Determination of sex and scrapie resistance genotype in preimplantation ovine embryos.

in Molecular Reproduction and Development (2009), 76(2), 183-190

The aim of this study was to test the accuracy of genotype diagnosis after pre-amplification of DNA extracted from biopsies obtained by microblade cutting of ovine embryos and to evaluate the viability of ... [more ▼]

The aim of this study was to test the accuracy of genotype diagnosis after pre-amplification of DNA extracted from biopsies obtained by microblade cutting of ovine embryos and to evaluate the viability of biopsied embryos after vitrification/warming and transfer to recipients. Sex and PrP genotypes were determined. Sex diagnosis was done by PCR amplification of ZFX/ZFY and SRY sequences after PEP-PCR while PrP genotype determination was performed after specific pre-amplification of specific target including codons 136, 154 and 171. Embryos were collected at Day 7 after oestrus. Blastocysts and expanded blastocysts were biopsied immediately after collection whereas compacted morulae were biopsied after 24 hr of in vitro culture. Eighty-nine biopsied embryos were frozen by vitrification. Fresh and vitrified whole embryos were kept as control. DNA of biopsies was extracted and pre-amplified. Sex diagnosis was efficient for 96.6% of biopsies and PrP genotyping was determined in 95.8% of codons. After embryo transfer, no significant difference was observed in lambing rate between biopsied, vitrified control and fresh embryos (54.5%, 60% and 66.6%, respectively). Embryo survival rate was not different between biopsied and whole vitrified embryos (P = 0.38). At birth, 96.7% of diagnosed sex and 95.4% of predetermined codons were correct. Lamb PrP profiles were in agreement with parental genotype. PEP-PCR coupled with sex diagnosis and nested PCR coupled with PrP genotype predetermination are very accurate techniques to genotype ovine embryo before transfer. These original results allow planning of selection of resistant genotype to scrapie and sex of offspring before transfer of cryopreserved embryo [less ▲]

Diagnosis and monitoring of pregnancy in sheep: reality and perspectives

in Productions Animales (2003), 16(2), 79-90

The present review aims at presenting different methods for pregnancy diagnosis. Since these methods are to be used in the herds, precocity, sensitivity, specificity, accuracy to predict pregnant and non ... [more ▼]

The present review aims at presenting different methods for pregnancy diagnosis. Since these methods are to be used in the herds, precocity, sensitivity, specificity, accuracy to predict pregnant and non-pregnant ewes, and the possibility to determine fetal numbers are carefully analysed. The progesterone assay is accurate as early as day 17th after fecundation; but the fertilization day must be precisely known. The pregnancy specific or associated glycoproteins (PSPB/PAG) assay is highly sensitive and specific at day 22 in blood and at day 32 in milk samples. However, this assay does not allow to predict the fetal number. The ultrasonography B-mode used on 30 days of gestation gives very good or excellent performance in terms of sensitivity and specificity depending on the practitioner. The method is also able to predict the real time fetal number. [less ▲]

Amélioration des méthodes de cryopréservation et de transfert d’embryons chez les petits ruminants

in Proceedings: 8e Rencontres autour des Recherches sur les Ruminants (2001)

ln small ruminants, the costs of embryo transfer is a main limiting factor to the use of this method. The use of ultra rapid techniques such as embryo vitrification and direct transfer may contribute to ... [more ▼]

ln small ruminants, the costs of embryo transfer is a main limiting factor to the use of this method. The use of ultra rapid techniques such as embryo vitrification and direct transfer may contribute to reduce a part of the costs and increase the use of embryo transfer in sheep and goats. ln order to evaluate the efficiency of these techniques, two experiments were performed. ln a first experiment the viability of vitrified/thawed embryos was compared to results obtained after transfer of fresh embryos in ewes or frozen embryos (with slow freezing method) in goats. The pregnancy raté at term as weIl as embryo survival rate did not differ significantly according to embryo treatment (in ewes : 72% and 60% for fresh embryos vs 72% and 50% for vitrified embryos; in goats : 69% and 55% for frozen embryos vs 48% and 39 % for vitrified embryos). ln a second experiment, the possibility to transfer the vitrified embryos or frozen embryos directly after thawing (without cryoprotectant removal and evaluation of the morphological status of the embryos) was tested by comparison with the standard technique of transfer of vitrified or frozen/thawed embryos (removal of cryoprotectant and morphological evaluation). No significant effect of the transfer method was observed on the pregnancy rate at term and embryo survival rate (in ewes/vitrified embryos : 67% and 49% for traditional transfer vs 75% and 53% for direct transfer ; in goats/ vitrified embryos : 23% and 15% for traditional transfer vs 38% and 26% for direct transfer ; in goats/frozen embryos : 74% and 45% for traditional transfer vs 71% and 57% for direct transfer) [less ▲]

Utilisation répétée des gonadotropines exogènes dans le contrôle de la reproduction: justifications et effets secondaires potentiels

in Annales de Médecine Vétérinaire (1998), 142(2), 373-396

Gonadotropins of pituitary origin (follitropin -FSH-, lutropin -LH-, human menopausal gonadotropin -hMG) or trophoblastic origin (human chorionic gonadotropin - hCG- also called pregnant urine ... [more ▼]

Gonadotropins of pituitary origin (follitropin -FSH-, lutropin -LH-, human menopausal gonadotropin -hMG) or trophoblastic origin (human chorionic gonadotropin - hCG- also called pregnant urine gonadotropin -pU-, equine chorionic gonadotropin -eCG- also called pregnant mare serum gonadotropin -PMSG-) are largely used to treat infertility or to better control sexuality in various species of mammals: bovine, canine, caprine, equine, feline, human,ovine, porcine, rabbit, ... This manuscript reports the discovery of these molecules, describes their biochemical characteristics and gives the main indications (e.g. oestrus induction and synchronization, ovulation, superovulation, fertility improvement, anoestrus treatment, receptivity induction) in animals and human. Litterature data reporting on effects and possible side effects of their repeated use are presented. Due to slight differences in amino acids sequences, carbohydrates contents and tridimentionnal structures, repeated use of exogeneous gonadotropins may induce immunological reactions leading to non specific reactions and lower biological response. A reflection concludes this revue, pointing out the positive influence of the philogenetic distance existing between the species origin of the gonadotropin (human or equine) and the treated one, on the intensity of the immune reaction. [less ▲]

Anti-PMSG in sheep subjected annualy to oestrus synchronisation

in Reproduction Nutrition Development (1997), 37

Estimation of the long-term consequences on reproduction performance of the oestrus synchronisation treatments that are annually applied to ewes was carried out on nine officially controlled dairy flocks ... [more ▼]

Estimation of the long-term consequences on reproduction performance of the oestrus synchronisation treatments that are annually applied to ewes was carried out on nine officially controlled dairy flocks in the Roquefort region of France. A hormonal treatment combining the insertion of a vaginal fluoro-gestone acetate (FGA) sponge for 14 days and the injection of about 500 IU of pregnant mare serum gonadotropin (PMSG) at withdrawal was applied to the ewes in seven of the nine flocks. The ewes in the two other flocks were used as controls. Blood samples were taken from each female just before the treatment (to test for the presence of residual antibodies) and 20 days after the PMSG injection. Anti-PMSG antibody binding rates were calculated for each blood sample. The residual binding rate increased with age and induce negative effects on the following years reproduction performances, ie, they increased the probability that the ewes would not become pregnant. [less ▲]