Double-Dose Artificial Insemination and Sperm Kinematics of Percoll Density Gradient-Sexed Semen in Holstein-Friesian Cattle

Abstract

This study aimed to evaluate the effect of sperm sexing using the Percoll density gradient centrifugation (PDGC) method on the kinematic, reproductive performance, and sex ratio of calves in Holstein-Friesian (HF) cattle. The study used three treatments: conventional semen (G0), PDGC sexed semen at 20%–60% (G1), and PDGC sexed semen at 20%–65% (G2). Kinematic variables were analyzed using a computer-assisted sperm analysis (CASA) system, while reproductive performance was evaluated based on non-return rate (NRR), conception rate (CR), calving rate (CvR), and sex ratio. Insemination in the sexed semen group was performed using a double-dose method to compensate for the potential decrease in viable sperm. The results showed that the PDGC method significantly affected several kinematic variables, particularly motility, distance average path (DAP), distance straight line (DSL), distance curved line (DCL), curvilinear velocity (VCL), straight-line velocity (VSL), average path velocity (VAP), linearity (LIN), straightness (STR), Wobble (WOB), amplitude of lateral head movement (ALH), and beat cross frequency (BCF) (p<0.05), reflecting changes in sperm movement dynamics due to the density-based separation process. However, there were no significant differences in reproductive performance variables between treatments, indicating that PDGC-based sexing does not significantly reduce in vivo fertilization capacity. The use of sexed semen increases the proportion of female offspring compared to conventional semen. Sexing sperm using the PDGC method is a relatively safe and applicable approach to increase the chances of female offspring without sacrificing reproductive efficiency, especially when combined with a double-dose insemination strategy.