P-262 Clinical viability of oocytes with micronuclei at fertilisation
Authors: Liyanage S , Johnson K , Leung V , Evans P , Stephens N , Sorby K 
Published: July 2024

Abstract

Study question
Does the presence of micronuclei at fertilisation impact on embryo potential?

Summary answer
Embryos displaying micronuclei at fertilisation exhibit decreased utilisation rates and lower euploidy rates, however are capable of resulting in clinical pregnancies.

What is known already
The appearance of two pronuclei defines normal fertilisation. With advances in imaging techniques, a range of pronuclei phenotypes can now be identified in more detail, including additional micronuclei. Micronuclei are smaller satellite structures appearing around the pronuclei. Previous studies have determined that embryos containing two primary pronuclei and one micronuclei are generally diploid and can develop to blastocyst stage. However, these studies were limited in sample size, did not have consistent imaging techniques and assessed only the presence or absence of micronuclei, not their size or number.

Study design, size, duration
Retrospective cohort analysis in a single, large IVF centre including all oocytes entering culture between January 2022 and December 2023 (n = 33,237). All embryos were cultured in Embryoscope+ using Vitrolife sequential media at 6% CO2 and 5% O2. ICSI and naturally inseminated oocytes were included. All embryos were cultured to day 5/6 before determining suitability for transfer, freezing and/or genetic testing.

Participants/materials, setting, methods
During routine daily annotation, embryologists assessed fertilised oocytes for the presence of micronuclei. This study re-examined all embryos annotated as displaying micronuclei and further assessed: number of micronuclei, their position (‘apposed’ or ‘distant’ from the pronuclei, and in the case of &gt; 1 micronuclei ‘clustered’ or ‘spread’) along with the area (μm2) of the two largest micronuclei and area (μm2) of primary pronuclei using Embryoscope+ software. Clinical pregnancy was defined as presence of a gestational sac.

Main results and the role of chance
Micronuclei were observed in 0.45% (n = 149/33,237) of all oocytes assessed. The majority of these presented as one additional nuclei 92.60% (n = 138), 7.38% (n = 11) showed two additional nuclei and 0.67%(n = 1) had multiple (more than 2) micronuclei. All micronuclei appeared apposing a primary pronucleus. In the event of multiple micronuclei, 82% were clustered whereas 9% was spread throughout the around the primary pronuclei. The average area of the largest micronuclei in each oocyte was 80.33μm2 ± 26.81.The mean area of all micronuclei was 76.80μm2, approximately 16% of the size of primary pronuclei measured, 465.09μm2.
The utilisation rates of embryos derived from micronuclei fertilisation were 28.18% (N = 42) compared to 49%, n = 4118 of 2PN derived embryos. All but one of these presented two primary pronuclei and one additional micronuclei, with the remaining embryo displaying two micronuclei. Twelve of these embryos were transferred (in a fresh stimulated cycle (n = 5) or a subsequent frozen embryo transfer cycle (n = 7), resulting in four clinical pregnancies, however only one ongoing clinical pregnancy.
PGT testing was performed on 19 embryos, with only two, 6.67% returning a euploid results, 68.42% resulting an aneuploid, of the remaining four embryos, 2 achieved a mosaic result and 2 received a no result.

Limitations, reasons for caution
As the occurrence of micronuclei is rare, only a limited number of embryos were available for analysis. Furthermore, this study was conducted in a large single laboratory and can not account for differences in culture conditions between laboratories. Inter-operator variability among initial fertilisation annotations could not be excluded.

Wider implications of the findings
By creating a definition of the micronuclei measurement, it provides safeguard for micronuclei to be assessed in culture. The data presented here can help guide clinical decisions where micronuclei are present.

Trial registration number
not applicable