M.C. Garcia

Effects of semen fractionation and dilution ratio on equine spermatozoal motility parameters

Two experiments were conducted to examine the effects of semen fractionation and dilution ratio on motility parameters of stallion spermatozoa. In Experiment 1, three ejaculates from each of three stallions were divided into sperm-rich (SR) and sperm-poor (SP) fractions to determine the difference in sperm concentration. Mean sperm concentration in SR fractions (349.5 x 10(6)/ml) was greater (P < 0.001) than that of SP fractions (96.9 x 10(6)/ml). In Experiment 2, three ejaculates from each of two stallions were divided into SR and SP fractions. Fifty percent of the original volume of SR fractions was combined with 50% of the original volume of SP fractions for each ejaculate to represent total ejaculates. SR and total ejaculates were diluted with skim milk-glucose semen extender as follows: 1) no dilution, or dilution to 2) 100 x 10(6)sperm/ml, 3) 50 x 10(6)sperm/ml, or 4) 25 x 10(6)sperm/ml. Semen samples were evaluated at 0.5, 3, 6, 12, and 24 h postejaculation (25 degrees C storage temperature) for percentages of total spermatozoal motility (TSM) and progressive spermatozoal motility (PSM). Mean TSM was greater (P < 0.05) in SR ejaculates than total ejaculates at 12 and 24 h postejaculation. Mean TSM of undiluted semen was lower (P < 0.05) than other dilution ratios over all periods. Mean TSM was greater (P < 0.05) at a 25 x 10(6)sperm/ml dilution ratio than a 50 x 10(6)sperm/ml dilution ratio at 12 and 24 h postejaculation, and greater (P < 0.05) than a 100 x 10(6)sperm/ml dilution ratio from 3 to 24 h postejaculation. Similar patterns were found for PSM. Collection of SR ejaculates and dilution to 25 x 10(6)sperm/ml improved longevity of spermatozoal motility.

Conditioned Suppression of Sexual Behavior in Stallions and Reversal With Diazepam

Sexual behavior dysfunction unaccompanied by detectable physical or endocrine abnormality is an important cause of reproductive failure among domestic stallions. Several authors have suggested that such dysfunction may be psychogenic, related to negative experience associated with intense handling and training. An experimental model of experience-related dysfunction was developed by exposing pony stallions to erection-contingent aversive conditioning. This resulted in rapid, specific suppression of sexual arousal and response similar to spontaneously occurring dysfunction. Subsequently, treatment with a CNS-active benzodiazepine derivative (diazepam) reversed these effects.

Comparison of two techniques for obtaining endometrial bacteriologic cultures in the mare

The endometria of 39 mares were cultured simultaneously using a swab guarded with a double cannula and distal, teflon plug and an unguarded swab with a single, open cannula. Sheep blood (5%) agar, Mac-Conkeys agar, and Sabourads agar media were innoculated with each swab. The presence of bacterial or fungal growth was determined after 24 and 48 hours of aerobic incubation at 37 C. There were significantly more plates that failed to yield bacterial or fungal growth when streaked with swab specimens obtained with the guarded cannula than when streaked with those obtained with the unguarded cannula. It was concluded that while culturing the endometrium of mares for bacteria or fungi, the use of a guarded instrument consisting of a double cannula with a closed end will result in the recovery of fewer contaminants; therefore, it will be more likely to result in a more accurate representation of uterine bacterial and fungal flora.

Gonadotropin Releasing Hormone (GnRH) Affects Precopulatory Behavior in Testosterone- Treated Geldings

Twelve pony geldings with (n = 6) and without (n = 6) testosterone replacement (200 micrograms/kg testosterone propionate in oil, SC every 48 hours) received either gonadotropin releasing hormone (GnRH; 25 micrograms SC every 3 hours) or control treatment. Sexual behavior was recorded during 4-minute exposure to an estrous mare, 3 times weekly for 2 weeks before treatment, 3 weeks during treatment, and 3 weeks after treatment had been discontinued. The group receiving testosterone and GnRH (n = 3) exhibited significantly greater flehmen response frequency and attention duration and significantly lower vocalization frequency and erection duration than the group receiving only testosterone (n = 3). GnRH had no apparent effect on sexual behavior in geldings without testosterone replacement (n = 3). These results support an hypothesis of testosterone-dependent, CNS-mediated effects of GnRH on precopulatory behavior in the stallion.

Comparison of pregnancy rates achieved with frozen semen using two packaging methods

The pregnancy rate achieved with frozen semen using two packaging methods was compared in two separate experiments. The sperm-rich fraction of the semen was collected and immediately extended in an egg-yolk based extender to a final concentration of 100 x 10(6) sperm/ml. Half of each ejaculate was packaged in 4.0-ml plastic straws and the other half in 10-to 12-ml flattened aluminum tubes. After detection of a 35-mm follicle, mares were inseminated daily until ovulation. Pregnancy was detected, and embryonic vesicle size was measured by ultrasonic echography Days 14 to 26 after ovulation. The one-cycle pregnancy rate achieved using both packaging methods was 50% overall (23 46 ), with 46% (12 26 ) and 55% (11 20 ) rates achieved, respectively, using straws and aluminum tubes. These rates were not significantly different.

Investigation of the representativeness of a single endometrial sample and the use of trichrome staining to aid in the detection of endometrial fibrosis in the mare

The uteri of five mares were removed and endometrial samples were procured from 12 specific locations in the uteri and the samples were processed and duplicate sections were stained with hematoxylin and eosin (H&E) or Massons trichrome stains. The samples were interpreted in a blind manner by one person, and pathologic changes were classified according to Kenney (1). An assessment of stromal fibrous connective tissue and focal periglandular fibrosis or fibrotic nests was made. There were no significant differences in luminal epithelial cell heights or in the occurrence and severity of stromal fibrous connective tissue, focal periglandular fibrosis, or lymphatic lacunae among locations (P > 0.05). There was an effect of location on the occurrence and severity of inflammation (P < 0.05). If only inflammation was considered in categorization, this would have resulted in changing the category in 9 of 60 samples. There was no increase in tendency for inflammation, fibrosis or lymphatic lacunae to occur in the horns versus the body of the uterus, nor of the dorsal versus the ventral uterus (P > 0.05). There was no effect (P > 0.05) of type of stain on the ability to detect incidence and severity of focal periglandular fibrosis. There was an effect (P < 0.05) of type of stain on the ability to detect the incidence and severity of stromal fibrous connective tissue. The use of the trichrome stain showed slightly increased distribution of stromal fibrous connective tissue.

Influence of dystocia on white blood cell and blood neutrophil counts in mares.

A retrospective study was done on total white blood cell (WBC) and blood neutrophil counts of 41 mares referred to one of two veterinary hospitals for correction of dystocia. The mares were 2 to 19 years of age and included draft, light, and pony breeds. The WBC and neutrophil counts were performed at varying intervals from time of admission to 10 d after delivery of the feti. Retrospective analyses of WBC and neutrophil counts from 10 normal foaling mares from two Pennsylvania breeding farms (Thoroughbred and Trakehner) and from 14 normal foaling pony mares were done as controls. Mean WBC (10446 +/- 2296 cells/mul) and neutrophil (6850 +/- 2136 cells/mul) counts on the day of delivery in mares with normal parturition were slightly elevated over values reported as normal in the literature. The mean blood cell counts gradually declined to 6124 +/- 1255 WBC/mul and 3692 +/- 409 neutrophils/mul on Day 2 postpartum and returned to normal baseline values by Day 3 postpartum (8868 +/- 2693 WBC/mul, 4298 +/- 1966 neutrophils/mul). No toxic neutrophils were present in mares with normal delivery. Mean WBC (11346 +/- 3298 cells/mul) was elevated on the day of delivery in mares with dystocia as a result of neutrophilia with a left shift (9297 +/- 3298 neutrophils/mul). An apparently faster decline occurred in WBC and neutrophil counts of mares with dystocia than in mares with normal delivery, until a marked leukopenia (3905 +/- 1292 WBC/mul) and neutropenia (1570 +/- 1340 neutrophils/mul) occurred on Day 3 postpartum. The leukopenia and neutropenia persisted until Day 5 postpartum. Toxic neutrophils were present in several mares with dystocia.

Comparison between two techniques for endometrial swab culture and between biopsy and culture in barren mares

The endometria of 77 barren mares was swabbed simultaneously using a swab guarded with a single cannula and distal, gelatin capsule (completely guarded swab - CGS) and a partially guarded swab (PGS) with an open cannula. Sheep blood (5%) agar plates were inoculated with each swab, while MacConkeys agar plates were inoculated with the swabs from 44 mares. The presence of bacterial or fungal growth was determined after 24 and 48 hours of aerobic incubation at 37 C. Organisms present were identified, counted, and categorized as saprophytic or pathogenic flora. The endometria of all mares were biopsied immediately following swabbing. Histologic evidence of inflammation in biopsy specimens was classified as (1) none, (2) slight, discrete, focal, and (3) slight or moderate, diffuse, widespread infiltration of inflammatory cells. The number of inflammatory cells migrating through the luminal epithelium was counted and averaged. There were significantly fewer CGS than PGS cultures that yielded growth at 24 and 48 hours of incubation after being streaked on blood agar and MacConkeys agar plates. There were fewer pathogenic bacterial or fungal colonies present at 48 hours of incubation on blood agar plates after being streaked with CGS as compared to PGS. There were no differences in the number of pathogenic bacterial or fungal colonies present at 24 hours of incubation on blood agar or at 24 and 48 hours of incubation on MacConkeys agar plates. There was no correlation between CGS or PGS culture of pathogens and severity of histologic inflammation. There was a positive correlation between culture of pathogens and number of inflammatory cells migrating through the luminal epithelium.

Bull sexual behavior tests: Stimulus cow affects performance

Abstract Sixty-four Santa Gertrudis field-breeding bulls were subjected to three series of modified Osborne-type sexual behavior tests in which one bull was exposed to one restrained stimulus cow in a small pen for 10 min. The individual stimulus cow was found to be a significant factor in copulatory, but not pre-copulatory, response of bulls. Within one series, a specific identified effect was that pre-ovulatory cows elicited significantly greater copulatory response than post-ovulatory cows.

Embryonic loss in pony mares induced by intrauterine infusion of Candida parapsilosis.

Pony mares which were detected pregnant by transrectal ultrasonography received a single intrauterine infusion of either sterile saline (control, n = 12 mares) or 10(6)Candida parapsilosis (treated, n = 12 mares) between Days 11 to 14 postovulation. Subsequent embryonic loss was studied by daily ultrasonography of the mares uterus, by serum progesterone levels, by endometrial swabs for cytologic and microbiologic examination and by endometrial biopsies that were taken after embryonic loss was detected. Significantly fewer (P<0.01) embryonic losses occurred in control than in treated mares (4 12 vs 12 12 ). The mean interval from intrauterine infusion until embryonic loss was 5.8 +/- 2.8 d for control mares (n = 4) and 2.1 +/- 0.2 d for treated mares (n = 12). Prior to embryonic loss, moderate to marked edema of the endometrial folds in 12 of 12 treated mares and free fluid in the uterine lumen of 5 of 12 treated mares were detected by ultrasonography. After embryonic loss, Candida parapsilosis was cultured from the uteri of 8 of 12 treated mares, and E . coli was cultured from the uteri of 2 of 4 control mares. Postloss endometrial smears had cytologic evidence of inflammation in 10 of 12 treated mares and 3 of 4 control mares. Intrauterine inoculation of C. parapsilosis consistently induced embryonic loss and may provide a basis to further study the relationship between endometritis and embryonic loss in mares.