38 QUAIL EGG YOLK IN CITRATE EXTENDER IS SUITABLE FOR CRYOPRESERVATION OF NGUNI BULL SEMEN

2016 ◽  
Vol 28 (2) ◽  
pp. 149
Author(s):  
M. M. Seshoka ◽  
M. L. Mphaphathi ◽  
K. S. Mafolo ◽  
M. Nkadimeng ◽  
Z. C. Raphalalani ◽  
...  
Keyword(s):  
Sperm Motility ◽  
Egg Yolk ◽  
Sperm Analysis ◽  
Chicken Egg ◽  
Bull Semen ◽  
Significant Difference ◽  
Computer Aided ◽  
Chicken Egg Yolk ◽  
Fresh Semen ◽  
Motility Rate

Traditionally, commercial hen egg yolk has been used in extenders or freezing media because of its easy availability. However, the use of quail egg yolk has not been used for preserving Nguni bull semen. The aim of the study was to compare the suitability of different quail egg yolk concentrations (5, 10, 15, and 20%) for cryopreserving Nguni bull semen. Semen was collected from 14 stud Nguni bulls with the aid of electro ejaculator. Collected semen samples were kept in a thermos-flask containing warm water at a temperature of 37°C and transported to the laboratory for further analyses. The sperm motility traits were evaluated using computer-aided sperm analysis prior extension. Semen samples were then randomly allocated into 5, 10, 15, and 20% of quail egg yolk and 20% concentration of chicken egg yolk (control) in citrate extender. The extender was supplemented with 12% of glycerol (Seshoka et al. 2012) as a cryoprotectant, and semen samples were diluted (1 : 2) and equilibrated for a period of 4 h at 5°C. After equilibration, semen samples were loaded into 0.25-mL straws, placed into a controlled rate programmable freezer, and stored in a LN tank (–196°C) until thawing. Frozen semen straws were thawed in a water bath at 37°C for 1 min. Thawed semen was evaluated for sperm motility traits using a computer-aided sperm analysis system. Data were analysed with ANOVA. A significant difference was recorded between fresh total sperm motility rate (99%) and frozen-thawed semen samples with either 5% (87.3%) quail or 20% (87.6%) chicken egg yolk citrate extender compared with 10% (92.6%), 15% (91.2%), or 20% (89.9%) quail egg yolk citrate extender. Moreover, fresh semen also resulted in a significantly higher progressive sperm motility rate (39.3%) as compared with frozen-thawed with 5% (26.2%) or 20% quail (28.5%) or 20% chicken (22.7%) egg yolk citrate extender. The results also demonstrated that the use of 10, 15, and 20% quail egg yolk in citrate extender yielded comparable results on total sperm motility with fresh semen as compared with 5% quail and 20% chicken egg yolk. In conclusion, quail egg yolk extender provided sufficient cryo-effectiveness to cryopreservation of Nguni bull semen.

53 COMPARISON OF 4 DIFFERENT CRYOPROTECTANTS ON FREEZING NGUNI BULL SPERMATOZOA EVALUATED BY COMPUTER-AIDED SPERM ANALYSIS

2014 ◽  
Vol 26 (1) ◽  
pp. 140
Author(s):  
M. M. Seshoka ◽  
M. L. Mphaphathi ◽  
T. L. Nedambale
Keyword(s):  
Sperm Motility ◽  
Artificial Insemination ◽  
Egg Yolk ◽  
Cleavage Rate ◽  
Sperm Analysis ◽  
Bull Semen ◽  
Significant Difference ◽  
Computer Aided ◽  
Freezing Curve ◽  
Motility Rate

Cryopreservation has been reported to damage approximately 40 to 50% of viable spermatozoa in bulls. It is critical to evaluate frozen-thawed spermatozoa with computer-aided sperm analysis (CASA) and find a suitable cryoprotectant for Nguni semen. The study was conducted to compare cryo-effectiveness of glycerol (GLY), dimethyl sulfoxide (DMSO), propanediol (PND), and ethylene glycol (EG) cryoprotectants at 12% concentrations during freezing of Nguni bull semen. Semen was collected from 18 stud Nguni bulls of proven fertility with the use of an electro-ejaculator. Collected semen samples were kept in a thermo flask (37°C) and transported to the laboratories. Semen samples were pooled and sperm motility rate was evaluated using CASA. Semen was then diluted (1 : 2; vol : vol) with egg-yolk citrate extender supplemented with either 12% GLY, DMSO, PND, or EG. Semen samples were equilibrated for 4 h at 5°C. After equilibration, samples were loaded into 0.25-mL straws and placed into the controlled rate programmable freezer using a customized freezing curve (from 5 to –5°C at 0.008°C min–1 and from –3 to – 130°C at 6°C min–1). Following thawing of semen, artificial insemination was conducted on 104 oestrus-synchronised Nguni cows. The IVF was also conducted on 120 oocytes to check the cleavage rate. Data were analysed using ANOVA. There was a significant difference (P < 0.05) between raw total sperm motility (94.70 ± 2.63) and frozen-thawed sperm total motility with GLY (77.80 ± 11.03), EG (20.35 ± 11.86), DMSO (15.68 ± 10.14), and PND (11.19 ± 11.27) groups. The pregnancy rate following artificial insemination was 75.9% and a total of 86.6% oocytes had cleaved after fertilization with frozen (12% GLY)/thawed semen. In conclusion, cryopreservation process reduced sperm motility and velocity rates, regardless of cryoprotectant. Egg-yolk citrate extender supplemented with 12% glycerol had recorded the highest post-thaw sperm motility rate.

scholarly journals Comparative study of chicken egg yolk and quail egg yolk in two chilled canine semen extenders

2020 ◽  
Vol 17 (4) ◽  
pp. 62-69
Author(s):  
S.A. Ubah ◽  
M. Sule ◽  
I.C. Chibuogwu ◽  
P.K. Columbus ◽  
K.O. Abah ◽  
...  
Keyword(s):  
Sperm Motility ◽  
Semen Quality ◽  
Egg Yolk ◽  
Chicken Egg ◽  
Significant Difference ◽  
Skimmed Milk ◽  
Mass Activity ◽  
The Difference ◽  
Chicken Egg Yolk

The aim of this work was to substitute chicken egg yolk with quail egg yolk in two semen extenders and to evaluate the quality of the extended canine semen following chilled storage. Semen was pooled from male dogs (n= 4) of about 18-months old and body weight of about 28 kg. Four extenders were tested: (1) tris buffered chicken egg yolk extender (2) tris buffered quail egg yolk extender, (3) skimmed milk chicken egg yolk extender and (4) skimmed milk quail egg yolk extender. Semen was diluted with corresponding extender in the ratio 1:4. The diluted semen samples were analyzed for motility, mass activity, viability, abnormalities percentage and pH for three consecutive days. There was no significant difference (P>0.05) between chicken egg yolk and quail egg yolk in either tris diluent or skimmed milk extender with respect to pH, mass activity and sperm motility. Samples stored in both the tris and skimmed milk-based extenders with quail egg yolk displayed greater viability than those in chicken egg yolk but the difference was not significant (P>0.05). Viability, mass activity and sperm motility decreased as treatment days increased in both chicken and quail egg yolk extenders. Results showed that a pH of 6.5 was maintained from day 0 to day 3. There was no difference in semen quality between chicken and quail egg yolk in either the tris diluent or skimmed milk extender (P> 0.05). It was recommended that quail egg yolk could be substituted for chicken egg yolk in the two canine semen extenders. Further modifications of the diluents with quail egg yolk might produce an improved result. Keywords: Canine, Chicken Chilled, Egg yolk, Extenders, Quail, Semen

76 THE EFFECT OF GLYCEROL CONCENTRATIONS IN EGG-YOLK CITRATE EXTENDER ON THE QUALITY OF CRYOPRESERVED NGUNI BULL SEMEN

2013 ◽  
Vol 25 (1) ◽  
pp. 185
Author(s):  
M. M. Seshoka ◽  
M. L. Mphaphathi ◽  
F. V. Ramukhithi ◽  
T. R. Netshirovha ◽  
C. Hlungwani ◽  
...  
Keyword(s):  
Sperm Motility ◽  
Rural Areas ◽  
Egg Yolk ◽  
Positive Control ◽  
Negative Control ◽  
Glycerol Concentration ◽  
Bull Semen ◽  
Frozen Semen ◽  
Thawing Process ◽  
Motility Rate

There are bull shortages in South African poor rural areas. Artificial-insemination technology could play a significant role on breeding emerging farmer’s cattle. The objective of this study was to compare glycerol concentrations (0, 4, 8, or 12%) during freezing of Nguni bull semen to conduct AI in different villages. Semen was collected by electro ejaculator from 2 Nguni bulls of known and proven fertility. Collected semen samples were kept in a thermo flask (37°C) and transported to the laboratories within 10 min after collections. Semen samples were pooled and evaluated by Sperm Class Analyser® and allocated randomly per treatment. Semen was then diluted (1 : 2 v:v) with egg-yolk citrate extender supplemented with either 0% (negative control), 4, 8, and 12% of glycerol concentration or AndroMed® (positive control). Semen samples were equilibrated for 4 h at 5°C. After equilibration period, samples were loaded into 0.5-mL straws and placed horizontally into the controlled rate (–5, –8, –10, –12, –15, –25, –35°C min–1) from 5°C until target temperature of –80°C is reached. The frozen semen straws were stored in a liquid nitrogen tank (–196°C) until thawing. Treatment means were separated using Fisher’s protected t-test least, and data are presented as mean ± SD. There was a significant differences (P < 0.05) between raw total sperm motility (83.3 ± 19.3) and frozen–thawed sperm with either 0% (0.0 ± 0.0), 4% (30.2 ± 13.4), 8% (47.9 ± 12.5), or 12% (61.5 ± 4.7) of glycerol and on AndroMed® (27.7 ± 17.8) group. Regardless of the glycerol concentrations used, the freezing-thawing process reduced (P < 0.05) the Nguni total sperm motility rate compared to uncryopreserved sperm (83.3 ± 19.3). In conclusion, egg-yolk citrate extender supplemented with 12% glycerol yielded a better (P < 0.05) total sperm motility rate (61.5 ± 4.7) as compared with the 0% (0.0 ± 0.0), 4% (30.2 ± 13.4), 8% (47.9 ± 12.5), and AndroMed® (27.7 ± 17.8) group. Further studies are required to test other levels of glycerol concentrations (>12%) on freezing Nguni semen and conducting AI.

163 EVALUATION OF VIABILITY OF BULL SEMEN COLLECTED BY ELECTRO-EJACULATION USING COMMERCIAL SEMEN EXTENDER AND 2 CULTURE MEDIA AT CONTROLLED ROOM TEMPERATURE

2017 ◽  
Vol 29 (1) ◽  
pp. 190
Author(s):  
A. M. Raseona ◽  
O. A. Ajao ◽  
L. D. Nethengwe ◽  
L. R. Madzhie ◽  
T. L. Nedambale ◽  
...  
Keyword(s):  
Sperm Motility ◽  
Assisted Reproductive Technologies ◽  
Culture Media ◽  
Reproductive Technologies ◽  
Sperm Viability ◽  
Progressive Motility ◽  
Bull Semen ◽  
Significant Difference ◽  
Semen Extender ◽  
Motility Rate

Preservation of semen is an important process to ensure that semen quality is sufficient for assisted reproductive technologies. The aim of this study was to evaluate the viability of bull semen collected by electro-ejaculation using commercial semen extender and 2 modified culture media stored at controlled RT (24°C) for 72 h. Two Nguni bulls were used for semen collection; after collection, the semen was evaluated macroscopically for volume, pH, and colour, and microscopically for sperm motility, viability, and morphology. Uncontaminated semen samples with progressive motility >70% and morphological defects <20% were pooled after collection before being aliquoted into 3 extenders, namely Triladyl, modified Ham’s F10, and TCM-199 culture media, at a dilution ratio of 1:4 and then stored at controlled RT (24°C). Sperm motility rate was analysed using the computer-aided sperm analyser after 0, 24, 48, and 72 h of storage. Sperm morphology and viability was performed after staining the sperm cells with spermac and nigrosine-eosin stain, respectively. The study was replicated 4 times and data were analysed using ANOVA. Triladyl had a higher sperm viability rate (41.3%) and total motility rate (96.3%) for 72 h (P < 0.01) compared with the 2 modified culture media, Ham’s F10 (26.5 and 86.8%) and TCM-199 (25.0 and 86.7%), respectively. However, Ham’s F10 had higher progressive motility rate (37.8%) as compared with the other extenders, TCM-199 (31.7%) and Triladyl (23.4). There was no significant difference (P > 0.05), in viability rate between Ham’s F10 (26.5%) and TCM-199 (25.0%). No significant difference (P > 0.05) in straight line velocity was observed for the three extenders. Furthermore, no significant difference was observed in total sperm abnormalities, except for reacted acrosomes and absent tails (P > 0.05), between the 2 Nguni bulls. Nguni semen can be preserved in Triladyl or modified Ham’s F10 and TCM-199 culture media stored at 24°C and stay viable for 72 h. Triladyl proved to be the best suitable extender of the 3 extenders, showing higher sperm viability and total motility rate as compared with Ham’s F10 and TCM-199 modified culture media.

scholarly journals PENGARUH PENGENCER KUNING TELUR AYAM DENGAN AIR KELAPA MUDA TERHADAP INTEGRITAS MEMBRAN PLASMA DAN ABNORMALITAS SPERMATOZOA DOMBA SAPUDI PENGARUH PENGENCER KUNING TELUR AYAM DENGAN AIR KELAPA MUDA TERHADAP INTEGRITAS MEMBRAN PLASMA DAN ABNORMALITAS SPERMATOZOA DOMBA SAPUDI

2020 ◽  
Vol 8 (2) ◽  
pp. 139
Author(s):  
Mayrena Ayu Cesaria ◽  
A.T.Soelih Estoepangestie ◽  
Suherni Susilowati ◽  
Tatik Hernawati ◽  
Sri Pantja Madyawati ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Egg Yolk ◽  
Coconut Water ◽  
Good Effect ◽  
Storage Duration ◽  
Significance Level ◽  
Chicken Egg ◽  
Significant Difference ◽  
Chicken Egg Yolk ◽  
And Storage

This study aimed to determine the influence of mixture diluter of chicken egg yolk and young coconut water on the integrity of plasma membrane and abnormalities of Sapudi sheep spermatozoa. The experimental design used in this study was a complete random method. The data was analysed using ANOVA, and if there is a significant difference then proceed with Duncan’s Multiple Range Test ( DMRT ) at significance level of 5%. Based on the result of the research, the influence of mixture diluter of chicken egg yolk and young coconut water was not significantly different ( p > 0,05 ) on the intact plasma membrane of spermatozoa on the first day until the fourth day. Interaction between treatment and storage duration had no effect on spermatozoa abnormality on the first day, second day and the fourth day. It can be concluded that mixture diluter of chicken egg yolk and young coconut water had a good effect on the plasma membrane and spermatozoa abnormalities up to the third day.

165 THE EFFECT OF INDIGENOUS CHICKEN EGG YOLK SOURCES AND TEMPERATURES ON SHORT-TERM PRESERVATION OF SOUTH AFRICAN INDIGENOUS GOAT SEMEN

2017 ◽  
Vol 29 (1) ◽  
pp. 191
Author(s):  
M. A. Bopape ◽  
T. L. Nedambale ◽  
C. M. Pilane ◽  
K. C. Lehloenya
Keyword(s):  
South African ◽  
Sperm Motility ◽  
Egg Yolk ◽  
White Leghorn ◽  
Short Term ◽  
Indigenous Chicken ◽  
Treatment Groups ◽  
Chicken Egg ◽  
Indigenous Goat ◽  
Chicken Egg Yolk

Egg yolk is a common constituent of semen extender and protects sperm against cold shock. Besides its protective characteristic, chicken egg yolk is mostly included in extenders for semen preservation due to its abundant availability. The aim of the study was to compare egg yolk sources from different indigenous chicken egg yolk sources and storage temperatures on short-term preservation of South African indigenous goat semen. Semen was collected from 8 South African indigenous goats with artificial vagina during the breeding season (autumn). From each of the 8 goats, 6 replicates were done. Semen samples were randomly allocated into 4 treatment groups of Tris-based extenders containing 20% of egg yolk from White Leghorn as a control, Ovambo, Potchefstroom Koekoek, and Venda chicken breeds. The extended semen samples were stored at 5 or 25°C for 48 h. Semen samples were evaluated for sperm motility using computer-aided sperm analysis sperm viability and morphology with fluorescence microscope at 0, 3, 24, and 48 h. Data was analysed with ANOVA using Stata® version 12 (StataCorp, College Station, TX, USA) statistical software to test the differences between the treatments. The total, progressive, and rapid sperm motility rates were higher in freshly collected semen (90.6, 42.7, and 33.0%, respectively) compared with treatment groups. Semen extended with Tris without egg yolk had higher total sperm motility rate at both 5°C (48.1; 51.1%) and 25°C (43.3; 53.3%) temperatures for 48 h. Semen extended with egg yolk from different egg yolk sources had no sperm motility from 24 h when stored at 25°C. Semen extended with Tris without egg yolk (48.1%) had higher sperm motility than Ovambo, Potchefstroom Koekoek, Venda, and White Leghorn (3.7, 0, 0, and 0.4%, respectively) when stored at 5°C for 48 h. However, sperm motility declined when storage increased. In conclusion, addition of egg yolk had no effect on preserving goat semen. However, Tris-based extender without addition of egg yolk preserved semen for longer period than semen extended with egg yolk regardless of egg yolk origin or chicken breed.

37 DOUBLE FREEZING AND THAWING OF NGUNI BULL SEMEN

2016 ◽  
Vol 28 (2) ◽  
pp. 148
Author(s):  
M. L. Mphaphathi ◽  
M. M. Seshoka ◽  
T. R. Netshirovha ◽  
Z. C. Raphalalani ◽  
T. C. Chokoe ◽  
...  
Keyword(s):  
Sperm Motility ◽  
Egg Yolk ◽  
Freezing And Thawing ◽  
Sperm Analysis ◽  
Treatment Groups ◽  
Bull Semen ◽  
Before And After ◽  
Repeated Freezing ◽  
Thawing Process ◽  
Marked Decline

Indigenous bulls semen are important for conservation programs. The objectives of this study were to evaluate the effects of repeated freezing and thawing on sperm motility characteristics. Semen was collected from 4 Nguni bulls by means of electro ejaculator and stored in a thermo flask (37°C). Sperm total motility, progressive and nonprogressive motility, and velocity were assessed using computer-aided sperm analysis before and after freezing. Semen was then diluted with egg yolk citrate extender (fraction A), then followed by 12% of glycerol + egg yolk citrate extender (fraction B, Seshoka et al. 2012). Diluted semen samples were equilibrated for 4 h at 5°C. After the equilibration period, samples were loaded into 0.25-mL straws and transferred into a controlled rate programmable freezer. After the target temperature of –130°C was reached, semen straws were stored in a LN tank (–196°C). After 3 months of storage, straws were thawed at 15°C (first and second freezing and thawing followed the same process) for 5 min and further evaluated post-thawed at 0 and 15 min during incubation at 15°C. Treatment means were separated using Fisher’s protected t-test least. No significant differences were recorded between the raw semen total sperm motility percentage (93.2%) and first frozen-thawed at 0 min (82.6%), with the total sperm motility rate recovery of 88.6%. In addition, there was a marked decline recorded in sperm total motility during the first frozen-thawed at 15 min (77.6%), second frozen-thawed at 0 min (31.3%), and second frozen-thawed at 15 min (30.1%; P < 0.05). The sperm curvilinear velocity and average path velocity was reduced following first frozen-thawed (P < 0.05) but remained constant and stable between the treatment groups (P > 0.05). In conclusion, the freezing-thawing process did not reduce the Nguni bull total sperm motility during the first freezing and thawing process, compared with raw semen. However, a drastic decline was recorded during the second freezing-thawing processes, compared with raw semen.

scholarly journals Freezing of rabbit semen after the addition of glycerol and dimethylsulfoxide in the extender

2018 ◽  
Vol 52 (4) ◽  
pp. 299 ◽  
Author(s):  
S. SAMOUILIDIS (Σ. ΣΑΜΟΥΗΛΙΔΗΣ) ◽  
K. SAOULIDIS (Κ. ΣΑΟΥΛΙΔΗΣ) ◽  
A. FOUKOS (Α. ΦΟΥΚΟΣ) ◽  
P. YPSILANTIS (Π. ΥΨΗΛΑΝΤΗΣ) ◽  
A. DEMERTZIS (Α. ΔΕΜΕΡΤΖΗΣ) ◽  
...  
Keyword(s):  
Litter Size ◽  
Sperm Motility ◽  
Egg Yolk ◽  
Tris Buffer ◽  
Cryoprotective Agents ◽  
Group I ◽  
Average Litter Size ◽  
Significant Difference ◽  
Group Ii ◽  
Fresh Semen

In this study, semen from 10 rabbits was frozen after it was diluted in two different extenders, A and B. Extender A consisted of Tris-buffer in which 20% of egg yolk and 3% of glycerol were added, while extender Β had the same composition, plus 3% of dimethylsulfoxide (DMSO). The semen of the buck exhibiting the best post-thaw motility in the two extenders, was used for the insemination of two corresponding groups (I and II) of 20 does each. The average sperm motility of fresh semen (77%) was significantly reduced (P<0.05) after it was frozen in the extender A (47%) and Β (54%). The post-thaw sperm motility in extender Β was significantly higher (P<0.05) than that in extender A. The percentage of the animals that gave birth in group II (60%) was increased compared to that of the animals in group I (50%), but not significantly (P>0.05). Also, no significant difference (P>0.05) was observed between the average litter size of group I (6,8±1,7) and group II (7,0±1,9). Thereafter, it is concluded that freezing rabbit semen in an extender that contains a combination of the cryoprotective agents glycerol and DMSO, in the proportion of 3% and 3%, respectively, results in a significant improvement of post-thaw sperm motility, while it doesn't affect significantly its fertility value neither the size of the litters.

16 COMPUTER-ASSISTED SPERM ANALYSIS SPERM PARAMETERS OF BOS TAURUS CRIOLLOS BULL SEMEN v. THOSE OF OTHER BOS TAURUS BREEDS IN ARGENTINA

2008 ◽  
Vol 20 (1) ◽  
pp. 88
Author(s):  
M. G. Lüssenhoff ◽  
G. Larraburu ◽  
R. Cavia ◽  
A. Garcia Guerra ◽  
G. M. Brogliatti
Keyword(s):  
Sperm Motility ◽  
Sperm Morphology ◽  
Bos Taurus ◽  
Hybrid Vigor ◽  
Computer Assisted ◽  
Sperm Analysis ◽  
Bull Semen ◽  
Significant Difference ◽  
Computer Assisted Sperm Analysis ◽  
Motility Parameters

Criollos is the oldest cattle breed in America and the world at large. The origin dates back to the first cattle brought by C. Columbus. They are well known for their toughness and longevity. Their genetic variability is another advantage to be taken into account in crossbreeding because it ensures high hybrid vigor. The objectives of the present study were to determine computer-assisted sperm analysis (CASA) motility parameters and sperm morphology of Criollos bull semen (brought from Parque Nacional Los Glaciares, Patagonia, Argentina) v. other Bos taurus semen. A total of 29 ejaculates of different adult bulls (Angus and Hereford) and 25 ejaculates of Criollos bulls were evaluated in an artificial insemination center between December 2004 and December 2005. Semen collection was done by electroejaculation (EE) in all breeds. Fresh semen was diluted in a semi-defined semen extender (Andromed; Minitüb, Tiefenbach, Germany) and frozen in an automatic freezer (Digicool, IMV, Paillette Crista, France). Stain morphology was done using eosin/nigrosin, and evaluated in an optic microscope with phase contrast. Parameters of volume, total concentration (CONC), motility %, motile progressive sperm %, velocity average path (VAP, μm s–1), velocity straight line (VSL, μm s–1), curvilinear velocity (VCL, μm s–1), and linearity (LIN, %) were determined by CASA (HTM-ceros 12.1, Berkley, CA, USA). The results obtained were analyzed statistically with a one-way ANOVA test and are summarized in Table 1. Results from the sperm motility analysis indicate that there were no significant differences (P > 0.05) in CONC, motility %, motile progressive %, or VSL between Criollos and other B. taurus bull semen. VAP and VCL rates were significantly higher in Criollos bull semen than in other B. taurus semen, and LIN was lower. Also, there was a significant difference in volume collected between both breeds. Regarding sperm morphology, Criollos bull semen is at the maximum limit acceptable for head defects (detached heads: 15%). These results suggest that Criollos bull semen is not different from other Bos taurus semen; however, it does show different velocity and linearity rates. Table 1. CASA sperm motility parameters for semen from Criollos v. other Bos taurus bulls This research was supported by Centro Genetico Bovino Eolia S.A.

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