Birth of puppies after intrauterine and intratubal insemination with frozen-thawed canine semen

The present study was performed to assess the fertility of frozen-thawed dog semen prepared by freezing with 6% glycerol and thawing at 70℃ for 8 sec, and to evaluate the least number of post-thaw spermatozoa necessary to achieve pregnancy by intrauterine or intratubal artificial insemination. It was found that the pregnancy rate of intrauterine artificial insemination was 100% using 6% glycerol buffer and thawing at 70℃ for 8 sec with 5 × 10⁷ spermatozoa. Even though the pregnancy rate (80%) and the whelping rate (24.5%) in the 5 × 10⁶ spermatozoa inseminated group were lower than those of the 5 × 10⁷ spermatozoa group, conception was confirmed with 5 × 10⁶ spermatozoa. Although the pregnancy rate of intratubal insemination was low (20%) with 4 × 10⁶ spermatozoa, this study is the first report to show the pregnancy rate of intratubal insemination with frozen-thawed ejaculated canine semen. In order to improve the pregnancy rate with intratubal insemination of canine spermatozoa, it is necessary to investigate the optimal insemination site of the uterine tube, the appropriate number of sperm, and the direct effect of buffer on oocytes.     

Efficiency of Repeated In Vivo Oocyte and Embryo Recovery After rhFSH Treatment in Rabbits

This study aims to assess the efficiency of in vivo oocyte and embryo recovery after a recombinant human FSH (rhFSH) treatment in rabbit does. Females were distributed in two experimental groups: donor does were treated with rhFSH (superovulation group) for 3 days prior to artificial insemination (embryo recovery) or ovulation induction (oocyte recovery) and does without treatment remained as the control group. Mature oocytes or embryos were collected with the laparoscopy technique 16 h after ovulation induction (oocytes) or 72 h after artificial insemination (embryos). Up to four recoveries were performed with each doe. Recovery efficiencies differed significantly between embryos (84%) and oocytes (58%). Yet, the recovery rates for the superovulation and control groups did not differ. The rhFSH group was associated with a significant increase (p < 0.05) in the number of oocytes and embryos recovered in comparison with the control group (10.2 ± 1.0 and 14.3 ± 1.2 vs 6.0 ± 2.7 and 8.4 ± 2.3 for oocytes and embryos, respectively). Results from this study indicate that repeated in vivo oocyte and embryo recovery from rhFSH superovulated does maximizes the number of oocytes or embryos collected from the same female.     

Relationship between the sperm count and the fertilization rate of ova ovulated from the contralateral ovary in intrauterine horn insemination in cats

Unilateral intrauterine horn insemination (UIUI) was carried out in cats, and we investigated the fertilization rate of ova ovulated from the contralateral ovary. Various numbers of sperm were used to inseminate the uterine horn on the side where ovulation was inhibited. The rates of conception were 1/11 (9.1%), 2/11 (18.2%), and 5/7 (71.4%) in the 2 x 10(6), 4 x 10(6), and 8 x 10 (6) groups, respectively. Furthermore, the fertilization rate was 70.7% in the 8 x 10(6) group. Thus, ova ovulated from the contralateral ovary were not fertilized or the fertilization rate was low in some cats even when UIUI was performed with a large number of sperm.     

Comparison of fertility on intrauterine insemination between cryopreserved ejaculated and cauda epididymal sperm in dogs

The fertility was compared between ejaculated and cauda epididymal sperm sensitized with prostatic fluid in dog after freeze-thawing using the fertility of ova from the contralateral ovary after injection (2 × 10(8) sperm) into dog uterus on the unilateral ovariectomized side, on the basis of the presence or absence of conception. No significant difference was observed in sperm quality after freeze-thawing between the two groups and conception rates were equivalent and low. Therefore, to achieve a high fertility by intrauterine insemination of canine frozen-thawed ejaculated and cauda epididymal sperm, intrauterine insemination on both sides is recommended, rather than insemination with a lot of sperm of the uterine horn on one side.     

Intrauterine and intravaginal insemination with frozen canine semen using an extender consisting of orvus ES paste-supplemented egg yolk tris-fructose citrate

Our previous report indicated that addition of Orvus ES Paste (OEP) to the extender of frozen canine semen protected acrosomes and maintained sperm motility after thawing. In this study, artificial insemination (AI) using the frozen semen was carried out. The frozen semen was prepared using egg yolk Tris-fructose citrate, and the final concentrations of glycerol and OEP were 7% (v/v) and 0.75% (v/v), respectively. AI was performed during the optimal mating period predicted from the peripheral plasma progesterone level. In intrauterine insemination (IUI), the bitches were laparotomized and 1 x 10(8) spermatozoa were infused into one of the uterine horns. In insemination of non-OEP supplemented semen, 3 x 10(8) spermatozoa were inseminated. In intravaginal insemination (IVI), 10-40 x 10(8) spermatozoa were inseminated. Conception was obtained in nine of 10 bitches (90.0%) that underwent IUI. The number of newborns was from 1 to 7 (mean 3.6 +/- 0.9). The mean ratio of the number of puppies to the number of ovulations in the inseminated uterine horn was 71.8%. The number of puppies did not exceed the number of ovulation in the inseminated uterine horn. Conception using non-OEP supplemented frozen semen was unsuccessful in all four bitches. In IVI, conception was not obtained in any of the six bitches that received insemination of 10 x 10(8) or 40 x 10(8) spermatozoa, but two of three bitches that received insemination of 20 x 10(8) spermatozoa were fertilized. It was shown that a high conception rate can be obtained by IUI using OEP-supplemented frozen canine semen. Developmenmt of a non-surgical method of IUI and a method of freezing canine sperm applicable to IVI is necessary.     

Reproduction of the Female Ferret (Mustela putorius furo)

The domestic ferret is a seasonally polyoestrous species. Females reach puberty at the age of 8–12 months. Females exhibit a constant oestrus between late March and early August if they are not bred. Increasing tumescence in the pink-coloured vulva is a sign of pro-oestrus. Oestrus can persist for up to 5 months, but once ovulation is induced, either pregnancy or pseudopregnancy ensues. Follicular development and atresia overlap in such a manner that there is a recent cohort of follicles available for ovulation whenever copulation might occur. Copulation may last from 15 min to 3 h, the average being 1 h. Ovulation is induced by pressure on the cervix associated with copulation. After sufficient LH release, the pre-ovulatory follicles mature and an average of 12 oocytes (5–13) per female are ovulated 30–40 h after copulation into the ovarian bursa. The ferret oocytes are most capable of being fertilized up to 12 h after ovulation, i.e. 42–52 h after copulation. Ferret oocytes ovulate at the metaphase of the second meiotic division (MII) embedded in three layers of corona radiata cells. Embryos enter the uterus over a period of several days starting on day 5 after mating. Between days 12 and 13 after mating, the embryos have become implanted in the endometrium. Implantation in the ferret is central with rapid invasion of the uterine epithelium by the trophoblast over a broad area that eventually becomes a zonary band of endotheliochorial placenta. Gestation length is 41 days (39–42 days). The domestic ferret gives birth to an average of eight kits (1–18 kits), which weigh 6–12 g at birth.     

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).     

Sperm migration in pigs after deep intrauterine and intraperitoneal insemination

Deep intrauterine insemination in pigs allows sperm deposition only into one uterine horn, but bilateral fertilization of oocytes occurs. How the sperm reach the contralateral oviduct remains disputable. The aim of this experiment was to study possible transperitoneal and/or transuterine sperm migration ways. Follicle growth and ovulation were induced in 24 peripubertal gilts with eCG and hCG 72 h after eCG. Endoscopic intrauterine insemination (IUI) was performed 32 h after hCG with 20 ml of extended semen (60 × 10(6) spermatozoa) as follows: Group CONTROL (n=8) received IUI into the right horn, and the left horn served as non-treated control; Group LIGATURE (n=8) received IUI into the right horn, and the left horn was closed by endoscopic double ligature close to the bifurcation; Group INTRAPERITONEAL (IPI; n=8) received IUI into the right uterine horn, the left horn was closed by double ligature and semen was deposited intraperitoneally at the surface of the left ovary. Genital tracts were removed 65-66 h after hCG, the oviducts were flushed and ova (n=299) were analyzed for fertilization and cleavage. Furthermore, the accessory spermatozoa count/oocyte was graded as 0, without spermatozoa, 1, <5 spermatozoa, 2, 5-50 spermatozoa, 3, 50-100 spermatozoa and 4, >100 spermatozoa. The results indicate that low dose IUI into one horn provides a lower grade of accessory spermatozoa in the contra-lateral side (1.6 vs. 2.8). No spermatozoa were found in ova flushed from oviducts of the ligated uterine horn, even after intraperitoneal insemination (P<0.05), and no fertilization occurred, respectively. Our results clearly indicate that after low dose IUI into one uterine horn, spermatozoa reach the contralateral oviduct via transuterine migration.     

Distribution and viability of spermatozoa in the canine female genital tract during post-ovulatory oocyte maturation

Background

Unlike other domestic mammals, in which metaphase-II oocytes are ovulated, canine ovulation is characterized by the release of primary oocytes, which may take 12 to up to 36 hours. Further 60 hours are needed for maturation to secondary oocytes which then remain fertile for about 48 hours. Oestrus takes 7 to 10 days on average and may start as early as a week before ovulation. This together with the prolonged process of post-ovulatory oocyte maturation requires an according longevity of spermatozoa in the female genital tract in order to provide a population of fertile sperm when oocytes have matured to fertilizability. Therefore the distribution and viability of spermatozoa in the bitch genital tract was examined during post-ovulatory oocyte maturation.

Methods

Thirteen beagle bitches were inseminated on the day of sonographically verified ovulation with pooled semen of two beagle dogs containing one billion progressively motile spermatozoa. Ovariohysterectomy was performed two days later (group 1, n = 6) and four days later (group 2, n = 7). The oviduct and uterine horn of one side were flushed separately and the flushing’s were checked for the presence of gametes. The oviducts including the utero-tubal junction and the uterine horns, both the flushed and unflushed, were histologically examined for sperm distribution.

Results

The total number of spermatozoa recovered by flushing was low and evaluation of viability was limited. Prophase-I oocytes were collected from oviduct flushing in group 1, whereas unfertilized metaphase-II oocytes were detected in group 2. From day 2 to day 4 after ovulation a significant decrease in the percentage of glands containing sperm (P<0.05) and a marked reduction of the mean sperm number in uterine horn glands were observed. A concomitant diminution of spermatozoa was indicated in the utero-tubal junction accompanied by a slight increase in sperm numbers in the mid oviduct.

Conclusions

Oocyte maturation to metaphase-II stage is accompanied by a continuous sperm detachment and elimination in the uterine horns. Entrance of spermatozoa into the caudal oviduct seems to be steadily controlled by the utero-tubal junction thus providing a selected sperm population to be shifted towards the site of fertilization when oocyte maturation is completed.     

Precocious estrus and reproductive ability induced by PG 600 in prepuberal gilts

A total of 29 SPF Large White prepuberal gilts (mean age 152 days at treatment) were examined for estrous and ovulatory responses after PG 600 treatment. After treatment, 85.2% of the gilts showed standing estrus within 6 days. Whereas the treatment-to-estrus interval and duration were 3.7 and 1.9 days respectively. As ovulation occurred on Day 5 to 6, appropriate timing of artificial insemination would be about 4 days after treatment. Fertility of gilts revealed to be excellent, giving rise to a high percentage of normal embryos, 85.3%. Meanwhile, development and growth of fetuses were mostly normal. Other reproductive performances recorded were: mean litter size 6.8; mean birth weight 1.26 kg; weaning-to-return estrus interval 5 to 8 days. In conclusion, PG 600 was found to be useful in inducing fertile estrus in prepuberal gilts, a result which will be of interest for commercial pig farms.     

Induction of Parturition with Aglepristone in Various Sized Bitches of Different Breeds.

The objective of this study was to confirm in various breeds of dogs the efficacy and safety of a parturition induction treatment described to be successful in Beagle dogs. Parturition was induced in seven various sized pregnant bitches of different breeds, with 15 mg aglepristone per kg at day 59–61 post-estimated ovulation day, followed 24 h later by 0.15 IU oxytocin per kg subcutaneous injections every 2 h. Two bitches were small-sized bitches (<10 kg), three bitches were large-sized bitches (30–40 kg) and two bitches were giant bitches (>40 kg). The results were compared to a control group (n = 6), in which bitches underwent a natural delivery in the same environmental conditions as the induced group. In the induced group, parturition was successfully induced in 7/7 bitches. The first pup in a litter was born on average 25.9 ± 3.29 h after aglepristone administration (21–30 h). Two of seven bitches from the small-sized group delivered some of their pups before the first administration of oxytocin. The mean duration of parturition was 9.6 ± 5.4 h vs 8.0 ± 4.8 h in the control group. The mean interval between two successive pups being delivered was 115.6 ± 82.8 min (34–265) vs 68.8 ± 24.5 min in the control group (p < 0.03). The mean weight at parturition did not differ significantly between the two groups. One litter of four Yorkshire Terrier pups in the induced group were premature at the time of birth and died between 19 and 29 h post-delivery. This study, although on a very limited number of dogs, confirms the efficacy of the aglepristone/oxytocin protocol to induce parturition in dogs.     

Consequences of Nitric Oxide Synthase Inhibition During Bovine Oocyte Maturation on Meiosis and Embryo Development

The importance of nitric oxide synthase (NOS) in bovine oocyte maturation was investigated. Oocytes were in vitro matured with the NOS inhibitor N^w- l -nitro-arginine methyl-ester (10⁻⁷, 10⁻⁵ and 10⁻³  m l -NAME) and metaphase II (MII) rates and embryo development and quality were assessed. The effect of l -NAME (10⁻⁷  m ) during pre-maturation and/or maturation on embryo development and quality was also assessed. l -NAME decreased MII rates (78–82%, p < 0.05) when compared with controls without l -NAME (96%). Cleavage (77–88%, p > 0.05), Day 7 blastocyst rates (34–42%, p > 0.05) and total cell numbers in blastocysts were similar for all groups (146–171 cells, p > 0.05). Day 8 blastocyst TUNEL positive cells (3–4 cells) increased with l -NAME treatment (p < 0.05). For oocytes cultured with l -NAME during pre-maturation and/or maturation, Day 8 blastocyst development (26–34%) and Day 9 hatching rates (15–22%) were similar (p > 0.05) to controls pre-matured and matured without NOS inhibition (33 and 18%, respectively), while total cell numbers (Day 9 hatched blastocysts) increased (264–324 cells, p < 0.05) when compared with the controls (191 cells). TUNEL positive cells increased when NOS was inhibited only during the maturation period (8 cells, p < 0.05) when compared with the other groups (3–4 cells). NO may be involved in meiosis progression to MII and its deficiency during maturation increases apoptosis in embryos produced in vitro . Nitric oxide synthase inhibition during pre-maturation and/or maturation affects embryo quality.     

Follicular Dynamics, Ovulation and Conception Rates in Bitches

Real-time ultrasound imaging was used in a clinical study to estimate the number of follicles of different sizes, ovulation and conception rates, and to study follicle dynamics following oestrus-induction of bitches.
Follicles were identified during late anoestrus (between 100 and 60 days prior to the pre-ovulatory LH surge) and there appeared to be a shift in the population from small follicles (1–3 mm in diameter) to large follicles (>4 mm diameter) approximately 2 days prior to the LH surge. Corpora lutea could be reliably identified although the majority were cavitated. High ovulation rates (97–100%) and pregnant rates (86–100%) were detected, and although the conception rate was approximately 70% it varied between 8 and 92%. Within the narrow range of the clinical population studied there were trends relating age to reproduction performance. Oestrus induction with a gonadotrophin regime appeared to result in large numbers of small follicles that did not ovulate, whilst when using cabergoline the number of small and large follicels and the number of copora lutea were similar to those of control cycles.     

Effect of Modified Eros Centre on Some Reproductive Parameters of the Sow

The effect of a modified eros centre on weaning to oestrus interval, follicle size, ovulation and farrowing rate and total born litter size was investigated. In modified eros centre 94.4% and in group housing 79.1% of the sows (p < 0.01) expressed oestrus within 10 days post‐weaning. Weaning to oestrus interval was shorter (p < 0.001) for sows kept in modified eros centre. The interval from onset of oestrus to the time of ovulation was longer for sows in group housing (p=0.05). The time of ovulation was negatively correlated (r=?0.50) with the interval from weaning to oestrus (p=0.005). The time of ovulation after onset of oestrus was significantly (p < 0.05) shorter for sows expressing oestrus within 2–4 days of weaning, compared with the animals that expressed oestrus between days 5 and 6 post‐weaning and was shortest for sows expressing oestrus after day 6 post‐weaning. Farrowing rate was not affected by a modified eros centre. Litter size tended to be smaller in group‐housed weaned sows (p=0.10). The timing of last artificial insemination relative to time of ovulation did not affect litter size (p > 0.10). The implication of these results is that a modified eros centre may improve some of the post‐weaning oestrous parameters of the sow.     

Supplementation with Calcium Salts of Linoleic and trans-Octadecenoic Acids Improves Fertility of Lactating Dairy Cows

Objectives were to evaluate effects of feeding a calcium salt rich in linoleic and trans -octadecenoic acids (LTFA) on synthesis of prostaglandin F_2α based on its metabolite (PGFM), uterine involution and pregnancy rates in lactating dairy cows. Five hundred and eleven Holstein cows were blocked according to parity, body condition score and milk yield in the previous lactation. Primiparous and multiparous cows were randomly assigned to one of the two treatments consisting of calcium salt (2% diet dry matter) of either palm oil (PO) or LTFA from 25 days prepartum to 80 days of lactation. Cows were time-inseminated at 70 ± 3 days postpartum. Feeding LTFA tended (p = 0.08) to decrease the incidence of puerperal metritis (15.1% vs 8.8%). Primiparous cows supplemented with LTFA showed larger increase in plasma PGFM concentration at day 1 postpartum (17018 vs 6897 p m ). Pregnancy rate after first insemination tended (p = 0.07) to be greater at 27 days after insemination (37.9% vs 28.6%), and was greater (p = 0.05) at 41 days after insemination (35.5% vs 25.8%) for cows fed LTFA compared with PO. These results indicate that unsaturated fatty acids fed in a rumen inert form have the potential to modulate reproductive events and improve pregnancy rates in lactating dairy cows.     

Direct and correlated responses to two-stage selection for ovulation rate and number of fully formed pigs at birth in swine

Our objectives were to estimate responses and genetic parameters for ovulation rate, number of fully formed pigs at birth, and other production traits following two-stage selection for increased ovulation rate and number of fully formed pigs. Eight generations of selection were practiced in each of two lines. One selection line was derived from a line that previously selected eight generations for an index to increase ovulation rate and embryonic survival (the IOL pigs). The other selection line was derived from the unselected control line of the index selection experiment (the COL pigs). The control line (C) was continued with random selection. Due to previous selection, Line IOL had greater ovulation rate (4.24 +/- 0.38 and 4.14 +/- 0.29 ova) and litter size (1.97 +/- 0.39 and 1.06 +/- 0.38 pigs) at Generation 0 of two-stage selection than did Lines COL and C. In Stage 1, all gilts from 50% of the largest litters were retained. Approximately 50% of them were selected for ovulation rate in Stage 2. Gilts selected for ovulation rate were mated to boars selected from the upper one-third of the litters for litter size. At Generations 7 and 8, differences in mean EBV for ovulation rate and litter size between Lines IOL and C were 6.20 +/- 0.29 ova and 4.66 +/- 0.38 pigs; differences between Lines COL and C were 2.26 +/- 0.29 ova and 2.79 +/- 0.39 pigs; and differences between Lines IOL and COL were 3.94 +/- 0.26 ova and 1.86 +/- 0.39 pigs. Regressions of line mean EBV on generation number were 0.27 +/- 0.07 ova and 0.35 +/- 0.06 pigs in Line IOL; 0.30 +/- 0.06 ova and 0.29 +/- 0.05 pigs in Line COL; and 0.01 +/- 0.07 ova and 0.02 +/- 0.05 pigs in Line C. Correlated responses were decreased age at puberty and increased number of pigs born alive, number of mummified pigs, prenatal loss, and individual and litter birth weight. Two-stage selection for ovulation rate and number of pigs per litter is a promising procedure to improve litter size in swine.     

The ovarian response and fertility in cows with acidosis induced by acetic acid]

The effects of acetic acid administered at an amount of 300 to 600 g (5 to 10 mol) to the rumen of breeding cows, were investigated on ovulation, conception and progesterone levels in the blood and milk of cows with cloprostenol-induced (Oestrophan Spofa) oestrus at a dose of 500 micrograms i.m. In the group of 15 cows exposed to the acetic acid load five cows got in calf after the first insemination (33.3%), and 12 cows (80.0%) after all inseminations in 37.6 days after cloprostenol administration, with the insemination index 1.67 (Tab. I). In the control group (five cows) four cows (80.0%) got in calf after the first insemination, in total all five breeding cows got in calf in 20.6 days after cloprostenol administration, with the insemination index 1.2. In the experimental group of 15 cows a clinical examination of ovaries on day 7 after insemination revealed ovulation disorders in eight cows, that means in 53.3% of the animals (Tab. II). No ovulation disorders were observed in the control group of five cows. Progesterone levels in the blood showed high variability (Tab. III). In the group of cows administered acetic acid they were by more than a half lower (1.49; 0.67; 1.53 per ml) on days 7, 14 and 21 after insemination in comparison with the control group (3.35; 2.5; 3.38 ng per ml). The average progesterone levels in milk (Tab. IV) were 1.27 and 1.53 on day 7, 6.74 and 7.27 on day 14 and 3.52 and 11.85 ng per ml on day 21, respectively, the higher values apply to the control. It was not possible to evaluate reliably from the progesterone levels in the blood and milk if ovulation took place and if the corpus luteum was developing (Tab. V and VI). The clinical control of ovaries on days 7 and 8 after oestrus and insemination was more reliable to determine the ovulation disorders than the progesterone determination in the blood and milk of cows.     

NORMAL CANINE PEDIATRIC GASTROINTESTINAL ULTRASONOGRAPHY

The normal sonographic appearance of the adult canine gastrointestinal tract has been described. Interpretation of abdominal ultrasonographic findings in puppies is difficult due to the lack of information on normal ultrasonographic findings. The gastrointestinal tract, jejunal lymph node size and the presence and appearance of abdominal fluid were investigated in 23 normal, 7–12-week-old Beagle puppies. The duodenal wall thickness was greater than in other parts of the gastrointestinal tract (mean 3.8 ± standard deviation [SD] 5 mm, range 3.2–4.8 mm). The mean stomach wall thickness was 2.7 ± SD 0.4 mm (range 2.2–3.7 mm), the mean jejunal wall thickness was 2.5 ± SD 0.5 mm (range 1.2–3.4 mm), and the mean colonic wall thickness was 1.3 ± SD 0.3 mm (range 0.7–2.0 mm). In addition, mean duodenal and jejunal mucosal layer thicknesses measured 2.7 ± SD 0.5 mm (range 2.0–3.8 mm) and 1.5 ± SD 0.4 mm (range 0.6–2.5 mm), respectively. Homogenous, hypoechoic jejunal lymph nodes were easily found and the mean thickness was 7.1 ± SD 2.2 mm (range 1.5–12.5 mm). A mild amount of anechoic free peritoneal fluid was present in all puppies.     

Ultrastructural Study of the Canine Zona Pellucida Surface During In Vitro Maturation

The aim of the present study was to compare the ultrastructure of the surface of the zona pellucida (ZP) of immature and in vitro matured dog oocytes using scanning electron microscopy (SEM). Bitch oocytes were collected after ovariohysterectomy; the ovaries were sliced and the released cumulus–oocyte complexes (COCs) were washed with phosphate buffered saline (PBS). The selected COCs were randomly allocated into three groups, two groups were processed after in vitro maturation at both 72 and 96 h and a third group was processed immediately at immature state in PBS medium. After that, oocytes were fixed, critical point dried and viewed by using SEM. The diameters of the outer holes of the ZP were measured on a total of 93 oocytes; the results were analyzed with anova . The mean diameters of holes were different between groups (p < 0.05): 0.69 ± 0.12, 1.56 ± 0.19 and 1.42 ± 0.27 μm, for immature and in vitro matured oocytes for 72 and 96 h, respectively. The difference in the hole sizes between immature and in vitro matured canine oocytes indicates that the ZP surface is related to oocyte maturity in canines.     

Reproductive response of ewes to immunization with Fecundin (ovandrotone)

Ewes of five crossbred genotypes (total of 221 ewes) were immunized with a commercial preparation of androstenedione-7HSA in DEAE dextran adjuvant in a 2-yr trial. In yr 1 ewes were allocated within genotype to a 2 x 2 application of immunization and premating flushing. In yr 2 they were allocated within previous Fecundin treatment into either Fecundin-treated or control groups. First-time immunization with Fecundin increased ovulation rate by .70 ova/ewe in yr 1 and by .85 ova/ewe in yr 2 (P less than .05). Ewes treated in both years produced fewer ova than those treated in the 2nd yr only (P less than .05), but they did not differ in litter size. Although Fecundin reduced conception to first mating, the overall conception rate was not affected. Litter size of treated ewes was increased by .19 and .38 lambs/ewe lambing (P less than .05) in yr 1 and yr 2, respectively. Fecundin treatment in yr 1 produced a carry-over effect in yr 2 by increasing litter size by .26 lamb/ewe (P less than .05) while reducing conception to first mating. The litter size of ewes with two ovulations was reduced by Fecundin treatment in yr 1 (P less than .05) but showed no consistent pattern in yr 2. Overall, ewes with two ovulations produced .7 to .8 more lambs than ewes with one ovulation, and ewes with three ovulations produced .4 more lambs than those with two ovulations. Ewes of different genotypes did not differ in their response to Fecundin.