In beef cattle, one common measure of reproductive performance, in a herd that uses estrous synchronization followed by embryo transfer (ET), is the number of females pregnant divided by the number of females synchronized and that received an ET (pregnancy/ET; P/ET). In general, P/ET measured at 30 days of expected pregnancy is in the 30 to 50% range [1], [2]. Part of the failure in pregnancies is attributed to sub-optimal estrous synchronization, specifically in timed-ET programs [3], [4], [5]. However, when ET is performed only in recipients that showed estrus followed by ovulation the P/ET is rarely greater than 50% [6], [7]. The outcome to each opportunity of pregnancy after ET is likely the composite result of the competence of the embryo for development and signaling to the mother, ability of the uterus to support embryonic development, and to be receptive to embryonic signals, the embryonic competence by uterine ability interaction, and other factors extrinsic to the pregnancy [8], [9]. It is likely that uterine receptivity varies among recipients and varies among pregnancy opportunities within an individual recipient.

Ascertaining uterine ability to support embryo development of an individual female is challenging, as it requires measuring the pregnancy outcome over repeated opportunities, on a short interval of time (to be able to minimize the effect of aging and confounding with season of the year) and while minimizing variation in other factors that influence pregnancy. Geary and co-authors [10] gave virgin heifers three or four subsequent opportunities to maintain a pregnancy after estrous synchronization and ET and classified the heifers according to the pregnancy outcomes as high fertile (pregnant in all opportunities), subfertile (pregnant in one opportunity) or infertile (never pregnant). Authors determined that endometrial and luminal molecular characteristics varied according to the heifer fertility classification [11], [12]. For example, subfertile heifers had a three-fold decreased expression of endometrial genes associated with regulation of extracellular matrix (ECM) structure and organization, cell adhesion, and cell movement that could alter conceptus adhesion to the luminal epithelium [12], [13]. These findings provided mechanistic clues for the variation in the ability of females to maintain pregnancy after ET. Variables such as those determined by endometrial gene expression are challenging to measure directly, without disturbing the ongoing pregnancy. An alternative, less invasive approach would be to measure systemic changes in maternal function associated with pregnancy. In this regard, expression of interferon-stimulated genes (ISGs) in peripheral blood mononuclear cells (PBMC) are a proxy for IFNT secretion by the conceptus, and serum concentrations of pregnancy-associated glycoproteins (PAGs) reflect placentation [14], [15], [16], [17]. There is a gap in knowledge regarding the relationship between the abundance of ISGs and PAGs and the fertility classification of embryo recipients.

Expression of ISGs in PBMCs or polymorphonuclear leukocytes has served as a marker for pregnancy status between days 18 and 21 of gestation. Classical ISGs (ISG15, RSAD2, OAS1) exhibited increased expression in pregnant versus nonpregnant cows [14], [15], [16], [18], [19]. The magnitude of ISG expression in blood immune cells was also associated with the parity and breed of the cow [16], [20]. Whether such magnitude of ISGs expression in PBMCs is associated with the fertility classification of cattle is still unclear. Studies examining the uterine transcriptome of heifers of varying fertility showed no difference in mRNA abundance of ISGs, despite fertile heifers exhibiting greater conceptus length compared to subfertile heifers [12], [21]. Furthermore, ISG expression has been used as a method of identifying pregnancy loss, but there have been conflicting results regarding differential expression between pregnant cows and those that experienced pregnancy loss [16], [22].

PAGs are produced by the trophoblast cells of the conceptus and have been identified as an indirect pregnancy specific marker used to identify pregnancy status in cattle. Since the first identification of PAGs, known as pregnancy specific protein B (PSPB) or PAG-1, over 20 separate PAG genes have been identified, all of which are pregnancy specific [23].Therefore, blood concentration of PAGs has been used as a method of pregnancy detection after day 28 of gestation [16], [24]. Among heifers with varying fertility, PAG concentrations remained similar on day 28 and 44 of gestation [10], [25]. Concentrations of PAG among cows sired by bulls classified as having high or low sire conception rate (SCR) were similar on day 33 of gestation [26]. However, serum and plasma PAG concentrations have been researched as a marker of embryonic/fetal viability or loss at various stages of gestation [27].

The main hypothesis of this study was that expression of ISGs in PBMCs and abundance of PAGs in serum is greater in ET recipients classified as fertile than ET recipients classified as subfertile. A secondary hypothesis was that expression of ISGs in PBMCs and abundance of PAGs in serum is greater in ET recipients that retain the pregnancy vs. recipients that lose the pregnancy. The objectives were to: (1) compare the expression of the ISGs, RSAD2 and ISG15, isolated from PBMCs 20 days after estrus in ET recipients classified by fertility and test the accuracy of ISGs to predict pregnancy attainment and pregnancy loss and (2), compare the concentrations of PAGs on day 25, 32, and 39 after estrus in ET recipients classified by fertility and test the accuracy of PAGs to predict pregnancy attainment and pregnancy loss.