Tubal superfetation following frozen–thawed single‐embryo transfers in two separate cycles: Case report and literature review

Superfetation is a very rare occurrence. In the context of assisted reproduction, it has been reported only as an intrauterine pregnancy after ovarian stimulation and/or embryo transfer in the presence of an undiagnosed ectopic pregnancy. Here we report a case of a 27‐year‐old anovulatory patient, gravida 1 para 1, who underwent two frozen–thawed single‐blastocyst transfers in separate cycles. The patient reported that 12 days after the first transfer, she had menstrual bleeding and stopped her estradiol and progesterone supplementation without undergoing a blood human chorionic gonadotropin (βhCG) test. At her request, a second cycle was immediately initiated, with endometrial thickness measuring 4 mm. Eleven days after the second transfer, the βhCG value was inappropriately high. A right tubal pregnancy corresponding to 8 gestational weeks was diagnosed. Laparoscopy revealed a prominent right tubal pregnancy in addition to a significantly smaller left tubal pregnancy. The discordant tubal pregnancies were confirmed histologically. To our knowledge, superfetation involving a second ectopic pregnancy coexistent with a first, contralateral ectopic pregnancy consequent to consecutive in vitro fertilization procedures has not previously been described in the medical literature. This case emphasizes the importance of routine βhCG testing after every IVF cycle, even if apparently unsuccessful.

3][4] The aim of the present study was to describe a case of bilateral tubal superfetation caused by two separate single frozen-thawed blastocyst transfers performed in two separate artificial cycles in an anovulatory patient.

| C A S E REP ORT
A 24-year-old female patient was admitted to the in vitro fertilization (IVF) unit of a tertiary medical center after 4 years of primary infertility.Clinical characteristics included body mass index 36 kg/m 2 , amenorrhea, hirsutism on physical examination, and multifollicular ovaries by sonography, all corresponding with the diagnosis of polycystic ovary syndrome (PCOS) according to the Rotterdam criteria. 5 At presentation to the emergency department, the patient had light vaginal bleeding but was otherwise asymptomatic.A right tubal pregnancy was diagnosed, with crown-rump length corresponding to 8 gestational weeks; there was no heartbeat.Laparoscopy confirmed the diagnosis (Figure 1a), and salpingectomy was performed.
A second pregnancy was also suspected in the left tube (Figure 1b).However, as the patient had signed a presurgical consent form only for unilateral salpingectomy, it was deemed safe and appropriate to perform left conservative salpingostomy.Histologic examination of the excised right fallopian tube (Figure 2a,b) demonstrated an ectopic tubal pregnancy, and the left salpingostomy specimen (Figure 2c) showed blood clots with scant chorionic villi, consistent with a younger left ectopic pregnancy.
The patient gave their free informed consent to the anonymous publication of this case.The Rabin Medical Center IRB committee decided that the publication of this case report did not require its approval.

| DISCUSS ION
Superfetation is a very rare event.Only one case of spontaneous superfetation involving an intrauterine gestation following a spontaneous ectopic pregnancy has been previously described. 3ART can overcome the natural barriers to superfetation, and there are reports of superfetation occurring under these condtions, with or without an additional spontaneous conception. 2,4Administration of exogenous gonadotropins or clomiphene citrate during pregnancy has been shown to result in follicular development and even the generation of potentially viable embryos. 6,7 the unique case presented here, both the first gestation and the second, coexistent, gestation were entirely iatrogenic, without any contribution of the patient's ovarian cycle.Transfer of a frozenthawed embryo under artificial endometrial preparation without follicular development in an anovulatory patient resulted in a tubal pregnancy that was undiagnosed due to human error.The pregnancy persisted despite the discontinuation of progesterone support for about 2 weeks and an extremely low systemic progesterone level (0.8 nmol/L).It did not interfere with the ectopic implantation of a second embryo in the contralateral tube in another artificial cycle, again without any follicular development.To the best of our knowledge, consecutive tubal superfetation has not been previously reported.
The patient described in this case underwent IVF treatment in one center and eventually had emergency surgery in another center which did not have access to her full reproductive record in real time.Therefore, informed consent was obtained only for unilateral salpingectomy, and surgery on the contralateral side, where the gestation was unexpected, had to be conservative and minimal, leaving the diseased tube in situ.
In addition to the uniqueness of this case, the findings stress the importance of two issues.Routine βhCG testing must be performed after each and every cycle, even if pregnancy seems unlikely and patient compliance is poor.In cases in which laparoscopy is performed due to a tubal pregnancy or pathologic findings in a patient undergoing IVF, the presurgical discussion and consent should be broad in order to cover unanticipated bilateral tubal pathologies.
Her endocrine profile was as follows: leutenizing hormone (LH) 10.6 IU/L, follicle-stimulating hormone (FSH) 6.4 IU/L, 17-hydroxyprogesterone 2.29 nmol/L, testosterone 2.6 nmol/L, androstenedione 11.7 nmol/L, and anti-Müllerian hormone 8.9 ng/mL.The male partner had a normal spermatogram.Hysterosalpingography demonstrated a normal uterine cavity, normal right tubal anatomy and spillage, and left distorted tubal anatomy with limited spillage but without hydrosalpinx.The patient did not respond to stimulation with clomiphene citrate or FSH up to 150 IU/day.She was referred for IVF in order to increase the FSH dose without inducing exaggerated multifollicular recruitment and multiple gestation.The first stimulation and oocyte pick-up (antagonist protocol, 300 IU/day, agonist-only trigger) resulted in the generation of 16 blastocysts.Following fresh (single) embryo transfer (ET), the patient conceived and gave birth to a healthy baby by spontaneous delivery at 40 weeks' gestation.The patient returned 3 years later for further attempts to conceive with the frozen blastocysts.She underwent four unsuccessful frozen-thawed embryo transfers (FETs), all with satisfactory artificial preparation of the endometrium.Hysteroscopy and antiphospholipid studies revealed no abnormalities.The fifth FET was performed with artificial preparation, and a single blastocyst was transferred.The patient reported that menstrual bleeding occurred 12 days later, and she stopped her estradiol and progesterone supplementation without undergoing a blood βhCG test.Immediately thereafter, at her request, another artificial endometrial preparation cycle was started.F I G U R E 1 Laparoscopic evaluation of the pregnancy.(a) Prominent right tubal pregnancy, diagnosed preoperatively, salpingectomy was performed.(b) Less prominent left tubal pregnancy, not diagnosed preoperatively; therefore, salpingostomy was performed.F I G U R E 2 Histologic evaluation of the fallopian tubes.(a) The right salpingectomy specimen showed products of conception at the distal end of the fallopian tube.Note the chorionic villi in the left side of the image, and the fallopian fimbriae with hemorrhage on the right.(b) At slightly higher magnification of the right salpingectomy specimen, chorionic villi are seen in the bottom left, with trophoblastic invasion into the fallopian tube wall in the top right.(c) The left salpingostomy specimen showed a small blood clot with scattered embedded chorionic villi (hematoxylin and eosin, original magnification a = 20×, b = 40×, c = 20×).Endometrial thickness at the time measured 4 mm.After 10 days of treatment with oral 17β-estradiol preparation, blood estradiol level was 1212 pmol/L (progesterone 0.8 nmol/L, LH 3.1 IU/L), and endometrial thickess increased to 9 mm.Progesterone was initiated, and another single blastocyst FET was performed.The patient did not present as instructed for her βhCG test, and only did so after receiving several reminders on day 11 after FET.The results showed a βhCG level of 20 815 IU/L.The patient was referred for immediate admission to the emergency department, but she complied only 2 days later and addressed the emegency department of another hospital adjacent to where she was staying at that time.