47 Comparative transcriptome analysis of embryos from beef oocyte donors fed endophyte-infected tall fescue

Endophyte-infected tall fescue consumption is connected to reproductive failures and early embryonic losses in beef cattle. Cows grazing endophyte-infected tall fescue (E+) have a 41% lower conception rate than those on endophyte-free (E−) pastures. Research indicates that the critical period for ergot alkaloids negatively affecting conception is around ovulation and the first 6 days of embryonic development. Therefore, this study aimed to understand the biological mechanisms underlying embryonic losses by analyzing the gene expression profiles of embryos from beef oocyte donors affected by fescue toxicosis. Twenty-four Simmental × Angus heifers were divided into two groups receiving different dietary treatments for a period of 45 days: 12 heifers received E+ seeds, while 12 heifers were fed E− seeds, mixed with their diet. E+ heifers received a daily dose of 20 μg of ergovaline per kilogram of bodyweight. Follicles were aspirated from each heifer for IVF to produce embryos at the end of the trial. In total, eight single embryos at Day 7 of development (n = 4 per group) underwent RNA extraction (0.58–0.71 ng mL⁻¹). RNA libraries were prepared with the SMARTer® Stranded Total RNA-Seq Kit v3 (Takara Bio Inc.), which adds a unique molecular identifier (UMI) for downstream data analysis. Reads were filtered, trimmed, and aligned to the bovine reference genome (ARS-UCD1.2). PCR duplicates were removed based on the UMIs, and then gene counts were estimated. Normalized gene counts were analyzed to detect differentially expressed genes (DEGs) between the two groups. The total number of embryos that developed into the blastocyst stage was lower in the E+ group (n = 9) compared with the E− group (n = 15). In addition, all E+ embryos reached blastocyst stage 6, while 14 of the E− embryos achieved expanded blastocyst stage 7, suggesting a slower development of E+ embryos. Out of the 13 107 expressed genes, 79 were differentially expressed between the two groups of embryos (FDR < 0.05), with all DEGs upregulated in embryos from E+ donor heifers. Gene enrichment analysis, performed using ShinyGO 0.80 (http://bioinformatics.sdstate.edu/go/), revealed molecular functions (FDR < 0.05) of DEGs (e.g. ANKRD1, APOE, AXL, C1QTNF3, CCN2, CDH11, CDK6, COL11A1, COL5A1, COL5A2, CSRP3, CTHRC1, DCLK1, F5, FOXJ2, GJA1, ITGAV, NECTIN2, PDGFC, PLXNA2, PROS1, RARRES2, RUNX1, TAGLN, and TMEM176B) that are involved in embryo development (gastrulation and formation of the primary germ layer) and cellular processes (proliferation, differentiation, motility, and migration). Response to stress and coagulation processes were also enriched. These results suggest that E+ heifers had altered maternal oocyte content that negatively affected early embryonic development. Altered gene expression of E+ embryos may lead to embryonic losses. Alternatively, to cope with the effects of fescue toxin, E+ embryos may trigger the expression of genes that play important functions in embryonic development. To our knowledge, this study is the first to report embryonic gene expression changes due to E+ exposure before fertilization in beef cattle. It may provide novel insights to mitigate the effects of fescue toxicosis on beef reproduction efficiency.