Embryo transfer: a comparative biosecurity advantage in international movements of germplasm.

This paper uses cattle as a model to provide an overview of the hazards involved in the transfer of in vivo-derived and in vitro-produced embryos. While scientific studies in recent decades have led to the identification of pathogens that may be associated with both in vivo- and in vitro-derived embryos, those studies have also been the basis of appropriate disease control measures to reduce the risks to a negligible level. These disease control measures have been identified and assessed by the International Embryo Transfer Society's (lETS) Health and Safety Advisory Committee, the expert body that advises the World Organisation for Animal Health (OIE) on matters related to the safety of embryo transfer. Through the OIE's processes for developing and adopting international standards, the disease control measures identified by the IETS have been incorporated into the Terrestrial Animal Health Code. The basic principles rely on the crucial ethical roles of the embryo collection team and embryo transfer team, under the leadership of approved veterinarians. Decades of experience, with nearly 10 million embryos transferred, have demonstrated the very significant biosecurity advantage that embryo transfer technology has when moving germplasm internationally, provided that the international standards developed by the IETS and adopted by the OIE are strictly followed.

Biosecurity and the various types of embryos transferred.

The aim of the present paper was to review some features related to the risk analysis of three types of embryos to be transferred, namely the in vivo derived, the in vitro produced and the cloned ones. For in vivo-collected embryos, a considerable number of experiments and scientific investigations have been performed and hundreds of thousands of embryos are transferred annually with no contamination of associated diseases. Provided that the code of practice such as that published by the International Embryo Transfer Society is strictly followed by the embryo transfer practitioners, the statement made some 17 years ago saying that the in vivo-derived embryo transfer was the safest way of exchanging genes remains entirely true, thanks to the professionalism of the embryo transfer industry. For the in vitro-produced embryos, some particular rules have to be followed because of specific risks for some pathogens to strongly adhere to the zona pellucida of such embryos. There are some means to monitor and control those effects, and the transfer of in vitro-produced embryos can also be a very safe way to exchange genes around the world. The third type of embryos, the cloned ones, is a quite different category and the risk analysis to be soundly made still needs a lot of investigations so as to characterize the potential risks if there are, in terms not only of disease transmission but also in terms of public health, zoonotic risks as well as those related to quality and safety of food. The problem in this regard, is more directly addressed for offspring of clones than to the cloned embryos themselves. Published data on this issue are increasing in numbers so that progress in that area is expected in the few years to come.

Health and Safety Advisory Committee (HASAC) of the International Embryo Transfer Society (IETS) has managed critical challenges for two decades.

The International Embryo Transfer Society (IETS) was founded in 1974. Early members used the society as a forum for the exchange of scientific and technical information relevant to a newly emerging embryo transfer industry. The impact that embryo transfer could have on the international trade of livestock genetics was clear by 1982, so the IETS commissioned the Import/Export Committee. The initial challenge for this Committee was to deal with concerns about disease transmission via embryo transfer. Many of the early concerns have been dispelled, but at the time they threatened the continued development of a fledgling industry. Over the past two decades, many new critical challenges have been met and managed by this Committee, which was recently renamed the Health and Safety Advisory Committee (HASAC). Assessing risks of animal disease transmission via reproductive technologies and establishing protocols for managing these risks are still major issues for HASAC. However, additional concerns have developed as views of the society changed and as novel applications of biotechnology in farm animals were identified. This paper is intended to chronicle some of the major changes and challenges that were managed by members of the HASAC and its Subcommittees from the early years of embryo transfer to the current millennium with technological advances in molecular biology.

Pregnancy-specific protein B (bPSPB) and progesterone monitoring of post-partum dairy cows with placental retention.

The relationship between placental retention, progesterone and pregnancy-specific protein B (bPSPB) was determined in 60 calving Holstein cows. The cows were divided into two groups with placental retention (WPR, n = 16) and no placental retention (NPR, n = 44). Every 4 days, until 60 days post-partum, blood samples were taken and the uteri were checked using ultrasonographv. The puerperal characteristics of NPR and WPR were as follows: mean days of abnormal vaginal discharge: 20.2+/-5 versus 35.6+/-7 (P < 0.01); mean intervals to uterine involution: 21.4+/-3.7 versus 27.6+/-7.6 days (P < 0.01); rate of endometritis: 25 versus 100% (P < 0.01). The mean numbers of oestrus cycles per cow were 1.75+/-0.5 versus 0.85-/+0.9 (P < 0.05) and the mean durations of the first oestrus cycle were 18+/-3.5 versus 16+/-2.1 days (P > 0.05). The mean intervals to first ovulation were 21.5+/-8.4 versus 35+/-19 days (P < 0.01). bPSPB blood concentrations were higher in the WPR group at calving with 955+/-170 versus 750+/-205 ng/ml (P < 0.01) and also during the first 32 days post-partum with 173.68+/-47.3 versus 131.0+/-29.2 ng/ml (P < 0.01). The mean bPSPB half-life was similar in the two groups: 6.9+/-2.5 versus 6.5+/-2.1 days (P > 0.05). In conclusion, it was found that placental retention was associated with a higher rate of endometritis, a lower number of cycles, longer interval to first post-partum ovulation and higher concentration of bPSPB at calving and during the post-partum period. The positive relationship between bPSPB concentrations and calf birth weight and their association with post-partum pathological events may be useful in monitoring animals presenting high concentrations at calving.

Identified and unidentified challenges for reproductive biotechnologies regarding infectious diseases in animal and public health.

The aim of the present paper is to review the known and theoretical risks for in vivo derived and in vitro produced embryos as well as for nuclear transferred or transgenic embryos in terms of animal diseases or diseases of public health consequence. For in vivo derived embryos, a considerable number of experiments and scientific investigations have resulted in recommended guidelines and procedures that ensure a high level of safety. The effectiveness of these measures has been validated by field experience with the safe transfer of several million embryos over the past three decades. In vitro produced embryos have several characteristics that differentiate them from the former, in particular a structure of the zona pellucida that results in a more frequent possible association of pathogens with the embryo. However, the guidelines prescribed by the IETS, the international standard setting body (OIE) and existing national regulatory frameworks are in place to minimize the risk of disease transmission. No specific public health risks have been identified to date with respect to in vivo or in vitro derived embryos. In regard to nuclear transferred and transgenic embryos, theoretical risks have been identified in relation to the potential effects on some intrinsic viruses such as endogenous retroviruses but very little targeted experimental work has been carried out on infectious diseases that could have adverse consequences on animal or human health. Although there has been no report of such adverse consequences associated with the limited number of animals produced to date by such reproductive technologies, a precautionary approach is warranted given the potential negative impacts and it would be prudent to restrict at this stage, the international movement of such "manipulated" embryos.

Hygienic aspects of storage and use of semen for artificial insemination.

The artificial insemination (AI) industry has developed over the last 50 years to the extent that it is used in almost every country in the world. One of the main factors contributing to its success is the confidence of the farmers that germplasm is not associated with pathogens, so that AI can be performed without risks. This has been achieved as a result of a considerable amount of research based on sound scientific data that has identified the major risk pathogens. A summary of these studies, given in this section, shows that despite the large number of agents that could be transmitted via the semen, there are cost-effective means to prevent such hazards. One of the basic rules is that the males should be housed in strictly protected semen collection centres (SCCs). Such centres should be approved by the veterinary authorities based upon specific criteria, which include special housing and operating specifications. This also includes specific means of monitoring the health of individual males through regular clinical examinations, assessment of semen and testings for various diseases. Two new challenges can now be identified, one relevant to so-called emerging diseases the impact of which on the status of the semen donors should always be assessed, and the second, relates to endangered genetic resources which may become extinct without active conservation programmes. The experience gained by the AI industry over the last 50 years should help to solve those problems. Currently, the use of semen derived from approved SCCs warrants their disease-free status.

Exposure of in vitro-produced bovine embryos to foot-and-mouth disease virus.

The aim of this study was to investigate whether foot and mouth disease virus (FMDV) interacts with in vitro produced (IVP) bovine embryos. One milliliter of a suspension of FMDV (2 x 10(7) TCID50/mL) was added to several batches of these embryos 7 d after in vitro fertilization, by which time they had either developed to the morula/blastocyst stage (n = 256) or degenerated (n = 260). Six experiments were performed in which developed or degenerated batches of embryos were incubated with FMDV for periods of 1 h (3), 2 h (2) or 4h (1). After this, the embryos were washed 10 times according to the International Embryo Transfer Society (IETS), then pooled and ground up to form a suspension, and assayed on cell cultures for FMDV. The cell cultures were observed daily for cytopathic effects for 3 d post exposure. In addition to the cell culture method, the polymerase chain reaction (PCR) technique was used to assay for the presence of the virus in the washing fluids. Assays for FMDV were also conducted on the first and second wash and on the pooled sample constituting the eight, ninth and tenth wash. With the exception of the second wash from a batch of embryos exposed to FMDV for 2 h, all samples of the first and second wash produced FMDV cytopathic effects, but none occurred with the pooled samples of the 8th, 9th and 10th wash. FMDV was also isolated from all but 1 of the batches of embryos after 1 h of incubation, from 1 of 4 batches after 2 h of incubation and from all batches after 4 h incubation. By contrast, the presence of virus could not be demonstrated by PCR based on the technique used here. These results show that 7 d old IVP bovine embryos can retain FMDV after washing, unlike in vivo-derived embryos, which do not appear to carry risks of FMDV transmission when washed according to IETS recommendations. Stricter controls are, therefore, necessary when using IVP embryos from cattle in a non-FMD-free zone in domestic or international trade.

Absence of response to oestrus induction and synchronization treatment is related to lipid mobilization in suckled beef cows.

Energy status, follicular growth, oestradiol and LH secretion were investigated in 17 suckled Charolais cows synchronised 59.0 +/- 3.6 days after calving with a 10 day ear implant containing 3 mg of Norgestomet. The cows received 3 mg of Norgestomet and 5 mg of oestradiol valerate by IM injection at implant insertion (day 0) and 600 IU PMSG at implant removal (day 10). They were artificially inseminated (AI) 48 and 72 h after implant removal. Energy status was assessed by measuring weekly plasma concentrations of non-esterified fatty acids (NEFA), beta hydroxy-butyrate (BHB), glucose and insulin 7 weeks before AI. Progesterone plasma concentrations were measured during the same period to assess the presence of a functional corpus luteum. Follicular growth was followed daily by ultrasonography from day -3 to day 13. Oestradiol secretion was measured on day -3, day 6 and day 10 from five hourly samples. Oestradiol and LH plasma concentrations were measured hourly from 29 to 48 h after implant removal for seven cows. Cows were checked for pregnancy by ultrasonography 45 days after AI. Pregnant cows (P) were compared with non-pregnant cows (NP) for energy status, follicular growth, and oestradiol secretion by split-plot ANOVA. Two cows (11.8%) were cyclic before treatment, seven ovulated after treatment (41.2%) and five were found pregnant 45 days after AI (29.4%). There was no difference in body condition score and body weight between P and NP cows on day 0 (2.5 +/- 0.2 and 685 +/- 24 kg vs 2.5 +/- 0.1 and 670 +/- 13 kg; P > 0.05). Mean plasma NEFA concentrations before treatment were significantly lower in P than in NP cows (218 +/- 29 mu eq/L vs 279 +/- 18 mu eq/L; P < 0.05). No significant differences between P and NP cows were found for BHB, glucose and insulin concentrations. P cows presented more medium sized follicles (5 mm < or = diameter < 10 mm) than NP females during the period of observation (2.65 +/- 0.19 vs 2.50 +/- 0.12; P = 0.05). Plasma oestradiol concentrations were not different between P and NP cows on day -3 (8.4 +/- 0.7 pg/mL vs 7.7 +/- 0.4 pg/mL, P > 0.05), day 6 (10.4 +/- 0.6 pg/mL vs 9.8 +/- 0.4 pg/mL, P > 0.05) but were higher in P than in NP cows on day 10 (10.9 +/- 0.6 pg/mL vs 7.8 +/- 0.4 pg/mL; P < 0.05). After implant removal, oestradiol secretion only increased in P cows and a LH peak occurred whereas no increases in oestradiol (11.0 +/- 0.4 pg/mL vs 6.3 +/- 0.3 pg/mL, P < 0.05) and LH (6.0 +/- 0.5 ng/mL vs 1.2 +/- 0.5 ng/mL, P < 0.05) secretion were observed in NP cows. The conclusion was that follicular growth, oestradiol secretion, ovulation and pregnancy rate after oestrus synchronisation treatment are related to mobilization of energy stores before treatment in suckled beef cows in the same body condition score.

Sources of variation of post-partum cyclicity, ovulation and pregnancy rates in primiparous Charolais cows treated with norgestomet implants and PMSG.

In this study, sources of variation of postpartum cyclicity, ovulation and pregnancy rates were analyzed for 723 primiparous suckled Charolais cows treated with combined norgestomet implants (Crestar) and 600 IU PMSG (Chronogest) injected at the time of implant removal. The cows were inseminated 48 and 72 h after implant removal. Cyclicity and ovulation rate were estimated by progesterone assay and pregnancy rate by ultrasonography. At time of implant insertion, difficulty of previous calving, body condition score (BCS, from 1 to 5), interval from calving to implant insertion and herd related factors were recorded and their effects analyzed by logistic regression models. Cyclicity, ovulation and pregnancy rates were, respectively, 14.7% (106/723 ), 67.1% (381/568 ) and 42% (303/722 ) and were affected by BCS, calving conditions and interval from calving to implant insertion (P values from < 0.01 to < 0.0001). For ovulation and pregnancy rate, an interaction between BCS and interval from calving to implant insertion was found (P < 0.01). No other main factor or interaction was found to be significant. Cyclicity rate was lower in BCS1 (score < 2.5) cows (9.6%) than in BCS2 (19.8%) or BCS3 (score > 2.5) cows (22.4%), and decreased as difficulty of calving increased (23.2, 13.6 and 10.1%, respectively, for calving conditions 1, 2 and 3 cows). Cyclicity rate increased with interval from calving to implant insertion (8.2, 10.2 and 19.5%, respectively, for interval from calving to implant insertion < 60 d, between 60 and 70 d and > 70 d). Similar trends were found for ovulation rate. Previous difficult calving conditions influenced pregnancy rate negatively (47.9, 43.8 and 32.5% for calving conditions 1, 2 and 3 cows, respectively; P < 0,005).

Influence of postpartum energy restriction on energy status, plasma LH and oestradiol secretion and follicular development in suckled beef cows.

Effects of postpartum energy restriction, parity and time after parturition on energy status (measured by glucose, insulin, non-esterified fatty acids (NEFAs) and beta-hydroxybutyrate), LH secretion and follicular growth were investigated in ten primiparous and nine multiparous suckled cows. Females were allocated by parity, body mass and body condition score at calving to diets supplying either 100% (CE, n = 10) or 70% (LE, n = 9) of energy requirements until day 70 postpartum. Metabolic parameters were measured every week from calving to day 70 postpartum. Blood samples were collected at intervals of 15 min for 10 h on day 30 and day 50 after parturition for LH measurement. Ovaries were examined between days 20 and 30 and days 40 and 50 postpartum by ultrasonography. Energy supply affected mean plasma concentrations of glucose (CE: 0.64 +/- 0.01 g l-1 versus LE: 0.61 +/- 0.01 g l-1; P < 0.05) and NEFA (CE: 168 +/- 17 mu eq l-1 versus LE: 309 +/- 18 mu eq l-1; P < 0.01) but by day 70 postpartum, glucose and NEFA concentrations were not significantly different between the two groups. LH pulse amplitude and frequency were not affected by energy supply (P > 0.10). However, at day 30 postpartum, LH pulse frequency was negatively correlated with plasma concentration of NEFA (r = -0.61; P < 0.01). Cows fed diets supplying 100% of energy requirements had more large follicles than did cows fed low energy diets (CE: 0.82 +/- 0.05 versus LE: 0.31 +/- 0.05; P < 0.05). The size of the largest follicle was greater in CE cows than in LE cows (CE: 10.2 +/- 0.1 mm versus LE: 8.7 +/- 0.2 mm; P < 0.05). Between 40 and 50 days postpartum, the size of the largest follicle was negatively correlated with NEFA concentration (r = -0.5; P < 0.05).(ABSTRACT TRUNCATED AT 250 WORDS)

[The use of gene amplification for the detection of the Aujeszky's disease virus in the sperm of boars]. / Utilisation de la technique d'amplification de gène pour la détection du virus de la maladie d'Aujeszky dans le sperme de verrat.

The aim of the present study was to specify the experimental conditions for the use of the polymerase chain reaction (PCR) in the detection of the pseudorabies virus in boar semen. Boar semen was artificially contaminated by Aujeszky's disease virus and submitted to the PCR. The seminal plasma fraction of the semen was found inhibitory for PCR. An incubation of the samples with proteinase K followed by an extraction of the viral DNA on a glass matrix was shown to be able to remove the inhibition. The method was simple, rapid and allowed the detection of around 370 viral DNA sequences per microliter of sample. The detection of the amplified DNA by a specific enzymatically labelled probe could improve the sensitivity of the method.

Effects of Trypanosoma congolense infection on the pituitary gland of Baoulé bulls: immunohistochemistry of LH- and FSH-secreting cells and response of plasma LH and testosterone to combined dexamethasone and GnRH treatment.

The effects of Trypanosoma congolense infection were investigated at the pituitary level on trypanosome resistant Baoulé bulls (aged 3-6 years), using immunohistochemistry of LH- and FSH-secreting cells and a combined dexamethasone and GnRH challenge. The pituitaries of two control and five naturally infected Baoulé bulls were removed after slaughter and the LH- and FSH-secreting cells were examined immunohistochemically, using specific polyclonal antibodies against beta LH and beta FSH. No significant impairment of the labelling and distribution of LH- and FSH-secreting cells was seen in infected bulls when compared with control animals. No parasites were found in the pituitary glands. Plasma LH and testosterone concentrations were determined in eight control and eight infected bulls by enzymeimmunoassay and radioimmunoassay techniques, respectively. Blood samples were collected at intervals of 30 min two times before and nine times after dexamethasone treatment (20 mg i.m.). GnRH (Busereline: 20 micrograms, i.m.) was injected 4.5 h later and samples were collected every 15 min for 180 min. After dexamethasone treatment, LH and testosterone concentrations declined dramatically in both groups. Four hours after treatment, the mean testosterone concentration for both groups was 0.44 ng ml-1. After GnRH injection, LH concentrations in the infected group increased rapidly to a mean maximum value of 30 ng ml-1 by 165 min. In contrast, the increase in LH concentration in non-infected bulls was more gradual and the mean maximum value, reached at the same time, was only 20 ng ml-1. Testosterone concentration increased rapidly and in a similar manner in both groups for the first 90 min (0.08 +/- 0.04 ng ml-1).(ABSTRACT TRUNCATED AT 250 WORDS)

Distribution of BHV1 in fractions of semen from a naturally infected bull.

The aim of the present study was to investigate the distribution of BHV1 (Bovine Herpesvirus 1) in the semen from a naturally infected bull and more particularly, to determine if there were viral particles associated with sperm that could interfere with in-vivo or in-vitro fertilization. Frozen semen from a single ejaculate that had been previously identified as being contaminated with BHV1 was used. The contents of 12 ministraws were mixed. Two aliquotes served as controls, the first was held at 0 degrees C in ice water while the second was held at 4 degrees C in the centrifuge during the procedure. The remaining semen was separated into seminal plasma and sperm cell fractions by centrifugation. The seminal plasma was kept at 0 degrees C until viral titration. The pellet was treated by 10 consecutive washes in PBS and by a trypsin treatment after the fifth wash. Subsequently, the last pellet was macerated to break the cell membranes. Aliquotes for viral titration were taken from all washing fluids, from pellets after the fourth, sixth and tenth washes, and from the last crushed pellet. These aliquotes were kept in ice water until titration. The virus was titrated on secondary cell cultures of fetal calf kidney. The titers were 4.3, 4.2 and 4.4 log TCID50/ml, respectively, in the 2 controls and the seminal plasma. Titers declined from 3.8 log to 0 from the first to the tenth wash. The titer was 2.2 log in the resuspended pellet of sperm cell fraction prior to trypsin treatment. No virus was detected in the sixth pellet, the tenth one or from the crushed cells. It was concluded that a significant proportion of the BHV1 particles was associated with the sperm cells. Ten washes and a trypsin treatment could remove the adsorbed virus. No viral particles were detected within the sperm cells.

[Value of adding antibiotics to frozen bovine semen: the example of mycoplasmas and campylobacters]. / Intérêt de l'addition des antibiotiques dans la semence congelée de bovins: l'exemple des mycoplasmes et des campylobacters.

Two studies have been conducted to control opportunistic pathogen bacteria in frozen bovine semen. The first one was reported by Shin et al. (1988) and was based on (a) an experimental in vitro contamination of raw bull semen with various types of bacteria, (b) adding several antibiotics, alone or in combination with different concentrations. The second experiment, performed in our laboratory, was designed to compare from 240 split ejaculates the effect of adding concentrated solutions of antibiotics to the raw semen vs control (ie addition of antibiotics in the extender). The combination of gentamicin (500 micrograms/ml)-tylosin (100 micrograms/ml)-linco-spectin (300/600 micrograms/ml) was found to be the most effective on the strains of bacteria tested. The addition of a concentrated solution of antibiotics to the raw semen was not found here to be of any advantage on the numbers of CFU. Furthermore, such a protocol contributed to significantly decrease the percentage of motile spermatozoa. In conclusion, opportunistic pathogens such as those here studied may be kept under control with the appropriate antibiotics and with no deleterious effect on semen parameters provided that they are added at the adequate concentrations in the diluents used for semen cryopreservation.

Health surveillance of the transfer of bovine embryos fertilised in vitro.

The production of embryos fertilised in vitro is part of the third generation of biotechnologies applied to reproduction and is now undergoing development and application in the field. International exchanges of these embryos have already commenced. However, there are certain associated health risks. In decreasing order of importance, these involve dangers inherent in the initial biological material from which gametes and various somatic cells vital to such production (follicular and tubal cells, etc.) are derived, then possible external contamination during the production process, and finally storage and transport. Although the research which has been conducted on the interaction between these embryos and pathogens is still rather limited, as the technique has been under development for only a short time, it is clear that the media in which the oocytes undergo maturation, the spermatozoa undergo capacitation or the embryos are cultured constitute excellent indicators of the potential microbiological or viral environment. This can be utilised in health surveillance of the manipulations. Recently, specific recommendations for making such exchanges safe have been inserted into the Office International des Epizooties International Animal Health Code. These recommendations are based on the concept of official approval of specific embryo transfer teams and examination of animals from which gametes are obtained.

[Pathogenic effects of Trypanosoma congolense on the testis of Baoulé bulls: quantitative and morphometric histology]. / Effets pathogènes de Trypanosoma congolense sur le testicule des taurins Baoulé: histologie quantitative et morphométrique.

The effect of Trypanosoma congolense on testis was studied in 53 trypano-resistant "Baoulé" bulls by quantitative histology and morphometry. The daily spermatozoa production per testis of control groups (n = 45) was 382 +/- 334 x 10(6) (m +/- sd) and the epididymis contained 0.6 +/- 1 x 10(9) spermatozoa in the caput, 0.3 +/- 0.3 x 10(9) in the corpus and 1.2 +/- 1.8 x 10(9) in the cauda. The infected bulls (n = 8) showed no significant difference (P > 0.05) when compared to the control despite their average low value. The morphometric analysis during infection revealed a significant (P < 0.05) decrease (32%) of total Leydig cell volume per testis, 4.4 +/- 0.9 cm3 for the control (n = 5) and 3.0 +/- 0.8 cm3 for infected bulls (n = 8). The number of round spermatids per Sertoli cell and the daily round spermatid production (DRSP) per testis were also significantly reduced in infected bulls when compared to controls (P < 0.05), 5.2 +/- 0.7 and 2.8 +/- 2 for round spermatid per Sertoli cell and 6.1 +/- 2.0 and 3.1 +/- 1.9 x 10(8) for DRSP. These observations indicate that Trypanosoma congolense infection alters the interstitial tissue and meotic divisions of germinal cells leading to low daily round spermatid production per gram of testis in "Baoulé" bulls.

Effect of PG F2 alpha treatment on conception rates of dairy cows treated with a GnRH agonist 12 to 14 days after Artificial Insemination.

The effect of a GnRH agonist (10 ug Buserelin) on conception rate was determined when injected into dairy cows 12 to 14 days after Artificial Insemination (AI). The following factors were taken into account: previous treatment prior to AI with Prostaglandin F2 alpha, clinical history recorded prior to AI since last calving, parity and milk yield. A number of 118 cows, from one large dairy herd, were involved in this study. A total of 210 AI's were performed, followed by 140 GnRH treatments and 70 saline injections. Reproductive events were then recorded. The rank of AI was equally distributed among the two groups. 153 AI's were preceded by a Prostaglandin F2 alpha (Dinoprost - 25 mg) treatment among which 103 were subsequently GnRH-treated and 50 were controls. Plasma progesterone concentrations were determined daily for 34 days after AI in 13 treated and 13 control cows. An Early Pregnoncy Diagnosis (EPD) from milk progesterone concentration was performed on Day 21 after AI in all cases. Post-AI GnRH agonist treatment resulted in a significant enhancement of the conception rate: 60% vs 44%; p < 0.01, respectively for treated and control animals. PG F2 alpha treatment prior to GnRH injection had a major influence on the conception rate (62% vs 40%; p < 0.01). No effect was seen (54 % vs 55 %) in non-PG F2 alpha treated females which were subsequently injected respectively with GnRH or saline after AI. Previous clinical reproductive disorders, parity and milk yield had no significant effect. In non-pregnant treated animals, GnRH agonist treatment resulted in an increased rate of heat detection on days 20 - 25 after AI (91 % vs 74 %) and a higher fertility rate on the following AI was seen (59 % vs 44 %; p < 0.05). In conclusion, GnRH agonist treatment 12 to 14 days after AI only enhanced the conception rate of females which had previously been treated with PG F2 alpha. In non-pregnant cows, this treatment had also a benficial effect on heat detection and improved the conception rate at the subsequent AI.

Pregnancy-specific protein B and progesterone concentrations in French Alpine goats throughout gestation.

The 34 French Alpine dairy goats originated from a single flock and were artificially inseminated 44 h after synchronization of oestrus. They were bled daily at the jugular vein from 15 to 27 days after AI. An early pregnancy diagnosis by RIA of progesterone concentration was performed 21 days after AI. In pregnant goats (greater than 1.5 ng progesterone/ml) daily sampling was extended until 30 days after AI and, from those, 9 were bled every 2 weeks until the end of pregnancy and at 50 and 63 days post partum. Pregnancy-specific protein B (PSPB) was also assayed. The kidding rate was 67.6% (23/34). PSPB concentrations (ng/ml) in pregnant goats were significantly different from those of non-pregnant goats at 24 days after AI (0.82 +/- 0.18 vs 1.78 +/- 0.19; mean +/- s.e.m.) and rose to 40 ng/ml at the end of pregnancy. From Day 25 and throughout gestation, females with 2 fetuses had higher PSPB concentrations than did those with a single fetus (P less than 0.05). In the 2 goats exhibiting late embryonic mortality according to progesterone concentrations, one had a PSPB profile very similar to those of pregnant goats until 30 days while the other did not show any elevation of PSPB concentration. It is concluded that PSPB profiles in goats are similar to those found in cows throughout pregnancy and that PSPB RIA may be useful for pregnancy diagnosis or diagnosis of late embryonic mortality.

Evaluation of bull semen fertility by homologous in vitro fertilization tests.

In vitro fertilization assays were performed to investigate their validity in evaluating artificial insemination (AI) bull fertility. A total of 1,532 oocytes, collected from ovaries at the abattoir, were subsequently used in a 4 x 6 x 2 factorial design: 4 doses of heparin added into the capacitation and fertilization medium (0; 0.05; 0.1 and 0.2 micrograms/ml), 6 different bulls with known on-field non-return (NR) rates (range: 64.6-75.3%) and 2 different ejaculates for each bull, collected within a approximately 1-month interval. Oocytes were considered fertilized when 2 pronuclei (or more) were seen in the ooplasm. Both the heparin dose and bull exerted a highly significant effect on the in vitro fertilization (IVF) rates which ranged, per oocyte group, from 30-80%; bull x dose of heparin interaction was significant (P less than 0.001). The 0.05 micrograms/ml dose of heparin was optimal for discriminating individual bulls. At that dose, the correlation coefficients between the bulls, NR rates and the IVF rates from each ejaculate (within-bull or the mean of two ejaculates), were highly significant (r = 0.83). The rates of polyspermy were also significantly influenced by bull and heparin dose, but there was no interaction. In conclusion, capacitation and fertilization in a modified Tyrode medium containing 0.05 micrograms/ml of heparin may be a valuable tool for evaluating AI bull fertility.

Preliminary report of familial thymic lymphosarcoma in Holstein calves.

Seventy-three cases of the thymic form of leukosis were found in Holstein calves in five departments of France over a period of five months. Most of the calves had been sired by the same bull. The calves were negative for specific antibodies to bovine leukaemia virus. Morphological studies including light and electron-microscopic cytology, and serological and virological studies of 14 of the cases suggest that the disease was transmitted genetically.