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Transmission of classical swine fever virus by artificial insemination during the 1997-1998 epidemic in The Netherlands: a descriptive epidemiological study.

In the course of the 1997-1998 CSF epidemic in the Netherlands, two semen collection centres (SCC) became infected. As an eradication strategy for an acute crisis situation, it was concluded that all semen of the boars at the SCCs collected and distributed in the risk period of 28 January to 7 March 1997 was potentially contaminated (suspect semen). As a consequence, a total of 1,680 pig herds, mainly located in the southern part of the Netherlands, were officially declared CSF suspect. The purpose of this study was to investigate whether infection of farms through contaminated semen played a significant role in the CSF epidemic. A total of 123 CSFV infected herds were identified, that had received suspect semen from one or both of the infected SCCs. In 87 out of these 123 infected herds, infection by way of artificial insemination (AI) could be excluded either according to the insemination information or the infection pattern observed. In only 21 herds, infection by way of AI was regarded as possible according to the insemination information and infection pattern. Owing to missing information, no conclusion could be drawn about the possibility of infection of 15 farms by way of AI. Thus, we conclude that at most 36 farms may have been infected through AI during the CSF epidemic in the Netherlands.

Animals↗

Reproductive behaviour in poultry: implications for artificial insemination technology.

1. Reproductive ability requires both endocrine and behavioural components. 2. Most reproductive behaviour is dependent upon the presence of sufficient circulating concentrations of the gonadal steroids, which in turn are synthesised and secreted in individuals who are in good reproductive condition. Mating behaviour patterns are thus not only essential for reproduction, but can provide excellent indices of the reproductive ability of an individual. 3. A number of factors can suppress or enhance reproductive behaviour in poultry, including management practices, flock social interactions, environmental variables, stressors, and disease. 4. Aspects of the regulation of reproductive behaviour and the endocrine control of reproductive processes in the male and in the female are reviewed in this paper. 5. An understanding of the impact of social and environmental stressors on reproductive physiology and behaviour is extremely important, both in order to improve breeding efficiency in natural mating systems and to facilitate the most effective application of artificial insemination technology.

Animals↗

Birth of offspring of pre-determined sex after artificial insemination of frozen-thawed, sex-sorted and re-frozen-thawed ram spermatozoa.

The fertility of ram spermatozoa cryopreserved prior to, and following, sex-sorting by flow cytometry was assessed after insemination of mature Merino ewes at a synchronised estrus. Ewes were inseminated with spermatozoa from three rams, split into four treatment groups: 50 x 10(6) motile non-sorted, frozen-thawed (Control50), 15 x 10(6) motile non-sorted, frozen-thawed (Control15), 15 x 10(6) motile sex-sorted, frozen-thawed (SF15) or 15 x 10(6) motile frozen-thawed, sex-sorted, re-frozen-thawed (FSF15) ram spermatozoa. Separation of SF15 and FSF15 treatments into X- and Y-chromosome-bearing populations was achieved using a high-speed sperm sorter. The percentage of ewes lambing after insemination was similar for Control15 (36/74; 48.6%), SF15 (35/76; 46.1%) and FSF15 (26/72; 36.1%) groups (P>0.05). A higher percentage of ewes produced lambs in the Control50 (38/70; 54.3%) than the FSF15 group (P<0.05). Fifty-one of the 55 (92.7%) lambs derived from fresh, sex-sorted frozen-thawed spermatozoa were of the predicted sex, as were 41/43 (95.3%) lambs derived from frozen-thawed, sex-sorted, re-frozen-thawed spermatozoa. This study demonstrated for the first time in any species that frozen-thawed spermatozoa, after sex-sorting and a second cryopreservation step, are capable of producing offspring of the predicted sex following artificial insemination.

Animals↗

Expression of TGF-beta1, TGF-beta2, TGF-beta3 and the receptors TGF-betaRI and TGF-betaRII in placentomes of artificially inseminated and nuclear transfer derived bovine pregnancies.

Bovine nuclear transfer pregnancies are characterized by a high incidence of placental abnormalities, notably, increased placentome size and deficiencies in trophoblast cell function and establishment of placental vasculature. Alterations in gene expression during placental growth and development may contribute to the appearance of large placentomes in pregnancies derived from nuclear transfer. The placenta synthesizes a number of cytokines and growth factors, including the transforming growth factor-betas (TGF-betas) that are involved in the establishment, maintenance and/or regulation of pregnancy. All forms of TGF-beta and their receptors are present at the fetal-maternal interface of the bovine placentome, where they are thought to play an important role in regulating growth, differentiation, and function of the placenta. Using real-time RT-PCR, we have examined the expression of TGF-beta1, TGF-beta2, TGF-beta3 and the receptors TGF-betaRI and TGF-betaRII in placentomes of artificially inseminated (AI) and nuclear transfer (NT)-derived bovine pregnancies at days 50, 100 and 150 of gestation. TGF-beta1, TGF-beta2 and TGF-beta3 mRNA expression increased by 2.0-2.8-fold, while TGF-betaRI and TGF-betaRII mRNA expression decreased by 1.7-2.0-fold in NT placentomes compared to AI controls at all gestational ages examined. These findings indicate that NT placentomes may be resistant to the growth suppressive effects of TGF-betas and could contribute to the placental proliferative abnormalities observed in NT-derived placentas. Alternatively, deficiencies in placentation may provide a mechanism whereby TGF-betas are dysregulated in NT pregnancies.

Activin Receptors, Type I↗

A comparison of progestin-based protocols to synchronize ovulation and facilitate fixed-time artificial insemination in postpartum beef cows.

The experimental objective was to compare pregnancy rates after fixed-time AI in postpartum suckled beef cows following administration of two progestin-based protocols to synchronize ovulation. Cows (n = 424) at three locations (n = 208, 122, and 92 per location) were stratified by age, BCS, and days postpartum (DPP) and assigned randomly to one of the two treatment protocols. The MGA Select-treated cows (MGA Select; n = 213) were fed melengestrol acetate (MGA, 0.5 mg x cow(-1) x d(-1)) for 14 d and carrier for 8 d, and then GnRH (100 microg i.m. Cystorelin; d 26) was injected 12 d after MGA withdrawal, and PG (25 mg i.m. Lutalyse) was administered 7 d after GnRH. Cows assigned to the 7-11 Synch protocol (7-11 Synch; n = 209) were fed carrier for 15 d and MGA for 7 d, and then injected with PG on d 22 (d 7 of MGA), GnRH on d 26, and PG again on d 33. Artificial insemination was performed at fixed times for cows in both treatments at 60 or 72 h after d 33 PG for 7-11 Synch and MGA Select groups, respectively. All cows were injected with GnRH (100 microg of i.m. Cystorelin) at AI. There was no treatment x location interaction for age (P = 0.90), BCS (P = 0.64), or DPP (P = 0.93), and the results were therefore pooled for the respective treatments (age [7-11 Synch, 5.5 +/- 0.2; MGA Select, 5.5 +/- 0.2], BCS [7-11 Synch, 5.7 +/- 0.1; MGA Select, 5.6 +/- 0.1], and DPP [7-11 Synch, 41.1 +/- 1.1; MGA Select, 42.1 +/- 1.1]). Blood samples were collected 8 and 1 d before MGA or carrier to determine pretreatment estrous cyclicity (progesterone >or=1 ng/mL; 7-11 Synch, 59/209 [28%]; MGA Select, 54/213 [25%]; P = 0.50) and again on d 33 PG to evaluate treatment response as a percentage of cows with progesterone concentrations in serum >or=1ng/mL (7-11 Synch, 184/209 [88%]; MGA Select, 177/213 [83%]; P = 0.15). Pregnancy rates resulting from fixed-time AI did not differ (P = 0.25) between treatments (7-11 Synch, 128/209 [61%]; MGA Select, 142/213 [67%]), nor did pregnancy rates (P = 0.77) at the end of the breeding season (7-11 Synch, 198/208 [95%]; MGA Select, 204/213 [96%]). These data indicate that pregnancy rates were comparable after fixed-time AI, following administration of the 7-11 Synch and MGA Select protocols. Both protocols provide opportunities for beef producers to use AI and eliminate the need to detect estrus.

Animals↗

Impact of cryopreservation on spermatozoa from infertile men: implications for artificial insemination.

The objective of this study was to evaluate the effect of cryopreservation-thawing on the quality of sperm from men with subfertile semen parameters. Twenty-seven patients with teratozoospermia, six of whom also had asthenozoospermia, were studied and their sperm parameters were compared to those of fertile donors (n = 71) in their fresh, post-thaw, and washed samples. After thawing, the percentage decrease in motility was significantly greater in patients than in donors, but the motility yield (post-thaw motility/prethaw motility) reached an average of 58% in the patient group vs. 68% in the donors (p = .02). No single characteristic of the fresh samples from patients or donors could significantly predict post-thaw outcome. For the patient group, however, multiple regression analysis provided a cutoff sperm concentration (50 x 10(6)/mL) and motility (40%) below which a very poor post-thaw recovery was obtained. The frozen-thawed-washed specimens had significantly higher velocity than the frozen-thawed samples, both in patients and donors. The results suggest that some patients with teratoasthenozoospermia yield acceptable sperm parameters after freezing-thawing-washing, and therefore these ejaculates could be used individually (or pooled) in artificial insemination.

Cryopreservation↗

Harnessing the biology of the oviduct for the benefit of artificial insemination.

Spermatozoa fulfil a single role, namely achieving syngamy by transporting the haploid genome to their counterpart gamete, the oocyte. Simple as this may seem, it is fraught with many difficulties, especially in the face of biological processes that enable females to select spermatozoa after they have mated multiply with several males. Conversely, the female reproductive tract sequesters a privileged sperm subpopulation in the oviductal isthmus for variable periods of time, releasing them when the time is opportune for fertilisation. Recent studies of sperm transport in the female reproductive tract suggest that these phenomena involve signalling dialogues between spermatozoa and the female reproductive tract environment. Opportunities for mutual signalling are immense but have received relatively little attention. The oviduct is an organ of crucial significance in modulating sperm function and may be one of the most important sites for determining many aspects of sperm selection and competition. The oviductal environment possesses the potential for enhancing sperm survival, suppressing and activating sperm motility as required, and responds to the arrival of spermatozoa by producing novel proteins. While the biological nature of the sperm-oviduct dialogue is interesting for its own sake, the mechanisms that govern these processes offer opportunities for the improvement of artificial insemination procedures. If oviductal proteins enhance sperm survival, they offer opportunities for the development of long-life semen diluents. Conversely, if we understood the basis of sperm selection we may be able to concentrate on identifying and using only the best sperm subpopulations for improved animal breeding efficiency.

Animals↗

Detection of Chlamydia trachomatis in semen of artificial insemination donors by the polymerase chain reaction.

OBJECTIVE: To assess the feasibility of detecting Chlamydia trachomatis in cryopreserved donor semen by a specific, direct polymerase chain reaction (PCR). DESIGN: Cryopreserved donor semen was tested for the presence of C. trachomatis by a specific PCR, directly applied to semen without prior DNA purification. SETTING: Tertiary care fertility center in a teaching hospital and university-based laboratory for molecular pathology. PARTICIPANTS: Cryopreserved semen from 30 donors was investigated. These semen samples had previously given negative results in cell culture for C. trachomatis. Two different ejaculates of each donor, cryopreserved with an interval of 2 years, were retrospectively analyzed. INTERVENTIONS: None. MAIN OUTCOME MEASURE: The presence of C. trachomatis as demonstrated by PCR. RESULTS: In 3 of 30 donors C. trachomatis was detected in both ejaculates, whereas in 2 additional donors only one of the two samples tested positive. Additional samples from 2 positive donors, together with samples from 3 negative donors, were studied more extensively, to test the reproducibility and reliability of PCR results. All ejaculates of the donors, previously positive for C. trachomatis by PCR, indeed appeared to be positive, whereas the samples of the negative donors remained negative. CONCLUSIONS: The direct PCR is a reliable, sensitive, and valuable method for detection of C. trachomatis in semen. The incidence of contamination of donor semen with C. trachomatis in the donor population in this study stresses the need for rigorous screening of donor semen before artificial insemination, preferably using a sensitive method such as the PCR.

Adult↗

Semen characteristics, cryopreservation, and successful artificial insemination in the Blue rock pigeon (Columba livia).

The present study was undertaken in the Blue rock pigeon (Columba livia) to evaluate the annual semen characteristics, to identify a suitable extender for semen short-term storage, to determine a protocol for cryopreservation of semen and finally to check whether intracloacal insemination would lead to the birth of a chick. Semen characteristics such as semen volume, sperm concentration, sperm motility, and percentage of normal spermatozoa were maximum during the monsoon season. TALP was observed to be the most suitable semen extender and the sperm survived best at 37 degrees C at a dilution of 1:100 in TALP. Further, cryopreservation studies on pigeon semen indicated that 8% DMSO with or without egg yolk (20%) proved to be a better cryoprotectant compared to glycerol and polyethylene glycol. In addition, the slow freezing protocol was better than the fast-freezing protocol and about 40% of the cryopreserved spermatozoa were motile following thawing. Computer-aided semen analysis indicated that pigeon spermatozoa were extremely active immediately after dilution in TALP and exhibited linear trajectories persisting up to 9h. But, with time there was a time-dependent decrease in the velocity parameters (VAP, VSL, and VCL). Cryopreserved spermatozoa following thawing also exhibited linear trajectories but had reduced velocity as evident from the significant decrease in VAP, VSL, and VCL. Further, artificial inseminations using fresh semen resulted in 45% fertilization and birth of a live chick.

Animals↗

Ultrastructural observations on in vivo fertilisation in the brushtail possum, Trichosurus vulpecula, following PMSG/LH superovulation and artificial insemination.

Information on the dynamics of gamete interaction in marsupials is very limited and not available for any species from the major Australian Order Diprotodontia which includes most of the more familiar animals such as kangaroos, possums and the koala. This study addressed this deficiency by examining the ultrastructure of in vivo fertilised eggs from common brushtail possums (Trichosurus vulpecula). Females were superovulated by treatment with 15 IU PMSG and then 4 mg porcine LH 3 days later, and inseminations were performed 910-13 h after LH) using epididymal spermatozoa. Between 33 and 39 h after LH injection females were killed, reproductive tracts excised and the oviduct ampulla segment flushed for eggs. Three of the six eggs examined were fertilised as judged by the presence of sperm remnants in the cytoplasm. On the basis of these eggs it was found that sperm penetration left a large hole in the zona pellucida (ZP), suggesting that sperm zona penetration occurs primarily by the enzymatic action of acrosomal enzymes. Sperm lying within the perivitelline space were lacking both an outer acrosomal membrane and the associated acrosomal contents, while both these structures were found on sperm embedded within the mucoid layer, which is consistent with induction of the acrosome reaction by binding to the ZP. Once inside the egg cytoplasm, the sperm head travelled only a short distance before chromatin decondensation occurred. Fertilised eggs showed signs of cytoplasmic activation including cytoskeleton association with apparently dividing mitochondria and prominent rough endoplasmic reticulum. Unfertilised eggs appeared to be undergoing degenerative changes and lacked any evidence of activation. This study was demonstrated that superovulation and laparoscopic intravaginal artificial insemination provide a system through which perifertilisation events in the possum and other monovular Australian marsupials can be examined experimentally.

Animals↗

Use of factor VII-deficient beagles bred by artificial insemination to evaluate mechanisms of thrombosis associated with prothrombin complex concentrates.

Artificial insemination with frozen semen was used to breed beagles homozygous for a deficiency in clotting factor VII, with plasma concentrations of 2-3% of normal adult beagle pooled plasma. The factor VII-deficient beagles were used to determine if factor VII activity has a role in the thrombogenic activity of prothrombin complex concentrates (PCCs) containing factors II, IX and X by evaluating changes in selected indicators of thrombogenicity. No differences were seen between the responsiveness of the factor VII-deficient and normal beagles, indicating that the thrombogenic activities of PCCs are mediated via effects on the intrinsic clotting system.

Animals↗

Fertility trial in Zebu cattle after a natural or controlled estrus with prostaglandin F2 Alpha, comparing natural mating with artificial insemination.

With the object of comparing reproductive efficiency obtained by natural mating and by artificial insemination (AI), not only following a natural estrus but also after an induced estrus with PGF2Alpha in Zebu cattle in the tropics, 244 adult cows were divided into 4 groups. Group I (N = 69) and Group III (n = 62) were injected with 25 mg of PGF2Alpha when a functional CL was found on rectal examination. Group I was inseminated and group III was served by natural mating, both groups within five days after injection. Groups II (n = 57) and IV (n = 56) were left untreated, group II being AI and group IV ran with a fertile bull for 22 days. Estrus detection was carried out only in the injected groups (I and III) for 15 minutes every three hours between 0600 and 1800. All information was analyzed by linear trigonometric models. The onset of estrus occurred on average 68.7 h after injection in group I and 59.5 h in group III. However only 46.3% and 54.8% of animals were detected in estrus in group I and III respectively, the difference being significant (P < 0.10). Conception rates were 18.6%, 29.8%, 19.3% and 33.9% for groups I, II, III, and IV respectively. A significant difference (P < 0.10) existed between the injected groups and the untreated ones.

Journal Article↗

Fertility of ewes following artificial insemination with semen frozen in pellets or straws, a preliminary report.

In two trials involving the artificial insemination of 194 ewes, the fertility of ram semen was examined following freezing, either in pellet form or in straws, and after storage in a chilled state (15 degrees C) for up to 16 hours. Estrus was synchronized in ewes by intravaginal sponge (MAP) treatment for 14 days. At sponge removal 600 IU PMSG was injected and the ewes received two inseminations 50 and 60 hours later. Fertility was assessed at lambing. In trial 1, the mean lambing rate of 52% (16/31) for semen frozen in pellets was higher than 29% (9/31) for semen frozen in straws but this difference was not significant. In trial 2, ewes inseminated with chilled semen and semen frozen in pellets had lambing rates of 83% (44/53) and 55% (44/79) respectively (P<0.001).

Journal Article↗

Artificial insemination using cryopreserved sperm in the silkworm, Bombyx mori.

We report in this paper that female moths artificially inseminated with cryopreserved sperm (-196 degrees C) could oviposit eggs when the sperm was preserved for 356days, and that the fertilization rate and the number of eggs laid were almost equivalent to those obtained in normally mated moths. The optimal cooling rate for sperm freezing was 5-65 degrees C/min for maintaining a high fertility of sperm. The simple and reliable method of cryopreservation was to put the semen first in a deep freezer at -80 degrees C and thereafter put them in liquid nitrogen. When female moths of 'white 2' egg-color mutant strain were inseminated with a mixture of frozen-thawed sperm from males of normal-colored egg strain and non-frozen sperm from males of the 'white 2', female moths deposited a majority of 'white 2' eggs and a very small number of eggs of normal color. The result shows that there was a competitive fertilization of sperm between the two strains of the silkworm, and that sperm fertility was reduced to a considerable extent by freezing at -196 degrees C. These results may contribute not only to basic studies on fertilization in Lepidoptera but also to the development of long-term preservation procedure of genetic resources by using cryopreserved sperm of Bombyx mori.

Journal Article↗

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

We compared conventional methods for laparoscopic and cervical artificial insemination (AI) to a transcervical AI procedure (Guelph System for Transcervical AI; GST-AI) for use with frozen semen in Merino ewes. The GST-AI procedure was performed by an experienced operator in Experiment 1 (771 ewes) and by 2 inexperienced operators in Experiment 2 (555 ewes). In Experiment 1, intrauterine insemination by GST-AI was achieved in 76% of the ewes. The pregnancy rate at Day 70 for ewes inseminated by laparoscopy (48%, 120 251 ) was higher (P<0.01) than for ewes inseminated by either intrauterine GST-AI (32%, 64 201 ) or cervical AI (9%, 24 256 ). The overall (intrauterine and intracervical) pregnancy rate for GST-AI was 26% (68 264 ) and was unaffected by depth of insemination within the cervix. Pregnancy rates were unaffected by ram or day of insemination. In Experiment 2, the operators achieved intrauterine inseminations by GST-AI in 43% (78 182 ) of the ewes, with a significant operator effect (P<0.01) on depth of cervical penetration. The pregnancy rate to intrauterine GST-AI (40%, 31 78 ) did not differ from that to laparoscopic insemination. The total pregnancy rate for GST-AI in Experiment 2 (19%, 34 182 ) was lower (P<0.05) than that for laparoscopic AI (39%, 72 187 ) but superior (P<0.05) to that for cervical AI (1%, 1 186 ). The GST-AI pregnancy rates were affected by depth of AI (P<0.01) and by operator (P<0.05). It is concluded that GST-AI is superior to cervical AI, and may have application in Merinos if cervical penetration rates can be improved.

Journal Article↗

Genetic correlation estimations between artificial insemination sire performances and their progeny beef traits both measured in test stations.

In France, beef traits of artificial insemination (AI) beef bulls are improved through the sequential selection for their own performances and for their male progeny performances, both being recorded in test stations. The efficiency of such programmes mainly depends on the genetic correlations between sire performances and progeny beef traits. Such correlations were independently estimated, using the multivariate REML (restricted maximum likelihood) method in a Limousin and a Charolais programme. In both breeds, high genetic correlations were observed between sires and progeny analogous morphology scores (from 0.64 to 0.82). Genetic correlations estimated between sires and progeny growth (from 0.41 to 0.70) were lower probably due to the difference of diet in central and progeny stations. Correlations between sire muscling scores and progeny skeletal frames (and vice-versa) were negative (from -0.05 to -0.58). The genetic correlations of sire traits with progeny dressing percentage (DPp) and carcass fatness score (CFp) were only low to moderate. These results show that the selection of bulls at the end of performance testing in test stations may be efficient in improving progeny growth and morphology. However, such a selection is insufficient in improving their dressing percentage and carcass composition.

Journal Article↗

The integration of the transfer of frozen embryos with artificial insemination in a commercial beef herd.

This trial was designed to examine the integration of the transfer of frozen embryos and artificial insemination (A.I.) in a commercial herd of beef cows. Embryos were collected nonsurgically from 15 superovulated cows and all embryos judged viable (n = 69) were frozen in 0.25 ml plastic A.I. straws. Glycerol was used as a cryoprotectant and the embryos were frozen with omission of the traditional "seeding" step. On three consecutive days immediately preceding the onset of a 60-day breeding season, 54 embryos were transferred to multiparous cows with calves at foot. The remaining 82 cows in the same herd were used as controls. Cows receiving embryos were maintained under observation, but not inseminated for 21 days, after which cows exhibiting signs of estrus were inseminated up to Day 60. Cows in the control group were inseminated at observed estrus throughout the 60-day period. The overall pregnancy rate for the experimental and control groups did not differ (77.8 % and 73.2 % respectively). Of the experimental cows, 24.1 % conceived to the embryo transfer, and 53.7 % to A.I. The mean calving to conception interval for the experimental cows was 96.3 days (median 101 days) and for the control cows it was 92.3 days (median 88 days). The calving pattern in the experimental cows was biphasic, calvings resulting from embryo transfer of A.I. accounting for the two peaks. Reconception rates in the following breeding season were not different between the embryo transfer or A.I. groups or between experimental and control herds. Weaning masses of calves born of embryo transfer were significantly higher than the other groups, but calves resulting from A.I. in the experimental herd were not different from calves in the control herd. It was concluded that the transfer of frozen embryos could be successfully integrated into an A.I. program with a limited breeding season without detrimentally affecting the overall reproductive performance of the recipient herd.

Journal Article↗