Effect of thawing method on bull sperm survival in ejaculates frozen in 4 ml and 8 ml volumes

Abstract

The aim of this study was to evaluate the effect of different thawing protocols (slow (P1), medium (P2), and fast (P3)) on percentage of motile sperm (MOT) and percentage of sperm cells with intact membranes (INT) in Holstein (4 bulls; 72 samples) and Czech Fleckvieh (4 bulls; 72 samples) semen frozen-thawed in 4 ml and 8 ml volumes. MOT was analysed in fresh semen, as well as immediately after thawing (T0) and 30 min after thawing (T30). INT was analysed using hypoosmotic swelling test (HOS test) in fresh ejaculate (FE), after diluting (DE), and at T0. The differences between FE parameters and frozen-thawed ejaculate parameters, expressing changes that occur during cryopreservation, were calculated. Apart from the effect of thawing protocol, the effect of breed and the effect of quality of FE expressed by MOT immediately after collecting were evaluated, too. Unlike thawing of semen cryopreserved in straws (0.25 and 0.5 ml), thawing using the slow protocol (P1) was the most appropriate (P < 0.05) for both observed volumes. There were found significantly higher MOT in the volume of 8 ml in both T0 and T30 and in the volume of 4 ml in T30 in samples thawed using P1 and P2. MOT in T0 was significantly affected by breed in samples frozen in 8 ml and in T30 in samples frozen in 4 and 8 ml. There were found no significant differences in INT in all reported volumes, however decrease of INT during cryopreservation was affected by breed.