Intraperitoneal insemination and retrograde sperm transport in dairy cows

To examine the efficiency of retrograde sperm transport following intraperitoneal insemination, live and dead spermatozoa were used at different concentrations, and sperm recovery from cervical mucus (0.5 ml) 2, 6, 12 and 24 h following insemination was evaluated. Forty lactating Friesian cows, in their second to fourth lactation period, were used in this experiment. Thirty-six cows received intraperitoneally either live or dead spermatozoa. Each group of six cows received one of three total sperm numbers of 30, 45 and 90 million. Four cows were inseminated with 90 million spermatozoa into the uterus and served as a control group. All cows were inseminated towards the end of oestrus. After intrauterine insemination sperm recovery declined, but motile and/or immotile spermatozoa were recovered from all cows at any time. In cows inseminated intraperitoneally, sperm was recovered from the cervix at 6-24 h when 90 million were inseminated. A greater number of spermatozoa was recovered after dead rather than after live sperm inseminations. Only immotile, intact or broken spermatozoa and tail-less heads were recovered after intraperitioneal insemination using either live or dead spermatozoa. No sperm was recovered for 30 and 45 million inseminations. Our results show that, following intraperitoneal insemination, there is passive sperm transport from the peritoneal cavity to the genital tract close to the time of ovulation, and suggest a higher sperm retention in the genital tract when live as opposed to dead spermatozoa are used.     

Unilateral intrauterine horn insemination of fresh semen in cats

The sperm count required were investigated to obtain a conception rate of 80% by unilateral intrauterine insemination (UIUI) of fresh semen in cats. The conception rates obtained by insemination before and after ovulation were also examined. Thirty-six female cats aged 1-7 years were used in the experiments, and the number of experimental cases was 44. Seven male cats aged 2-12 years from which semen could be collected by the artificial vagina method were used. In artificial insemination, 100 iu x 2 or 250 iu of hCG was administered on days 2-4 of estrus, and sperm were introduced into the uterine horn with a greater number of ovulations (or mature follicles) 15, 20 and 30 hr after hCG administration by laparotomy. The inseminated sperm counts were 2 x 10(6) (Exp. 1). 4 x 10(6) (Exp. 2), and 8 x 10(6) (Exp. 3). As a result, ovulation was induced in 42 of 44 cases (induction rate: 95.5%) regardless of the dosage of hCG. Conception was obtained by UIUI in two of 16 animals (conception rate: 12.5%) in the Exp. 1, five of 16 animals (31.3%) in Exp. 2, and eight of 10 animals (80.0%) in Exp. 3. Regarding the relationship between the ovulation state at insemination and conception, the conception rate obtained by insemination before ovulation was clearly higher than that obtained by insemination after ovulation (p<0.05). Regarding the number of kits compared to the number of ovulations on the inseminated side, the percentages of cases in which the number of kits exceeded the number of ovulations on the inseminated side were similar in all groups inseminated with a different number of sperm. It is therefore necessary to investigate conception rates obtained by bilateral insemination to increase the fertility rate. Based on the above findings, it was shown that the sperm count required for fertilization by UIUI is 8 x 10(6).     

Distribution of spermatozoa in the genital tract of artificially inseminated heifers

Eight heifers were artificially inseminated in the uterine body with 160×10⁶ spermatozoa frozen in French mini-straws. The heifers were slaughtered 2 (n = 4) or 12 (n = 4) h after insemination and spermatozoa were recovered by flushing defined segments of the reproductive tract. The efficiency of the method was checked in different ways. There was a slight underestimation of the number of recovered spermatozoa. This underestimation was randomly distributed among heifers and genital tract segments.The total number of spermatozoa recovered was higher at 2 than at 12 h (14.6 vs 0.6 % of the total number inseminated). Most spermatozoa were found in the vagina both at 2 and 12 h after insemination and in greater number at 2 h. In uterus there was a slight decline in the number of spermatozoa recovered at 2 versus 12 h after insemination. The number of spermatozoa recovered from the oviducts were similar at 2 (89.6 × 10³) and 12 h (71.5 × 10³) after insemination. At 2 h spermatozoa were found in all parts of the oviduct with the majority located in the utero tubal junction, whereas at 12 h the most were recovered from isthmus.More spermatozoa were recovered from the left than from the right side of the tract in 6 of the 8 heifers. Only in 1 heifer were the majority of spermatozoa found in the oviduct ipsilateral to the follicle bearing ovary.     

Sperm interaction with the female reproductive tract

Sperm transit in the female tract is a critical event for the success of fertilization. From their deposition in the vagina to final migration in the oviduct, sperm pass through the different compartments of the genital tract in which they encounter different environments. The cervix and the uterotubal junction (UTJ) are two barriers with different relative importance according to the species. The protein composition, the degree of glycosylation and the hydration of the cervical mucus change during the oestrous cycle. Several sperm surface proteins are associated with their migration through the cervical mucus and the UTJ. Data regarding the interaction of sperm with secretions of the epithelial tissue lining the different compartments of the female genital tract during the sperm transit are reviewed, with a particular emphasis on the migration of sperm through the cervix.     

Producing progeny from endangered birds of prey: treatment of urine-contaminated semen and a novel intramagnal insemination approach.

Wild raptors brought into an ex situ environment often have poor semen quality that is further compromised by urine contamination. Generally, it is believed that in birds, artificial insemination into the cloaca or caudal vagina of females requires large doses of high-quality spermatozoa to maximize fertility. In an effort to define and overcome some of the challenges associated with reproduction in wild raptors, the objectives of this study were to 1) evaluate the frequency, impact, and remediation of urine contamination in fresh ejaculates for the purpose of maintaining sperm motility and viability in vitro, and 2) develop a deep insemination method that allows low numbers of washed sperm to be placed directly into the magnum to increase the probability of producing fertilized eggs. The species evaluated include golden eagle (Aquila chrysoetos), imperial eagle (A. adalberti), Bonelli's eagle (Hiernaetus fasciatus), and peregrine falcon (Falco peregrinus). Semen samples were collected and pooled by species, and a minimum of 25 pooled ejaculates per species were evaluated for urine contamination, pH, sperm viability, and sperm motility; the samples were either unwashed or washed in neutral (pH 7.0) or alkaline (pH 8.0) modified Lake's diluent. Female golden eagles and peregrine falcons were inseminated via transjunctional, intramagnal insemination with washed spermatozoa from urine-contaminated samples. Urine contamination occurred in 36.8 +/- 12.8% (mean +/- SEM) golden eagle, 43.1 +/- 9.1% imperial eagle. 28.7 +/- 16.1% Bonelli's eagle, and 48.2 +/- 17.3% peregrine falcon ejaculates. The pH in urine-contaminated semen samples ranged from 6.48 +/- 0.3 to 6.86 +/- 0.2, and in noncontaminated samples it ranged from from 7.17 +/- 0.1 to 7.56 +/- 0.1. Sperm viability and motility were reduced (P < 0.05) in all species for unwashed vs. washed sperm after 30 min incubation at room temperature. Two peregrine falcon chicks and one golden eagle chick hatched after intramagnal insemination. This study demonstrates that urine contamination, a common and lethal acidifier in manually collected raptor ejaculates, can be circumvented by immediate, gentle seminal washing. Furthermore, these processed sperm, when deposited by transjunctional intramagnal insemination, can produce live young.     

Effect of unilateral cornual insemination upon fertilization rate in superovulating and single-ovulating cattle

In Exp. 1, 21 first-service cattle and seven repeat-breeder cattle, averaging 4.7 infertile services, were brought into estrus and superovulated by treatment with follicle-stimulating hormone and prostaglandin F2 alpha. At insemination, semen was deposited in the greater curvature of one uterine horn, about midway between the utero-cervical junction and the utero-tubal junction. Cattle were necropsied 2 to 7 d after estrus and ova were recovered and examined. The fertilization rate for first-service cows was 74% of 362 intact ova and for repeat-breeders, 43% of 128 intact ova (P less than .001). Fertilization rate in first-service cows was 81% on the side of semen deposition and 68% on the opposite side (P less than .01); the rates in repeat-breeders were 54% and 32% (P less than .025). Differences between sides were due mostly to four cows that averaged 93% fertilization on the side of semen deposition and 19% on the opposite side. The proportion of fertilized ova with accessory sperm (17%) did not differ between sides of the reproductive tract. In Exp. 2, 60 first-service and 32 repeat-breeder cows in natural estrus had semen deposited in the uterine body or in the greater curvature of one uterine horn, either on the side of impending ovulation or on the opposite side. At necropsy, 55 ova were recovered from first-service cows, of which 42 (76%) were intact and 13 (24%) were ruptured or fragmented. Of the 42 intact ova, 41 (98%) were cleaved. From the 32 repeat-breeders, 30 ova were recovered, of which 26 (87%) were intact and 4 (13%) were ruptured; 23 of the 26 intact ova (88%) were cleaved. Site of semen deposition had no significant effect on either fertilization rate or number of accessory sperm in either type of cow. First-service cows averaged more accessory sperm (40) than did repeat-breeders (19, P less than .01). Overall results indicated that sperm deposited deep in one uterine horn fertilized ova nearly as frequently in the opposite oviduct as in the adjacent oviduct except in 14% of superovulating cattle.     

Fertility assessment through heterospermic insemination of flow-sorted sperm in cattle

The ability to assess fertility of bovine sperm accurately and rapidly would be very useful for research and applications to the cattle industry. Sperm motility and other in vitro tests of sperm normality are only partially correlated with fertility, and lengthy breeding trials are expensive and time consuming. Heterospermic insemination by mixing sperm from more than one male provides an in vivo method to assess relative fertility among bulls that can be economical and rapid. Sperm that had been flow-sorted and cryopreserved from four groups of four bulls were inseminated in all combinations of three bulls within groups into nonsuperovulated heifers or superovulated heifers. Embryos were collected nonsurgically between d 13.5 and 20 following estrus and evaluated for paternity by genotyping. Following determination of paternity, a heterospermic index was created for each bull using a maximum likelihood function. These indices ranged from 0.22 +/- 0.15 to 2.43 +/- 0.43 (mean = 1.00, with a higher value indicative of greater fertility). In all four groups, either the high- or low-fertility bull was identified (P < 0.05) using a total of 25 to 36 genotypable embryos from nonsuperovulated heifers. The heterospermic rankings of bulls were similar for single and superovulated heifers for one group of bulls, but dissimilar for a second group. Heterospermic insemination followed by genotyping of embryos proved to be efficacious for rapidly ranking fertility of flow-sorted sperm from bulls when females were not superovulated, but results were less clear when females were superovulated.     

Conception rates in early postpartum ewes bred naturally or by intrauterine insemination

Ewes were treated with a medroxyprogesterone acetate (MAP) sponge for 8 d followed, at sponge removal, with 500 IU pregnant mare serum gonadotropin (PMSG) at d 30, 40 or 50 (d 0 = lambing) to induce estrus. Dry and lactating ewes were divided into equal numbers at each postpartum day and bred at estrus. Conception rates and number of accessory sperm were determined by flushing the oviducts 3 d after mating and examining the recovered ova. There was no effect (P greater than .05) of lactational status on conception rates. Conception rates increased (P less than .05) from d 30 (10%) to d 40 (45%) and from d 40 to d 50 (80%). There were fewer (P less than .05) ova with accessory sperm (5/26) in d-30 ewes compared with d-40 (10/27) or d-50 (12/24) ewes. In Exp. 2, ewes were assigned to two groups after receiving PMSG on d 30: 1) mated naturally or 2) inseminated during laparotomy near the uterotubal junction (UTJ). Dry and lactating ewes were divided evenly within each of the two treatments. Oviducts were flushed and ova were examined for cleavage. The conception rate was 60% in ewes that were inseminated in the UTJ vs 10% in ewes mated to rams (P less than .05). Lactational status had no effect on results. In conclusion, conception rates in postpartum ewes treated with MAP sponge and PMSG increased from postpartum d 30 to d 50 with natural breeding, and d-30 conception rates were increased over natural mating by insemination into the uterine horn near the UTJ.     

Improvement by ergonovine of sperm transport, fertilization and pregnancy rates in ewes in natural or prostaglandin-induced estrus

Eight experiments were conducted with 451 ewes to test effects of ergonovine, prostaglandin F2 alpha (PGF2 alpha) and phenylephrine on sperm transport and fertility. In most experiments, ewes were mated at estrus and necropsied 2 or 3 h later. Sperm were flushed from the oviducts, uterus and anterior, middle and posterior thirds of the cervix and counted. Various doses of PGF2 alpha or phenylephrine given im at mating caused no significant increase in sperm numbers in any segment of the tract 2 h later. Three different dose levels of ergonovine were given im to ewes in natural estrus 1 h after mating and ewes were necropsied 3 h after mating. Doses of .2 and 1.0 mg were ineffective, but .5 mg increased sperm numbers about 10-fold in the oviducts and uterus. When given im at the time of artificial insemination, .6 mg of ergonovine increased the fertilization rate at 3 d from 5/25 in control ewes to 12/25 (P less than .05). In three experiments with ewes in PGF2 alpha-induced estrus, .6 mg of ergonovine increased sperm numbers in the cervix and uterus at 3 h after mating and in the uterus and oviducts at 23 h, near ovulation. Other ewes were artificially inseminated in the external cervical os and one-half of the ewes were given .6 mg of ergonovine im; ewes not returning to estrus were laparotomized at 22 to 26 d and embryos removed. After insemination during natural estrus with .2 ml of semen, pregnancy rates were 14/25 for control ewes and 15/25 for ergonovine-treated ewes; after insemination during natural estrus with .1 ml of semen, 6/35 and 18/35 (P less than .005); after insemination during PGF2 alpha-induced estrus with .2 ml of semen, 7/60 and 12/60. Fertilization and pregnancy rates combined were 32/145 (22%) for all control ewes and 57/145 (39%) for ergonovine-treated ewes (P less than .005).     

Reproductive criteria of beef bulls during and after exposure to increased ambient temperature

Sixteen yearling Angus bulls were randomly assigned to one of two temperature-controlled chambers to determine the effects of elevated ambient temperature on body functions and semen characteristics. After 8 wk adjustment at 23 C, eight heat-stressed bulls were exposed to 35 +/- 1 C for 8 h and 31 +/- 1 C for 16 h during each 24-h period, and eight control bulls were maintained at 23 +/- 1 C for 8 wk. Then all bulls were exposed to 23 C for 8 wk. Bulls were fed so that both control and stressed bulls gained at similar rates (.58 kg/d). Semen was collected with an artificial vagina twice weekly before, during and after heat stress. During treatment, the respiratory rate of stressed bulls was greater (P less than .001) than that of control bulls (54.2 +/- 1.5, 29.9 +/- 1.5 breaths/min, respectively). Rectal temperatures were increased (P less than .01) from 38.2 +/- .1 to 38.7 +/- .1 C and water consumption was increased by 35% in stressed bulls when compared with controls. Semen volume was not altered by treatment, but percentage of motile sperm decreased (P less than .01) in stressed bulls by 2 wk after the start of heat treatment. Sperm motility of stressed bulls returned to normal values 8 wk after the end of heat treatment. Similarly, the percentage of aged acrosomes on sperm from stressed bulls increased (P less than .01) by the second week of treatment and remained greater than that of controls throughout the stress period.(ABSTRACT TRUNCATED AT 250 WORDS)     

Fertilization and blastocyst development in oocytes obtained from prepubertal and adult pigs

Polyspermic fertilization and embryo quality are important issues for the in vitro production of pig embryos. We hypothesized that oocyte donor (prepubertal gilt vs. sow) affects polyspermy and blastocyst development in vitro and that the sexual maturity of the oocyte donor affects the response to sperm concentration in the fertilization medium. In Exp. 1, oocytes of sows and gilts were mounted and stained 12 h after insemination to provide fertilization data. In Exp. 2, putative embryos were developed in vitro to 144 h post-insemination before mounting. In both experiments, cumulus-oocyte complexes (COC) were collected from ovaries of prepubertal gilts and adult sows. Sperm were added after maturation of COC for 40 to 44 h. Sperm from two boars at 0.5 to 5.0 x 10(6) sperm/mL was used for insemination. More (P < 0.01) monospermic fertilizations were observed in oocytes derived from gilts than for oocytes from sows. There were fewer (P < 0.02) penetrated sperm per fertilized oocyte in oocytes from gilts compared with sows. There were effects of semen donor (boar) on the percentage of monospermic (P < 0.01) and polyspermic (P < 0.002) fertilizations, and on the number of penetrated sperm/fertilized oocyte (P < 0.02). In Exp. 2, cleavage and blastocyst formation was evaluated at 2 and 6 d postinsemination, respectively. More (P < 0.001) blastocysts developed from sow-derived oocytes than from gilt-derived oocytes. More (P < 0.05) total cells per blastocyst were observed in embryos from sow-derived oocytes than from gilt-derived oocytes. Semen donor affected the percentage of oocytes cleaving (P < 0.02), and a boar x sperm concentration interaction affected (P < 0.05) the incidence of blastocyt formation. Results indicate that sexual maturity of the donor is not responsible for the high incidence of polyspermy in porcine in vitro fertilization. However, blastocyst development is improved by the use of oocytes from sows rather than from prepubertal gilts.     

CASA Assessment of Kinematic Parameters of Ram Spermatozoa and their Relationship to Migration Efficiency in Ruminant Cervical Mucus

Sperm motility is an indicator of male fertility because of its importance for sperm migration through the female genital tract and for gamete interaction at fertilization. This study analyses the relationship between computer assisted semen analysis (CASA) motility patterns and sperm migration of rams in ruminant cervical mucus. In experiment 1, spermatozoa extended with sperm analysis medium (SAM) and seminal plasma were compared in terms of motility. In experiment 2, 56 semen samples were collected either with artificial vagina (AV) or electroejaculator to be compared in terms of motility performance. In experiment 3, 104 ejaculates collected by AV from 26 males were analysed via the CASA system to characterize their motility patterns. In experiment 4, ejaculates from pairs of rams (20 rams in total) were simultaneously assessed for mucus migration (ovine, caprine, bovine) and motility patterns to evaluate the correlations between both parameters. Semen collected by AV and extended in SAM allows the most reliable assessment for sperm motility. Ram spermatozoa move fast and follow a linear trajectory compared with other ruminants. Continuous line velocity (VCL) and average path velocity (VAP) are the only sperm kinematic parameters that presented significant positive correlations with the ability to migrate in sheep cervical mucus (p < 0.05). Continuous line velocity, VAP, straight line velocity and linearity are highly significantly related with migration efficiency in goat cervical mucus (p < 0.01) and only lateral head displacement is negatively related to sperm migration in bovine cervical mucus (p < 0.05). These results suggest that specific kinematic parameters confer the ability of spermatozoa to colonize and migrate through epithelial mucus with different rheological properties.     

The effect of time of artificial insemination on fertilization status and embryo quality in superovulated cows

Thirty nonlactating Holstein cows were superovulated to determine the effect of artificial insemination time on fertilization status and embryo quality. During the luteal phase of the estrous cycle, cows were administered 38 mg FSH-P in a 4-d descending dose regimen. Luteolysis was induced with two injections of prostaglandin on the last day of FSH-P treatment. All cows were continuously monitored for behavioral estrus using the HeatWatch estrus detection system. All cows were inseminated once with one .5-mL straw (50 x 10(6) sperm) at either 0 (n = 10), 12 (n = 10), or 24 h (n = 10) after the first standing event. The elapsed time (mean +/- SD) from the first prostaglandin dose to the first standing event was 39.4 h +/- 7.7 h. The (mean +/- SD) duration of behavioral estrus was 13.2 h +/- 4.1 h. The (mean +/- SD) number of standing events was 27 +/- 17. Five hundred twenty-nine embryos and ova were recovered nonsurgically 6 d after insemination. Fertilization rates were 29 (0 h), 60 (12 h), and 81% (24 h) (P < .01). Percentages of excellent and good, fair and poor, and degenerate embryos were not different (P > .05). Percentages of embryos with accessory sperm were 5 (0 h), 8 (12 h), and 41 (24 h) and differed between the 0 and 24 h and the 12 and 24 h inseminations (P < .01). Artificial insemination of superovulated, nonlactating Holstein cattle 24 h after onset of estrus increased fertilization rate and percentage of embryos with accessory sperm compared with insemination at 0 or 12 h after onset of estrus. Embryo quality was not affected by time of insemination.     

Relation of heat resistance tests and sperm survival to pregnancy and fertility in sows

Sperm of 28 boars of the Landrace breed was evaluated with the help of the thermoresistance test (TRT). The principle of the TRT is that the sample of fresh sperm is left in a 38 degrees C bath for 120 min and after that time the activity of the sperm is evaluated. 308 ejaculates were evaluated in this way and 1342 sows were inseminated by these ejaculates after a short storage. The highest conception rate was stated after insemination with the sperm whose activity was above 50% at the end of the TRT. 366 of 400 sows became pregnant, i.e. 82.43%, whereas out of 370 sows inseminated with sperm whose activity was 0-30% at the end of the TRT only 269 conceived, i.e. 72.70%; difference was statistically highly significant. No statistically significant difference was recorded between the fertility (number of piglets per one litter) and the TRT results. A significant dependence was observed between the sperm survival after 72 hours and conception (P less than 0.05). The same significant relation was found between survival within 72 hours and TRT in fresh ejaculates. This justified the recommendation to introduce the TRT among the test used for evaluation of the quality of fresh boar ejaculate.     

Distribution of Live and Dead Spermatozoa in the Genital Tract of Gilts at Different Times After Insemination

Twenty-four gilts were inseminated pair-wise with live or dead spermatozoa from the same ejaculate. The insemination dose was 100 ml undiluted semen containing, on average, 19×10⁹ spermatozoa. The gilts were slaughtered 1, 2, 6 and 12 h after insemination. The numbers of spermatozoa were counted in the uterus, uterotubal junction and in four equally long segments of the oviduct, called I–IV, with a haemocytometer. IV was adjacent to the uterotubal junction. The numbers of spermatozoa recovered in the uterus diminished significantly during the first 12 h after insemination. From gilts inseminated with live spermatozoa more spermatozoa were recovered in the uterotubal junction than from gilts inseminated with dead spermatozoa. Two h after insemination spermatozoa were recovered in all oviducts. Significantly more live than dead spermatozoa were recovered in Segments III and IV of the oviduct, regardless of time. In gilts inseminated with live spermatozoa the sperm count in Segment I varied significantly with time, being hiigest 2 h after insemination. At 6 and 12 h there were no distinct differences in the distribution of live spermatozoa between the various oviduct segments. The numbers of spermatozoa recovered in the oviduct were at these times apparently related to the sperm depots in the uterotubal junction.     

Hysteroscopic insemination of mares with low numbers of nonsorted or flow sorted spermatozoa

The objectives of this study were 1) to compare pregnancy rates resulting from 2 methods of insemination using low sperm numbers and 2) to compare pregnancy rates resulting from hysteroscopic insemination of 5 x 106 nonsorted and 5 x 106 spermatozoa sorted for X- and Y-chromosome-bearing populations (flow sorted). Semen was collected with an artificial vagina from 2 stallions of known acceptable fertility. Oestrus was synchronised (June to July) in 40 mares, age 3-10 years, by administering 10 ml altrenogest orally for 10 consecutive days, followed by 250 microg cloprostenol i.m. on Day 11. All mares were given 3000 iu hCG i.v. at the time of insemination to induce ovulation. Mares were assigned randomly to 1 of 3 treatment groups: mares in Treatment 1 (n = 10) were inseminated with 5 x 10(6) spermatozoa deposited deep into the uterine horn with the aid of ultrasonography. Mares in Treatment 2 (n = 10) were inseminated with 5 x 10(6) spermatozoa deposited onto the uterotubal junction papilla via hysteroscopic insemination. Mares in Treatment 3 (n = 20) were inseminated using the hysteroscopic technique with 5 x 10(6) flow sorted spermatozoa. Spermatozoa were stained with Hoechst 33342 and sorted into X- and Y-chromosome-bearing populations based on DNA content using an SX MoFlo sperm sorter. Pregnancy was determined ultrasonographically at 16 days postovulation. Hysteroscopic insemination resulted in more pregnancies (5/10 = 50%) than did the ultrasound-guided technique (0/10 = 0%; P<0.05) when nonsorted sperm were inseminated. Pregnancy rates were not significantly lower (P>0.05) when hysteroscopic insemination was used for sorted (5/20 = 25%) and nonsorted spermatozoa (5/10 = 50%). Therefore, hysteroscopic insemination of low numbers of flow sorted stallion spermatozoa resulted in reasonable pregnancy rates.     

黑白花公牛冻精受精力的检测—穿透仓鼠卵的试验

本试验用SPA法(Sperm Penetration Assay)即精子穿透分析法对8头黑白花公牛冻精的受精力进行检测,以预测各公牛精液受精力的水平。结果:每卵表面平均精子数为3.2～9.5个,每卵内精子数为1.6～3.4个,被穿透的卵子比例即穿透率为34.9～65.7％,具雄原核的卵子比例即原核卵率为16.0～32.4％。这些数值与各公牛冻精授精500多头母牛的效果作对照,8头公牛平均80(70～90)天的不返情率为56％(45.8～71.1％),不返情率与雄原核卵率,穿透率呈高度正相关,相关系数分别为 0.82和 0.92。 另一组试验结果表明:授精时间对穿透率特别是对雄原核形成有很大影响。授精2小时后观察,没有雄原核形成,只有附着于卵子表面的精子;授精4小时后,精子已通过卵黄膜进入卵内,并开始形成雄原核;授精6小时后形成发育良好的雄原核。 这些观察结果有助于利用这项技术在生产上对公牛精液的受精力进行客观评定,为选择后备公牛提供重要的生理指标,特别是对评定冷冻精液的品质具有一定的参考价值。并为精子顶体反应、受精机理、体外受精、遗传学和胚胎学等领域的研究提供新的手段。     

Distribution of Spermatozoa and Embryos in the Female Reproductive Tract after Unilateral Deep Intra Uterine Insemination in the Pig

The present study was performed to investigate the number of either the spermatozoa or the embryos in the reproductive tracts of sows after unilateral, deep, intra uterine insemination (DIUI). Two experiments were conducted, 10 sows were used in experiment I and eight sows were used in experiment II. Transrectal ultrasonography was used to examine the time when ovulation took place in relation to oestrus behaviour. The sows were inseminated with a single dose of diluted fresh semen 6-8 h prior to expected ovulation, during the second oestrus after weaning. In experimental I, five sows were inseminated by a conventional artificial insemination (AI) technique using 100 ml of diluted fresh semen, containing 3000 x 10(6) motile spermatozoa and five sows were inseminated by the DIUI technique with 5 ml of diluted fresh semen, containing 150 x 10(6) motile spermatozoa. The sows were anesthetized and ovario-hysterectomized approximately 24 h after insemination. The oviducts and the uterine horns on each side of the reproductive tracts were divided into seven segments, namely ampulla, cranial isthmus, caudal isthmus, utero-tubal junction (UTJ), cranial uterine horn, middle uterine horn and caudal uterine horn. Each segment of the reproductive tracts was flushed with Beltsville thawing solution (BTS) through the lumen. The total number of spermatozoa in the flushing from each segment were determined. In experimental II, eight sows were inseminated by the DIUI technique using 5.0 ml diluted fresh semen containing 150 x 10(6) motile spermatozoa. The sows were anesthetized 61.1 +/- 12 h after insemination (48-72 h) and the embryos were flushed from the oviduct through the proximal part of the uterine horn. It was revealed that, in experimental I, the spermatozoa were recovered from both sides of the reproductive tract in the AI-group, and from unilateral side of the reproductive tract in the DIUI-group (three sows from the left and two sows from the right sides). The number of spermatozoa recovered from the reproductive tracts was higher in the AI- than the DIUI-group (p < 0.001). In experiment II, fertilization occurred in five of eight sows (62.5%) after DIUI. The number of ova that ovulated were 16.4 +/- 2.6 per sow and the embryos numbering 11.4 +/- 2.3 per sow were recovered from both sides of the reproductive tract. In conclusion, the spermatozoa given by DIUI could be recovered from only one side of the reproductive tract of sows at approximately 24 h after DIUI via the flushing technique. However, embryos were found in both sides of the oviducts and the proximal part of the uterine horns 48-72 h after insemination, indicating that the fertilization occurred in both sides of the oviducts.     

Spiramycin concentrations in plasma and genital-tract secretions after intravenous administration in the ewe

Uterine infections are associated with reduced fertility in ruminant species. Spiramycin is a macrolide antibiotic potentially active against most of the microorganisms isolated from secretions of infected genital tracts. The present work investigated the ability of systemically administered spiramycin to enter genital secretions, by determining the disposition kinetics of the antibiotic in both plasma and uterine genital secretions. Five healthy ovariectomized ewes were given a single intravenous (i.v.) injection of spiramycin, at a dose of 20 mg/kg. Plasma and genital secretion samples were collected at predetermined intervals for 5 days post-injection. Blood was collected from the jugular vein while mucus was obtained by inserting polyurethane sponges into the vagina. The spiramycin concentration peak in genital-tract secretions was obtained 2.53 +/- 0.63 h after the i.v. administration. The mean residence time was significantly longer (P less than 0.01) in the mucus (18.31 +/- 3.24 h) than in plasma (6.99 +/- 2.53 h). An average mucus to plasma ratio of 7.87 +/- 3.00 was calculated from the area under concentration-time curves covering the period under study. These data indicate that after systemic administration to ewes, spiramycin is rapidly found in genital-tract secretions, at concentrations which are sufficiently high and persistent to suggest its use in the treatment of post-partum uterine infections.     

Artificial insemination with frozen epididymal sperm in cats

Artificial insemination with frozen cauda epididymal sperm was performed in cats. Sperm were transmigrated from the epididymides in 10 male cats. The mean sperm motility and viability were 67% and 82.5%, respectively, and 11.6 x 10(7) sperm were recovered. The mean sperm motility after thawing was 24.0%. Eleven female cats received unilateral intrauterine insemination of 5 x 10(7) sperm, and the conception rate was 27.3% (3/11). This was the first case of conception obtained with frozen epididymal sperm in cats.