Approaches to Identify Pregnancy Failure in Buffalo Cows.

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.

Plasmatic Profile of Pregnancy-Associated Glycoprotein (PAG) during Gestation and Postpartum in Sarda and Lacaune Sheep Determined with Two Radioimmunoassay Systems.

This study was carried out to determine ovine pregnancy-associated glycoprotein (oPAG) levels in the plasma of Sarda and Lacaune ewes throughout gestation and in the first month postpartum, using two heterologous radioimmunoassays (RIA-706 and RIA-srPool) and to study the correlations between PAG levels and fetal gender and number. On Day 18 of pregnancy, PAG concentrations were detected in 90.1% and 80.8% of Sarda pregnant ewes with RIA-706 and RIA-srPool, respectively; and in 90% and in 75% of Lacaune pregnant ewes with RIA-706 and RIA-srPool, respectively. From Day 30, PAG concentrations were detected in all pregnant ewes by using both RIA methods. In the postpartum period, the PAG concentrations in Sarda ewes decreased rapidly reaching minimal levels (<1 ng/mL) on day 28 using both RIA-706 and RIA-srPool. In Lacaune ewes, PAG-706 levels were higher than PAG-srPool from parturition until the last day of observation (Day 28 postpartum). It was also observed that mean concentrations were higher in multiple than in single pregnancies in Sarda and Lacaune ewes. Moreover, due to the rapid disappearance of PAG concentration following lambing, a cut-off limit in postpartum animals was not required as a means for detecting a new pregnancy.

Characterization of CD79αcy+ cells in placentas from ruminants.

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.

Purification of pregnancy-associated glycoproteins from late-pregnancy Bubalus bubalis placentas and development of a radioimmunoassay for pregnancy diagnosis in water buffalo females.

BACKGROUND: Pregnancy-associated glycoproteins (PAGs) were first described as placental antigens present in the blood serum of the mother soon after implantation. Here, we describe the purification of several pregnancy-associated glycoproteins from water buffalo placenta (wbPAGs). A specific radioimmunoassay (RIA) was developed for early pregnancy diagnosis in buffalo species. RESULTS: Amino-terminal microsequencing of immunoreactive placental proteins allowed the identification of eleven wbPAGs sequences [Swiss-Prot accession numbers: P86369 to P86379]. Three polyclonal antisera (AS#858, AS#859 and AS#860) were raised in rabbits against distinct wbPAG fractions. A new RIA (RIA-860) was developed and used to distinguish between pregnant (n=33) and non-pregnant (n=26) water buffalo females. CONCLUSIONS: Our results confirmed the multiplicity of PAG expression in buffalo placenta. In addition, the RIA-860 system was shown to be sensitive, linear, reproducible, accurate and specific in measuring PAG concentrations in buffalo plasma samples from Day 37 of gestation onwards.

Double radial immunodiffusion as a tool to identify pregnancy-associated glycoproteins in ruminant and nonruminant placentae.

Pregnancy-associated glycoproteins (PAGs) are antigens synthesized in the superficial layers of the ruminant trophoblast. Initially, they were identified either as proteins released into the maternal bloodstream (where they have applications in pregnancy diagnosis) (PAG1) or as molecules binding to the LH receptor (PAG2). In this study, double radial immunodiffusion was used to test the ability of antisera raised against different PAG molecules (bovine, ovine and caprine) to react with placental extracts from nonruminants (rabbit, cat, mouse, pig, and wild pig) and ruminants (cow, ewe, and goat). Placental extracts from all nonruminants tested except rabbit reacted with anti bovine PAG2 (anti-boPAG2). Extracts of ruminant placentas reacted with different antisera, confirming the expression of various PAG molecules. According to the time at which the placentas were collected (early or middle pregnancy), the reaction differed as regards the thickness, position, and number of precipitation lines, suggesting that PAG expression varies as pregnancy progresses. Bos indicus and Bos taurus placental extracts exhibited different reactions with anti-boPAG2: a single precipitation line in the former case and two lines in the latter. This suggests differential expression of boPAG2 related glycoproteins in these two subspecies.