Oocyte activation is initiated when a fertilising spermatozoon delivers sperm-borne oocyte-activating factor(s) into the oocyte cytoplasm. Candidates for oocyte activation include two proteins, phospholipase Cζ1 (PLCZ1) and postacrosomal WW-binding protein (PAWP; also known as WBP2 N-terminal like (WBP2NL)). We localised PLCZ1 and WBP2NL/PAWP in stallion spermatozoa and investigated the PLCZ1 content and sperm parameters as well as cleavage after intracytoplasmic sperm injection (ICSI). PLCZ1 was identified as 71-kDa protein in the acrosomal and postacrosomal regions, midpiece and principal piece of the tail. Anti-WBP2NL antibody identified two WBP2NL bands (~28 and ~32kDa) in the postacrosomal region, midpiece and principal piece of the tail. PLCZ1 and WBP2NL expression was positively correlated (P=0.04) in sperm heads. Flow cytometry evaluation of PLCZ1 revealed large variations in fluorescence intensity and the percentage of positively labelled spermatozoa among stallions. PLCZ1 expression was significantly higher in viable than non-viable spermatozoa, and DNA fragmentation was negatively correlated with PLCZ1 expression and the percentage of positively labelled spermatozoa (P<0.05). The use of equine sperm populations considered to have high versus low PLCZ1 content resulted in significantly higher cleavage rates after ICSI of bovine and equine oocytes, supporting the importance of PLCZ1 for oocyte activation.

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We retrospectively analysed data from heterospermic and homospermic boar semen for motility and morphology during a 2‐year period. Homospermic doses were also evaluated for viability, acrosome integrity, DNA fragmentation, osmolality and pH. Additionally, we investigated the effect of temperature upon arrival and the agreement between viability and motility as evaluating tool. We observed lower (p < .05) total motility (TM) and normal sperm morphology within summer and fall. Conversely, lower (p < .05) progressive motility (PM) was found at the beginning and end of each year. Viability and acrosome integrity were reduced (p < .05) in summer months but not exclusively, suggesting that samples could be compromised by transport temperature. Sperm DNA fragmentation was <6% with a small variation. Medium osmolality and pH slightly changed (p < .05). Sperm count was not source of variation on sperm parameters. Sample temperature upon arrival correlated with PM and VSL (p < .05), while motility was reduced <12°C (p < .05). Homospermic doses were less affected by season and arrival temperature, having better parameters (p < .05) than contemporaneous heterospermic samples but influenced by genetic line (p < .05). We found a high agreement between viable acrosome‐intact sperm and TM, especially when TM was ≥80%. Our data verify the improvement of sperm quality during time as sperm count/dose does not affect quality, but season effect persists regardless of ejaculate selection at the stud. Homospermic doses exhibited better parameters than heterospermic doses, seemingly being more resilient to temperature variations, suggesting that selection for sperm quality within boars selected by growth traits can improve the product quality.