Conjoined twins post single embryo transfer of a euploid embryo in a frozen embryo transfer cycle.

A mid-40s female G3P2012 patient presented with secondary infertility of 1 year duration in her second marriage. The patient’s history included one pregnancy loss, treated with dilation and curettage as well as two caesarean sections from a previous marriage. Infertility workup accounted for an advanced maternal age and the presence of a uterine isthmocele. Husband's sperm analysis revealed oligoasthenospermia. Prior to her presentation in our clinic, the patient underwent three cycles of ovarian stimulation (OS) for IVF/ intracytoplasmic sperm injection (ICSI), but due to premature ovulation, arrested embryos and aneuploid embryos respectively, no previous embryo transfer was performed. In our clinic, the patient underwent two OS cycles with subsequent two failed frozen embryo transfer (FET) cycles. In a hormonal replacement therapy approach, a single euploid embryo, grade 5AB, was transferred on the sixth day of luteal phase support. 10 days post embryo transfer, the patient did her first bhCG test which turned out to be positive. The first value documented was 482.2 mIU/mL. As per the standard of practice of our clinic, the patient was asked to repeat the bhCG to document proper hormonal rise. The repeat bhCG test done 7 days after the initial test was 6141 mIU/mL. At 7 weeks+2 days, a transvaginal ultrasound revealed a single intra-uterine gestational sac (GS) with one fetal pole and one fetal heart. The crown to rump length measured 9.1 mm, the yolk sac was 4.5 mm, and the GS measured 14.8 mm. The patient was then discharged to follow up with her regular obstetrician at 7 weeks+2 days. At 11 weeks, the patient was diagnosed with thoracophagus conjoined twins with a single heart in a different centre as shown in figure 1 .

Due to the poor prognosis, the patient decided on a pregnancy termination. The patient was seen in the clinic 4 months after the termination. She was shaken from her experience, and as a result, after extensive counselling she preferred not to proceed with another treatment at that point in time. The patient consented to use her data anonymously. An informed consent was signed by the patient.

Elective single embryo transfers (eSETs) are being advocated to be the standard of care in patients undergoing in vitro fertilisation (IVF) treatments because of a better safety profile, especially concerning the reduction of high order pregnancy rates. However, eSET still carries a risk of multiple gestation. Of the largest registries published, the Japanese society sets the frequency of embryo splitting post-eSET at 1.36% with the rate of monozygotic twin and triplet pregnancies being 0.9%–3.1% and 0.048%, respectively. 1 Conjoined twins are even a rarer finding with less than 20 cases documented in literature after eSET transfers. 2 The exact mechanism of incomplete embryo splitting leading to conjoined twins is still vague with no clear predisposing factors being identified. The rarity of the conjoined twins in assisted reproductive technology (ART) adds to the complexity of finding the supporting data to explain the hypothesis related to embryo splitting. Hence, describing and publishing rare cases adds to the literature available in the hope of completing the pieces of the puzzle and providing plausible explanations of the unusual splitting. Thus, we present a case of thoracophagus conjoined twins after an eSET of a euploid embryo. To the best of our knowledge, it is the first case of conjoined twins in the setting of a euploid embryo transfer to be reported.

Conjoined twins are a form of incomplete monozygotic twinning (MZT). In natural conception, conjoined twins’ phenomena are thought to originate either from partial embryo separation at 13–15 days post fertilisation, from secondary fusing of already separated MZT, or from an initial duplication of early primordia. 3 5 In ART, based on a recent meta-analysis, it is believed that blastocyst stage embryos are at increased risk of undergoing some type of embryo fission, 6 despite having contradicting data especially in fresh cycles. 7 It could be assumed that advances in IVF laboratory settings, the use of improved culture media with antioxidant activity as well as embryologists’ training explain the decrease in the OR of embryo splitting from 2.2 to 1.7 when comparing the periods of 2007–2010 and 2010–2014. 1 8 The method of insemination, whether IVF or ICSI, has also been suggested as potential contributing factors. In the recent meta-analysis, Busnelli et al concluded that IVF was more associated with MZT than ICSI, 6 contradicting the suggested theory of mechanical manipulation of the zona pellucida as a risk factor for splitting. 8 Lower inner cell mass (ICM) quality (B or C) is also suspected to be a risk factor, as weak intercellular connections are thought to cause the ICM fission documented by time-lapse imaging. 9 Another study 10 found that higher ICM grade was associated with embryo splitting, which contradicts this finding. It was also proposed that compromised oocyte quality might be a risk factor for embryonic instability. Scott et al postulated that oocyte ageing might interfere with the formation of a proper cellular axis and cytoplasmic folding in the secondary oocyte’s pre or during the fertilisation phase. 11 Opposing data exists concerning the effect of performing embryo biopsy for preimplantation genetic testing. While Verpoest et al 12 and a recent meta-analysis 6 concluded that embryo biopsies do not increase the risk of monozygotic twinning, Sellers et al in a recent publication revealed that blastocyst biopsies could increase the twinning rates. 13 The latter finding requires further evaluation. Time-lapse imaging has been used extensively in recent years to improve reproductive outcomes through improved embryo selection. The time-lapse images could not, however, reveal a specific developmental mechanism that could consistently explain the abnormal embryonic fission. Sutherland et al , as previously stated, detected ICM duplication on time-lapse imaging in a monochorionic tri-amniotic triplet case after FET. 14 Grøndahl et al ’s assessment of the images did not reveal a possible cause for the parapagus conjoined twin. 2 Detailed descriptions of the images revealed that the observed findings (asymmetric first cleavage of the blastomere) were not specific for post-implantation abnormalities. A blind analysis of embryoscope images in MZT resulting from fresh embryo transfers revealed a significant association between a shortened S3 division time and embryo splitting. The analysis included the division images of 85 embryos, 22 of which resulted in MZT (p = 0.006). In the same study, ICM A grading was found to be associated with the splitting event. 10 Given that these effects were observed in fresh embryo transfers, their applicability to FETs might be limited. The lack of additional studies on this topic, as well as the rarity of the events, complicates the explanation of the predisposing factor for complete and incomplete splitting events which finally lead to conjoined twins. Conjoined twins have a poor obstetrical outcome which usually necessitates termination of pregnancy, a procedure that causes major emotional and physical distress for patients. Hence, a more comprehensive analysis of the diagnosed conjoined multiple gestation cases post embryo transfers might help identify common risk factors. Therefore, we reviewed and summarised the available literature, and the overview is presented in table 1 . In total, 28 case reports were identified; however, none of them included a case of conjoined twins after the transfer of a euploid embryo. Tables1 2 compare the conjoined twin cases resulting from FETs. Seven cases were identified; however, no straightforward risk factors could be highlighted due to the rarity of the condition as well as the lack of full documentation in the case reports published. Drawing conclusions on the various causes and predisposing factors for incomplete embryo splitting in the current situation is challenging due to the lack of a standardised framework for case reports as well as the absence of critical information in some research. GnRH : gonadotropin releasing hormone, hCG; human chorionic gonadotropin, hMG: human menopausa gonadotropin,rFSH:recombinantfollicularstimulatinghormone ET, embryo transfer; GS, gestational sac; ICSI, intracytoplasmic sperm injection; IVF, in vitro fertilisation; N/A, not applicable; NS, not specified. GnRH : gonadotropin releasing hormone, hCG; human chorionic gonadotropin, hMG: human menopausa gonadotropin ET, embryo transfer; GS, gestational sac; ICSI, intracytoplasmic sperm injection; IVF, in vitro fertilisation; N/A, not applicable; NS, not specified. It was a long-anticipated pregnancy. When we heard the heartbeat at our first ultrasound, we were all over the moon. That’s why when the doctor to whom I was referred to follow up my pregnancy informed me about the abnormality in the ultrasound, I couldn’t believe it. I wanted answers which he couldn’t really provide me with. We were heartbroken. The decision to terminate this pregnancy that we’ve waited for so long was one of the most difficult decisions of my life. Single embryo transfers can still result in twin gestations and rarely in conjoined twins. The exact mechanism of incomplete embryo splitting remains largely unknown. Proper unified documentation of conjoined twins’ cases post in vitro fertilisation might help in pinpointing the culprit leading to incomplete splitting.