Abstract
BackgroundThe outcome of cervical artificial insemination (AI) with frozen-thawed semen in sheep is limited by the inability of sperm to traverse the cervix of some ewe breeds. Previous research has demonstrated that cervical sperm transport is dependent on ewe breed, as sperm can traverse the cervix in greater numbers in some higher fertility ewe breeds. However, the molecular mechanisms underlying ewe breed differences in sperm transport through the cervix remain unknown. In this study, we aimed to characterise the cervical transcriptome of four European ewe breeds with known differences in pregnancy rates following cervical AI using frozen-thawed semen at the follicular phase of a natural oestrous cycle. Cervical post mortem tissue samples were collected from two Irish ewe breeds (Belclare and Suffolk; medium and low fertility, respectively) and from two Norwegian ewe breeds (Norwegian White Sheep (NWS) and Fur; high fertility compared to both Irish breeds) at the follicular phase of a natural oestrous cycle (n = 8 to 10 ewes per breed).ResultsHigh-quality RNA extracted from biopsies of the mid-region of the cervix was analysed by RNA-sequencing and Gene Ontology (GO). After stringent filtering (P < 0.05 and FC > 1.5), a total of 11, 1539 and 748 differentially expressed genes (DEGs) were identified in Belclare, Fur and NWS compared to the low fertility Suffolk breed, respectively. Gene ontology analysis identified significantly enriched biological processes involved in muscle contraction, extracellular matrix (ECM) development and the immune response. Gene co-expression analysis revealed similar patterns in muscle contraction and ECM development modules in both Norwegian ewe breeds, which differed to the Irish ewe breeds.ConclusionsThese breed-specific biological processes may account for impaired cervical sperm transport through the cervix in sheep during the follicular phase of the reproductive cycle. This novel and comprehensive dataset provides a rich foundation for future targeted initiatives to improve cervical AI in sheep.
Highlights
The outcome of cervical artificial insemination (AI) with frozen-thawed semen in sheep is limited by the inability of sperm to traverse the cervix of some ewe breeds
We identified that the use of exogenous hormones for oestrous synchronization did not affect the O-glycan composition in the high fertility ewe breeds, but it did in the other ewe breeds
Results showed no clear segregation between the two Irish ewe breeds, whereas comparing Fur and Norwegian White Sheep (NWS) against Suffolk showed a distinct grouping as illustrated on the principal component analysis (PCA) plot (Fig. 1)
Summary
The outcome of cervical artificial insemination (AI) with frozen-thawed semen in sheep is limited by the inability of sperm to traverse the cervix of some ewe breeds. We aimed to characterise the cervical transcriptome of four European ewe breeds with known differences in pregnancy rates following cervical AI using frozen-thawed semen at the follicular phase of a natural oestrous cycle. While fertilisation rates following laparoscopic AI with frozen-thawed semen were similar between Suffolk and Belclare ewes, both fertilisation rates and accessory sperm number following cervical AI with frozen-thawed semen was significantly higher in Belclare than the Suffolk ewes. This demonstrates that frozen-thawed sperm can traverse the cervix in greater numbers in some ewe breeds around the time of ovulation [5]. The migration of sperm through the cervix appears to be the critical limiting factor for the success of cervical AI, especially when frozen-thawed semen is used
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