Failure of isoxsuprine HCl to increase sheep fertility after cervical versus laparoscopic AI using chilled semen at different durations

The present study aimed to investigate the fertility of ewes artificially inseminated with three different methods using a synthetic semen extender, AndroMed. The three methods of artificial insemination (AI) were cervical AI with fresh-diluted or frozen-diluted semen at observed estrus, and an intrauterine AI with frozen-thawed semen. A total of 80 ewes were treated with a controlled internal drug release (CIDR) containing 0.3 g progesterone per device for 12 days. In Experiment 1 (26 Suffolk ewes), superovulation was induced with 20 mg follicle-stimulating hormone and 250 IU equine chorionic gonadotropin (eCG) two days and one day before CIDR removal, respectively, during the non-breeding season. In Experiment 2 (54 Suffolk and Suffolk crossbred ewes), an intramuscular injection of 500 IU eCG was administered one day before CIDR removal to synchronize estrus and ovulation during the breeding season. In Experiment 1, fresh-diluted or frozen-thawed semen was deposited into the cervical orifice after estrus detection, and an intrauterine AI with frozen-thawed semen was performed by laparoscopy at a fixed-time basis without estrus detection. Embryos were recovered by uterine flushing 6 days after AI, and the rates of recovered, fertilized (cleaved) ova and embryos at the morula or blastocyst stage were compared among the three AI methods. In Experiment 2, the pregnancy rates after the three AI methods were compared. In Experiment 1, the rates of recovered ova were not significantly different among the three AI methods (52.5-56.7%). The rate of fertilized ova (81.0%) by laparoscopic AI with frozen-thawed semen was significantly higher compared with cervical AI of fresh-diluted (25.5%) or frozen-thawed (3.5%) semen, but the rate of embryos at the morula or blastocyst stage (17.6%) was significantly lower than that of the cervical AI with fresh-diluted semen (69.2%). The rates of ewes yielding fertilized ova were not significantly different among the three groups (44.4, 11.1 and 62.5% for cervical AI with fresh-diluted and frozen-thawed semen and intrauterine AI with frozen-thawed semen). In Experiment 2, the pregnancy rate of ewes intrauterinally inseminated with frozen-thawed semen (72.2%) was significantly higher than those of ewes inseminated cervically with fresh-diluted (5.5%) or frozen-thawed (0.0%) semen. The present results showed that acceptable fertilization and pregnancy rates could be obtained by an intrauterine AI with frozen-thawed semen using a synthetic semen extender (AndroMed), but not sufficient by the cervical AI with either fresh or frozen semen.

Transcervical artificial insemination of Australian Merino ewes with frozen-thawed semen

The present study was conducted in crossbred ewes to compare the efficiency of different artificial insemination techniques with natural tupping following fixed time artificial insemination using fresh semen. Crossbred ewes (n=29) were randomly divided into three groups with 10 animals in natural tupping (NT) and laparoscopic artificial insemination (LAI) groups and nine animals in cervical artificial insemination (CAI) group. Ewes in all the treatment groups were subjected to the same estrus induction protocol: insertion of intravaginal progesterone sponges for 10 days followed by an intramuscular injection of 500 IU eCG at the time of sponge withdrawal. Immediately after removal of sponges, animals of NT group were kept with proven breeding rams up to 72 h. Fixed time cervical or laparoscopic insemination was done at 48 h after sponge removal. The insemination was repeated 12 h later in CAI group. The pregnancy and lambing rates were significantly higher in NT group (90%, 90%), than CAI group (55.5%, 55.5%) and LAI group (20%, 10%). The prolificacy rate was significantly higher in CAI group (180%) than NT group (111.1%) and LAI group (100%). The serum progesterone concentration was higher in pregnant ewes on day 10, day 17 and day 35 than non-pregnant ewes. The fixed time cervical insemination following intravaginal progesterone sponges for 10 day+eCG protocol resulted in better pregnancy rate and prolificacy rate in crossbred ewes during breeding season.

This study aimed to determine the cost of lamb produced from different artificial insemination te chniques in Santa Ines sheep. The experiment was conducted on a 3 x 2 factorial arrangement, on a randomized block design. One hundred and two females were divided in three groups: cervical artificial insemination with fresh semen - AI or frozen-thawed semen - AIFT or laparoscopic artificial insemination - LAI. On the determination of the total cost of the produced lamb, variable and fixed costs were considered. The costs from the labor suffered a great technical variation among the techniques as 9.4% in IAC, 17.7% in IAF e 26.4% in IAL. The expenses with hormonal treatment were higher when compared to other components corresponding to 65.8, 54.0 and 28.7% to IAF, AIFT and LAI, respectively. The total cost of the lamb changed according to the artificial insemination technique used, as the IAF cost was 4.4 times lower than IAC cost and 2.2 times lower than IAL cost. The IAF presents lower cost per produced lamb when compared to the other techniques. The IAC, due to low fertility rates achieved, was economically unfeasible, while the IA by laparoscopy, in spite of the satisfactory fertility rate, contributes to increase the cost of lamb produced.

Factors influencing the success of vaginal and laparoscopic artificial insemination in churra ewes: a field assay

Laparoscopic insemination with frozen-thawed semen is currently used for planned matings in the Sarda breeding programme. In order to find a fast and less intrusive artificial insemination (AI) method that could replace laparoscopic insemination, a field comparison of laparoscopic and transcervical techniques was carried out on 200 mature Sarda ewes. After AI, ewes were assigned to teaser and fertile rams for 2 months. Return rates and cumulative (AI + natural mating) lambing rates were recorded over three subsequent 23-day periods. Lambing rates to AI were significantly different (P < 0·01), and were 62% and 7% respectively for laparoscopic and transcervical AI. Cumulative lambing rates after two further 23-day periods of natural mating were no longer significantly different (P > 0·05) and reached 82% and 74% respectively. Ewes with body condition scores at AI higher than 2·75 showed better overall reproductive performance, but not higher pregnancy rate to AI. Plasma cortisol concentrations, sampled twice, before and after AI, were higher (P < 0·01) in the last sample, suggesting a stress response to insemination. Cortisol levels after AI were lower (P < 0·01) for ewes submitted to transcervical rather than laparoscopic insemination (P < 0·01). However, cortisol levels after AI were no greater than those recorded when ewes were restrained in a milking yoke different from that usually employed. Laparoscopic AI was confirmed as the most suitable technique for insemination offrozen semen in the Sarda breeding scheme.

Artificial insemination (AI) is a valuable tool for infusing genetic variation into stagnant populations, especially using frozen-thawed spermatozoa, and for propagating physically or behaviorally incompatible individuals. Within zoological institutions, AI has received growing interest for genetic management of endangered felids, provided that AI success can be optimized for applied usage. Traditionally, laparoscopic AI in cats involves treatment with equine chorionic gonadotropin (eCG) followed 80-85h later by human CG (hCG) to induce follicular development and ovulation, with subsequent bilateral sperm deposition into the uterine lumen. However, hCG, a large glycoprotein, may remain in circulation for several days post-injection, generating undesirable secondary ovulations. Uterine AI also requires relatively high numbers of spermatozoa to achieve sperm transport through the uterotubal junction and fertilization within the oviduct. Furthermore, sperm recovery from male cats frequently is poor, limiting the number of spermatozoa available for AI. Alterations in the AI protocol, using short-acting porcine luteinizing hormone (pLH) as the ovulatory signal and oviductal AI for sperm deposition, could improve fertilization success while requiring fewer spermatozoa. Our objectives in this study were to assess pregnancy and fertilization success in cats treated with one of two gonadotropin regimens (eCG/hCG vs. eCG/pLH) and inseminated laparoscopically at two sperm deposition sites, the uterus (UT) and the contralateral oviduct (OV). Sixteen female domestic cats were randomly assigned to either eCG (100 IU)/hCG (75 IU) or eCG/pLH (1000 IU) treatment groups. All 16 females ovulated following gonadotropin treatment and were inseminated with low sperm numbers (1x106 motile sperm/site; 5 µl volume) in one uterine horn and one contralateral oviduct using freshly collected semen from a different male for each site. Semen samples were obtained from two males of proven fertility via artificial vagina. Pregnant females were spayed at 20-21 days post-AI and recovered fetuses assessed for paternity using genetic analysis. Comparing gonadotropin regimens, similar numbers of females became pregnant following eCG/hCG (75%, 6/8) versus eCG/pLH (63%, 5/8). The number of corpora lutea (CL) at AI was similar between regimens, but hCG treatment increased the number of CLs at day 20 post-AI. Although hCG and pLH treatments produced similar numbers of normal fetuses, implantation abnormalities (e.g. empty gestational sacs, malformed placentae) were observed in the hCG, but not pLH, group. Overall, 11 (of 16; 69%) females became pregnant (5 cats from OV AI only, 2 from UT AI only, and 4 from both sites). In comparing insemination sites, more fetuses resulted from OV AI (36/49; 73%) than UT AI (13/49; 27%). To assess capacity for term pregnancies, three additional females were treated with eCG/pLH and inseminated in one oviduct and the contralateral uterine horn. All three females became pregnant (2-3 fetuses each) and healthy kittens were produced. In summary, laparoscopic oviductal AI with low sperm numbers in eCG/pLH-treated females resulted in high pregnancy and fertilization percentages in domestic cats. These findings suggest that this technique may have value for propagating endangered nondomestic cat species. Our recent success in using oviductal AI in eCG/pLH-treated ocelots (Leopardus pardalis) to produce a healthy ocelot kitten supports this cross-species applicability. (platform)

Artificial insemination in sheep has vastly increased in popularity as preserved semen is readily available. Laparoscopic artificial insemination (LAI), a surgical procedure, involves deposition of semen directly into the uterus using a laparoscope. We have reviewed LAI, compared benefits to natural breeding and transcervical artificial insemination, described procurement, and provided a cost benefit analysis. LAI is an advanced assisted reproductive technique with several benefits, including higher pregnancy rates requiring fewer sperm per breeding than vaginal or transcervical artificial inseminations. Although LAI involves a higher start-up cost compared to other techniques, it is an economically feasible option. With the continual development of new equipment, techniques, and semen preservation technologies, LAI can enhance herd genetics without investment in expensive genetically superior studs and maintaining them. LAI with an appropriate estrus synchronization protocol results in higher numbers of superior quality lambs per ewe for higher profit margin and higher lambing rates for producers, making LAI a profitable service for a veterinary practice.

Artificial insemination (AI) was the first important biotechnology applied to improve the genetics of farm animals. It allows the rapid and massive diffusion of desirable characteristics of males with high productive potential. We describe the different types of estrus induction and synchronization techniques and the use of the AI with fresh, chilled or frozen semen. Through the adequacy of the protocols of estrus synchronization and AI to the different production systems, the efficient use of reproductive techniques is possible, reaching acceptable pregnancy rates. Summary of reproductive results obtained using cervical and laparoscopic AI are presented.

Reproductive performance in gilts following applications of different insemination doses and techniques

Adult East Freisan crossbred (n = 220) and Chios ewes (n = 105) were divided into four groups and inseminated with chilled semen, which had been stored for 7, 13, 20 or 26 h at 5°C. Unilateral intrauterine insemination (50 x 10 6 spermatozoa in 0.25 ml) was performed with the aid of a laparoscope. Inseminations were carried out 48 – 52 h after pessary removal (30 mg FGA) without detecting estrus. The lambing rates after intrauterine insemination with chilled semen were found to be similar in East Fresian crossbred (40%) and Chios ewes (30%). Intrauterine insemination with chilled semen stored up to 26 h resulted in similar lambing rates; whereas, fertility of Chios ewes tended to decline with increased holding time of chilled semen. From this study, it is concluded that decreasing the storage time of chilled semen at 5°C improves pregnancy in Chios ewes and that East Fresian crossbred ewe’s conception rates to intrauterine insemination with chilled semen was relatively higher than Chios ewes. Key words: Dairy sheep, chilled semen, time of insemination, lambing rate.

Ovarian sensitivity to exogenous gonadotropins was assessed in the cheetah (Acinonyx jubatus) and clouded leopard (Neofelis nebulosa) to help optimize artificial insemination (AI). Eighteen female cheetahs were used on 29 occasions and were given i.m. injections of 100, 200, or 400 IU eCG and 100 or 250 IU hCG 80 h later. Twenty-three female clouded leopards were treated i.m. on 27 occasions with 25, 50, 75, 100, 200, or 400 IU eCG followed 80 h later with 75, 140, or 280 IU hCG. Ovaries were examined laparoscopically at 43-48 h after hCG in cheetahs and 39-50 h in clouded leopards. All gonadotropin dosages stimulated ovarian activity in both species, but ovulation success and corpus luteum (CL) morphology varied (p < 0.05) with treatment. For both species, the highest and intermediate eCG dosages resulted in ovulation in a high proportion (72-100%) of females. The lowest eCG dosage, although capable of stimulating follicular development, compromised ovulation and resulted in few (< 26%) postovulatory females. For each species, small CL (2-4-mm diameter) were observed with the highest and lowest eCG dosage, and large CL (5-8-mm diameter) were associated with intermediate eCG dosages. Aged CL (10-12 mm diameter) were observed in 4 of 23 (17.4%) clouded leopards with no prior male exposure, indicating occasional spontaneous ovulation. Nineteen laparoscopic intrauterine AI procedures were performed in eCG/hCG-treated postovulatory cheetahs. Eighteen AI procedures were conducted in eCG/hCG-treated postovulatory clouded leopards. Six of the 13 cheetahs (46%), all in the 200-IU eCG/100-IU hCG group, became pregnant, in contrast to none of the clouded leopards. This study has revealed differences in ovarian activity in two wild felid species as a result of changes in exogenous gonadotropin dosage. Because of this dose-effect response, this comparative approach is necessary to identify a gonadotropin regimen that can mimic "normalcy." Even then, the relatively high AI success in the cheetah compared to the clouded leopard suggests that factors other than ovarian response can dictate the efficiency of assisted reproduction in this taxon.

Fertility following artificial insemination (AI) programs in sheep is affected by many variables. The aim of this study was to identify extrinsic (year, season), intrinsic (age and breed) and AI procedure related (number of AIs, synchronization protocols, semen preservation method) factors that influence fertility after laparoscopic AI in sheep. Data from 159 laparoscopic AI procedures were categorized according to year (from 2013 to 2017), season (Winter, Spring, Summer and Autumn), breed (Dorper, White Dorper, Texel, Cross-breed), age in months (≤12, 13-24, 25-36, 37-48, ≥49), estrus synchronization protocol (short-term or long-term), cumulative number of AIs (1 to 4) and semen preservation method (frozen or fresh). Frequency analysis using chi-square test was used. Year, age, number of AIs and synchronization protocols did not influence fertility after laparoscopic AI in sheep. Fertility was higher (P&lt;0.05) in sheep inseminated in spring (64.28%) and summer (54.16%) when compared to winter (36.92%) and autumn (32.14%). Dorper breed fertility (58.02%) was higher (P&lt;0.05) than White dorper (25%) and Cross-breed (30.23%). There was a lower (P&lt;0.05) pregnancy rate in sheep inseminated with frozen semen (38.27%) than sheep inseminated with fresh semen (53.84%). In conclusion, season, breed and semen preservation method can influence fertility in sheep after laparoscopic AI.

This study consisted of two experiments. In experiment 1, stress responses of sheep which were restrained either in a laparoscopy cradle or a roll-over cradle were compared. The results of this experiment indicated that restraint in roll-over cradle is less (P &lt; 0·05) stressful than that in a laparoscopy cradle when assessed in terms of the elevation and duration of both the mean heart rate and plasma cortisol responses. Experiment 2 compared the stress responses of sheep subjected to restraint in a laparoscopy cradle, restraint in a laparoscopy cradle with intrauterine artificial insemination (AI) by laparoscopy, minimal restraint with cervical AI or restraint in a roll-over cradle plus foot-trimming. All treatments resulted in significant elevations in both heart rate and plasma cortisol concentrations (F &lt; 0·001). The peak heart rate was significantly (P &lt; 0·05) higher in ewes subjected to cervical AI than in those subjected to intrauterine insemination, with other treatments intermediate. The peak cortisol response did not differ among different treatments. The duration over which both the mean heart rate and -plasma cortisol concentrations remained significantly elevated above pre-treatment concentrations did not differ among treatment groups. The results of this study suggest that while restraint using a laparoscopy cradle is more stressful than that using a rollover cradle, the stress inflicted by intrauterine insemination by laparoscopy itself is no greater than that due to restraint using the laparoscopy cradle alone, cervical AI or the management practice offoot-trimming using a rollover cradle.

Uterine tone influences fertility of Merino ewes following laparoscopic artificial insemination