• Korean Journal of Animal Reproduction
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• v.26 no.2
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• pp.119-124
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• 2002

This study was carried out to investigate the effects of packing materials of frozen boar semen to improve reproductive performance efficiency in pig. Boars were raised at Swine Artificial Insemination Center in National Livestock Research Institute, Sunghwan, Chungnam, Korea. We compared packing protocols for frozen boar semen among 5$m\ell$ maxi-straw, 5$m\ell$ cryogenic-vial, and aluminum-pack. Cryogenic-vial packing material showed similar sperm characteristics compared with maxi-straw packing material when the sperm was frozen above 15cm from liquid nitrogen and thawed at 52$^{\circ}C$ for 190 seconds. We investigated different thawing times to find out the optimal condition of freezing and thawing protocol with cryogenic-vial. Freezing above 15cm from liquid nitrogen and thawing at 52$^{\circ}C$ for 190 seconds were the optimal protocol compared with 120 and 150 seconds. However, normal acrosome rates did not show any differences among thawing times. Post-thawing results of maxi-straw in water at 52$^{\circ}C$ for 45 seconds had better total motility and curve linear velocity than those of cryogenic-vial in water 52$^{\circ}C$ for 190 seconds. However, there were no differences on straightness and normal apical ridge of sperm between maxi-straw and cryogenic vial. Non-return rate, farrowing rate and litter size of sows inseminated with frozen boar semen of commercial farms were higher in the maxi-straw than cryogenic-vial, but there were no significant differences between maxi-straw and cryogenic-vial. In conclusion, there were no significant differences between maxi-straw and cryogenic-vial and so, we may replace cryogenic-vial packing method instead of maxi-straw packing method by improvement of freezing and thawing rate.

• Reproductive and Developmental Biology
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• v.33 no.1
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• pp.25-28
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• 2009

This study was carried out to investigate the general characteristics, such as volume, pH, sperm motility and sperm concentration of the semen collected from Beagle dogs (age $24{\sim}48$ months, weight $10{\sim}15\;kg$) by using the method of digital manipulation of the penis, and the effect of preservation temperature and time on motility of fresh semen. Multiple ejaculates were collected from 4 male Beagles. The average volume, pH, motility and sperm concentration of the second fraction (contained with small volume of the third fraction) per ejaculation were $2.94{\pm}0.24(SD)\;ml$, $6.43{\pm}0.42(SD)$, $97.04{\pm}3.50(SD)%$ and $1.67{\pm}0.23(SD){\times}10^8\;cells/ml$, respectively. Average semen volume per ejaculate, semen pH, sperm motility and sperm concentration of the first fraction from the ejaculate were $1.24{\pm}0.20(SD)\;ml$, $6.03{\pm}0.26(SD)$, $1l.30{\pm}4.02(SD)%$ and $7.25{\pm}1.02(SD){\times}10^5\;cells/ml$. Those of second fraction were $2.52{\pm}0.32(SD)\;ml$, $6.32{\pm}0.31(SD)$, $96.25{\pm}3.52(SD)%$ and $2.35{\pm}0.35(SD){\times}10^8\;cells/ml$. Those of third fraction were $2.71{\pm}0.27\;(SD)\;ml$, $6.52{\pm}0.20(SD)$, $95.65{\pm}2.78(SD)%$ and $5.72{\pm}0.29(SD){\times}10^7\;cells/ml$. Motility of semen was higher at $17^{\circ}C$ preservation temperature than $5^{\circ}C$ or $36^{\circ}C$ during preservation period. When preservation temperature was $17^{\circ}C$, motility was $96.54{\pm}2.05(SD)%$ at 1 h, $90.20{\pm}3.90(SD)%$ at 6 h, $89.05{\pm}2.01(SD)%$ at 12 h, $78.21{\pm}3.50(SD)%$ at 18 h, $45.24{\pm}6.25\;(SD)%$ at 24 h and $30.75{\pm}17.24(SD)%$ at 30 h, respectively.

• The Korean Journal of Malacology
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• v.26 no.3
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• pp.235-244
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• 2010

Ultrastructural characteristics of the testis and spermatogenesis of Crassostrea gigas were investigated by Transmission and Scanning Electron microscope observations. The testis is a diffuse organ consisting of branching acini containing differentiating germ cells in a variety of stages. The acinus is surrounded by an intermitent layer of myoepithelial cells andis divided into subcompartments that are partially separated by pleomorphic accessory cells which remain in close contact with germ cells until late stages of development. these accessory cells contain a large quantity of glycogen particles and lipid droplets in the cytoplasm. Therefore, it is assumed that they are involved in the supplying of the nutrients for germ cell development, while any phenomena associated with phagocytosis of undischarged, residual sperms by lysosomes could be find in the cytoplasm of the accessory cells. The morphology of the spermatozoon has a primitive type and is similar to those of other bivalves. Mature spermatozoa consist of broad, cap-shaped acrosomal vesicle, subacrosomal material (containing axial rod embedded in a granular matrix), a oval nucleus showing deeply invaginated anteriorly, two triplet substructure centrioles surrounded by four spherical mitochondria, and satelite fibres appear to the distal centriole and plasma membrane. Spermatozoa of C. gigas resemble to those of other investigated ostreids. In particular, the anterior region of the acrosomal vesicle is transversely banded. It is assumed that differences in this acrosomal substructure are associated with the inability of fertilization between the genus Crassostrea and other genus species in Ostreidae. Therefore, we can use sperm morphology in the resolution of taxonomic relationships within the Ostreidea. The spermatozoon is approximately $42-47{\mu}m$ in length including an oval sperm nucleus (about $0.91{\mu}m$ in length), an acrosome (about $0.42{\mu}m$ in length) and tail flagellum ($40-45{\mu}m$). The axoneme of the sperm tail flagellum consists of nine pairs of microtubules at the periphery and a pair at the center. The axoneme of the sperm tail shows a 9 + 2 structure. These morphological charateristics of acrosomal vesicle belong to the family Ostreidae in the subclass Pteriomorphia.

• Journal of Embryo Transfer
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• v.18 no.2
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• pp.143-149
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• 2003

This study was carried out to cryopreserve and to investigate characteristics of semen in Hanwoo. Semen was obtained from bulls selected by Daekwanryeong Branch station. Semen was collected each morning of the experiment, placed in water jacketed tubes at 37$^{\circ}C$, and trans-ported to the research laboratory within 10 minutes. Semen was extended with Egg yolk-glycerol extender to contain 50${\times}$10$^{6}$ sperm/ml. Semen was cooled over a 6h period in water jacketed tubes from about 25 to 5$^{\circ}C$, Egg yolk-glycerol extender was added in one step at 5$^{\circ}C$. Semen was aspirated into 0.5ml straws, which were sealed with powder. Egg yolk-glycerol extender, which is used in Hanwoo sperm frozen and stored, semen from 13 Hanwoo bulls collected, the postthawed percentages of motile sperm were 65.7%. In semen characteristics of Hanwoo bulls, number of bulls volume are 5.7 ml and total cell count are 975${\times}$10$^{6}$ m1 ejaculate.

• Asian-Australasian Journal of Animal Sciences
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• v.19 no.12
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• pp.1716-1721
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• 2006

The present study was conducted to observe the effect of osmolality of glycerolated TEST-yolk glycerol extenders on post-thawing sperm kinematics of ram spermatozoa of the native Malpura breed maintained in a semi-arid tropical environment. Good quality semen obtained from adult rams was pooled, split and diluted to 1,000 million spermatozoa per ml in complete TEST-yolk-glycerol extenders of 900, 1,200, 1,500 and 1,800 mOsm/kg osmolality. Diluted semen samples were loaded in 0.25 ml straws and cooled down to $-125^{\circ}C$ freezing temperature at the rate of $-25^{\circ}C$ per minute under controlled conditions before plunging into liquid nitrogen for storage. The thawing of straws was performed at $50^{\circ}C$ in a water bath for 10 seconds and sperm kinematics of the frozen-thawed spermatozoa were assessed by a computer-assisted sperm analysis technique. Osmolality of diluent had no significant effect on post-thawing % motility, % rapid, % medium and % slow moving frozen-thawed spermatozoa but significantly (p< 0.05) affected the % linearity and % straightness. The post-thawing % motility and % rapid motile spermatozoa were highest in samples extended in diluent of 1,500 mOsm/kg osmolality and lowest in 900 mOsm/kg. The curvilinear velocity of spermatozoa was significantly (p<0.05) higher for samples extended in 1,800 mOsm/kg, compared to those in 900 and 1,200 mOsm/kg, but the effect was not significantly different to those extended in diluent of 1,500 mOsm/kg osmolality. The study indicated that ram spermatozoa could tolerate a wide osmolality range for dilution in the complete TEST-yolk-glycerol extender for their cryosurvival. The highest recovery of motile spermatozoa following thawing was achieved in samples extended in the TEST-yolk-glycerol diluent of 1,500 mOsm/kg osmolality.

• Journal of Veterinary Clinics
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• v.26 no.6
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• pp.647-649
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• 2009

An orchiopexy was performed in an 18-month old adult English bulldog with bilateral cryptorchidism. One month postoperatively, the dog was twice-treated with GnRH (50 ug/kg) at an interval of 2 weeks. Semen was collected and evaluated before and after surgery. Fertility was determined by artificial insemination. No spermatozoa were observed before orchiopexy and 2 months postoperatively. However, 6 live sperm were detected 4 months postoperatively and normal sperm characteristics (except sperm concentration) were present 7 months postoperatively. A female bulldog, inseminated with the semen from the bulldog 8 months postoperatively, delivered 6 offsprings. Spermatogenesis and spermatozoa with fertilizing capacity were recovered by postpubertal orchiopexy and GnRH therapy in a bilateral cryptorchid dog.

• Clinical and Experimental Reproductive Medicine
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• v.50 no.4
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• pp.270-276
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• 2023

Objective: This study investigated the clinical and laboratory factors associated with the presence of dysmorphic oocytes in intracytoplasmic sperm injection (ICSI) cycles. Methods: The study involved 200 ICSI cycles, performed from 2020 to 2021, that yielded at least one mature oocyte. Clinical characteristics and ovarian stimulation methods were compared between 68 cycles with at least one dysmorphic oocyte (the dysmorphic group) and 132 cycles with normal-form oocytes only (the non-dysmorphic group). Dysmorphic oocytes were characterized by dark cytoplasm, cytoplasmic granularity, cytoplasmic vacuoles, refractile bodies in the cytoplasm, smooth endoplasmic reticulum in the cytoplasm, an oval shape, an abnormal zona pellucida, a large perivitelline space, debris in the perivitelline space, or an abnormal polar body. Results: The ages of the women, indications for in vitro fertilization, serum anti-Müllerian hormone levels, and rates of current ovarian endometrioma were similar between the dysmorphic and non-dysmorphic groups. In both groups, the three ovarian stimulation regimens, two types of pituitary suppression, and total gonadotropin dose were employed similarly. However, the dual-trigger method was used more frequently in the dysmorphic group (67.6% vs. 50%, p=0.024). The dysmorphic group contained significantly more immature oocytes and exhibited significantly lower oocyte maturity (50% vs. 66.7%, p=0.001) than the non-dysmorphic cycles. Within the dysmorphic group, significantly lower oocyte maturity was found in the cycles using a dual-trigger, but not in those with a human chorionic gonadotropin trigger. Conclusion: ICSI cycles with dysmorphic oocytes are closely associated with reduced oocyte maturity. This association was observed exclusively in dual-trigger cycles.

• Journal of Animal Science and Technology
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• v.52 no.5
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• pp.407-412
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• 2010

The objective of this study was to determine the effects of dietary supplementation of whole garlic powder (WGP) on semen characteristics and blood antioxidant level in boars. For this study, nine Duroc boars of 12 months age were used. Semen and blood samples were collected for 13 weeks, once in each week. The boars were fed the basal diet (BD; control) or BD supplemented with 3% WGP. There were no significant differences in the semen volume and sperm concentration between control and WGP group on all collection weeks. However, total sperm number per ejaculate was higher in the WGP group than that in the control group on collection weeks 6, 7 and 8 (P<0.05). Also, on collection weeks 5, 6, 7 and 8, mean of total ejaculated sperm numbers per boar were significantly higher in the WGP group compared to control group (P<0.05). On the other hand, ejaculation frequency per boar (boar's libido) and total ejaculated sperm number per boar were significantly increased in the WGP group compared to the control group, respectively (P<0.05). Although there was no difference in polyphenol level in seminal plasma between two treatment groups, polyphenol level in blood serum was significantly higher in the WGP group on collection weeks 9, 12 and 13 (P<0.05). These results indicate that dietary supplementation of 3% WGP improves boar libido and semen productivity such as ejaculation frequency per boar, total sperm number per ejaculate, mean of total ejaculated sperm number per head, and elevate the blood level of antioxidant (polyphenol) in boar serum.

• Korean Journal of Animal Reproduction
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• v.26 no.3
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• pp.275-289
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• 2002

The objective of this study was to develop an in vitro assessment of sperm fertilizing capacity of bulls and investigate the factors influencing sperm function and characteristics of frozen-thawed bovine spermatozoa. in vitro fertilization (IVF), the evaluation of motility and normal morphology, HOST (hypoosmotic swelling test), Ca-ionophore induced acrosome reaction, luminol and lucigenin-dependent chemiluminescence for the measurement of reactive oxygen species (ROS), the measurement of malondialdehyde formation for the analysis of lipid peroxidation (LPO), and the evaluation of DNA fragmentation using the method of 747-mediated nick end labelling (TUNEL) by flow cytometry were performed in frozen-thawed bovine spermatozoa. Correlations between the rates of fertilization, blastocyst formation after IVF and the values of respective assays were investigated. 1. IVF rate and blastocyst formation rate averaged 64.4% and 34.3% for spermatozoa from high -fertility bull group and averaged 18.5% and 6.2% for spermatozoa from low-fertility bull group, respectively. There were significantly different between two bull groups. Sperm motility and percentage acrosome reaction averaged 79.0% and 66.2% for spermatozoa from high-fertility bull group and averaged 40.7% and 22.9% for spermatozoa from low-fertility bull group, respectivitely. There were not different between two bull groups. 2. Luminol depenent chemiluminescence, LPO and DNA fragementation averaged 6.4, 2.0 nmol and 2.6% from spermatozoa from high-fertility bull group and averaged 6.5, 3.1 nmol and 7.4% for spermatozoa from low-fertility bull group, respectively. There were significantly different between two bull groups. There was no significant difference in lucigenin dependent chemiluminescence between two bull groups. 3. Fertilization rate was positively correlated with motility and the rate of Ca-ionophore induced acrosome reaction, but negatively correlated with the frequency of luminol-dependent chemiluminescence, the rate of LPO, and the percentage of sperm with DNA fragmentation. There was no correlation between fertilization rate and the percentage of swollen spermatozoa, normal morphology, and the frequency of lucigenin-dependent chemiluminescence. 4. Blastocyst formation rate was positively correlated with the rate of Ca-ionophore induced acrosome reaction, but negatively correlated with the frequency of luminol-dependent chemiluminescence, the rate of LPO, and the percentage of sperm with DNA fragmentation. There was no correlation between blastocyst formation rate and motility, the percentage of swollen spermatozoa, normal morphology, and the frequency of lucigenin-dependent chemiluminescence. In conclusion, these data suggest that ROS significantly impact semen quality. The assays of this study may provide a basis fur improving in vitro assessment of sperm fertilizing capacity.

• Journal of Life Science
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• v.27 no.11
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• pp.1276-1289
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• 2017

This study isolated and purified the antimicrobial peptide McSSP-31 from an acidified hemocyte extract of a Mytilus coruscus. The antimicrobial peptide was purified by using a $C_{18}$ reversed-phase high-performance liquid chromatography (HPLC). The peptide was determined to be 3330.549 Da by matrix assisted-laser desorption ionization time-of-flight mass spectrophotometer (MALDI-TOF/MS). The N-terminus of a 14 amino-acid sequence was identified as P-S-P-T-R-R-S-T-S-R-S-K-S-R by Edman degradation method. The acquired sequence showed a 93% similarity with the sperm-specific protein Phi-1, which is from M. californianus. The identified open-reading frame (ORF) of peptide was 306 bp encoding 101 amino acids, which was analyzed by rapid amplification of cDNA ends (RACE), cloning and sequencing analysis. We compared the full sequence with other known proteins that reveal the sperm-specific protein Phi-1 (93.5%) of M. californianus. Synthesized antimicrobial peptide (McSSP-31) showed antibacterial activity against gram-positive bacteria including B. subtilis, S. mutans, S. aureus and gram-negative bacteria including E. coli, K. pneumoniae, P. mirabilis, P. aeruginosa and fungi, C. albicans. Also, synthesized peptide showed strong antibacterial activity against antibiotic-resistant strains, including S. aureus. The cytotoxicity of the peptide was determined by using the HUVEC human cell line. The peptide did not exhibit any significant cytotoxic effects on the normal human cell line, and it had very low hemolytic activity with flounder hemoglobin. The results demonstrated that peptide purified from the hemocyte of a M. coruscus exhibits antibacterial activity against various bacteria and has the potential to be an alternative antibiotic agent.