Clinical efficacy of nutraceutical diet for cats with clinical signs of cutaneus adverse food reaction (CAFR).

Food allergies and food intolerances are clinically difficult to discriminate. Most often, along with cutaneous adverse food reactions or CAFR, they are classified as adverse food reactions, whose causes are numerous, including toxic compounds. Eighteen indoor-housed domestic cats with evident clinical symptoms related to CAFR (drooling, back and neck intense itching, neck eczema, chronic conjunctivitis and stomatitis) involving skin lesions were studied. Cytological evaluations of ear, skin and gingival swabs revealed an increased turnover of keratinocytes while the oxytetracycline ELISA determination showed an unexpected high amount of oxytetracycline in all cats at the first visit. All cats were then randomly assigned to receive a standard (SD group) or a nutraceutical diet (ND group) for 60 days. In the ND group a significant reduction of the mean serum concentration of oxytetracycline, pruritus intensity and skin lesion severity (**p<0.01, ***p<0.001, and ***p<0.001, respectively) was observed after 60 days, and associated with a significant improvement in the clinical picture. Although a direct correlation between oxytetracycline presence within cat sera and CAFR-related symptoms has never been described, this study highlights the benefit of a specific nutraceutical diet supplementation in improving clinical symptoms and skin lesions in cats with CARF.

Relationship between circulating anti-Müllerian hormone (AMH) and superovulatory response of high-producing dairy cows.

The main objective of this study was to evaluate the relationship between circulating anti-Müllerian hormone (AMH) and superovulatory response of dairy cows. Holstein cows (n=72) were milked twice daily and housed and fed individually in tiestalls. All animals were synchronized and flushed at 70±3 d in milk (DIM), near peak production (39.6kg/d). Blood samples for AMH analysis were collected at 3 different stages of a synchronized estrous cycle [at a random stage (40±3 DIM), proestrus (50±3 DIM), and diestrus (57±3 DIM)]. Body weights were measured weekly from calving until embryo collection. Statistical analyses were performed with Proc CORR and Proc GLIMMIX of SAS (SAS Institute Inc., Cary, NC). The 3 AMH samples from individual cows were correlated and not influenced by day of cycle. Surprisingly, AMH tended to be negatively correlated with body weight loss from calving to embryo collection (r=-0.22). More importantly, average AMH was highly associated (r=0.65) with superovulation response (number of corpora lutea on the day of the flush, CLN), total structures collected (r=0.48), and total transferable embryos (r=0.37), but not percentage of fertilized embryos (r=-0.20) or degenerate embryos (r=0.02). When cows were classified into quartiles (Q) of circulating AMH (Q1=0.01 to 82.6pg/mL; Q2=91.1 to 132.5pg/mL; Q3=135.3 to 183.8pg/mL; Q4=184.4 to 374.3pg/mL), we observed a >2-fold difference between first and fourth AMH quartiles in superovulation response (CLN: Q1=12.0±1.5; Q2=14.7±2.0; Q3=17.2±1.2; Q4=25.6±1.5) and embryo production. In conclusion, circulating AMH concentration was strongly associated with superovulation response, and evaluation of AMH could be used to identify cows with greater responses to superstimulation and thus improve efficiency of superovulation programs in dairy cows.

Transgenic expression of human CD47 markedly increases engraftment in a murine model of pig-to-human hematopoietic cell transplantation.

Mixed chimerism approaches for induction of tolerance of solid organ transplants have been applied successfully in animal models and in the clinic. However, in xenogeneic models (pig-to-primate), host macrophages participate in the rapid clearance of porcine hematopoietic progenitor cells, hindering the ability to achieve mixed chimerism. CD47 is a cell-surface molecule that interacts in a species-specific manner with SIRPα receptors on macrophages to inhibit phagocytosis and expression of human CD47 (hCD47) on porcine cells has been shown to inhibit phagocytosis by primate macrophages. We report here the generation of hCD47 transgenic GalT-KO miniature swine that express hCD47 in all blood cell lineages. The effect of hCD47 expression on xenogeneic hematopoietic engraftment was tested in an in vivo mouse model of human hematopoietic cell engraftment. High-level porcine chimerism was observed in the bone marrow of hCD47 progenitor cell recipients and smaller but readily measurable chimerism levels were observed in the peripheral blood of these recipients. In contrast, transplantation of WT progenitor cells resulted in little or no bone marrow engraftment and no detectable peripheral chimerism. These results demonstrate a substantial protective effect of hCD47 expression on engraftment and persistence of porcine cells in this model, presumably by modulation of macrophage phagocytosis.

Progesterone production in mares and echographic evaluation of the corpora lutea formed after follicular aspiration.

Ultrasound-guided follicular aspiration was performed in 26 Criollo crossbred mares, followed by the evaluation of ultrasonographic images of the Corpus luteum (CL) that was formed after puncture of follicles of different diameters (Group 25-29 mm; Group 30-35 mm and Group >35 mm). Serum progesterone (P(4) ) concentrations were measured to determine CL function. The size of the CL was measured and the CL was classified based on the following echoscore: 1- anechoic tissue; 2- poorly defined luteal structure with low echogenicity; 3- echogenicity analogous to a luteal structure. The proportion of aspirated follicles that formed a functional CL (based on P(4) concentration) 8 days after aspiration was 57.1% (4/7; CL size 25-29 mm), 75.0% (6/8; CL size 30-35 mm) and 72.7% (8/11; CL size >35 mm), respectively (p > 0.05). The echographic scores of aspirated follicles (indicating the presence or absence of a CL) were consistent with serum P(4) concentrations (p < 0.0001). Of 26 aspirations, 18 resulted in luteal function confirmed by increased progesterone concentrations ([P(4) ] > 1.0 ng/ml); 17 of these mares (94.4%) had an echoscore (2-3) compatible with luteinization (p = 0.0372). Eight days after aspiration, serum [P(4) ] > 2.0 ng/ml was associated with high (p = 0.0056) CL echoscore (3) in 15 of 17 mares (88.2%). The echoscore used in this study was valuable as a screening test to detect the presence of a functional CL after aspiration. An echoscore of 3 served as a practical and efficient method to confirm luteinization.

Canine and feline pregnancy loss due to viral and non-infectious causes: a review.

Among the causes for pregnancy loss, viruses and non-infectious factors are among the most important. In both dogs and cats, research and clinical evidence provide proof that there is an increasing incidence of pregnancy loss associated with infectious diseases like herpesvirus, as well as the presence of toxicants or chemicals in the animal's diet and environment. Endocrine causes must be taken into consideration when dealing with pregnancy loss. This review will cover the most recent knowledge regarding viral and non-infectious of pregnancy losses in the dog and cat.

Mucometra, cystic endometrial hyperplasia, and pyometra in the bitch: advances in treatment and assessment of future reproductive success.

Pyometra is a common reproductive disorder which affects nearly one fourth of all female dogs before they reach 10 y of age. An association between pyometra and the most common uterine disease of the bitch, cystic endometrial hyperplasia, has been established, as the latter allows commensal bacteria originating from the vagina to proliferate in the uterus at the end of estrus. The progressive degenerative process in the development of cystic endometrial hyperplasia is usually proposed as the initiating lesion for pyometra in bitches; this is mediated by progesterone and potentially aggravated by estrogens. However, a separate process caused by local uterine irritation to trophoblastic reaction and bacterial proliferation has been recently proposed as an alternate mechanism leading to the development of pyometra. Pyometra is clinically distinct in pathogenesis, signs, treatment and prognosis from postpartum metritis or mucometra. Treatment of pyometra has historically involved ovariohysterectomy, however, during the last 10 y, numerous effective treatments have been proposed to treat both open and closed cervix pyometra with good success and future fertility. Among the treatments available, the use of repeated low doses of prostaglandins alone or in association with either dopamine agonists or progesterone-receptor antagonists has been demonstrated to be a viable alternative for valuable breeding dogs.

Endocrinology of pregnancy in the dog: a review.

Pregnancy regulation in the dog is not yet fully elucidated. Since plasma progesterone concentrations are similar in pregnant versus non-pregnant animals, it is a poor reflection on CL function and progesterone metabolism. Increased progesterone secretion by the CL in pregnant animals follows implantation and relaxin secretion by the feto-placental units. Progesterone is absolutely required to maintain pregnancy and no placental sources of progesterone have been identified. Pregnancy can be artificially maintained by progesterone administration. Prolactin secretion appears to be increased in response to the increase in relaxin production and occurs independent of estrogen production by the CL. The respective roles of LH, FSH and prolactin are still unclear, with considerable conflicting evidence among studies. However, it appears that prolactin is absolutely required, whereas LH is either permissive or facilitates CL function during pregnancy. Pre-implantation events are still poorly defined in the bitch, and no embryonic factors have been isolated or purified, preventing early pregnancy diagnosis. Parturition occurs following luteolysis, which results from the release of prostaglandin F(2alpha), which begins 36h prepartum in a process similar to that observed in other species. The role of estrogens at the time of parturition remains undefined.

Retrograde ejaculation occurs in the dog, but can be prevented by pre-treatment with phenylpropanolamine: a urodynamic study.

Retrograde ejaculation is partial or total propulsion of semen from the posterior urethra into the urinary bladder; it is well characterized (and relatively common) in humans, with only a few reports in animals. Our objectives were to determine whether retrograde flow of semen occurred during ejaculation in mature dogs with normal fertility, and to determine the effects of phenylpropanolamine on this phenomenon (dose-titration, switch-back study). Retrograde ejaculation and urethral pressure profile measurements were evaluated (double-blind) in six dogs after 5 days of oral treatment with phenylpropanolamine (0, 2, 4, or 8 mg/kg); all dogs received all treatments (at 2-week intervals). The number of sperm in the urine was determined before and after each manual sperm collection. Urethral pressure profiles were obtained three times during each procedure. In the absence of phenylpropanolamine, sperm were present in the bladder after semen collection in all dogs (number varied significantly among individuals). The mean (+/-S.D.) number of sperm in the bladder was 17.0+/-5.0, 18.5+/-1.2, 5.1+/-5.0, and 4.8+/-0.1 x 10(6) sperm for 0, 2, 4, and 8 mg/kg, respectively (no significant difference between dogs given 4 or 8 mg/kg, but both were significantly lower than those given 0 or 2mg/kg). This reduction was significantly correlated to the increase in mean urethral pressure at the level of the sphincter (39 cm versus 59 cm H(2)O in placebo-treated dogs versus those given 8 mg/kg). In conclusion, we confirmed that retrograde ejaculation occurred during the ejaculatory process in normal dogs, and we demonstrated that phenylpropanolamine (4 or 8 mg/kg once daily for 5 days before collection) increased urethral pressure and reduced the number of sperm voided into the bladder during ejaculation.

Validation of a serum immunoassay to measure progesterone and diagnose pregnancy in the West Indian manatee (Trichechus manatus).

The objective was to validate a high-sensitivity chemiluminescent assay of serum progesterone concentrations for pregnancy diagnosis in manatees. Assay analytical sensitivity was 0.1 ng/mL, with mean intra- and inter-assay coefficients of variation of 9.7 and 9.2%, respectively, and accuracy had a mean adjusted R(2) of 0.98. Methods comparison (relative to Siemen's Coat-A-Count RIA) demonstrated r=0.98, Deming regression slope of 0.95, and an intercept of 0.01. Based on ROC analysis, a progesterone concentration >or=0.4 ng/mL was indicative of pregnancy. Assay results were not significantly altered by two freeze-thaw cycles of samples. Characteristic progesterone concentrations during pregnancy were Months 1-4 (1.7-4.7 ng/mL), 5-8 ( approximately 1.0 ng/mL), and 10 and 11 (0.3-0.5 ng/mL), whereas two late-pregnant females with impending abortion had progesterone concentrations of 0.1 ng/mL. Among pregnant females, maximum progesterone concentrations occurred in autumn (3.9+/-1.8 ng/mL), and were greater during all seasons than concentrations in non-pregnant females (0.1-0.2 ng/mL). Progesterone concentrations were also significantly higher in pregnant females than in non-pregnant females and males. This highly sensitive, specific, and diagnostic assay will be valuable for monitoring pregnancy and abortion in manatees.

Addition of different concentrations of prostasome-like vesicles at neutral or slightly alkaline pH decreases canine sperm motility.

Prostasome-like vesicles (PV) are components of the canine prostatic fluid during ejaculation and despite some enzymatic activities have been identified recently, their functions in the reproductive events in this species are still poorly understood. In this study, we evaluated at neutral or slightly alkaline pH, the effects of different concentrations of purified PV on sperm cell motility characteristics and the impact on the short- and long-term preservation of preserved semen maintained at room temperature. Two different experiments were performed. In the first experiment, purified PV were added at increasing concentration (1.25, 2.5, 5, 10, and 20 µL equivalent to 1, 2, 4, 8, and 16 µg of protein, respectively) to aliquots of 100 µL of preserved semen maintained at 22 °C at the following time points: 0, 30, 60, 120, 240, and 480 min. Computer-assisted sperm motility characteristics and pH were assessed three times at each time points, for each sample and for every concentrations. In the second experiment, the purified PV were added with the same methods as described above but only at time 0. Sperm motility characteristics and pH were assessed over the time. This study showed how the addition of purified PV to preserved semen affects negatively (p < 0.05) at neutral and alkaline pH, both total and progressive motility in a concentration depending manner. Furthermore, prostasome addition was demonstrated to change the quality of sperm movement which may represent a mechanism facilitating sperm cells attachment to the uterine epithelium and facilitating energy preservation before fertilization.

Serum concentration dynamic of energy homeostasis hormones, leptin, insulin, thyroid hormones, and cortisol throughout canine pregnancy and lactation.

Pregnancy and lactation represent critical periods of canine reproductive life and different hormones are required to maintain homeostasis and the correct energy requirements. The aim of this study was to evaluate leptin, insulin, triiodotyronine (T3), thyroxin (T4) and cortisol serum concentrations to better describe the canine maternal energy metabolism. Twelve Bloodhound bitches that had at least one litter and were considered to be healthy submitted blood samples every 15 days starting from day 0 (ovulation) throughout pregnancy up to the end of lactation (day 120). We found that leptin concentrations increased from day 0 to reach maximum value at day 45 (P < 0.001), decreased at day 60 (P < 0.05) and rose again at day 105 and day 120 (P < 0.05 and P < 0.01 respectively). Insulin concentrations significantly increased during pregnancy peaking at day 60 (P < 0.05). After parturition dropped and reached progressively lower values similar to those at day 0 at the end of lactation. During pregnancy Body Weight was in low correlation with leptin (r = 0.32, P < 0.01) and there was no correlation with insulin (r = 0.13, P > 0.1); moreover, no correlation was found during lactation. T3 concentrations showed a U-shaped trend with a significant reduction at day 75 (P < 0.05). T4 concentrations fluctuated without any significant change (ranging from 27.0 ± 1.6 nmol/l to 32.1 ± 1.9 nmol/l). Cortisol concentrations ranged within the reference values (minimal value at day 30 (49.8 ± 6.3 nm/L) and maximal value at day 45 (72.5 ± 16.1 nm/L)). Our study states that canine pregnancy and lactation evoke many hormonal changes necessary to mother and fetuses. In particular, this is the first report regarding the pattern of leptin during whole pregnancy and lactation. The present work lays the groundwork for further studies on endocrine homeostatic mechanisms using the dog as a model for human medicine.

Long-term motility and fertility conservation of chilled canine semen using egg yolk added Tris-glucose extender: in vitro and in vivo studies.

The effects of medium exchange on motility parameters of chilled canine semen preserved in egg yolk Tris-glucose (EYTG) extender were analyzed over a 27-d period. Semen extender was exchanged at three time points (Days 11, 21 and 27) after collection, when motility parameters were demonstrated to significantly decrease from parameters observed at semen preparation (Day 0) or at day of previous extender exchange. In the absence of medium exchanges, motile spermatozoa were observed up to Day 16 (mean +/- S.D. 1.5 +/- 0.3% of motile spermatozoa). A stimulation of the different semen motility parameters was observed after extender exchange. Semen extender exchange at Day 11 allowed conservation of motility until Day 21, compared to 16 d in the absence of extender exchange. At Day 21, when spermatozoa appeared immobile or dead, a second extender exchange was performed, allowing the extension of motility conservation up to Day 27. The third extender exchange, performed at Day 27, was no longer associated with motility stimulation. Glucose content in the medium decreased slowly over time; a concomitant decrease in pH was also observed. No changes in osmolarity were observed over time. To verify the fertility of long-term conserved chilled semen, two groups of 10 bitches were inseminated either once (Group 1) or twice at 48-h intervals (Group 2) intra-vaginally with semen conserved chilled for a mean of 9 +/- 1.8 d. Out of the 10 bitches inseminated once, 5 became pregnant, versus 7 in the group of animals inseminated twice. The present study reports the possibility to extend the conservation of chilled canine semen up to 3 wk with conservation of good fertility for at least 10 d. The role of energetic substrate and pH alteration is postulated and the classically accepted relation of semen motility/viability is raised.

Improving in vitro maturation and pregnancy outcome in cattle using a novel oocyte shipping and maturation system not requiring a CO2 gas phase.

The present work evaluated the benefit of a novel shipping and maturation medium (SMM) not requiring a CO2 gas for maturation and subsequent embryonic development of slaughterhouse and ovum pickup (OPU) bovine cumulus-oocyte complexes (COCs). Four experiments were conducted. In experiment 1, COCs were maturated for 18 hours in SMM and then incubated for 6 hours in, or 24 hours in a conventional system (control). Experiment 2 compared maturation for 24 hours in SMM versus 24 hours in the control. Experiment 3 compared three different incubation temperatures (37 °C, 38 °C, and 38.5 °C) for COCs maturation in SMM. In experiment 4, COCs obtained from 166 OPU sessions (representing two dairy and two beef breeds) in two locations (Wisconsin and California) were matured in SMM or control and evaluated relative to embryo production and pregnancy rates. Frozen semen was used for all experiments. The results for experiment 1 showed that the blastocyst rate and total embryo production rate (TE, Day-7 morulae plus all blastocysts) were higher for SMM than those in the control. However, no differences were observed for cleavage rate or blastocyst stage. In experiment 2, the blastocyst rate and TE were higher for SMM than those in the control; however, there was no difference for cleavage rate, total cell number, blastocyst stage. In experiment 3, the cleavage rate was similar, but the blastocyst rate and TE were greater for 38.5 °C than those for 38.0 °C and 37.5 °C. For experiment 4, Wisconsin OPU-derived COCs had a greater cleavage rate, blastocyst rate, TE, and blastocyst stage for SMM versus control. There were no breed effects. For the California trial, OPU-derived COCs matured in SMM had similar cleavage and pregnancy rates at Day 35 but greater blastocyst rates and transferred embryos per session than the control, which resulted in 2.2 more pregnancies per OPU session. Holstein COCs had superior embryonic development but similar pregnancy compared with Jersey. We conclude that COCs matured in SMM had greater oocyte competence than the control. Also, maturation at 38.5 °C in SMM was optimal for embryonic development. In summary, SMM resulted in greater embryonic development, similar pregnancy rates, but higher pregnancies per OPU session than the conventional maturation system.

Coexpression of human growth hormone-releasing factor 1-37-like and alpha-melanotropin-like immunoreactivities in neurones of the rat lateral dorsal hypothalamus.

Antibodies recognizing the 29-37 sequence of the human somatocrinin specifically stain a large population of interneurones located in the lateral dorsal hypothalamus. Staining comparisons revealed that these perikarya also contain alpha-MSH-like immunoreactivity. The neurones exhibiting human GRF1-37-like immunoreactivity correspond to the system previously shown to present alpha-MSH-like and rat CRF-like immunoreactivities.

Studies on factors influencing human plasma alpha-MSH.

The various physiological effects of alpha-MSH, mainly on the CNS and on pigmentation in animal models, are well documented in the literature. Only a few investigators have confirmed similar properties in the human. However, the possible physiopathological role played by this hormone in human melanoma is still poorly defined. In order to approach this subject in a manner as complete as possible, we have performed, during the past four years, three different series of experiments: 1) alpha-MSH measurements in plasma samples from: a. melanoma and other cancer patients, b. whole body UVA irradiated healthy adults, c. circadian rhythm determinations in melanoma patients and in healthy male adults; 2) alpha-MSH measurements in human melanoma tumours; 3) alpha-MSH receptor expression on human melanoma cells in culture involving: a. alpha-MSH radio-binding assays and b. tyrosinase assay. Our results so far show 1) increased alpha-MSH levels in melanoma patients' plasma, alpha-MSH responsiveness to UVA stimulated skin, large immunoreactive alpha-MSH content in melanoma metastases and an alpha-MSH circadian rhythm in some individuals different from cortisol; 2) alpha-MSH receptor expression in melanoma cells could be increased by various effectors able to stimulate melanogenesis.

In vivo investigation of luteal function in dogs: effects of cabergoline, a dopamine agonist, and prolactin on progesterone secretion during mid-pregnancy and -diestrus.

The role of prolactin on luteal function in dogs was investigated in vivo. The function of prolactin in mid-luteal phase was compared in pregnant and nonpregnant dogs. A dopamine agonist, cabergoline, known for its prolactin secretion inhibitory effects, was injected subcutaneously at a dose of 5 micrograms/kg body weight in five pregnant and five nonpregnant Beagle bitches. Mean plasma prolactin and progesterone were dramatically suppressed for 4 to 5 days after injection in both groups when compared with control pregnant and non-pregnant animals, whereas no effect on luteinizing hormone (LH) secretion was observed. The decline in plasma progesterone occurred after that in prolactin, suggesting plasma progesterone was impaired by inhibition of prolactin secretion. These results confirm the luteotropic importance of prolactin in pregnant bitches, and also demonstrate its importance in luteal phase of the nonpregnant dog. Second, to demonstrate that the effects of cabergoline were mediated by prolactin inhibition and not by a direct action on the corpus luteum, concomitant administration on Day 30 of cabergoline and prolactin (375 micrograms i.v. twice daily on Days 30 and 31) or cabergoline and LH (750 micrograms i.v. twice daily on Days 30 and 31) was affected in two groups of five pregnant animals each. Results showed that only prolactin was able to reverse the negative effects of cabergoline on circulating progesterone. This confirms the indirect mode of action of the dopamine agonist, cabergoline on corpus luteum function. Third, further investigation on the precise luteotropic role of prolactin was made by IV injection of 375 micrograms pure canine prolactin twice daily in five pregnant bitches on Days 30 and 31, and in five pregnant bitches on Days 40 and 41. No direct stimulatory effect of prolactin on plasma progesterone secretion occurred. Nor was there a noticeable effect on plasma LH secretion. These results suggest that prolactin is unable to directly stimulate progesterone secretion by the corpus luteum of pregnancy. The results of this study suggest that prolactin is an essential luteotropin in the dog from mid-luteal phase in both pregnant and nonpregnant animals. However, it appears to act by sustaining corpus luteum lifespan and function rather than by direct stimulatory effects on progesterone secretion.

Evaluation of the "Hamilton Thorn computer-based automated system" for dog semen analysis.

An objective evaluation of semen is warranted to assess the canine male fertility and to select appropriate techniques and extenders for semen preservation. With conventional microscopic evaluation, the subjectivity of the analysis makes any comparison of results difficult. In the present study, we validated the Hamilton Thorn computer-aided semen analyzer (HTR-IVOS10 analyzer) for objective assessment of canine semen. A description of fertile canine motility parameters using this analyzer is reported. Semen analysis at 38 degreesC is found to be more optimal and accurate than 30 degreesC. The Makler chamber was preferred to the Cell-vu, which induced a decrease of all semen motility parameters. The repeatability of the measures was good with intra-and inter-assay coefficients of variation below 10% and 20%, respectively, for most of measured parameters. An overestimation of semen concentration, increasing with dilution of semen, was observed when HTR-IVOS10 results were compared with the classical manual Makler cell evaluation. A significant decrease in semen motility parameters was recorded when high semen dilutions were used. Generated from the analysis of 42 mature fertile male beagle dogs, a description of semen motility parameters using the CASA system is presented to serve as reference when comparing Beagle ejaculates both in clinical and experimental studies.

Long-term preservation of chilled canine semen: effect of commercial and laboratory prepared extenders.

The present study was conducted to evaluate chilled semen conservation over time in 3 commercial and 4 laboratory prepared extenders, including a new Tris-glucose extender. The beneficial effect of adding egg yolk to these media was also analyzed. The effects of these extenders on motility and acrosome reaction were characterized objectively using a computer-aided semen analyzer and the chlortetracycline staining, respectively. No significant differences were observed when comparing the different commercial extenders without egg yolk, but addition of egg yolk improved all motility parameters significantly (preservation of 50% of motility was observed at 3.2+/-1, 2.9+/-0.5, 2.3+/-0.5, 8.5+/-0.2, 5.4+/-1.1, 5.2+/-0.4 d, for Biladyl, green extender and fresh-phos extenders without and with egg yolk, respectively). Motility parameters were best preserved in egg yolk supplemented Biladyl extender with a mean percentage of 86.3+/-10.5 motile spermatozoa after 7 d at 4 degrees C. Efficacy of egg yolk-supplemented commercial extenders on sperm motility at 4 degrees C was (in decreasing order) as follows: Biladyl > green extender > fresh-phos. However, high quality motility and the percentage of motile spermatozoa were highest with some of the laboratory prepared extenders: a 50% conservation rate of motile spermatozoa was observed following the use of supplemented egg yolk extenders. These are classified in decreasing order as follows: Tris-glucose (13+/-1 d) > Tris-fructose (9.7+/-0.6) > EDTA (4.+/-0.6 d) > Tris-bes (3.6+/-1.1 d). A low concentration of motile spermatozoa was still observed in the Tris-glucose egg yolk extender 16 d after collection, clearly demonstrating the importance of the medium and the beneficial effect of egg yolk on sperm motility of 4 degrees C chilled semen. Similar effects of extender were observed for acrosome reactions. Egg yolk clearly had a protective effect reducing acrosome reactions significantly in all media tested as follows: the highest acrosome losses were observed in the fresh-phos and EDTA extenders without egg yolk; the lowest rate was observed with Tris-glucose supplemented egg yolk extender. In conclusion, at 4 degrees C, egg yolk extender best-protected sperm motility parameters. Differences in osmolarity between the extenders in terms of substrate related to sperm metabolic activity may explain the optimal results obtained using egg yolk-supplemented Tris-glucose extender, which preserved motility and acrosome integrity in chilled dog semen. These results indicated that good quality dog spermatozoa could be preserved for up to 10 d.

Validation of the sperm quality analyzer (SQA) for dog sperm analysis.

In the present study, a simple and inexpensive unit (the Sperm Quality Analyzer-SQA), was evaluated for dog sperm analysis. Our objective was to propose a cheap, accurate and convenient device to be used in veterinary practices involved with dog fertility assessment and artificial insemination. The device was tested by analyzing repeatability and accuracy at different sperm concentrations and motility characteristics. The Sperm Motility Index (SMI), a numeric index provided by the SQA, was compared with the results obtained using a computer-aided sperm analyzer (Hamilton Thorn IVOS 10). The correlation between SMI and some sperm parameters as well as predictive values of the SMI were established. The dog sperm data provided by the SQA were consistent and repeatable (coefficient of variability below 10% for all concentrations tested). The SMI was significantly dependant on motile sperm concentration and a positive significant correlation was established for the different motile sperm concentrations from a concentration of 25 x 10(6) up to over 200 x 10(6) cells/mL. Zero motility did not affect SMI because non-motile cells, regardless of their concentration, do not cause any fluctuations in the optical density (OD). Over the tested 200 x 10(6) cells/mL value, a correlation still could be observed but it was not statistically significant, possibly because of a saturation of the system. In dog semen, the correlation is better between SMI values and the number of motile spermatozoa than with the overall motile concentration. Based on this observation, a predictive value was given to the SMI allowing for a sorting of dog ejaculates in 3 sperm categories (SMI <100, 100250) each characterized by a range of sperm number and motility. If a positive correlation between the SMI categories and fertility has been demonstrated in humans, such a correlation needs to be established in dogs.

Radiographic contrast medium for uterine insemination in the bitch, and its effect upon the quality and fertility of fresh dog semen.

Four ejaculates were collected from each of 6 adult male beagle dogs. The second fraction was divided into 3 aliquants which were then diluted with physiological saline, an isoosmolar solution of sodium/meglumine diatroate, and an iso-osmolar solution of iohexol. The diluted samples were incubated at 39 degrees C and evaluated at 0, 60, 90, 120, 240 and 360 min after dilution. A variety of assessments was made, including, spermatozoal motility, spermatozoal morphology, and acrosorne status. The practicality of using 1.0 ml of iso-osmolar contrast medium combined with radiographic examination was evaluated as a method of confirming accurate placement of a transcervical uterine catheter by injecting contrast after positioning the catheter in 4 beagle bitches. The effect of the procedure on fertility was assessed using 5 greyhound bitches which were inseminated with fresh semen and in which pregnancy was monitored using diagnostic B-mode ultrasound imaging. There was no significant difference between physiological saline and the sodium/meglumine diatroate solution upon semen quality, while the iohexol solution produced a significant reduction in spermatozoal motility and morphology. No adverse clinical effects were observed when contrast medium was administered into the uterus to either group of bitches. A subjective assessment of radiographic quality showed that the sodium/meglumine diatroate solution, which contained twice the iodine concentration of the iohexol solution, produced significantly greater radiopacity and was radiographically more useful than the iohexol solution. The sodium/meglurnine diatroate solution had no adverse effect upon the fertility of dog semen, and all bitches that were inseminated with this technique conceived and maintained the pregnancy to term. Litter size was considered to be normal for the breed. Small volumes of an iso-osmolar solution of sodium/meglumine diatroate may be useful for ensuring correct placement of transcervical catheters prior to artificial insemination in the bitch.