocyst or hatching blastocyst in simple media like m-KRB. These results meant in vitro devleopment of 1-cell stage mouse embryo neded complex media like Ham's F-10 which contained abundant protein components. In case of quality control for water, in vitro fertilization program. observation of in vitro development of 2-cell mouse embryos up to blastocyst or hatching blastocyst media such as m-KRB would be efficatious in detecting the difference of water quality.

• Korean Journal of Fisheries and Aquatic Sciences
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• v.51 no.4
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• pp.420-425
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• 2018

We characterized egg development and investigated the effects of temperature on embryonic development time, in the convict cichlid Amatitlania nigrofasciata. Fertilized eggs of this species have an ovoid shape (horizontal axis, $1.56{\pm}0.02mm$; vertical axis, $1.26{\pm}0.04mm$) and smooth translucent chorion surrounded by a layer of mucous secretion. At $27{\pm}0.5^{\circ}C$, the first cleavage, blastula, gastrula and 16-somite stages of A. nigrofasciata eggs began at 1.5, 4.83, 14 and 40 hours after fertilization, respectively. We measured the development rate of embryos at $12-33^{\circ}C$ and found that the time period from fertilization to hatching was 94 hours at $24^{\circ}C$, 64 hours at $27^{\circ}C$ and 50 hours at $30^{\circ}C$. At temperatures of $12-21^{\circ}C$, all fertilized eggs died before hatching and those incubated at $33^{\circ}C$ died immediately after hatching.

• Environmental Analysis Health and Toxicology
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• v.25 no.3
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• pp.241-251
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• 2010

Crucian carp (Carassius auratus) has been used as the sentinel species for POPs (Persistent Organic Pollutants) monitoring in aquatic environment. However, there is little information for dioxin toxicity and especially, early life stage toxicity in crucian carp have been never carried out. In this study, we investigated several toxic effects for 2,3,7,8-tetrachlorodibenzo-p-dioxin (2,3,7,8-TCDD) in fertilized egg obtained by natural fertilization from crucian carp. The embryos at 3 h post-fertilization (hpf) were treated with 0.039, 0.156, 0.625, and 2.5 (${\mu}g/L$) TCDD by waterborne exposure for 60 minutes and changed with fresh water 2 times per day. Fertilized eggs started hatching at 51 hpf and TCDD exposed embryo showed decrease of hatching rate in a dose-dependent manner at 75 hpf. Pericardial edema was continuously observed in larvae exposed to TCDD from hatching start time (51 hpf), followed by the onset of mortality. In addition, AhR-related gene, CYP1A was clearly increased by TCDD in a dose dependent manner. These results indicated that fertilized eggs obtained from crucian carp have the TCDD related gene regulation and a distinct TCDD developmental toxicity syndrome by TCDD exposure. Therefore, we suggested that early life stage test in crucian carp could be used as test methods on dioxins toxicity.

• Ocean and Polar Research
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• v.42 no.4
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• pp.303-311
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• 2020

In this study, the embryonic development and hatchability of eggs fertilized by the reciprocal hybrids of kelp grouper (Epinephelus bruneus) and red-spotted grouper (E. akaara) were evaluated with the goal of establishing a novel hybrid with enhanced growth and viability during the farming period in the temperate waters of Korea. The fertilization rates were lower for hybrids than for maternal purebreds and were significantly higher in the red-spotted grouper ♀ × kelp grouper ♂ hybrid (RGKG, 89.61 ± 1.58%) than in the kelp grouper ♀ × red-spotted grouper ♂ hybrid (KGRG, 74.82 ± 4.23%, p < 0.05). Unlike the fertilization rates, the hatching rates of fertilized eggs were similar between hybrids and maternal purebreds and did not differ significantly between KGRG and RGKG (72.74 ± 3.60% vs. 75.23 ± 2.20%, respectively, p > 0.05). The embryonic development of the hybrids was similar to that of maternal purebreds; however, irregular cleavage and asymmetric blastoderm were noticeable in the developing eggs of KGRG hybrids. The deformity rates of newly hatched larvae were higher in hybrids than in maternal purebreds and were significantly higher in KGRG than in RGKG (17.47 ± 1.28% vs. 7.11 ± 0.54%, respectively, p < 0.05). These results demonstrate the potential to produce viable larvae from these two hybrids. Although the production efficiency of KGRG was lower than that of RGKG, the fertilization, hatching, and deformity rates make both hybrids useful for further comparative studies regarding economic aspects.

• The Korean Journal of Malacology
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• v.31 no.3
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• pp.179-186
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• 2015

The Pacific oyster Crassostrea gigas oocytes were exposed to 4 cryoprotectants, dimethyl sulfoxide (DMSO), ethylene glycol (EG), methanol, or polyethylene glycol (PEG), each with 4 four concentrations (5, 10, 15, and 20%) and for 10, 20, 30 or 40 minutes for permeation. The oocytes were then fertilized, using normal sperm of the species. Fertilization and hatching rates were clearly influenced by cryoprotectant species and their concentration and permeation time. Overall, they decreased as concentrations and permeation time of cryoprotectants increased with optimum results at concentrations of 5-10% and a permeation time of 10 minutes. Larval abnormalities, a sign of the chemical damage, were a representative phenotype which was higher at a higher concentration and longer duration of the chemicals. In conclusion, best result was from 5% DMSO exposure for 10-20 minute permeation.

• Journal of Aquaculture
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• v.12 no.1
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• pp.39-45
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• 1999

In order to obtain the basic biological data for artificial seedling production of sea cucumber, Stichopus japonicus, the influence of water temperature and air dry for spawning induction, egg development and larvae rearing was investigated during the period from April, 1995 through September, 1995. Spawning induction rate by the water temperature was 6.0~17.5% and air dry was responsed 1.4~4.0%. Number of eggs spawned of Stichopus japonicus were $50~500\times10^4$ individuals, the fertilization and hatching rate were ranged 84.0~96.0%, 71.4~84.6% respectively. The fertilized egg of Stichopus japonicus appeared mean diameter of $154{\mu}m$. At a constant water temperature of $23^{circ)C$, it become 4 tell stage from fertilization after 2 hours 10 minutes, hatching larvae after 14 hours half, auricularia larvae after 3 days, doliolaria larvare after 11 days and pentactula larvae after 15 days ready for settlement. The suitable food in the larvae reared for 17 days after fertilization were shown the best growth and survival in the larvae food of Chaetoceros calcitrans. Optimum density for larvae rearing were maintained of the larval density lower than 2 individuals/ml.