Table 2.
Comparison of different methodologies for mitochondria DNA quantification and results from published articles
| Ref. | Sample size | Settings | nDNA method | mtDNA method | Result | Comments |
|---|---|---|---|---|---|---|
| Fragouli 2015 [12] |
379 embryos D3: 39 D5: 340 |
Single center |
D3: aCGH (24Sure, Illumina) D5: 302 → aCGH 38 → NGS (MiSeq or HiSeq, Illumina) |
RT-PCR (n = 341) NGS (n = 38) (PGM, Life Technologies) |
The relative quantity of mtDNA was significantly higher in old age, aneuploid, and non-implanted blastocysts, and cleavage stage in young age patient. Overall implantation rate: 38% (16/42); “Normal” mtDNA implantation rate: 59% All euploid blastocysts with mtDNA levels above the threshold failed to implant (100%, n = 14) |
Relative mtDNA threshold: 0.003 |
| Diez-Juan 2015 [13] |
270 embryos D3: 205 D5: 65 |
Single genetics laboratory | aCGH (24Sure, BlueGnome) | RT-PCR |
Mitochondrial score (Ms): mtDNA/nDNA Day 3: < 34 (MsA): IR = 59% (n = 51) 34–52 (MsB): IR = 44% (n = 52) 52–97 (MsC): IR = 42% (n = 50) > 97 (MsD): IR = 25% (n = 52) Ms > 160 (n = 22) IR = 0 Day 5: < 18.19 (MsA): IR = 81%(n = 16) 18.19–24.15 (MsB): IR = 50% (n = 16) 24.15–50.58 (MsC): IR = 62% (n = 16) > 50.58 (MsD): IR = 18% (n = 17) > 60 (n = 7): IR = 0 |
Less is better |
| Ravichandran 2017 [14] | 1505 euploid blastocysts | Multi-center | aCGH (24Sure, Illumina) or NGS (VeriSeq, Illumina) |
qRT-PCR 16s ribosomal RNA MajArc probe sequence |
Relative mtDNA levels (normalization against nuclear DNA sequence) Overall implantation rate: 65.6% (185/282) “Normal” mtDNA implantation rate: 74.3% Negative predictive value: 100% (n = 33) * elevated mtDNA embryos: 9.2% (139/1505) |
Relative mtDNA threshold: 0.0004 for 16s ribosomal RNA; 0.00035 for the MajArc sequence |
| Victor 2017 [16] |
1396 blastocysts Euploid: 775 Aneuploid: 621 |
Single center |
NGS (MiSeq, Illumina) qPCR validation: |
NGS (n = 1396) (MiSeq, Illumina) qPCR (n = 150) |
No difference regarding ploidy, age, and implantation potential (P vs. NP: 101/140 via NGS; 49/51 via qPCR) |
With correction factors tailored to gender and chromosomal status |
| Treff 2017 [17] |
374 blastocysts DET with 69 singletons |
Single center | qPCR | qPCR |
No systematic difference in relative mtDNA quantity between implanted and non-implanted embryos. MtDNA copy number was negatively correlated with oocyte age. Embryos biopsied on day 5 were more likely to have higher quantities of mtDNA compared with biopsied on day 6. Greater blastocyst expansion scores were correlated with lower mitochondrial content. Higher quality trophoblast and ICM were similarly correlated with lower mtDNA quantity. |
Highlight the importance of patient-specific variables |
| Fragouli 2017 [15] | 199 euploid blastocysts | Single center | NGS (VeriSeq, Illumina) | qPCR |
Ongoing pregnancy rate: Normal/low levels of mtDNA: 64% (121/190) Elevated mtDNA levels: 0% (0/9) * elevated mtDNA embryos: 4.5% (9/199) |
Blinded prospective non-selection study |
| Santos 2018 [23] | 1641 blastocysts | Single center | NGS (PGM, Thermo Fisher) | NGS (PGM, Thermo Fisher) | The probability of mtDNA content above the median increased by 6% when BMI increased; lowered by 39% when P4 increased, and increased by 42% in aneuploid blastocysts. | Potential variables that affect the mtDNA content |