Cock Semen Research Articles

105 CRYOPRESERVATION OF VENDA COCK SPERMATOZOA: EFFECT OF CRYOPROTECTANT FOLLOWING ANALYSIS BY COMPUTER-ASSISTED SPERM ANALYSIS

The choice of ideal permeable cryoprotectant for cock semen equilibration and freezing is critical. The aim of this study was to compare three different cryoprotectants [dimethyl sulfoxide (DMSO), ethylene glycol (EG), and propanediol (PND)] following cryopreservation. The abdominal massaging technique was used for semen collection from 5 Venda cocks. Individual ejaculates were diluted with modified Kobidil+ (mK+) extender (fraction A) at a ratio of 1:2 and equilibrated for 2 h at 5°C. Semen samples of 5 μL were taken at 0, 60, and 120 min and evaluated by CASA for spermatozoa motility parameters (rapid, medium slow, static, NPM and PM) and velocity. Semen was further diluted at 1:1 volume ratio with mK+ supplemented with 8% DMSO, EG and PND (fraction B), and equilibrated for additional 2h at 5°C and evaluated at 60 and 120 min for motility and velocity. Cooled semen were then transferred into 0.25-mL straws and placed into a programmable freezer. The temperature of the chamber was decreased in a stepwise manner, from 5°C to -20°C, at the rate of 1°C/min until it reached the target temperature. The straws were exposed to liquid nitrogen (LN2) vapor and then plunged into LN2 (-196°C). The semen straws were stored into LN2 tank at -196°C. After 3 months of storage, semen straws were thawed at 5°C and evaluated by CASA for spermatozoa motility and velocity. Data were analyzed by ANOVA. There were no significance differences between DMSO and EG regarding the survival and motility rate of frozen/thawed semen; however, these parameters were lower compared with the fresh semen. The PND was not a suitable cryoprotectant to cryopreserve Venda spermatozoa. In conclusion, ethylene glycol was found to be a suitable cryoprotectant to cryopreserve spermatozoa of South African Venda cocks. Department of Agriculture Forestry and Fisheries, NRF,ARC, Department of Science & Technology.

Osborne Selection Index and Semen Traits Interrelationships In Rhode Island Red and White Breeder Cocks

2 Abstract: Three hundred and sixty-one cocks from five hatches, one week apart, were used in this study. The birds which were made up of 203 birds from strain A (male line) and 158 birds from strain B (female line) were subjected to semen collection using the massage technique. The ejaculates were then subjected to both physical and chemical evaluations for semen quality analysis. The parameters considered were semen volume, semen colour, sperm progressive motility, sperm concentration, total sperm per ejaculate, concentration of live sperm and percent abnormal sperm. Results showed that the mean values for all the parameters lie within the acceptable range reported for normal cock semen. Moderate to high heritability estimates for most of the semen traits were also observed. The least square means (±SE) for semen volume, sperm progressive motility, sperm concentration, total sperm per ejaculate and concentration of live sperm cells obtained in this study were 0.42±0.02 ml, 73.46±2.04%, 1.47±0.15x10 /ml, 64.15±5.67x10 /ml 9 9 and 86.45±2.63%. The heritability estimates obtained were 0.55±0.03 for semen colour, 0.45±0.08 for semen volume, 0.83±0.04 for sperm progressive motility, 0.52±0.06 for sperm concentration, 0.33±0.02 for total sperm count, 0.46±0.03 for concentration of live sperm cells respectively. High and positive genetic correlations between Osborne Selection Index and semen volume, semen colour, sperm concentration as well as with concentration of live spermatozoa were also obtained. The lowest value (0.008±0.010) of phenotypic correlation obtained was for total sperm per ejaculate and the highest value (0.066±0.027) was for semen volume. Therefore the genetic correlation between Osborne Selection Index and most semen traits were positive, hence, selection of males on the index values, currently been practiced in NAPRI, will not bring about any deterioration in semen quality. On the other hand, due to significantly negative genetic correlation between Osborne index and abnormal sperms, it will indirectly improve the semen quality of both lines which in turn may yield better fertility in the Rhode Island flock.

Development of Two Stages Cock's Semen Extender for Room Temperature Storage at Laboratory

A simple cock’s semen dilutor (two-stage) with minimum ingredients was developed in the present study. The stage-1dilutor was aimed to reduce the motility and metabolism during storage and the stage-2 dilutor was to be added just prior to insemination to revive the motility to facilitate the entry of spermatozoa into sperm host gland. Initially, the semen was diluted 1 : 1 with the stage-1 dilutor having lower pH that reduced the motility and metabolism. Subsequently, after 24h, the diluted semen was mixed slowly with another dilutor stage-2 at 1 : 2 ratios to revive the motility of the spermatozoa to normal level. Phosphate buffer was the base for both storage (stage-1) and recovery (stage-2) dilutors with a pH of 6.5 and 7.5, respectively. In stage-1 dilutor sodium glutamate to minimize metabolism of spermatozoa and antibiotics to check bacterial growth were used whereas these were absent in stage-2 dilutor. Glucose was added as energy source in the dilutor of both stages. The osmolarity of stage-1 and stage-2 dilutors was 346 and 327Osm/kg H20, respectively. This dilutor can be used to store the cock’s semen at room temperature (17-21°C) for 24h with sustained motility. This new approach for two-stage dilutor of cock’s semen storage at room temperature for extended period could be a useful A.I. tool under farm condition.

Studies on the Short-Term Preservation of Cock's Semen

The aim of this research was to investigate the effects on sperm motility and semen pH of diluent and preservation temperature during the short-term preservation of cock's semen. During the research, semen was collected from cock's by manual massage method twice a week. The spermatological characteristics of samples which were collected from cocks were assessed and ejaculate volume (ml), sperm motility (%), sperm concentration (X109/ml), the percentage of abnormal sperm (9), the percentage of dead sperm (9) and semen pH were recorded as averaged 0.4±0.1, 85.0±2.8, 3.0±0.2, 5.4±0.7, 9.0±0.5 and 7.0±0.1 respectively. Diluted semen with Ringer, Milk powder, Avian and Physiological salt solution was stored at 5^0C and 18^0C. Sperm motility (%) of semen held at 5^0C after 7 hours was averaged 56.2±2.2, 63.8±2.3, 73.5±1.3, 43.3±2.7 for diluted semen respectively, but the motility of undiluted semen (control group) was found nil at the end of this time. Sperm motility (%) of semen held at 18^0C after 7 hours was recorded as 24.0±3.0, 32.2±4.4, 46.0±3.5 and 36.2±3.1 for diluted semen with the same diluents respectively, but the sperm motility of control group was found nil. pH values of semen held at 5^0C after 7 hours were recorded as averaged 6.0±0.0, 7.0±0.1, 6.9±0.1, 7.3±0.1 for diluted semen with Ringer, Milk powder, Avian and Physiological salt solution respectively and 8.1±0.1 for undiluted semen. pH values of semen held at 18^0C after 7 hours were found average 6.1±0.1, 6.8±0.1, 6.9±0.1, 7.5±0.1 for diluted semen with the same diluents respectively and 8.1±0.2 for undiluted semen. With this study, it was concluded that Avian diluent was superior to the other diluents.

Effects of Two Different Extenders on the Storage of Cock Semen at +5¼C

Total 12 cocks kept in individual cages were used in this study. Two times a week, total 24 ejaculates were collected from each cock and semen was pooled for spermatological evaluations. Spermatological tests were semen volume, mass activity, spermatozoa motility, spermatozoa concentration, live-dead spermatozoa rate and abnormal spermatozoa rate. The values of these characters were 3.29 ± 1.19 ml, 3.29 ± 0.62 (+), 85.83 ± 6.19 %, 4.34 ± 1.69 x 109/ml, 83.34 ± 6.43 %, 11.83 ± 0.96 % respectively. After these tests semen samples were divided into two equal parts and each was extended with 2 different extenders [Extender A= 80% milk base (90 parts skimm milk + 10 parts egg albumen) + 20% fructose solution (5% w/v) and, Extender B= 80% milk base + 10% fructose solution (5% w/v) + 10% sodium sulphate solution (2% w/v)]. The temperature of semen samples were decreased to +5^0C and were kept in refrigerator. Motility, live-dead spermatozoa ratio and morphological evaluations of the semens were repeated immediately when the temperature reached +5^0C (0. hour), at 6th, 24th, 48th and 96th hours at +5^0C. The evaluation of the results and statistics showed that extender B caused a decrease in motility from 85.83 ± 6.19% to 76.25 ± 10.34% (p In conclusion we can say that decrease of spermatological qualities of the semen extended and kept at +5^0C were unimportant at 0. hour. Extender B was the more suitable extender to dilute and keep the semen at +5^0C for short time use.

Correlations between the activities of semen acid phosphatase and Ca +-dependent ATPase and age in different breeds of cocks

1. 1. The present work aims to find a biochemical criterion for evaluating the evolution of sperm according to age through the study of the ATPase activity from the spermatozoa and the acid phosphatase from the seminal plasma of cocks from three different breeds. 2. 2. The optimal parameters of action of the cock semen acid phosphatase and the Ca 2+-dependent ATPase from the spermatozoa were studied. 3. 3. The substrate specificity of the semen acid phosphatase and its inhibition by tartrate, fluoride, metavanadate, molybdate and Hg 2+ were also studied. 4. 4. The two enzymes were determined from the Sussex, Golden Cornish and Plymouth Rock breeds at different ages. 5. 5. The data lead to the conclusion that some properties of bird spermatozoa are less influenced by breed while the acid phosphatase activity, secreted in the ductus deferens is a breed characteristics.

Effect of Carbon Dioxide on Fertility of Cock Semen Incubated In Vitro at 41 C

The effect of various atmospheric CO2 concentrations on fertility of cock semen incubated in minimum essential medium (MEM) for 6 hr in vitro at 41 C was evaluated. Fertility following intravaginal insemination was 54, 87, and 78% for semen incubated in .03, 10, and 15% by volume CO2, respectively. No differences in hatchability of fertile eggs were observed between the semen treatments. Oxygen consumption values monitored at the beginning and end of the holding period were similar for semen incubated in 10% atmospheric CO2 but decreased for semen incubated in .03 and 15% CO2.

Comparison of Diluents for Holding Cock Semen Six Hours at 41 C

Beltsville Poultry Semen Extender (BPSE) and Lake's Diluent A (LDA) were compared with minimum essential medium (MEM) for their ability to maintain the fertilizing capacity of cock semen held 6 hr at 41 C. Motility significantly declined from the beginning to the end of the holding period for semen in BPSE and LDA. Only in LDA, however, were the number of live spermatozoa significantly reduced. Although there were no differences in oxygen (O2) consumption measured at 1 and 6 hr for semen in BPSE and MEM, a significant reduction in O2 consumption was observed between these time periods for semen in LDA. Fertility of semen held in MEM (90.3%) was significantly higher than the unstored control semen (82.9%) and semen held in either BPSE (3.5%) or LDA (1.9%). No differences in hatchability of fertile eggs were observed between the semen groups.

Preservation of the Fertilizing Capacity of Cock Semen Incubated In Vitro at 41 C

Cock spermatozoa were incubated for 6 and 24 hr at 41 C with minimum essential medium (MEM), tissue culture fluid (TCF) removed from tissue cultured cells and in the presence of tissue cultured cells (TCC). The TCC were obtained from whole 9-day-old chick embryos. Fertility following intravaginal artificial insemination was 75, 77, and 80% for semen incubated at 41 C for 6 hr in MEM, TCF, and TCC, respectively. Fertility of semen incubated for 24 hr was approximately 50% for all three treatment groups. The MEM alone was as beneficial as TCF or TCC in maintaining in vitro sperm survival. These observations provide a method for maintaining in vitro the fertilizing capacity (as evaluated by intravaginal insemination) of large numbers of cock spermatozoa under conditions of temperature comparable to those of the hen's oviduct.

Survey of the Methods for Long Term Storage of Poultry Eggs

Mostly fertile eggs which were collected from White Leghorn laying hens inseminated artificially with 0.02ml cock semen per hen and infertile eggs which were collected from another flock of White Leghorn laying hens were analyzed for moisture, crude protein, crude fat and crude ash concentration. No difference was observed in the compositions in egg white and yolk between the two groups except the case of crude protein in egg white in which the concentration was higher in the infertile eggs. No difference was observed between the two groups in the preservation of interior quality of the eggs stored for 1, 5, 10, 20, 30, 40, 60, 80, and 100 days either in an unairconditioned room or in a box kept at 25°C.

The Influence of Short Term Exposure to Two Different Environmental Temperatures on Electrolyte Concentrations of Fowl Semen

The concentration of some of the electrolytes found in fowl semen changed when birds were transferred from a temperature environment of 19° C. to either 8° or 30° C.The pH of the semen obtained from males transferred to the 30° C. environment significantly decreased as opposed to a significant increase in the pH of the semen obtained from males transferred to the 8° C. environment. Sodium and potassium concentrations significantly increased in whole semen in both transfer groups, while magnesium increased significantly only in whole semen from males transferred to the 8° C. environment.Sodium increased in seminal plasma when the males were transferred from the 19° C. environment to the 8° C. environment. The data indicate that an abrupt change to either a warmer or colder environment causes a change in electrolyte concentration of cock semen.

Effect of Some Cryophylactic Agents on Turkey Spermatozoa

INTRODUCTIONTHE potential advantages of long-term preservation of animal germ cells by freezing have long been recognized. A cursory survey of the literature shows that there is a great deal of species specificity in effect of various treatments on spermatozoa survival. Glycerol appears to be the most useful cryophylactic agent investigated to date, but unfortunately the problem of species specificity precludes using the technique for turkey semen that was so successfully used in freezing bovine semen.Some advances have been made in the technology of frozen cock semen. Dialysis to remove glycerol found necessary by Polge (1951) to restore fertilizing capacity, was found not to be necessary if intrauterine artificial insemination was employed with glycerolated semen either fresh (Allen and Bohr, 1955) or frozen, (Allen, 1958) but the frozen semen inseminated produced no better fertility (25%) than dialyzed semen inseminated intravaginally. Washing with centrifugation, followed by intravaginal insemination produced at…

Effect of Osmotic Pressure of Semen Extenders on the Fertility and Hatchability of Turkey Eggs

A VERY interesting development in the theory of semen diluent formulation was suggested by Wales and White (1961) who observed that hypertonicity of the medium reduced the detrimental effects of high dilution. Yamane et al. (1962) claimed that seminal plasma is not isotonic with the spermatozoan midpiece. Although the osmotic pressure of the seminal plasma approximates 323 m Osm, he claims the midpiece suffers damage from plasmolysis when the osmotic pressure is less than about 500 m Osm. This damage results in a significant increase in bent midpieces (with sperm head lying back along the tail). Yamane et al. (1962) states that this abnormality is the result of hypotonicity of seminal plasma, and, since the tonicity of diluents has been adjusted to that of seminal plasma, this means the diluents are also hypotonic to the midpiece. He stored cock semen in a yolk-citrate glucose solution with an O.P. of 550…

Differentiation of Live from Dead Spermatozoa in Cock Semen

THE laboratory evaluation of semen quality often involves the determination of the percentage of the spermatozoa which are dead. Opal blue and eosin (Lasley, 1944; Emik and Sidwell, 1947; Warren et al., 1952), erythrosin (Warren et al., 1952), nigrosin and eosin (Cooper and Rowell, 1958; Chatterjee et al., 1967), bromphenol blue and nigrosin (Kamar, 1959; Chatterjee et al., 1967), congo red and nigrosin (Chatterjee et al., 1967), and aniline blue and eosin (Shaffer and Almquist, 1948) stains have been employed to differentiate live and dead spermatozoa. Results have been variable and the techniques are not well adapted for the rapid evaluation of large numbers of samples. A detailed review of these and related techniques has been given by Salisbury and Van Demark (1961).In preliminary studies with various stains and staining techniques, trypan blue wet mounts appeared to give good results in live-dead spermatozoa differentiation. The technique was simple, fast,…

Serological Studies on the Artificial Insemination of Domestic Fowl

These experiments were undertaken to clarify the effect of isoimmune and heteroimmune sperm agglutinin on the respiration and the mobility of cock spermatozoa.1. Fresh heteroimmune rabbit serum against cock spermatozoa has more harmful effect on the respiration and mobility of cock spermatozoa than isoimmune serum has.2. Inactivation (heating at 56°C for 30mins.) of immune sera did not decrease their sperm agglutinating ability, but decreased their harmful effect on the respiration of the spermatozoa.3. When inactivated immune sera were used, the respiration of the sperm decreased only when antisera of high titer were added to the semen. However, when the antisera of low titer were added, the respiration increased.4. The result of the preservation test of cock semen adding the sperm agglutinin showed the increased mortality of sperm within 2 days, in comparison with the case of adding the normal hen serum. Inactivation of either immune or normal sera showed a good effect for the preservation of cock semen.

Effects of Irradiation on the Metabolism of Spermatozoa

Most of the work done on irradiation of spermatozoa has dealt primarily with their fertilizing capacity and the morphology of the eggs fertilized by the irradiated spermatozoa (1-5). Only a few reports (5-7) are available with respect to radiation effects on the metabolism of spermatozoa, and these reports indicate that the metabolic activity of spermatozoa is highly radioresistant. For instance, Kosin (5) found that in vitro irradiation of cock semen with doses up to 10,000 r did not affect the oxygen uptake or the aerobic and anaerobic glycolysis of spermatozoa, whereas their fertilizing capacity was impaired at 231 r. Furthermore, Mann (6) mentioned that ram semen irradiated at 100,000 r and examined immediately after exposure had normal motility, fructolysis, and adenosine triphosphate content. Wu and Prince (7) have reported that ionizing radiation as high as 320,000 r did not affect the oxygen uptake of bull spermatozoa when respiratory measurements were taken 3 hours after irradiation. The present work is an attempt to throw some light on the role played by ionizing radiation on certain metabolic aspects of bull spermatozoa. A preliminary account of the present investigation has already been reported (8).

Assessment of Cock Semen Quality by Determination of the Change in its Initial pH over a Given Period of Time

METHODS for assessing the fertilizing capacity of fowl semen have been limited. Spermatozoan motility is used widely as an indicator of sperm quality in large animals and man. Shaffner and Andrews (1957) studying fowl sperm motility, found significant correlations between the latter and the fertilizing capacity of the semen. In recent large scale fertility studies on commercial breeding farms practicing artificial insemination, the authors have found highly significant correlation between sperm motility and the number of fertilized eggs (unpublished data).On the other hand, it has to be admitted that the assessment of semen motility, yields results which are only partially accurate, as they are based on subjective estimation, and data recorded by different workers can seldom be reliably compared.Other methods for the evaluation of semen quality (Nigrosin-eosin staining technique, resazurin or methylene blue reduction time, etc.) are successfully used in laboratory studies and as routine procedures in many…

METABOLISM OF TESTOSTERONE BY SEMEN AND THE EFFECT OF TESTOSTERONE ON OXIDATIVE METABOLISM OF SPERMATOZOA.

SUMMARYWhen testosterone‐4‐C14 was incubated with ram, bull, dog, and cock semen, 6–11 p.c. was converted to androstenedione‐C14. Testosterone could not be shown to affect oxidative metabolism of ram, bull, or dog spermatozoa.

Effect of chloride ions on cock spermatozoa.

The respiration rate, rate of fructolysis and incidence of abnormal spermatozoa were studied in cock semen immediately after dilution (1:10) and after storage at 5°C. Tyrode's solution was the basic diluent, and the effect of replacement of chloride by either glutamate or phosphate was investigated. Replacement by glutamate resulted in a linear reduction of respiration and fructolysis rates of fresh semen and of the fructolysis rate of stored semen. Respiration rate of stored semen decreased nonlinearly. Replacement of chloride by phosphate had no significant effect on the metabolic rates of either fresh or stored semen. Chloride ions caused an increase in the incidence of abnormal sperm after storage. When glutamate replaced chloride this increase was reduced, whereas in phosphate replacement there was evidence of a synergistic effect between chloride and phosphate in causing abnormalities. Differences in metabolic rates could not be accounted for by changes in the incidence of dead or abnormal sperm.

Some Effects of Diluting Cock Semen

THE term “dilution effect” has been used frequently in the literature but no consistent definition has been adopted. Many writers have used it to denote adverse effects which dilution has on sperm survival, but Bishop (1954), working with bovine spermatozoa, defined it in a wider sense to denote any changes in sperm physiology which result from dilution. Effects of dilution have been discussed in reviews by Emmens and Blackshaw (1956) and Austin and Bishop (1957). Primary effects include some, and possibly all, of the following: decrease in sperm density, “mechanical” damage during the process of mixing, introduction of substances toxic to spermatozoa and dilution of nutritive substances necessary for sperm survival; metabolic changes especially in oxygen uptake, changes in permeability of the cell wall and dilution of protective substances have also been observed. Some, if not all of these effects, if sufficiently severe, will result in a decrease in fertilising …