Defining an Optimal Range of Centrifugation Parameters for Canine Semen Processing

SIMPLE SUMMARY: Processing canine semen is necessary for the preparation of cooled shipments or cryopreservation. Our goal was to define an acceptable range of centrifugation parameters (gravitational (g) force and time) without a severe negative impact on semen quality. We hypothesized that higher g forces (900 g vs. 400 g or 720 g) and longer treatment (10 min. vs. 5 min) improve sperm recovery rates yet causes a larger magnitude of decline in semen parameters over a 48 h cooling period. Initial raw semen evaluations served as each dog’s own control. Sperm recovery rates post-centrifugation were similar between treatment groups. Sperm plasma membrane integrity, motility and morphology were not different between treatment groups but declined over time. In conclusion, a range of 400–900 g for 5–10 min centrifugation provides clinically viable semen quality after up to 48 h of cooled storage in dogs. ABSTRACT: Our objective was to determine a clinically relevant range of centrifugation parameters for processing canine semen. We hypothesized that higher gravitational (g) force and longer time of centrifugation would result in improved spermatozoa recovery rate (RR) but poorer semen quality. Cooled storage under standard shipping conditions was used as a stressor to evaluate long-term treatment effects. Individual ejaculates collected from 14 healthy dogs were split into six treatment groups (400 g, 720 g, and 900 g for 5 or 10 min). Sperm RR (%) was calculated post-centrifugation, and plasma membrane integrity (%, Nucleocounter(®) SP-100™), total and progressive motility (%, subjective and computer-assisted sperm analysis), and morphology (%, eosin-nigrosin staining) were assessed on initial raw semen (T0), post-centrifugation (T1), and 24 h (T2) and 48 h (T3) after cooling. Sperm losses were minimal, and RRs were similar across treatment groups (median >98%, p ≥ 0.062). Spermatozoa membrane integrity was not different between centrifugation groups at any time point (p ≥ 0.38) but declined significantly during cooling (T1 vs. T2/T3, p ≤ 0.001). Similarly, total and progressive motility did not differ across treatments but declined in all groups from T1 to T3 (p ≤ 0.02). In conclusion, our study showed that centrifugation within a range of 400 g–900 g for 5–10 min is appropriate for processing canine semen.