Factors influencing oocyte recovery during ultrasound-guided follicle aspiration in mares: A postmortem study

The objectives of this study were to investigate the influence of different OPU variables (vacuum and injection pressures during follicle aspiration and flushing, needle twisting to scrape follicles, number of follicle flushes and type of flushing media) on oocyte recovery rate (ORR) and morphology. Overall, 120 postmortem excised ovaries were processed in 62 replicates (1.9 ± 0.7 ovaries per replicate), with a total of 1336 follicles punctured and aspirated (11.1 ± 6.2 follicles per ovary) by ultrasound-guided follicle aspiration resembling the OPU procedure from live mares. The aspiration pressure (50 vs. 300 mmHg) did not influence (P > 0.1) ORR (57.1 and 55.1 %, respectively) but the higher pressure (1.9 mL/s aspiration flow) resulted in the recovery of more denuded oocytes than the lower pressure (0.8 mL/s) (66 % vs 42 %, P < 0.05). High injection pressure to flush follicles (1.5 mL/s) was associated with lower (P < 0.05) ORR (31 %) and higher loss of oocytes in the leaked fluid (10 % of aspirated follicles) than lower pressures (37 % and 47 % ORR for 0.3 and 0.8 mL/s injection flow rates, respectively). The number of flushes per follicle influenced (P < 0.05) the ORR: 28.8, 46.1, 42.7, 43.0 and 64.2 % for x0, x1, x3, ×6 and ×10 flushes groups. More oocytes (P < 0.05) were lost in the leaked fluid outside the ovary in groups ≥ ×3 (5-7%) than in ×0 and ×1 flushes (<1 %). The ORR was superior following scraping of follicle wall by twisting the needle (55.2 %) than no scraping (42.9 % ORR), while the type of flushing media (0.9%NaCl + heparin vs commercial flushing media with PVA) did not influence ORR. In conclusion, follicle scraping by needle twisting and follicle flushing during 10 times yielded the highest oocyte recovery. Repeated follicle flushing with high injection pressure was associated with a higher loss of oocytes outside the ovary in the leaked fluid.