Impacts of double biopsy and double vitrification on the clinical outcomes following euploid blastocyst transfer: a systematic review and meta-analysis.

study questionCompared to the 'single biopsy + single vitrification' approach, do 'double biopsy + double vitrification' or 'single biopsy + double vitrification' arrangements compromise subsequent clinical outcomes following euploidy blastocyst transfer? SUMMARY ANSWER: Both 'double biopsy + double vitrification' and 'single biopsy + double vitrification' led to reduced live birth/ongoing pregnancy rates and clinical pregnancy rates. WHAT IS KNOWN ALREADY?: It is not uncommon to receive inconclusive results following blastocyst biopsy and preimplantation genetic testing for aneuploidy (PGT-A). Often these blastocysts are warmed for re-test after a second biopsy, experiencing 'double biopsy + double vitrification'. Furthermore, to achieve better workflow, IVF laboratories may choose to routinely vitrify all blastocysts and schedule biopsy at a preferred timing, involving 'single biopsy + double vitrification'. However, in the current literature, there is a lack of systematic evaluation of both arrangements regarding their potential clinical risks in reference to the most common 'single biopsy + single vitrification' approach. STUDY DESIGN, SIZE, DURATION: A systematic review and meta-analysis were performed, with the protocol registered in PROSPERO (CRD42023469143). A search in PUBMED, EMBASE, and the Cochrane Library for relevant studies was carried out on 30 August 2023, using the keywords 'biopsy' and 'vitrification' and associated variations respectively. Only studies involving frozen transfers of PGT-A tested euploid blastocysts were included, with those involving PGT-M or PGT-SR excluded. PARTICIPANTS/MATERIALS, SETTING,

methodsStudy groups included blastocysts having undergone 'double biopsy + double vitrification' or 'single biopsy + double vitrification', with a 'single biopsy + single vitrification' group used as control. The primary outcome was clinical pregnancy, while secondary outcomes included live birth/ongoing pregnancy, miscarriage, and post-warming survival rates. Random effects meta-analysis was performed with risk ratios (RR) and 95% CIs were used to present outcome comparisons. MAIN RESULTS AND THE ROLE OF CHANCE: A total of 607 records were identified through the initial search and nine studies (six full articles and three abstracts) were eventually included. Compared to 'single biopsy + single vitrification', 'double biopsy + double vitrification' was associated with reduced clinical pregnancy rates (six studies, n = 18 754; RR = 0.80, 95% CI = 0.71-0.89; I2 = 0%) and live birth/ongoing pregnancy rates (seven studies, n = 20 964; RR = 0.72, 95% CI = 0.63-0.82; I2 = 0%). However, no significant changes were seen in miscarriage rates (seven studies, n = 22 332; RR = 1.40, 95% CI = 0.92-2.11; I2 = 53%) and post-warming survival rates (three studies, n = 13 562; RR = 1.00, 95% CI = 0.99-1.01; I2 = 0%) following 'double biopsy + double vitrification'. Furthermore, 'single biopsy + double vitrification' was also linked with decreased clinical pregnancy rates (six studies, n = 13 284; RR = 0.84, 95% CI = 0.76-0.92; I2 = 39%) and live birth/ongoing pregnancy rates (seven studies, n = 16 800; RR = 0.79, 95% CI = 0.69-0.91; I2 = 70%), and increased miscarriage rates (five studies, n = 15 781; RR = 1.48, 95% CI = 1.31-1.67; I2 = 0%), but post-warming survival rates were not affected (three studies, n = 12 452; RR = 0.99, 95% CI = 0.97-1.01; I2 = 71%) by 'single biopsy + double vitrification'. LIMITATIONS, REASONS FOR CAUTION: All studies included in this meta-analysis were retrospective with varying levels of heterogeneity for different outcomes. Not all studies had accounted for potential confounding factors. Only one study reported neonatal outcomes. WIDER IMPLICATIONS OF THE

findingsOur data indicated adverse impacts of 'double biopsy + double vitrification' and 'single biopsy + double vitrification' on clinical outcomes following euploid blastocyst transfers. Patients should be carefully consulted about the risks when offered such approaches. The biopsy process should be carried out as carefully and competently as possible to minimize an inconclusive diagnosis. STUDY FUNDING/COMPETING INTEREST(S): R.W. is supported by a National Health and Medical Research Council Emerging Leadership Investigator Grant (2009767). There is no other external funding to report. All authors report no conflict of interest. REGISTRATION NUMBER: CRD42023469143.