Effects of postpartum diseases on antral follicle count and serum concentration of Anti-Müllerian hormone in dairy cows.

The antral follicle count (AFC) and Anti-Müllerian hormone (AMH) concentration are validated markers for ovarian reserve in cattle, but their use as fertility markers is controverse. Here we assessed the effects of postpartum diseases on AFC and AMH concentration, as well as the influence of parity and breed on these parameters. Cows (n = 513, mostly Holstein Friesian and Brown Swiss, parity 3.0 ± 1.8) underwent a single ultrasonography examination 28-56 days after parturition and categorized as having low (n ≤ 15 follicles), intermediate (n = 16-24 follicles), or high (n ≥ 25 follicles) AFC based on objective video analysis of recorded sequences. Blood samples for AMH determination were collected at the time of examination and animals divided into low (< 0.05 ng/ml) and high AMH (≥ 0.05 ng/ml) group, respectively. No effects of postpartum diseases or breed on either AFC or AMH groups could be observed. There was a strong interaction between parity and AFC, primiparous cows having less follicles (13.6 ± 6.2 vs. 17.1 ± 7.0, P < 0.001) than pluriparous cows. The AFC did not affect reproductive parameters or productivity of the cows. In comparison, pluriparous cows with high AMH concentration had shorter calving to first service (86.0 ± 37.6 vs. 97.1 ± 46.7 days, P < 0.05) and calving to conception (123.8 ± 51.9 vs. 135.8 ± 54.4 days, P < 0.05) intervals, but lower milk yield (8440.3 ± 2292.9 vs. 8927.9 ± 2192.5 kg, P < 0.05) compared to cows with low AMH. In conclusion, no effect of postpartum diseases on AFC or AMH concentration of dairy cows could be observed. However, an interaction between parity and AFC, as well as associations of AMH with fertility and productivity in pluriparous cows, were demonstrated

Dynamic regulation of the transcriptome and proteome of the equine embryo during maternal recognition of pregnancy

During initial maternal recognition of pregnancy (MRP), the equine embryo displays a series of unique events characterized by rapid blastocyst expansion, secretion of a diverse array of molecules, and transuterine migration to interact with the uterine surface. Up to date, the intricate transcriptome and proteome changes of the embryo underlying these events have not been critically studied in horses. Thus, the objective of this study was to perform an integrative transcriptomic (including mRNA, miRNAs, and other small non-coding RNAs) and proteomic analysis of embryos collected from days 10 to 13 of gestation. The results revealed dynamic transcriptome profiles with a total of 1311 differentially expressed genes, including 18 microRNAs (miRNAs). Two main profiles for mRNAs and miRNAs were identified, one with higher expression in embryos ≤5 mm and the second with higher expression in embryos ≥7 mm. At the protein level, similar results were obtained, with 259 differentially abundant proteins between small and large embryos. Overall, the findings demonstrated fine-tuned transcriptomic and proteomic regulations in the developing embryo associated with embryo growth. The identification of specific regulation of mRNAs, proteins, and miRNAs on days 12 and 13 of gestation suggested these molecules as pivotal for embryo development and as involved in MRP, and in establishment of pregnancy in general. In addition, the results revealed new insights into prostaglandin synthesis by the equine embryo, miRNAs and genes potentially involved in modulation of the maternal immune response, regulation of endometrial receptivity and of late implantation in the mare

Uterine extracellular vesicles as multi-signal messengers during maternal recognition of pregnancy in the mare

In contrast to other domestic mammals, the embryo-derived signal(s) leading to maternal recognition of pregnancy (MRP) are still unknow in the mare. We hypothesize that these embryonic signals could be packed into uterine extracellular vesicles (uEVs), acting as multi-signal messengers between the conceptus and the maternal tract, and contributing to MRP. To unveil these signals, the RNA and protein cargos of uEVs isolated from uterine lavages collected from pregnant mares (P; day 10, 11, 12 and 13 after ovulation) and cyclic control mares (C; day 10 and 13 after ovulation) were analyzed. Our results showed a fine-tuned regulation of the uEV cargo (RNAs and proteins), by the day of pregnancy, the estrous cycle, and even the size of the embryo. A particular RNA pattern was identified with specific increase on P12 related to immune system and hormonal response. Besides, a set of proteins as well as RNAs was highly enriched in EVs on P12 and P13. Differential abundance of miRNAs was also identified in P13-derived uEVs. Their target genes were linked to down- or upregulated genes in the embryo and the endometrium, exposing their potential origin. Our study identified for first time specific molecules packed in uEVs, which were previously associated to MRP in the mare, and thus bringing added value to the current knowledge. Further integrative and functional analyses will help to confirm the role of these molecules in uEVs during MRP in the mare

Oviductal extracellular vesicles miRNA cargo varies in response to embryos and their quality

BACKGROUND: Increasing evidence points to an active role of oviductal extracellular vesicles (oEVs) in the early embryo-maternal dialogue. However, it remains unclear whether oEVs contribute to the recognition of the presence of embryos and their quality in the oviduct. Hence, we examined whether the molecular cargo of oEVs secreted by bovine oviduct epithelial cells (BOEC) differs depending on the presence of good (≥ 8 cells, G) or poor (< 8 cells, P) quality embryos. In addition, differences in RNA profiles between G and P embryos were analyzed in attempt to distinguish oEVs and embryonic EVs cargos. METHODS: For this purpose, primary BOEC were co-cultured with in vitro produced embryos (IVP) 53 h post fertilization as follows: BOEC with G embryos (BGE); BOEC with P embryos (BPE); G embryos alone (GE); P embryos alone (PE); BOEC alone (B) and medium control (M). After 24 h of co-culture, conditioned media were collected from all groups and EVs were isolated and characterized. MicroRNA profiling of EVs and embryos was performed by small RNA-sequencing. RESULTS: In EVs, 84 miRNAs were identified, with 8 differentially abundant (DA) miRNAs for BGE vs. B and 4 for BPE vs. B (P-value < 0.01). In embryos, 187 miRNAs were identified, with 12 DA miRNAs for BGE vs. BPE, 3 for G vs. P, 8 for BGE vs. GE, and 11 for BPE vs. PE (P-value < 0.01). CONCLUSIONS: These results indicated that oEVs are involved in the oviductal-embryo recognition and pointed to specific miRNAs with signaling and supporting roles during early embryo development

Structural variant-based pangenome construction has low sensitivity to variability of haplotype-resolved bovine assemblies

Advantages of pangenomes over linear reference assemblies for genome research have recently been established. However, potential effects of sequence platform and assembly approach, or of combining assemblies created by different approaches, on pangenome construction have not been investigated. Here we generate haplotype-resolved assemblies from the offspring of three bovine trios representing increasing levels of heterozygosity that each demonstrate a substantial improvement in contiguity, completeness, and accuracy over the current Bos taurus reference genome. Diploid coverage as low as 20x for HiFi or 60x for ONT is sufficient to produce two haplotype-resolved assemblies meeting standards set by the Vertebrate Genomes Project. Structural variant-based pangenomes created from the haplotype-resolved assemblies demonstrate significant consensus regardless of sequence platform, assembler algorithm, or coverage. Inspecting pangenome topologies identifies 90 thousand structural variants including 931 overlapping with coding sequences; this approach reveals variants affecting QRICH2, PRDM9, HSPA1A, TAS2R46, and GC that have potential to affect phenotype

Structural variant-based pangenome construction has low sensitivity to variability of haplotype-resolved bovine assemblies

Advantages of pangenomes over linear reference assemblies for genome research have recently been established. However, potential effects of sequence platform and assembly approach, or of combining assemblies created by different approaches, on pangenome construction have not been investigated. Here we generate haplotype-resolved assemblies from the offspring of three bovine trios representing increasing levels of heterozygosity that each demonstrate a substantial improvement in contiguity, completeness, and accuracy over the current Bos taurus reference genome. Diploid coverage as low as 20x for HiFi or 60x for ONT is sufficient to produce two haplotype-resolved assemblies meeting standards set by the Vertebrate Genomes Project. Structural variant-based pangenomes created from the haplotype-resolved assemblies demonstrate significant consensus regardless of sequence platform, assembler algorithm, or coverage. Inspecting pangenome topologies identifies 90 thousand structural variants including 931 overlapping with coding sequences; this approach reveals variants affecting QRICH2, PRDM9, HSPA1A, TAS2R46, and GC that have potential to affect phenotype

Effect of oxytocin infusion on luteal blood flow and progesterone secretion in dairy cattle

The objective of this study was to investigate the effects of oxytocin infusion on corpus luteum (CL) function during early to mid-diestrus by measuring luteal size (LS) and luteal blood flow (LBF) along with plasma levels of progesterone (P4) and prostaglandin metabolites (13,14-dihydro-15-keto-prostaglandin F2α, PGFM). On day (D) 7 of the estrus cycle (D1 = ovulation), seven cows received 100 IU of oxytocin (OXY) or placebo (PL) following a Latin square design. LS and LBF increased in both groups over time and no differences were observed between the groups. PGFM did not differ either within the groups over time or between the groups at any time point. P4 of the OXY group was higher compared to that of the the PL group 360 min after the infusion (p = 0.01) and tended to be higher at the time points 450 min, 48 h, and 72 h (all p = 0.08). Results from this study support the hypothesis that OXY is not directly involved in the mechanism(s) governing blood flow of the CL and has no remarkable effects either on luteal size or P4 and PGFM plasma levels. Further investigation is needed to elucidate the role of OXY in CL blood flow during early and late luteal phases

Rapid Accumulation of Polymorphonuclear Neutrophils in the Corpus luteum during Prostaglandin F2α-Induced Luteolysis in the Cow

Prostaglandin F2α (PGF2α) induces luteolysis within a few days in cows, and immune cells increase in number in the regressing corpus luteum (CL), implying that luteolysis is an inflammatory-like immune response. We investigated the rapid change in polymorphonuclear neutrophil (PMN) numbers in response to PGF2α administration as the first cells recruited to inflammatory sites, together with mRNA of interleukin-8 (IL-8: neutrophil chemoattractant) and P-selectin (leukocyte adhesion molecule) in the bovine CL. CLs were collected by ovariectomy at various times after PGF2α injection. The number of PMNs was increased at 5 min after PGF2α administration, whereas IL-8 and P-selectin mRNA increased at 30 min and 2 h, respectively. PGF2α directly stimulated P-selectin protein expression at 5–30 min in luteal endothelial cells (LECs). Moreover, PGF2α enhanced PMN adhesion to LECs, and this enhancement by PGF2α was inhibited by anti-P-selectin antibody, suggesting that P-selectin expression by PGF2α is crucial in PMN migration. In conclusion, PGF2α rapidly induces the accumulation of PMNs into the bovine CL at 5 min and enhances PMN adhesion via P-selectin expression in LECs. It is suggested that luteolytic cascade by PGF2α may involve an acute inflammatory-like response due to rapidly infiltrated PMNs

Relationship between Clinical Uterine Findings, Therapy, and Fertility in the Mare

Persistent breeding-induced endometritis (PBIE) is a major cause of subfertility in horses and the susceptibility is increased by several factors. The aim of this study was to determine the effects of clinical uterine findings and PBIE therapies, respectively, on pregnancy rate in mares. The analysis included records from 220 mares (390 cycles) inseminated at an artificial insemination (AI) center in Switzerland. Gynecological examinations were performed repeatedly before and after AI to determine cervical tone, uterine edema, and intrauterine fluid accumulation. Pregnancy rate was lower (p 0.05) regarding uterine edema. Intrauterine fluid accumulation reduced pregnancy rate (p = 0.002). Oxytocin administration had beneficial effects on pregnancy rate (p = 0.015), especially for barren mares, while uterine lavage did not have any effect (p > 0.05). The results show that cervical tone and intrauterine fluid accumulation, but not its degree, are useful parameters for assessment of fertility in mares. Oxytocin treatment improved pregnancy rates in mares with PBIE while uterine lavage had a limited effect

Transrectal Doppler sonography of uterine blood flow during the first two weeks after parturition in Simmenthal heifers

Transrectal Doppler sonography was used to evaluate uterine blood flow during the first two weeks after parturition in six primiparous Simmental cows. The uterine blood flow was evaluated on the day of parturition (Day 0), once daily from Days 1 to 8 and then every other day until Day 14. Blood flow was quantified by determining the diameter (D), the time-averaged maximum velocity (TAMV), the pulsatility index (PI) and the blood flow volume (BFV) of the uterine arteries ipsilateral and contralateral to the formerly pregnant uterine horn. During the first four days after calving D, TAMV and BFV declined (ipsilateral: TAMV 70%, BFV 87%, contralateral: D 47%, BFV 84%; p ＜ 0.05), while PI increased (ipsilateral 158%, contralateral 100%; p ＜ 0.05) distinctly. Between Days 4 and 14 only the ipsilateral D (12%) and the BFV of both arteries (ipsilateral 5%, contralateral 8%) decreased (p ＜ 0.05). Blood flow variables were very strongly correlated with each other (r ＞ ±0.75, p ＜ 0.05), with negative correlations with PI and positive correlations with all other investigated factors. Overall, this study revealed characteristic changes in uterine perfusion during the first two weeks after parturition in cows that were pronounced during the first four days postpartum