The response to buck effect (BE) was studied in Beni Arouss goats during anoestrus (AS) and breeding season (BS). Prior to AS, bucks were exposed to artificial long days during 75 days followed by a natural photoperiod. Goats of group 1 were treated for 11 days with 20 mg of fluorogestone acetate (FGA) combined to 300 IU of eCG and 50 µg of cloprostenol injected 48 h prior to FGA-removal. Goats of group 2 and 3 were subjected to the same treatments, but eCG injection was replaced by a sexually active buck introduced 0 (group 2) or 48 hours (group 3) before FGA-removal. During the AS, 77% of goats induced by the BE showed an oestrus at [60–74] hr following FGA-removal for group 3 and [71–77] hr for group 2 (p < .05). In group 1, 77% of goats displayed an LH surge [24–40] hr after FGA-removal and 67% developed a luteal phase, but no LH surge or luteal response was detected in groups 2 and 3. During the BS, oestrus response reached 100% in goats synchronised with the BE ([22–68] hr in groups 2 and 3). In all groups, 84% of the goats displayed an LH surge at [30–70] hr post-treatment and 69% displayed luteal phase within 3–8 days post-treatment. After 11–15 days, the occurrence of ovulations followed by normal luteal phase was raised in all groups (91%). It was concluded that photostimulated bucks failed to induce and synchronise ovulation in goats previously treated with 20 mg of FGA and 50 µg of cloprostenol during anoestrus.Highlights The response to the buck effect was studied in Beni Arouss goats during anoestrus and breeding season. The use of sexually active bucks to induce and to synchronise oestrus and ovulation in goats treated with 20 mg of FGA and cloprostenol during the anoestrus season is not efficient in comparison with an hormonal treatment based on administration of FGA, eCG and cloprostenol. The same protocol appears as an adequate alternative for oestrus and ovulation synchronisation during the breeding season.

Mature granulated trophoblast binucleate cells (BNC) have been found in all ruminant placentas examined histologically so far. BNC are normally fairly evenly distributed throughout the fetal villus and all their granules contain a similar variety of hormones and pregnancy associated glycoproteins (PAGs). Only the Giraffe is reported to show a different BNC protein expression, this paper is designed to investigate that. Gold labelled Lectin histochemistry and protein immunocytochemistry were used on deplasticised 1 μm sections of a wide variety of ruminant placentomes with a wide range of antibodies and lectins. In the Giraffe placentomes, even though the lectin histochemistry shows an even distribution of BNC throughout the trophoblast of the placental villi, the protein expression in the BNC granules is limited to the BNC either in the apex or the base of the villi. Placental lactogens and Prolactin (PRL) are present only in basally situated BNC: PAGs only in the apical BNC. PRL is only found in the Giraffe BNC which react with many fewer of the wide range of antibodies used here to investigate the uniformity of protein expression in ruminant BNC. The possible relevance of these differences to ruminant function and evolution is considered to provide a further example of the versatility of the BNC system.

Transmission of bluetongue (BT) virus serotype 8 (BTV-8) via artificial insemination of contaminated frozen semen from naturally infected bulls was investigated in two independent experiments. Healthy, BT negative heifers were hormonally synchronized and artificially inseminated at oestrus. In total, six groups of three heifers received semen from four batches derived from three bulls naturally infected with BTV-8. Each experiment included one control heifer that was not inseminated and that remained BT negative throughout. BTV viraemia and seroconversion were determined in 8 out of 18 inseminated heifers, and BTV was isolated from five of these animals. These eight heifers only displayed mild clinical signs of BT, if any at all, but six of them experienced pregnancy loss between weeks four and eight of gestation, and five of them became BT PCR and antibody positive. The other two infected heifers gave birth at term to two healthy and BT negative calves. The BT viral load varied among the semen batches used and this had a significant impact on the infection rate, the time of onset of viraemia post artificial insemination, and the gestational stage at which pregnancy loss occurred. These results, which confirm unusual features of BTV-8 infection, should not be extrapolated to infection with other BTV strains without thorough evaluation. This study also adds weight to the hypothesis that the re-emergence of BTV-8 in France in 2015 may be attributable to the use of contaminated bovine semen.

The aim of this work was to find the best strategy to diagnose pregnancy failures in buffalo. A total of 109 animals belonging to a buffalo herd subjected to a synchronization and artificial insemination (AI) program were enrolled in this study. Blood samples were collected at days 0, 14, 25, 28 and 40 after AI for the determination of progesterone (P4) and pregnancy-associated glycoproteins (PAGs) by the radioimmunoassay (RIA) method. Transrectal ultrasonography was performed on day 25, 28 and 40 after AI to monitor pregnancy. The animals included in the data analysis were assigned ex post in pregnant (n = 50) and mortality (n = 12) groups. By ultrasonography, the predictive sign of mortality was the heartbeat. At day 25, the PAGs concentration was significant in predicting embryonic mortality with respect to ultrasonography and P4, at the cut-off of 1.1 ng/mL. At day 28, either PAGs, at a cut-off of 2.2 ng/mL, or ultrasonography, with no detection of heartbeat, were highly predictive of embryonic mortality. PAGs were the best marker (p < 0.05) for predicting embryonic mortality between 25 and 40 days of gestation in buffalo. Its utilization as a diagnostic tool can influence management decisions in order to improve farm reproductive management.

The aim of this study was to further develop and extensively describe a surgical technique in order to realise long-term fetal blood sampling in the bovine species. Eleven Holstein-Friesian 6-to 8-month pregnant cows (4-10 years old) were used for this study. Gestational age on the day of surgery varied from approximately 180 days (n = 1) to 240 days (minimum: 232 days, maximum 252 days; n = 10). The fetal medial tarsal artery was catheterised in pregnant cows with a polyvinyl catheter in dorsal recumbency under general anaesthesia. Although 5 out of 11 operations (45.5%) performed between 232 and 252 days of gestation were lost due to different causes mainly associated with peritonitis and septicaemia, the mean interval between operations and calvings was 42.5 days (between 27 and 95 days). It is important to emphasise that a well-trained surgical team is needed for bovine fetal cannulation in order to be able to decrease the risk factors during the operations. Due to the fact that after 5 unsuccessful cases none of the pregnancies were lost, this skill can be reached, and our technique can enable bovine fetal blood sampling for long-term endocrinological and physiological investigations before and during parturition.

Twenty-four Colombian Creole, Romney Marsh, Hampshire and Corriedale ewes raised under high-altitude conditions in a non-seasonal country were used to determine and to characterize the onset of puberty and of regularity ovarian function since the age of 3months. They underwent blood collection for determination of progesterone and monthly weight assessment. Oestrus was assessed daily by a teaser ram. In all breeds, age at onset of puberty ranged from 5.4 to 6.9 months and age at onset of regular ovarian function ranged from 7.4 to 8.6 months. Colombian Creole showed a higher body development at onset of puberty: 73.5 ± 8.3% versus 56.2 ± 7.4 in Romney Marsh, 58.8 ± 10.4 in Hampshire, and 57.3 ± 8.0 in Corriedale (p < 0.05), as well as a higher relative daily weight gain (+17%, p < 0.05). A negative correlation between daily weight gain and age at onset of puberty was established. Progesterone at onset of puberty and of regularity did not differ between breeds, but characterization of the luteal phase by the progesterone area under the curve (P4-AUC) revealed at both time points significantly lower values for Creole lambs (p < 0.05). Decreased P4-AUC paralleled a higher proportion of short oestral cycles observed prior to onset of regularity in Creoles, whereas an increased proportion of extra-long cycles were recorded in Romney Marsh (p < 0.05). These results establish first reference data for economically important breeds raised under tropical conditions. In comparison with other breeds, Colombian Creole requires a higher body development to achieve puberty and that an important proportion of short characterizes its prepubertal period cycles.

This study was designed ability using antibody against buffalo PAG to detect concentration of pregnancy-associated glycoprotein (PAG) in bovine plasma samples. Successful purification of buffalo PAG molecules with high enrichment degree by using Vicia villosa aûnity chromatography was recently reported. In this research, the RIA system using novel anti buffalo-PAG was used for bovine pregnancy diagnosis. In a total 437 blood sampling of dairy cow in Belgium were used. Three PAG-RIA systems were undertaken to measure PAG concentration in bovine routine blood samples: AS#497 (or RIA-1) using rabbit antisera against purified bovine PAG67kDa, AS#706 (or RIA-2) using rabbit antisera against caprine PAG55+62kDa and new AS#859 (or RIA-3) using rabbit antisera against buffalo PAG59.5-75.8kDa). The measured PAG concentrations were similar for the systems As#497 and As#859; although the dilution was shown parallel, the measured PAG concentrations were significantly different between systems As#706 and As#859. On the other hand, the coefficients of correlations between concentrations obtained by the use of As#859 and As#497 were highly comparable to those between As#859 and As#706 or As#497 and As#706. In conclusion, the present study clearly shows that antisera against buffalo PAG can be used to develop new PAG-RIA system for bovine pregnancy diagnosis with high comparable results.

Primiparous and multiparous lactating crossbred dairy cows with a mature corpus luteum and a follicle with >10 mm in diameter were treated with cloprostenol. Those cows that showed oestrus within 5 days after treatment were inseminated (Group P). The other cows (Group PG) were treated with GnRH 2 days after cloprostenol treatment and timed artificial insemination (AI) was performed on the consecutive day, or were inseminated (Group G) after detected oestrus and treated with GnRH immediately after AI. The control cows (Group C) after detected oestrus were only inseminated. All of the AIs using frozen semen were done between 6 and 7 a.m. while the ultrasonographic examinations after AI were performed between 4 to 6 p.m. The ovaries of each cow were scanned by means of transrectal ultrasonography from the day of AI until ovulation. Daily blood samples were collected for progesterone measurements. The ovulation and pregnancy rates among the groups changed between 84.6% and 95.5%, as well as 44.4% and 60%, respectively, however the differences were not statistically significant. All the cows were evaluated according to date of ovulation after AI and the pregnancy rate was 55.4% (Group 1: ovulation occurred between AI and 9-11 h after AI), 54.5% (Group 2: ovulation occurred between 9-11 h and 33-35 h after AI) and 35.5% (Group 3: ovulation occurred between 33-35 h and 57-59 h after AI), respectively. There was a trend (P=0.087) for 2.2 greater odds of staying open among cows inseminated between 33 to 35 h and 57 to 59 h before ovulation compared to cows inseminated within 9 to 11 h before ovulation. If ovulation occurred before AI, the pregnancy rate was only 22.2%, therefore determination of optimal time for AI is of great importance.

Previous work carried out to characterise different immune cells in ruminant placentas found strong CD79αcy nuclear labelling in cells histologically resembling trophoblast cells. In the attempt to characterize this cell population, placentomes collected from cattle, sheep and water buffaloes were examined by immunohistochemistry with single and double labelling using monoclonal antibodies (mAb) against B lymphocytes and trophoblast cells. Most CD79αcy + cells co-expressed placental lactogen or cytokeratin and were CD21 and MHC class II negative strongly suggesting they do not have a B cell origin. However, a potential immunological role of these cells cannot be ruled out and it is currently unknown if the findings described may have an impact on physiological knowledge, health, and or diseases pathogenesis in ruminants.

This study was conducted to characterise pregnancy-associated glycoprotein (caPAG) in peripheral plasma during gestation and postpartum periods of nulliparous and multiparous does with one or two foetuses using a caPAG specific two-step sandwich ELISA system. Earliest time-points for detection of pregnancy and foetal number with appropriate cut-off values were identified. Plasma samples from 15 pregnant (multiparous: n = 8; nulliparous: n = 7; during pregnancy and postpartum period) and six non-pregnant (during oestrous cycle) goats were collected and analysed. Mean caPAG concentration was greater than the threshold for pregnancy detection (S-N = 0.40) on d22, peaked on d45 and remained unchanged until parturition. From d45 until parturition, caPAG concentration in multiparous does with two foetuses was 1.4 to 1.8 fold greater (P < 0.001) than those with one foetus. For the ELISA, 0.83 (S-N) was the most appropriate cut-off to differentiate does with two from those with a single foetus with an overall sensitivity and accuracy of 88.9% and 84.7%, respectively. Circulating caPAG concentration in multiparous goats was greater (P < 0.05) compared with nulliparous goats during the early pregnancy and postpartum periods. After parturition, caPAG concentrations markedly decreased and were basal within 14 days postpartum. In conclusion, using the caPAG specific ELISA, results indicated there were unique gestational and postpartum profiles for caPAG concentrations that are affected by number of foetuses and parity of the doe. The marked decrease in concentration of caPAG following parturition indicates there would not be compromising of the detection of subsequent pregnancies in goats using this technique.

The efficacy of eight combinations of fluorogestone acetate (FGA, 20 or 40 mg as intravaginal device during 11 days), equine chorionic gonadotropin (eCG, 300 or 500 UI injected 48 hr before FGA removal) and prostaglandin F2α (cloprostenol, 0 or 50 μg injected 48 hr before FGA removal) aiming at induction and synchronization of oestrus and ovulation was evaluated during the anoestrus season in spring and during the breeding season in autumn in adult Beni Arouss goats. Oestrous behaviour was recorded between 12 and 60 hr after FGA removal. Blood samplings allowing to assess onset of the pre-ovulatory LH surge and increase of progesterone as sign of an active corpus luteum were performed, respectively, between 20 and 60 hr and 3, 5, 8 and 15 days after FGA removal. No season-related differences (spring vs. autumn) were observed for oestrous response (95% vs. 93%), pre-ovulatory LH surge (94% vs. 84%) and luteal response after 3-8 and 11-15 days post-treatment (respectively 92% vs. 66% and 92% vs. 98%). The onset of oestrus (21 [13-53] vs. 32 [12-54] hr) and LH surge (26 [20-60] vs. 38 [22-60] hr) occurred significantly later in autumn. FGA (40 vs. 20 mg) in autumn significantly delayed the onset of oestrus (36 [16-54] vs. 23 [12-47] hr) and LH surge (44 [26-58] vs. 33 [22-60] hr). Significant treatment-related differences were recorded for onset of LH surge (earliest for 20 mg FGA, 300 IU eCG, 50 μg PGF2α ) and onset of luteal phase (latest for 40 mg FGA, 300 IU eCG, 50 μg PGF2α ). In conclusion, the hormone combinations tested appeared equally effective in terms of oestrous and ovulation rates. Season has influenced significantly the onset of oestrus and LH surge, and the high dose regimen of FGA delayed the ovarian response in autumn.

: The main objective of this study was to assess PAG2 mRNA expression in maternal blood cells at the peri-implantation period in water buffalo; moreover, we wanted to evaluate the earliest time in which PAG-2 could be detected in maternal blood. Thirty-two lactating buffaloes artificially inseminated (AI) were utilized. Blood was collected at Days 0, 14, 18, 28, 40 after AI (AI = day 0). Pregnancy was diagnosed by ultrasound at Days 28 and 40 post AI. Out of 32 buffaloes, 14 were pregnant (P group) and 18 were not pregnant (NP group). The plasma PAG-2 threshold of 1.0 ng/mL in the P group was reached at day 40 post AI. PAG2 mRNA expression differed between the P and NP groups, and was either evaluated in Peripheral Blood Mononuclear Cells (PBMC) or Polymorphonuclear Leukocytes (PMN), starting from day 14. However, both the estimated marginal means and multiple comparisons showed that PAG2 mRNA expression was higher in PMN than PBMC. In the present study, PAG-2 appeared in the blood (40 Days post AI), and an early expression of PAG2 mRNA at Day 14 post AI was also observed. Although further research is undoubtedly required, PAG2 mRNA in peripheral blood leukocytes could be using to better understand the role that PAGs play during pregnancy in buffalo.

This is the first time that PAG determination using two different antisera raised against PAG molecules purified from both caprine (RIA-706) and bubaline placentas (RIA-860) is reported in water buffalo. Ninety-eight buffalo cows, belonging to a buffalo herd subjected to a synchronization and artificial insemination (AI) programme, were enrolled in this study. Blood samples were taken on days 0 (AI), 23, 25, 28, 30 and 45. Pregnancy was confirmed by ultrasonography on days 28 and 45. The blood of 20 buffaloes that had calved was tested every five days from the day of calving until day 50 postcalving. Differences in PAG concentrations were observed between pregnant and nonpregnant buffaloes starting from day 23 post AI using both RIA-706 and RIA-860 (p < 0.001). However, estimated mean concentrations of PAG measured by RIA-706 were higher than RIA-860 (p < 0.001) and Bland-Altman analysis showed biases ranged from 0.0 ng/ml at day 23 to 0.79 ng/ml at day 28 post AI. Moreover, RIA-706 showed greater sensitivity and accuracy both at 23 and 25 days of pregnancy. RIA-706 and RIA-860 decreased below 1 ng/ml from 40 and 30 days postpartum, respectively, suggesting that PAG are better recognized by the antisera raised against the caprine PAG in the postpartum period also. This is essential when using PAG as an appropriate marker of early pregnancy after postpartum for detecting new pregnancies. The results of this study show that the ability of RIA systems to recognize early PAG could be improved using antisera raised against PAG molecules isolated from caprine placenta.

This study investigates for the first time mRNA pregnancy-associated glycoprotein 2 (PAG-2) expression in blood cells during early pregnancy in water buffalo. The PAGs constitute a large family of glycoproteins expressed in the outer epithelial layer of the placenta in eutherian species. All PAGs are not concomitantly expressed throughout pregnancy; some of them are expressed in the earlier phases, whereas others appear later and are expressed over a shorter period. Twenty-one lactating buffaloes were analyzed-17 females were synchronized with PRID and artificially inseminated (AI), whereas four females were synchronized but not inseminated (control group). Blood was collected at Days 0, 18, 28, 40, and 75 from AI (AI = Day 0). Expression of PAG-2 mRNA in blood samples was measured with real-time polymerase chain reaction. Pregnancy diagnosis was performed on Day 28 (D28) and Day 40 (D40) after AI by ultrasonography (US) and by PAG-1 RIA method. The females diagnosed pregnant at D28 and confirmed pregnant at D40 were defined as D28(+)D40(+) group; the females diagnosed pregnant at D28 but not confirmed pregnant at D40 were defined as D28(+)D40(-) group; and the females that were diagnosed as nonpregnant on either days were defined as D28(-)D40(-) group. PAG-2 mRNA at Day 0 was not observed in any groups. The D28(+)D40(+) group showed the highest expression, starting on Day 18 and increasing progressively up to Day 75. PAG-2 mRNA was also expressed on Day 18 in both D28(+)D40(-) and D28(-)D40(-) groups, but their levels were lower than those of D28(+)D40(+) group and almost constant over time. PAG-2 mRNA was never detected in the control group. The significant difference in the expression of PAG-2 mRNA between the D28(+)D40(+) group and the D28(-)D40(-) group, starting from Day 18, suggests that these animals might have conceived, but have experienced early embryonic loss; therefore, the PAG-2 mRNA was still present in blood circulation although at lower levels, as found in the D28(+)D40(-) group. In conclusion, this study shows that PAG-2 mRNA can be detected in peripheral maternal blood cells earlier than circulating PAG-1 molecules and could be useful for studies on early pregnancy and embryonic mortality. (C) 2016 Elsevier Inc. All rights reserved.

For the first time in literature this study describes the pregnancy-associated glycoprotein (PAG) profile of buffalo cows during gestation and the post-partum period using antiserum raised against PAG-molecules purified from buffalo placenta (AS#860).Ninety-eight buffalo cows, belonging to a buffalo herd subjected to a synchronization and artificial insemination (AI) program, were enrolled in this study. Blood samples were taken on days 0 (AI), 23, 25, 28, 30 and then biweekly until the end of pregnancy. Pregnancy was confirmed by ultrasonography on days 28 and 45, and by rectal palpation from day 60 onwards. Blood samples were suspended for the non-pregnant cows on day 45, while the blood of 20 buffaloes that had calved was tested every five days from the day of calving until day 50 post-calving. A cut-off value of 1.0 ng/mL was used in order to discriminate between pregnant and non-pregnant buffaloes. We used Linear Mixed models after Log(x+1) transformation to analyse the PAG concentrations.Fifty-two buffalo cows had become pregnant out of 98 synchronized (53%) and 46 remained nonpregnant (47%) as shown by ultrasonography and the PAG analysis. Significant differences (P < 0.001) in PAG concentrations were observed between the pregnant and non-pregnant buffaloes from day 23 as the PAG of the non-pregnant cows was always close to zero. Conversely, the PAG of the pregnant cows increased progressively from day AI until day 105 post-insemination and then stabilized until the end of pregnancy. Regarding pregnancy diagnosis, the sensitivity of PAG-RIA 860 system (ability of the test to correctly identify pregnant buffalo) ranged from 23% on day 23-98% on day 28 post AI; the specificity (ability to correctly identify non-pregnant buffaloes) was 100% throughout the sampling period. PAG progressively decreased from parturition to day 25 post-partum; from day 30 post-partum, the concentrations fell below 1 ng/mL and were close to 0 on the last day of observation (50 d post-partum).In conclusion, our results showed that RIA-860 is highly accurate for diagnosing pregnancy in buffaloes starting from day 28 of gestation. Furthermore, the rapid disappearance of PAG concentration after calving means that a cut-off limit in post-partum for detecting a new pregnancy is not required. (C) 2017 Published by Elsevier Inc.

L’efficacite d’un traitement de maitrise des cycles oestraux a ete appreciee sur deux races de zebus en elevage semi-intensif du Burkina Faso. L’essai a permis de determiner les proportions d’oestrus induits et de fertilite sur 170 femelles zebus Azawak et zebu Goudali traitees aux implants de norgestomet. Le traitement au norgestomet a consiste a la pose d’un implant a l’oreille pour une duree de 10 jours. Cette pose a ete suivie de deux injections de 2 ml (500 μg) de prostaglandine (Eustrumate) et 2 ml (400 UI) de PMSG (folligon) respectivement aux jours 8 et 10. Le critere majeur de venu en oestrus a ete l’acceptation du chevauchement. Les inseminations ont ete pratiquees en double 48 heures et 72 heures apres la fin du traitement. Le taux moyen d’oestrus induit a ete de 92,9%. Le delai moyen d’apparition de l’oestrus a ete de 33,15 ± 4,35 heures avec des variations entre types raciales. La duree moyenne de l’oestrus a ete de 11,7 ± 1,9 heures. Le taux moyen de fertilite a ete de 31,1%. Mot cles: zebu, oestrus induit, insemination artificielle, norgestomet English Abstract The efficiency of a estrous induced treatments was monitored on two zebu race in semi-intensives farms of Burkina Faso. The assay determined the rate of induced estrus and fertility on 170 zebu Azawak and zebu Goudali cows treated with norgestomet implant. The treatment Azawak zebu cows received Norgestomet ear implant for 10 days associated with estradiol valerate (2ml), and followed with PGF 2α ( 500 μg) and PMSG (400 UI) im injections respectively at days 8 and 10. Inseminations were practiced in double 48 hours and 72 hours after the end of the treatment. The proportion of estrus induced was 92.9% and was significantly affected (P < 0.05) by hormonal treatment. The elapsed time of estrus display was 33.15 ± 4.35 hours with variations between zebu types. The average length of estrus was 11.7 ± 1.9 hours. The average rate of fertility was of 31.1%. Keywords: zebu, estrus induced, artificial insemination, norgestomet