Background: Sperm quality and the number of sperm introduced into the uterus during artificial insemination (AI) are pivotal factors influencing pregnancy outcomes. However, there have been no reports on the relationship between sperm concentration at AI and sperm quality in Hanwoo cattle. In this study, we examined sperm quality and pregnancy rates after AI using sperm inseminated at different concentrations. Methods: We evaluated the motility, viability, and acrosomal membrane integrity of sperm at different concentrations (10, 15, 18, and 20 million sperm/straw) in 0.5-mL straws. Subsequently, we compared the pregnancy rates after AI with different sperm concentrations. Results: After freeze-thawing, sperm at the assessed concentrations showed similar viability and acrosomal membrane integrity. After AI, cattle in the 10 million group had significantly lower pregnancy rates compared to those in the 18 and 20 million groups. Conversely, there were no statistically significant variances observed between cattle in the 10 and 15 million groups. Conclusions: Sperm at concentrations of 10, 15, 18 and 20 million per straw exhibited comparable motility, viability, and acrosomal membrane integrity. However, a concentration of at least 18 million sperm per straw is required to achieve a consistent rate of pregnancy rate in Hanwoo cattle after AI.

In the present study, we examined the effect of straw size on spermatozoa motility, viability, acrosome integrity, mitochondrial membrane potential, and plasma membrane integrity after freezing-thawing. Hanwoo semen was collected from three bulls and diluted with an animal protein-free extender, divided into two groups, namely, 10 million spermatozoa in 0.25 mL and 20 million spermatozoa in 0.5 mL straw, and cryopreserved. In Experiment 1, the motility and motility parameters of the frozenthawed spermatozoa were evaluated. After freezing-thawing, the spermatozoa motility parameters fast progressive, straight line velocity, and average path velocity were compared between the 0.25 mL straw and 0.5 mL straw groups. They were 35.2 ± 1.0 and 32.3 ± 0.7%, 34.6 ± 0.7 and 31.8 ± 0.5 μm/s, 51.4 ± 1.3 and 47.1 ± 1.1 μm/s, 0.25 mL straw and 0.5 mL straw groups, respectively. In Experiment 2, the viability, acrosome membrane integrity, and mitochondrial membrane potential of the frozen-thawed spermatozoa were assessed. After freezing-thawing, the percentages of spermatozoa with live, intact acrosomes and high mitochondrial membrane potential were compared between the in 0.25 mL straw and 0.5 mL straw groups. They were 48.0 ± 2.6% and 35.6 ± 2.8% between the 0.25 mL straw and 0.5 mL straw groups. In Experiment 3, the plasma membrane integrity of frozen-thawed spermatozoa was compared. After freezingthawing, the plasma membrane integrity was higher for the in 0.25 mL straw group than the 0.5 mL straw group. They were 62.0 ± 2.2 and 54.1 ± 1.3% between the 0.25 mL straw and 0.5 mL straw groups. In conclusion, our results suggest that freezing semen in 0.25 mL straw improves the relative motility, viability, and acrosomal, mitochondrial membrane potential, and plasma membrane integrity of Hanwoo bull spermatozoa.

This study was conducted to examine the oocyte recovery efficiency through having an OPU session once and twice a week. Also, the oocyte recovery efficiency was examined by using OPU after two and three months of rest period. Six cows were used for oocytes collection and were randomly divided into two groups. In experiment 1, OPU sessions were conducted once and twice a week to collect oocytes. The collected oocytes between once and twice OPU groups were classified into four groups (grade 1, 2, 3 and 4) according to the quality of cumulus cells and ooplasm. Based on the result, the percentage of collected oocytes per aspirated follicle number was similar between once and twice OPU session groups (65.5 ± 1.9 and 68.7 ± 1.4 vs.). However, the percentage of grade 1 oocytes from the twice OPU session group was significantly high compared with that of the once a week OPU session group (25.3 ± 0.9 and 32.5 ± 1.2% vs. once and twice session group, respectively, p < 0.05). In experiment 2, the group with three months of rest period tended to have a high percentage of collected oocyte compared with the group with two months of rest period (64.6 and 70.9% vs. 2 and 3 months rest group, respectively, p = 0.62). The percentage of grade 4 in the group with three months of rest period was significantly low compared with the group with two months of rest period group (27.3 and 36.5% vs. two and three months rest group, respectively, p = 0.05). In conclusion, twice a week OPU session is suitable for collection of high quality oocytes by using OPU, and three months of rest period is needed for the recovery of oocyte quality of a donor cow.

In this study, two epididymal spermatozoa recovery methods in relation to sperm number, motility, viability and acrosome reaction were examined. Seven bulls were castrated and 7 testicles with epididymides were transferred to the laboratoy. Epididymis in each bull was randomly used for flushing and mincing methods with semen extender (Optixcell, IMV, France). The recovered spermatozoa with adjusted sperm concentration to 40 × 106 cells/mL was diluted with optixcell and cryopreserved. In experiment 1, the difference in the total number of spermatozoa using flushing and mincing methods was insignificant (2570.0 and 2505.2 × 106 cells/mL, respectively). For experiment 2, the percentage of motile spermatozoa and motility parameters between flushing and mincing methods were studied through the use of sperm class analyzer after frozen-thawing. The percentage of total motile sperm between flushing and mincing methods was almost the same with 89.5±12.8 and 91.4±7.9%, respectively. The same is the case with experiment 3 wherein the viability and acrosomal integrity of frozen-thawed epididymal spermatozoa by flushing and mincing was insignificantly different. The results from the study showed that both flushing and mincing methods can be used for epididymal spermatozoa recovery in bull.

Serum metabolites were analyzed to investigate relationship of pregnancy and non-pregnancy Hanwoo cows. Totally, 251 Hanwoo cows were used in the present study. Grazing was carried out for 5 months in the pasture. In barn feeding, concentrate 3.0 Kg (TDN 68%, CP 14%) and rice straw 6 kg(TDN 50%, CP 6.5%) were fed. For artificial insemination (AI), progesterone-supplying device (CIDR) was introduced to vagina of Hanwoo cows and 2.0 mL of GnRH. One week after introduction, CIDR was removed and 5.0 mL of PGF2α was injected intramuscularly. After 2.5 day, AI was accompanied by a 2 mL of GnRH intramuscular injection and a second AI was carried out 3.5 day. The pregnancy diagnosis was confirmed by rectal palpation about 90 days after AI. Blood samples were collected from the jugular vein after 3 hours of feeding. Analysis of serum metabolites was performed on six types of metabolites: glucose(mg/dl), cholesterol(mg/mL), BUN(mg/dl), AST(U/L), ALT(U/l), and nonessterified fatty acids(NEFA, uEq/L). The metabolic profile test was analyzed by analyzer (Hitachi, 7020, Japan). Pregnant and non-pregnant groups showed serum metabolites as follows. In 60 pregnant group: Glucose 88.9 ± 2.5, Cholesterol 149.8 ± 4.9, BUN 16.9 ± 0.4, AST 99.1 ± 2.6, ALT 35.9 ± 0.9 and NEFA 326.7 ± 15.7. In 43 non-pregnant cow group: Glucose 89.2 ± 3.3, Cholesterol 165.9 ± 4.6, BUN 17.4 ± 0.6, AST 108.9 ± 0.6, ALT 37.8 ± 1.0 and NEFA 419.2 ± 32.8. Cholesterol, AST and NEFA levels in non-pregnant cows were significantly higher than those in pregnant cows (P<0.05). In sum of grazing and barn feeding group was totally 148 cows. Seventy nine pregnant cows showed high glucose and low NEFA levels compared to 69 non-pregnant cows (P<0.05). In conclusion, pregnant group showed high level of glucose and low level of cholesterol, NEFA. Further study needed to obtain more accurate level of metabolites in serum for pregnant and non-pregnant cows.

In this study, we examined number, motility and plasma membrane integrity of spermatozoa from six regions of epididymis in bull. Six testicles with epididymides were castrated from six bulls (mean±standard error, age of days = 441.3±9.6, body weight (kg) = 367±8.4, scrotal circumference (cm) = 30.7±0.4) at Hanwoo Research Institute, NIAS and transported to laboratory within 1 hour. Testicular weight, length, width and circumference were recorded. Epididymis in each bull was randomly used for recovery of spermatozoa. Epididymis was divided into six regions: efferent duct (ED), caput, corpus, proximal cauda (Pcauda), distal cauda (Dcauda) and vas deferens (VD). In experiment 1, we examined sperm number of each region of epididymis. Each region of epididymis contained different number of spermatozoa: ED (37.8±15.7 × 106cells/ml, 8.2%), caput (93.6±18.8 × 106cells/ml, 20.2%), corpus (33.0±8.5 × 106cells/ml, 7.1%), Pcauda (104.2±23.5 × 106cells/ml, 22.5%), Dcauda (180.5±32.5 × 106cells/ml, 39.0%) and VD (14.0±5.0 × 106cells/ml, 3.0%). In experiment 2, sperm motility of each epididymal region was examined by computer assisted sperm analysis (SCA, MicroOptic) system. Sperm motility was divided into 4 groups (fast progressive, slow progressive, non-progressive and immotile) based on WHO guideline. Percentages of fast progressive of Pcauda and Dcauda (11.0±2.3 and 15.4±3.6%) were significantly higher than that of ED, Caput, Corpus and VD which is 0.1±0.1, 1.5±0.6, 1.9±0.7 and 0.3±0.2%, respectively (p<0.05). In experiment 3, percentage of intact plasma membrane spermatozoa of each regions were examined by hypoosmotic swelling test. Percentages of intact plasma spermatozoa were not significantly different among six regions of epididymis: ED, caput, corpus, Pcauda, Dcauda and VD which is 68.0±8.6, 74.0±5.3, 68.5±6.2, 70.8±5.5, 71.0±5.8 and 64.6±10.8%, respectively. In conclusion, in the present study, we found out distribution, motility and plasma membrane integrity of spermatozoa from six regions of epididymis in Hanwoo bull. These results will be contributed to basic research about spermatozoa transportation and characters in epididymis of bull.

In this study, we examined sperm penetration and blastocyst developmental rate of oocytes to determine fertilizability of cauda epididymal spermatozoa in Hanwoo bull. One testicle with epididymides were castrated from one Hanwoo bull (14 months of age) and transported to laboratory. Spermatozoa recovered from cauda epididymis by mincing with semen extender (Optixcell, IMV, France) and cryporeserved in liquid nitrogen tank until use. As control, frozen Hanwoo semen was used. Cumulus oocyte complexes (COCs) were collected from follicles (2-8 mm) of slaughtered ovaries and 10 to15 COCs were matured in 50μl droplet with M-199 media supplemented with 10% fetal bovine serum, 10μg/ml FSH, 10μg/ml LH, 10μg/ml EGF for 22 to 24 hours in a humidified atmosphere of 5% CO2 in air. After maturation of COCs, matured COCs were co-incubated with cauda epididymal spermatozoa in 100μl droplet in modified Brackett and Oliphant media supplemented with 2.5 mM theophylline for 12 or 18 hours under 5% CO2 in air. Sperm concentration was adjusted to 5 × 106cells/ml. After IVF for 18 hours, presumptive zygotes were cultured in modified synthetic oviductal fluid with 1mM glutamine, 12 essential amino acids, 10 μg/ml insulin under 5% CO2, 5% O2 in air. In experiment 1, we examined sperm penetration rate at 12 hours of IVF of frozen-thawed epididymal sperm. Total penetration rate among cauda epididymis and control were not significantly different (mean±standard error, cauda epididymis and control vs. 49.7±11.3 and 54.4±12.8%) In experiment 2, cleavage and blastocyst development rate were evaluated at day 2 and day 8 after IVF for 18 hours. Cleavage rate among cauda epididymis and control was similar different (cauda epididymis and control vs. 81.2±3.4 and 82.7±2.5%). However, blastocyst developmental rate of cauda epididymis group was significantly higher than that of control group (cauda epididymis and control vs. 24.4±1.6 and 12.2±2.8%, p<0.05). In conclusion, cauda epididymal spermatozoa in Hanwoo bull has high fertilizability and embryo development. Cauda epididymal sperm can be used as an alternative to ejaculated frozen sperm in vitro.

In this study, we examined total number, motility and plasma membrane integrity of epididymal spermatozoa from cauda epididymis of bull after preservation at 4ºC. Totally, 23 testicles were castrated from 23 bulls (mean±standard error, age of days = 426.0±7.3, body weight (kg) = 379.7±8.4, scrotal circumference (cm) = 31.0±0.4) at Hanwoo Research Institute, NIAS, and transported to laboratory and preserved on 1, 4 and 6 days at 4 ºC. As control, epididymal spermatozoa recovery from 7 testicles was conducted after transportation to laboratory immediately. In experiment 1, we compared total number of spermatozoa among groups. Total number of spermatozoa from epididymis was not significantly on different preservation day of 0, 1, 4 and 6 which is 1778.0±304.7, 1824.8±343.9, 1228.4±91.7, 1201.8±178.6×106 cells/ml, respectively). In experiment 2, we examined spermatozoa motility and motility parameters (VCL (μm/s), VSL (μm/s), VAP (μm/s), LIN (%)) by computer assisted sperm analysis (SCA, MicroOptic) system. Percentage of motile on 0 and 1 day (88.9±5.2 and 85.8±6.1) was significantly higher than that on 4 and 6 days (32.6±6.5 and 34.3±8.25). Percentage of VCL (μm/s) on 0 and 1 day (93.5±7.6 and 83.0±14.9) was significantly higher than that on 4 and 6 days (36.6±5.1 and 39.5±5.5) (p<0.05). Percentage of VSL (μm/s) on 0 day (28.0±2.1) was significantly higher than that on 1, 4 and 6 days (20.2±3.0, 9.0±2.0 and 8.5±1.6, p<0.05). Percentage of VAP (μm/s) on 0 and 1 days (49.4±3.8 and 41.3±6.6) was significantly higher than that on 4 and 6 days (18.2±3.0 and 19.3±2.8, p<0.05). Percentage of LIN (%) on 0 day (30.7±2.6) was significantly higher than that on 4 and 6 days (23.4±2.7 and 21.1±1.0, p<0.05). Motility of spermatozoa was divided into 4 groups (fast progresive, slow progressive, non-progressive and immotile) based on WHO guideline. Percentage of fast progressive on day at 0 was significantly higher than that on 1, 4 and 6 days (0, 1, 4 and 6 days vs. 19.8±1.9, 10.2±1.1, 2.6±1.0 and 2.3±1.2%, respectively). In conclusion, cauda epididymal spermatozoa should be recovered within one day after preservation at 4 ºC to recover high quality of epididymal spermatozoa in Hanwoo bull

사람과 어류에게 감염증을 유발하는 Edwardsiella sp., Streptococcus sp. 및 Vibrio sp. 의 해양 유해세균에 대한 해조류 추출물의 항균 활성을 조사하였다. 연구에 사용된 4종의 식용 해조류 중에서 감태 MeOH 추출물이 본 연구에서 사용 된 6종의 모든 해양 유해세균에 대해 넓은 범위의 항균 활성을 나타내었다. 감태의 MeOH 추출물의 유기용매 분획 추출물들 중에서, EtOAc 분획 추출물이 가장 높은 항균활성을 나타내었으며 6종의 해양 유해세균에 대하여 128 μg/mL에서 256 μg/mL의 MIC 값을 나타내었다. 또한, HPLC 분석에 의해 감태 EtOAc 분획 추출물에 phlorotannin 화합물인 dieckol이 다량 존재하고 있는 것이 확인 되었다. 결론적으로, 감태 추출물의 phlorotannin 화합물이 여러 유해세균에 대한 강한 항균활성을 나타내는 것처럼, 해양 유해세균에 대해서도 강한 항균활성을 나타내는 것으로 판단된다.

실내 공기질 관리의 중요성이 대두되면서 쾌적한 실내환경에 도움을 주는 공기청정 기능과 습도 조절 기능을 동시에 갖춘 제습기와 공기청정기가 각광받고 있다. 하지만 오랜 기간 동안 공기정화제품을 사용하게 될 시에는 필터가 오염되어 본연의 기능을 상실하게 되는 것으로 알려져 있지만 이에 대한 구체적인 연구나 보고는 드문 편이다. 이에 본 연구에서는 가정과 사무실 등에서 사용한 공기정화제품을 수거하여 주요 부위별 미생물 오염도 및 주요 오염 미생물들을 분석하였다. 그 결과, 4 종류의 공기정화제품에서 오염도가 높은 부위는 필터부위, 물이 직접 닿는 부위 및 공기가 외부로부터 직접적으로 들어오는 입구부위 등에서 미생물학적 오염도가 가장 높았다. 하지만 공기정화제품은 사용하는 환경과 조건에 따라서 미생물학적 오염도 및 오염 미생물의 성상은 각각 다르게 나타났다. 하지만 이들 공기정화제품들에는 공통적으로 Staphylococcus sp., Micrococcus sp. 그리고 Bacillus sp.의 세균과 Cladosporium sp. 및 Penicillium sp.의 진균이 공통적으로 오염되어 있는 우점종인 것으로 분석되었다.

Bacteriocins from lactic acid bacteria have attracted much attention in recent years because of their useful worth in increasing safety and extending shelf life of foods. These substances show an inhibitory effect against some food spoilage bacteria and food-borne pathogens. The inhibitory effect of the bacteriocin produced by lactic acid bacteria against Listeria monocytogenes(L. monocytogenes) was examined in this study. The culture supernatants of 5 kinds of bacteria among the 10 kinds of tested lactic acid bacteria had the inhibitory activity against Listeria sp., various Gram positive and Gram negative bacteria. Bacteriocin produced by Lactobacillus plantarum(Lact. plantarum) LMG 7945 was the most active toward L. monocytogenes. Bacteriocin production of the Lact, plantarum LMG 7945 cultured on MRS broth was increased late logarithmic phase over early stationary phase. This bacteriocin was stable at heat treatment and acidic pH relatively; The activity was retained after heating at 121 for 15min and was active in the pH range of 2-4 but was lost above pH 5.