Testicular Volume and Daily Sperm Output in Ragusano Donkeys

A careful evaluation of daily sperm output (DSO) and seminal parameters is indicative of the potential fertility of the animal and at the same time is useful for maximization of the reproductive activity. The aim of this article was to evaluate the seminal parameters, testicular volume (TV) calculated by ultrasound measurements, DSO, and any eventual relationship between these in Ragusano donkey jackasses. Eight Ragusano donkey stallions underwent a morphometric evaluation of the testes by ultrasound to calculate the TV, and daily semen collection for 10 consecutive days. TV ranged from 250 to 500 cm³ and no significant differences could be observed between the left and right testes. Only ejaculates from 4 to 10 days were analyzed, given that there was evidence that the sperm extragonadal reserve interfered with the total number of sperm in ejaculates during the early days. DSO in the Ragusano donkey ranged from 7 to 23 × 10⁹ sperms. The seminal parameters showed high values of motility, viability, and low abnormality percentages. Age, TVs, and sperm parameters were not significantly correlated. The correlation between TV and DSO was poor (P = .1) in this study and this may suggest that TV and DSO might simply not be highly correlated in the donkey.     

Sperm morphology in Estonian and Tori breed stallions

The standard procedure for assessing the breeding potential of a stallion includes the parameter total number of spermatozoa classified as morphologically normal. This study investigated sperm morphology of fresh semen in randomly chosen Estonian (E, n = 8) and Tori (T, n = 7) breed stallions with proven fertility. Two ejaculates were examined from each stallion. An aliquot from each ejaculate was fixed in 1 mL formol-saline immediately after collection and examined with phase-contrast microscope at a magnification 1000x for all types of morphological abnormalities. Furthermore smears were prepared and stained according to Williams (carbolfuchsin-eosin) for a more detailed examination of the sperm heads with light microscope at a magnification 1000x. Analysis of variance was applied to the data, and results are presented as LSmeans (+/- SE). One T stallion that had a disturbance in the spermatogenesis and one 22-year-old E stallion were not included in the analyses. The T stallions had on average 57.5 +/- 4.1% and the E-stallions 74.4 +/- 3.8% morphologically normal spermatozoa (p = 0.012). In 4 of 7 T stallions and 7 of 8 E stallions both ejaculates had > 50% morphologically normal spermatozoa. There was a significant difference between breeds in mean percentage of proximal droplets (17.3 +/- 2.7% and 2.9 +/- 2.5% for T and E stallions, respectively; p = 0.003).     

Sperm Production and Sperm Morphology of Swedish Warmblood Stallions

The present study investigated daily sperm output and sperm morphology of fresh semen in eight Swedish Warmblood stallions aged 5–8 years. They were used for artificial insemination, and their fertility during the breeding season of semen collection exceeded 60% per cycle. One ejaculate of semen was collected daily for 10 consecutive days from each stallion. The gel-free volume was measured, and the sperm concentration was assessed with a Bürker chamber. The volume of gel-free fraction was multiplied by the sperm concentration to give the total number of spermatozoa (TSN). Sperm morphology was examined in ejaculates collected on days 2, 5 and 10. An aliquot from each ejaculate was fixed in 1 ml formol–saline immediately after collection and examined under a phase-contrast microscope (magnification 1000×) to assess morphological abnormalities. Furthermore smears were prepared and stained according to Williams (carbolfuchsin–eosin) for a more detailed examination of the sperm heads under a light microscope (magnification 1000×). Analysis of variance was applied to data. Total spermatozoa number decreased progressively during the first 8 days of collection, and daily sperm output (DSO) was calculated as mean TSN of collections on days 8–10, being 6.4 × 10⁹ spermatozoa. The overall percentages of morphologically normal spermatozoa in ejaculates collected on days 2, 5 and 10 were above 70%, being significantly lower in ejaculate 2 (68.6%) compared with ejaculates 5 and 10 (72.9% respectively 75.3%).     

Lateral Branches of the Testicular Artery Affect Testicular Shape in Adult Stallions

The objective of this study was to investigate the arterial patterns of the stallion testis in relation to testicular shape. Two hundred and fifty-one stallion testes were evaluated for the presence of the lateral branches of the testicular artery. Seven specimens had their testicular arteries filled with latex milk, fixed in 70% alcohol, and dissected. Two hundred six specimens (82%) had a single testicular artery and no lateral branches; 39 testes (16%) had one lateral branch of the testicular artery; and six testes (2%) had two lateral branches of the testicular artery each. The lateral branches of the testicular artery obtained from the adult stallions, more than 5 years old, were associated with distinct lateral bulging, giving them a pear-like shape, whereas similar vascular pattern in young colts, less than 1 year old, did not cause similar shape change. Five distinct patterns of the branching of the testicular artery were determined. We concluded that the lateral branches of the testicular artery are present in approximately 20% of stallion testes. This anatomic pattern is associated with a lateral bulge that develops slowly over several years and is associated with a change in testicular shape from an ellipsoid in colts to a pear-like shape in adult stallions.     

Comparison of Apoptotic Cells Between Cryopreserved Ejaculated Sperm and Epididymal Sperm in Stallions

The development of a reliable technique to freeze epididymal semen would provide a unique opportunity to preserve valuable genetic material from unexpectedly lost stallions. The aim of this study was to compare the apoptotic indices of sperm obtained from ejaculate, sperm recently recovered from the epididymides (EP), and sperm recovered from epididymides stored at 5°C for 24 hours (EP-stored). For the first category, two ejaculates from seven stallions were collected and then submitted to cryopreservation using an egg yolk-based extender. One week after the last semen collection, the stallions were submitted to bilateral orchiectomy, and sperm from one of the cauda epididymis was harvested immediately after castration (EP). The remaining testicle was stored in a passive refrigeration container at 5°C for 24 hours before the cauda epididymal sperm was harvested (EP-stored). Sperm harvesting from the epididymis for EP and EP-stored was performed by retrograde flushing of the caudal portion of the epididymis using a skim milk-based extender. The recovered sperm was then cryopreserved using the egg yolk-based extender. Sperm motility parameters were studied by computer-assisted semen analysis, and apoptosis was estimated by measuring caspase activity and membrane phospholipid translocation using epifluorescence microscopy. The samples were evaluated immediately (0 hour) and 8 hours after thawing. At 0 hour, no differences in sperm parameters were observed among the groups, but after 8 hours, significant statistical differences were observed in sperm motility parameters and plasma membrane integrity among the treatment groups. In addition, viable cells with no apoptotic signs were more prevalent in EP and EP-stored, suggesting that epididymal sperm is less sensitive to the cold shock caused by sperm cryopreservation.     

How to Perform and Interpret Testicular Fine Needle Aspiration in Stallions

Although several methods of testicular biopsy have been proposed previously, testicular fine needle aspiration (FNA) has proved to be the simplest, the most rapid, inexpensive, and overall the least invasive technique for obtaining testicular biopsies. Testicular FNA is indicated for fertility investigations in stallions with oligozoospermia or azoospermia. It is also used for differential diagnosis of testicular enlargement. After sedation, the stallion’s testis is punctured to obtain testicular parenchyma samples containing cells mainly from the seminiferous epithelium. The material obtained is used to perform smears which are analyzed for identification and quantification of germ cells and Sertoli cells. The results are based on the presence of the cell types found in the smears and the proportions of Sertoli cells per germ cells. In addition to being a very useful diagnostic tool, testicular FNA is also used for follow-up examinations, as it is minimally invasive.     

Post-thawing Sperm Quality in Chilean Purebred Stallions: Effect of Age and Seasonality

In this work, we investigated the influence of age and seasonality on sperm motility and DNA fragmentation in post-thawing semen from Chilean Purebred Stallions (CPS), a horse breed presenting the oldest genealogy record in South America with an interesting reproductive industry. Despite that semen from aged CPS is frozen all year round, there is a lack of studies characterizing the breed semen freezability in accordance with age and seasonality. Twenty fertile CPS were grouped into the young group, the middle group, and the aged group. Ten ejaculates from each stallion were obtained by using an artificial vagina during summer (December) and winter (July) and directly frozen. Subsequently, the frozen semen was thawed and analyzed by a computer-assisted semen analysis and flow cytometer assessing progressive motility, mean velocity, and DNA fragmentation spermatozoa. Kruskal–Wallis test and Pearson’s correlation were used to determine statistical differences among groups and correlation among variables (P ≤ .05). Both spermatozoa motility traits decreased progressively in accordance with age and seasonality, showing the lowest values in the aged group during winter and the highest values in the young group during summer. Deoxyribonucleic acid fragmentation increased significantly in accordance with age and seasonality being highest in the aged group during winter and lowest in the young group during summer. Post-thawing sperm quality showed a negative correlation with the age of the stallions and a positive correlation with the normal sperm morphology before freezing. These results allow the conclusion that age and seasonality are important factors that need to be considered during the selection of CPS for reproductive programs.     

Phenotypic Correlations of Stride Traits and Body Measurements in Lipizzaner Stallions and Mares

Besides being well known for their use in classical dressage, Lipizzaner horses are today becoming more and more popular for use in driving sport, with Croatia as the largest breeder in the world. To estimate their phenotypic predisposition for driving sport, this research analyzes the correlation between body measurements and traits of length and speed of walk and hand-led trot in 18 stallions (LS) and 53 mares (LM) by using a digital camera. Statistical analysis of the data concerning phenotypic correlations among the indicated traits has revealed the potential of documenting a large number of structural exterior body characteristics and quantitative measurements in Lipizzaner horses, and technical equipment has been found to be useful in research of stride traits. Higher withers height (WH) achieved through breeding was found to be negatively correlated with the number of strides per second during walk and trot, but positively correlated with walk stride length, especially in LS. Higher values for chest girth (CG) and cannon bone circumference (CBC) were found to be positively correlated with speed, as well as with stride length in LS, and negatively correlated with the number of strides per second during walk and trot in both LS and LM. Because for many years the goal of breeding Lipizzaners in Croatia has been to obtain a bigger body format and longer stride, leading to better results in a driving sport, these findings confirm breeding goals in selection work guided toward better stride length and extended stride as the most desirable traits for driving sport.     

The Effects of Oral Altrenogest on Hormonal,Testicular, and Behavioral Profiles of Two-Year-Old Stallions

Altrenogest is frequently administered to young stallions in the equine industry as an off-label use to suppress sexual/aggressive behavior. In this study, 2-yr-old Quarter Horse stallions in early-performance training were administered altrenogest (0.044 mg/kg BW daily) to determine the effects on sexual/aggressive behavior, testicular parameters, and steroid hormone profiles. Horses were randomly assigned to treatment (n = 5) and control (n = 5) groups. The treatment group was administered a daily oral dose of 0.044 mg/kg BW daily for 67 d; the control group was given a daily oral sham dose of corn oil for 67 d. No significant differences were found between treatment groups in BW or body condition scores. Altrenogest had no effect on any behavioral parameters or seminal parameters measured. Averages of total scrotal width (TSW) were lower (P<0.03) for treatment animals at the end of treatment period (d 67). By d 157 mean TSW for both treatment and control stallions were not different. Both were significantly higher (P<0.03) than pretrial values at d 157. Comparisons of testicular weight (d 157) between control and treatment groups were not different. Histological analysis of testicular tissue revealed no significant difference for the average number of spermatids per seminiferous tubule. Altrenogest treatment reduced serum estrogen levels (estradiol 17-β) by d 67 (P<0.02); however, estrogen levels returned to pre-treatment values after a 90-d recovery. Serum testosterone values were not affected by treatment. Further research is needed to determine dose and age effects of altrenogest in the stallion.     

BMY热带肉牛屠宰测定报告

热带牛（BMY）生长快，产肉性能好。在完全放牧条件下。18月龄体重可达300kg以上，18月龄和30月龄牛的平均屠宰率达55.53%，平均净肉率为46.30%。分别比云南黄牛提高18.10和21.92个百分点，优质切块占胴体重的39.30%，且抗蜱耐热，适应性广，不仅能适应炎热的气候，而且在冰天雪地的环境中表现也较好。     

不同品种公羔初情期前睾丸生长及LH水平的差异

本试验测定了18头湖羊公羔及考力代20头公羔90日龄前睾丸生长和血浆LH水平。结果表明,湖羊睾丸的重量、直径在60和90日龄均显著大于考力代(P<0.05)。湖羊血浆LH水平从初生到90日龄均高于考力代。血浆LH水平与睾丸重量、直径均呈正相关。     

The Effects of an Intermittent Regimen of Altrenogest on Behavioral,Hormonal, and Testicular Parameters of Three-Year-Old Stallions

To suppress sexual and aggressive behavior in performance stallions, many trainers administer oral altrenogest in an off-label regimen of short-term, repetitive dosages despite the absence of any research to examine possible side effects. In this study, nine 3-yr-old Quarter-Horse stallions were initially blocked by sire and randomly allotted to two groups, treatment (n = 5) and control (n = 4). The treatment group received a total of three 10-d (d 1 to 10, 21 to 30, and 41 to 50) regimens of 0.088 mg/kg BW Regu-Mate® (Intervet, Millsboro, DE) followed by 10 d of nontreatment (d 11 to 20, 31 to 40, and 51 to 60). Following this 60-d period, there was a 36-d recovery period in which no treatments were administered. No significant changes were found between the groups in BW or body condition score (BCS). Of the testicular parameters analyzed, scrotal width (SW) and gross testicular parameters were unaffected by the altrenogest; however, mean spermatid counts were decreased in the testicular biopsies of the treated animals (P<0.001). Although no treatment differences were found for either serum estradiol or testosterone concentrations, there was a significant time × treatment interaction for estradiol (P<0.002). No significant differences were observed between the groups for any of the sexual and aggressive behavioral parameters analyzed from the standardized teasing tests at any point of the study. Nevertheless, large variations existed between individual stallions. Further research is needed that focuses on dosing and age effects to determine if offlabel uses of altrenogest designed to control behavior are compromising future fertility.     

Trypan Blue/Giemsa Staining to Assess Sperm Membrane Integrity in Salernitano Stallions and its Relationship to Pregnancy Rates

Aim of this study was to test the reliability of Trypan blue/Giemsa staining to evaluate sperm membrane integrity, acrosomal intactness and morphology in stallion to verify whether it could be applied in vitro as useful tool for sperm fertilizing ability. Fertility data on inseminated mares were collected to evaluate the relationship of sperm quality to pregnancy rates. Forty‐one ejaculates were collected from 3 stallions of Salernitano Horse Breed and evaluated for gross appearance, volume, visual motility and membrane integrity with Trypan blue/Giemsa staining and thirty‐five mares were inseminated during the breeding season from April to July. Differences among stallions were found in volume, sperm concentration (p < 0.05) and visual motility (p < 0.01). A decrease in sperm motility, concentration (p < 0.05) and total sperm number was found in June–July (p < 0.01). Live sperm with intact acrosome (LSIA) and proximal droplets (PD) were lower (p < 0.01) in June–July, while acrosome reacted sperm (ARS) percentage increased (p < 0.05). No fertility differences were found among stallions with an average fertility per cycle of 44.6% and a pregnancy rate of 68.6%. Higher percentages of LSIA were found in the ejaculates used to inseminate mares that became pregnant vs those used in mares not pregnant (p < 0.05). The significance of LSIA as test variable to verify the reliability of Trypan blue/Giemsa staining was confirmed by Receiver operating characteristic ROC analysis and the sensitivity of the test was 85% at a cut‐off value of 48% LSIA. Trypan blue‐Giemsa showed to be an accurate method that can be applied on field to evaluate sperm membrane integrity and to identify poor‐quality ejaculates.     

Cryopreservation of Piau‐Breed Wild Boar Sperm: Assessment of Cooling Curves and Centrifugation Regimes

This study aimed to assess the effects of different cooling curves and centrifugation regimes used in cryopreservation protocols on the post‐thaw viability of Piau‐breed wild boar (Sus scrofa) sperm using in vitro assessment tests. Two centrifugations (800  g for 10 min and 2400  g for 3 min) and two cooling curves (conventional cooling using nitrogen vapour – freezing 1 and automated cooling using a programmed freezing machine – freezing 2) were tested. Therefore, the treatments were divided into M3 – centrifugation at 2400  g for 3 min and freezing 2; M10 – centrifugation at 800  g for 10 min and freezing 2; R3 – centrifugation at 2400  g for 3 min and freezing 1; and R10 – centrifugation at 800  g for 10 min and freezing 1. No significant differences (p > 0.05) between treatments occurred post‐thawing regarding the total sperm motility means recorded. The mean values of the different treatments were not different from each other regarding the supravital staining (SV), hypo‐osmotic test (HO), sperm–egg binding assay or sperm morphology. This study showed that both the cooling curve and the centrifugation regime affected the quality of post‐thaw sperm, and centrifugation for shorter times and cooling curves using automated cooling are the most suitable for minimizing sperm injury.