The effect of day 5 blastocyst assessment timing on live birth prediction and development of a prediction algorithm.

RESEARCH QUESTION: Does variation in day 5 assessment timing confound live birth prediction using snapshot blastocyst morphology and is it possible to develop a numerical prediction algorithm? DESIGN: Retrospective multicentre cohort study including 4851 autologous oocyte single day 5 fresh embryo transfers performed at 11 Monash IVF clinics between 2016 and 2020. Repeat cycles of the same patients were excluded to avoid clustering effects in regression analysis. RESULTS: Hours post insemination (HPI) at day 5 assessment (115.9 ± 2.6 h) significantly correlated with blastocyst developmental stage (r = 0.118, P < 0.001). Independent association (expressed as adjusted odds ratio [aOR] and 95% confidence interval [CI]) was identified between live birth and HPI (aOR 0.950, 95% CI 0.925-0.976, P < 0.001) after accounting for blastocyst morphology and a range of patient/cycle characteristics. Algorithms were constructed using four significant live birth predictors: HPI at day 5 assessment, blastocyst developmental stage (aOR 1.347, 95% CI 1.217-1.491, P < 0.001), morphological grade (aOR 1.314, 95% 1.197-1.443, P < 0.001) and maternal age (aOR 0.922, 95% CI 0.907-0.936, P < 0.001). Receiver operating characteristic (ROC) analysis showed consistent predicting performance of algorithms via five-fold cross-validation, with similar area under the ROC curve (AUC 0.718, 0.715, 0.720, 0.712, 0.726, P < 0.001, respectively, in development subsets; and AUC 0.718, 0.731, 0.709, 0.741, 0.684, P < 0.001, respectively, in validation subsets). A score (ranging from 0.1 to 4.7) calculator based on the final algorithm was subsequently created. CONCLUSIONS: Day 5 assessment timing is a confounding factor for live birth prediction using snapshot blastocyst morphology. A numerical algorithm incorporating day 5 assessment HPI, blastocyst morphology and maternal age can be developed for live birth prediction.

Morphometric and morphokinetic differences in the sperm- and oocyte-originated pronuclei of male and female human zygotes: a time-lapse study.

PURPOSE: To study the morphometric and morphokinetic profiles of pronuclei (PN) between male and female human zygotes. METHOD(S): This retrospective cohort study included 94 consecutive autologous single day 5 transfer cycles leading to a singleton live birth. All oocytes were placed in the EmbryoScope + incubator post-sperm injection with all annotations performed retrospectively by one embryologist (L-SO). Timing parameters included 2nd polar body extrusion (tPB2), sperm-originated PN (tSPNa) or oocyte-originated PN (tOPNa) appearance, and PN fading (tPNF). Morphometrics were evaluated at 8 (stage 1), 4 (stage 2), and 0 h before PNF (stage 3), measuring PN area (um2), PN juxtaposition, and nucleolar precursor bodies (NPB) arrangement. RESULTS: Male zygotes had longer time intervals of tPB2_tSPNa than female zygotes (4.8 ± 0.2 vs 4.2 ± 0.1 h, OR = 1.442, 95% CI 1.009-2.061, p = 0.044). SPN increased in size from stage 1 through 2 to 3 (435.3 ± 7.2, 506.7 ± 8.0, and 556.3 ± 8.9 um2, p = 0.000) and OPN did similarly (399.0 ± 6.1, 464.3 ± 6.7, and 513.8 ± 6.5 um2, p = 0.000), with SPN being significantly larger than OPN at each stage (p < 0.05 respectively). More male than female zygotes reached central PN juxtaposition at stage 1 (76.7% vs 51.0%, p = 0.010), stage 2 (97.7% vs 86.3%, p = 0.048), and stage 3 (97.7% vs 86.3%, p = 0.048). More OPN showed aligned NPBs than in SPN at stage 1 only (44.7% vs 28.7%, p = 0.023). CONCLUSION(S): Embryos with different sexes display different morphokinetic and morphometric features at the zygotic stage. Embryo selection using such parameters may lead to unbalanced sex ratio in resulting offspring.

Live birth in a complete zona-free patient: a case report.

OBJECTIVE: To report a live birth from a patient with complete zona-free oocytes due to abnormal zona production and to reveal full time-lapse blastocyst development footage of its originating embryo. METHODS: A 34-year-old woman presented with a history of failed fertilization via standard in vitro fertilization insemination and a potential absence of zona pellucida. A total of 3 intracytoplasmic sperm injection cycles were undertaken with all oocytes collected being zona-free. Embryos created in the initial 2 cycles were cultured in the G1+/G2+ sequential media in a benchtop incubator. During the final successful cycle, the culture strategy was shifted to single step media (G-TL) in an Embryoscope+ incubator. RESULTS: The first 2 attempts led to a biochemical pregnancy or no blastocyst available for transfer. In the third cycle, 13 out of 24 collected oocytes were subjected to injection, with 4 being normally fertilized. Two blastocysts were subsequently formed, in which one was cryopreserved and the other transferred. A live baby girl (1570g) was subsequently delivered at 34 weeks of gestation by cesarean section. CONCLUSION: Live birth can be achieved for patients with zona production deficiency. Adjustment in ovarian stimulation and subsequent embryo culture strategies may have potentially contributed to the success of the 3rd cycle.