398 Subclinical Lipopolysaccharide Exposure During Oocyte Maturation Disrupts Early Bovine Embryonic Development
Authors: Castro Briza , Candelaria Juliana , Austin Molly , Shuster Charles , Gifford Craig A , Denicol Anna C , Gifford Jennifer A Hernandez 
Published: November 2023

Abstract
Reduced conception rates and infertility are associated with bacterial infections in cattle. Chronic and acute gram-negative bacteria release of lipopolysaccharide (LPS) endotoxin elicits robust immune responses capable of disrupting folliculogenesis, steroidogenesis, oocyte competence, and embryo development. However, the effects of subclinical or non-detectable infections on oocyte competence and subsequent embryo development remain to be fully elucidated. Therefore, the objective of this study was to investigate varying concentrations of LPS on oocyte and embryo competence. Bovine oocytes were collected from slaughterhouse-derived ovaries and incubated with vehicle control or increasing doses of LPS (0, 0.01, 0.1, and 1 µg/mL) for 21 h during in vitro maturation. Oocytes (n = 252 in 3 replicates) were fixed and stained for evaluation of nuclear maturation. A subset of oocytes (n = 153 in 3 replicates) was fertilized and cultured in a time-lapse incubator where individual well images were captured every 5 min for 8 d for video compilation and subsequent analysis. A set of embryos from LPS-matured oocytes (n = 1,126 in 5 replicates) were cultured for 8 d to evaluate blastocyst cell counts (n = 163 in 5 replicates). This evaluation included total, trophectoderm, and inner cell mass cell numbers. Results demonstrate no significant difference among 0.01, 0.1, or 1 µg/mL LPS treatment groups and the percent of oocytes at metaphase II (MII), meiosis I (MI), or germinal vesicle (GV) when compared with vehicle-treated controls after 21 h maturation (P = 0.44, 0.26, and 0.90, respectively). Likewise, no differences were detected among 0.01, 0.1, or 1 µg/mL LPS treatment groups and presumptive zygotes that did not cleave after fertilization (P = 0.84), those that cleaved but arrested at the 2-cell stage (P = 0.50), 4-cell (P = 0.76), prior to morula (P = 76), or morula stage (P = 0.60). However, the percentage of blastocysts derived from oocytes matured in 0.01 µg/mL LPS tended to decrease compared with vehicle-treated controls (11.38 ± 8.74 and 25.45 ± 2.73%, respectively, P = 0.09). Similarly, the proportion of oocytes that developed to the blastocyst stage was greater in vehicle-treated controls (25.45%) compared with embryos derived from oocytes matured in 0.1 (5.92 ± 3.62% P = 0.03) and 1 µg/mL (6.55 ± 3.62%, P = 0.03) LPS. Lipopolysaccharide treatment did not affect total blastocyst cell count (P = 0.73), as indicated by the number of cells of the trophectoderm (P = 0.45), and inner cell mass (P = 0.30). Therefore, these data suggest that although nuclear maturation was not affected by low LPS concentrations, LPS-matured oocytes that subsequently underwent in vitro fertilization, experienced decreased competence to develop to the blastocyst stage. Therefore, non-detectable or subclinical bacterial infections have the capability to decrease embryonic development in cattle.