Cooled-transported epididymides for donkey semen cryopreservation

In the event of death or unforeseen castration for medical reasons, epididymal semen harvesting represents the last opportunity to preserve valuable jacks’ genetics. Objective was to compare 2 semen cryopreservation methods (centrifugation versus noncentrifugation) on donkey cooled-shipped epididymides. We hypothesized that both methods would result in equivalent postthaw semen parameters. Experiment 1: donkeys (n = 7) housed at Bureau of Land Management in Tucson, AZ were surgically castrated. Testes and epididymides were placed in a plastic bag containing skim milk-based extender (25 ml, Botusemen, Botupharma, Scottsdale, AZ), and cooled-shipped overnight in an Equitainer to University of Illinois for semen cryopreservation. Each pair of epididymides was submitted to retrograde flushing with 5 - 10 ml with a cooling extender Botugold (Botupharma), or a similar volume of Botucrio (Botupharma). Botugold group samples were submitted to cushion-centrifugation (1000 g × 20 minutes), supernatant was discarded, and pellet was resuspended at 100 x106 sperm/ml in Botucrio. Botucrio group samples were resuspended to a similar final concentration. Semen from both groups was loaded in 0.5 ml straws, cooled at 5C for 20 minutes, placed 5 cm over LN 2 for 15 minutes, and then plunged in LN 2 for storage. Experiment 2: donkeys (n = 20) housed at Peaceful Valley rescue in San Angelo, TX were surgically castrated and epididymides dissected away from testes and each pair was shipped similar to Experiment 1, except that a different shipping container was used (Botuflex, Botupharma). Each pair was submitted to retrograde flushing with either Botugold or Botucrio and processed exactly as in Experiment 1. For both experiments, 1 straw for each group was thawed at 38C for 1 minute before sperm motility parameters assessments with CASA. Data were analyzed with R, using a mixed model Tukey’s as post hoc. Motility parameters were affected by shipment method (p < 0.05) but not by cushion centrifugation (p > 0.05) processing (Table). Donkeys had poor postthaw sperm motility compared to what was expected for donkey ejaculates subjected to cryopreservation. This was apparently the first study involving epididymal semen in donkeys.

Keywords: Epididymis, storage time, testis

This manuscript was originally published in the journal Clinical Theriogenology Vol 12(3) Sept 2020.  Clinical Theriogenology is the official journal of the Society for Theriogenology (SFT) and the American College of Theriogenologists (ACT).  This content has been reproduced on the IVIS website with the explicit permission of the SFT/ACT.