High proportion of zygotes with multiple pronuclei increase the embryo multinucleation rate during conventional IVF.

The incidence of multiple pronuclei (≥ 3PN) zygotes and blastomere multinucleation was found to be elevated in the presence of increased estradiol (E) levels and a greater number of retrieved oocytes. This implies a potential link between the incidence of multinucleation at the two-cell stage (MN2) and a higher proportion of ≥ 3PN zygotes. We aimed to investigate the effect of high proportion of ≥ 3PN zygotes on MN2 incidence during conventional in vitro fertilization (C-IVF) by using time-lapse monitoring. This study included 1195 patients from January 2020 to December 2022. The patients were categorized into three groups: Group 1 comprised patients with no ≥ 3PN zygotes (n = 422), Group 2 included those with 0-25% ≥3PN zygotes (n = 617), and Group 3 consisted of patients with more than 25% ≥3PN zygotes (n = 156). The MN2 rate, types of MN2 and clinical outcomes were compared among the three groups. Our data indicated that the MN2 rate was significantly lower in groups 1 and 2 compared to group 3 (18.33 versus 25.62%; p < 0.001 and 19.45 versus 25.62%; p < 0.001). The MN2 embryos exhibited similar rates of high-quality embryos (42.27 versus 43.50 versus 40.67%; p = 0.401) and available embryos (84.96 versus 84.04 versus 83.21%; p = 0.460) rates among the three groups. There were no significant differences in the proportion of MN2 with different types among the three groups (p > 0.05). The embryos displaying binucleated at the two-cell stage in one blastomere (2BI1) and true multinucleated at the two-cell stage in one blastomere (2MULTI1) showed significantly higher blastocyst formation rates compared to embryos exhibiting true multinucleated at the two-cell stage in both blastomeres (2MULTI2) (59.50 versus 45.40%; p < 0.001 and 59.40 versus 45.40%; p < 0.001). In conclusion, the occurrence of MN2 events might be associated with high proportion of ≥ 3PN zygotes incidence. The types of MN2 had significant reference value when selecting embryos for transfer during the cleavage stage.