Eicosanoid-immunometabotypes reveal genotypic links to cervicovaginal pH in progestagen synchronised South African Dohne Merino ewes.

Metabolites can be detected in various bodily fluids such as blood, urine and cervicovaginal secretions. Cervicovaginal fluid has distinct characteristics such as colour and pH that are affected by the underlying health of an ewe and her associated metabolic and endocrine processes dedicated towards maintaining homeostasis particularly after stress is experienced. This study profiled distinct metabolomic phenotypes defined by the pattern and abundance of eicosanoid mediators that reflect the underlying state of Dohne Merino ewes' immune activation and metabolic regulation based on their comparative cervicovaginal metabolome and pH before and after the administration of an intravaginal pessary using high resolution liquid chromatography mass spectrometry. Further, single nucleotide polymorphisms (SNPs) were analysed using genome-wide association analysis (GWAS) to correlate genomic effects of cervicovaginal pH. Twenty ewes were allocated to each treatment group defined by a CIDR-, sponge- or injection-based estrus synchronisation protocol and their cervicovaginal fluid sampled using FLOQSwabs. This study matched three features to metabolites of the KEGG Ovis aries eicosanoid synthesis and breakdown pathway and found a significant effect of sampling point, prior to and after pessary administration, on cervicovaginal pH (p < 0.05). Sponge-based estrus synchronisation significantly increased cervicovaginal pH compared to the CIDR-based group (p < 0.05) but not compared to the control group (p > 0.05), where CIDR-based and control group ewes did not differ significantly from each other (p > 0.05). Ten SNPs were correlated to cervicovaginal pH with GWAS results visualised using Manhattan, QQ, principal component analysis and volcano plots where effect sizes (β) ranged from -0.5 to 1.5, minor allele frequencies (MAF) from 0.23 to 0.49 and the genomic inflation factor λ was 1.053. SNPSs were mapped to genes regulating mucosal inflammatory signalling, epithelial integrity, and metabolic stress responses, offering a biologically plausible mechanism through which genetic variation interacts with device-induced disruptions to cervicovaginal homeostasis to produce significantly higher vaginal pH in sponge-treated ewes. This study advocates for the continued robust characterisation of metabotypes for phenotypes relevant to livestock production and advances the conceptual framework of immunometabolism by incorporating pH as a critical axis of regulation within reproductive mucosal environments of ewes.