Use of a new computer-assisted sperm analyzer for the assessment of motility and viability of dog spermatozoa and evaluation of four different semen extenders for predilution

摘要

The aims of this study were to evaluate a new computer-assisted sperm analyzer (CASA; SpermVision, Minitub, G) for the analysis of canine spermatozoa, and to suggest suitable semen diluents that should influence motility and viability parameters to the lowest extent. For these purposes, the sperm-rich fractions from 40 ejaculates were diluted with either saline (S), PBS, autologous prostate secretion (third fraction, PROST) or a modified TRIS buffer (TRIS; [Pena, A., Johannisson, A., Linde-Forsberg, C., 1999. Post-thaw evaluation of dog spermatozoa using new triple fluorescent staining and flow cytometry. Theriogenology 52, 965-980]). Viability was investigated both objectively with SYBR-14/PI in SpermVision and subjectively with carboxyfluoresceindiacetate (CFDA) fluorescence stains by optical microscopy. For all diluents and for most parameters, the intra- and inter-assay coefficients of variation were below 10% and 20%. Effects after dilution with TRIS and PROST were similar, but only after dilution with TRIS, total motility significantly correlated with estimated motility in raw semen (P<0.01, R=+0.514). After dilution, estimated and measured motility values were positively related, but correlated significantly with PBS only (P<0.01). After dilution with PBS and S, all motility parameters were significantly lower in comparison with PROST and TRIS, and after dilution with PBS, significantly more membrane damages occurred than with TRIS, S and PROST (P<0.05, <0.05 and <0.01). In samples diluted with TRIS, membrane integrity of spermatozoa determined with SYBR-14/PI and CFDA staining correlated significantly (R=+0.686; P<0.01). Concerning the location of membrane damages, no differences appeared after dilution with different media. In conclusion, SpermVision proved to be an accurate analyzer for the objective assessment of both motility and viability of canine spermatozoa. TRIS and PROST were the media with the lowest influences on parameters of canine semen, and are therefore recommendable for the use in SpermVision.