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Background: The intracytoplasmic sperm injection (ICSI) technique has low efficiency in cattle. This has mainly been attributed to the oocyte activation failure due to oocyte and/or sperm factors. Aim: Our aim was to evaluate the effect of conventional ICSI and Piezo-ICSI with bull or human sperm on bovine oocyte activation and embryo development and to assess its relationship with the phospholipase C zeta (PLCɀ) activity of both species. Methods: In vitro matured bovine oocytes were randomly divided into five groups and were fertilized as follows: conventional ICSI using bovine sperm with chemical activation (control), conventional ICSI using bovine sperm, Piezo-ICSI using bovine sperm, conventional ICSI using human sperm, and Piezo-ICSI using human sperm. PLCɀ activity was determined in bull and human sperm samples. Results: Within the groups using bull sperm, the oocytes fertilized by conventional ICSI had the lowest values of 2 pronuclei (PN) formation and cleavage, Piezo-ICSI increased both percentages and ICSI + chemical activation presented the highest 2 PN, cleavage, and blastocyst rates (p < 0.05). Within the groups using human sperm, the oocytes fertilized by Piezo-ICSI presented higher 2 PN and cleavage rates than those activated by conventional ICSI (p < 0.05). Piezo-ICSI with human sperm increased bovine oocyte activation as much as conventional ICSI + chemical activation with bovine sperm (p < 0.05). Higher values of PLCɀ activity were found in human sperm compared with bovine sperm (p < 0.05). Conclusion: Our results suggest that the higher stability of the bovine sperm in combination with its relatively low content of PLCɀ impairs bovine oocyte activation after ICSI. [ABSTRACT FROM AUTHOR]