Successful monozygotic twin delivery following in vitro maturation of oocytes retrieved from a woman with polycystic ovary syndrome: case report

The incidence of monozygotic twinning (MZT) appears to be increasing within the field of assisted reproductive technology (ART), although the factors contributing to the phenomenon are still far from being identified. On the contrary, in vitro maturation (IVM) of oocytes is becoming more accepted and more and more babies have been born worldwide using this procedure. Assessing its safety and impact on monozygotic twinning (MZT), and following up the health of these babies, is essential. We report here a first case of successful monozygotic (MZ) twin delivery following IVM. The patient was a 28-year-old Japanese female, referred to the IVF clinic for primary infertility. Several previous cycles of ovarian stimulation had resulted in ovarian hyperstimulation syndrome (OHSS). The patient received norethisterone-mestranol to initiate the menstruation, and oocyte retrieval was performed 36 h after hCG. A total of 22 immature oocytes were obtained. Following incubation for 24 h in IVM medium, 50% of the oocytes were matured to the metaphase II (MII) stage. Nine oocytes were fertilized after ICSI with the husband's sperm. Three day 3 embryos were transferred into the uterus on the fourth day following oocyte retrieval. Three weeks after embryo transfer, a single gestational sac was visualized in the uterus. At 7 weeks of gestation, two fetal poles with cardiac activity were seen in the single gestational sac. Serial ultrasound examinations revealed a MZ, monochorionic diamniotic pregnancy. After intensive perinatal monitoring, two healthy male infants were delivered by Caesarean section at 35 weeks of gestation.     

Concurrent IVF and spontaneous conception resulting in a quadruplet pregnancy.

Blastocyst transfer of just one or two embryos has been used to help limit the number of high-order gestations. In this case report we describe the occurrence of a quadruplet pregnancy after the transfer of only two blastocysts during IVF. Sonographic examination showed four fetuses and what appeared to be quadriamniotic/quadrichorionic sacs, suggesting that a concomitant spontaneous conception had occurred. Definite confirmation of zygosity was obtained by genetic testing using DNA microsatellite polymorphism determinations after the birth of one boy and three girls at 32 weeks gestation. Although this event has not been reported previously, the possibility of its occurrence should be kept in mind. IVF patients with patent Fallopian tubes should be cautioned against intercourse late in their controlled ovarian stimulation, especially if they would decline multifetal reduction.     

Early transvaginal embryo aspiration: a safer method for selective reduction in high order multiple gestations.

Assisted reproduction technologies and ovulation induction for treatment of infertility continue to cause high order multiple gestations. Increased perinatal morbidity and mortality, as well as maternal morbidity, may complicate these pregnancies. Selective fetal reduction, an acceptable therapeutic approach in these cases, is usually performed at or after the ninth week of gestation, with KCl injected in the vicinity of the fetal heart, and is associated with a total pregnancy loss rate of 11.7%. We report our experience with 90 women who underwent early (mean 7.5 weeks gestation, range 7. 0-8.0 weeks) transvaginal selective embryo aspiration. The mean number of viable embryos before and after reduction was 3.5 and 2.1 respectively. Six (6.7%) pregnancies were lost before 24 gestational weeks. One miscarriage occurred at the tenth gestational week. The other five pregnancies were aborted at 17.3-21.6 weeks gestation. Additional interventions were performed in three of these pregnancies: genetic amniocentesis in two cases and cervical suture in one case. In the subset of 39 patients with>/=4 embryos, only one (2.6%) pregnancy loss was recorded. This loss rate is significantly lower (P < 0.05) than the 15.3% loss rate in patients with >/=4 fetuses calculated from other work. Four (4.4%) other pregnancies were complicated by premature delivery (25-28 weeks gestation). Mean gestational age of delivered pregnancies in our series was 35.7 weeks. In conclusion, early transvaginal embryo aspiration is a simple and relatively safe method for multiple pregnancy reduction. The overall pregnancy loss rate associated with early embryo aspiration is similar to that of procedures performed at later gestational age, but is significantly lower when the initial number of embryos is four or greater.     

The probability of multiple births when multiple gestational sacs or viable embryos are diagnosed at first trimester ultrasound

The live birth outcome when multiple gestational sacs were diagnosed at first trimester ultrasound was reviewed in 227 twin, 43 triplet and five quadruplet pregnancies. When two gestational sacs were present, the probability of delivering twins was 63% for maternal age less than 30 and 52% for maternal age greater than or equal to 30. With three gestational sacs, the probability of a triplet birth was 45% for maternal age less than 30 and 18% for maternal age greater than or equal to 30. When two viable embryos were present, the probability of a twin birth was 90% for maternal age less than 30 and 84% for maternal age greater than or equal to 30. With three viable embryos, the probability of a triplet birth was 90% for maternal age less than 30 and 44% for maternal age greater than or equal to 30. Two gestations resulting from ovulation induction with clomiphene citrate were more likely to result in twin delivery at term, compared to spontaneous twin gestations (P = 0.012). These findings may be useful in the treatment and management of patients when multiple gestations are diagnosed early in pregnancy.     

Infertility: CASE REPORT Astonishing fertility from a single oocyte recovery

A 36 year old patient underwent in-vitro fertilization (IVF)and embryo transfer. Three embryos were transferred to her uterusand eight spare embryos were cryopreserved. The patient conceiveda quadruplet pregnancy and delivered two boys and two identicaltwin girls by Caesarean section at 30 weeks gestation. The couplesubsequently donated their cryopreserved embryos to the embryodonation programme for the use of other couples. Two agonadalpatients received these cryopreserved-thawed embryos, each ina hormone replacement cycle. Both conceived, one has safelydelivered twins and the other conceived a twin pregnancy, oneof the fetuses has vanished and the otter is progressing normally.The overall embryo implantation rate was seven out of nine (78%).     

Embryo reduction in multifetal pregnancies using saline injection: comparison between the transvaginal and the transabdominal approach.

A total of 30 patients with multifetal pregnancies, all resulting from treatment with superovulatory agents or assisted reproductive techniques, underwent embryo reduction. All patients had three or more fetuses (one sextuplet, two quintuplets, seven quadruplets and 20 triplets). The procedure was carried out using intra-embryonal injection of 0.9% sodium chloride solution. Embryo reduction was carried out via the transabdominal approach in 10 patients, performed at 11-12 weeks of gestation, and via the transvaginal route in 20 other patients, at 8-10 weeks of gestation. In the transabdominal group, one patient aborted following repeated attempts at embryo reduction while the other nine gave birth to healthy newborns (eight twins and one triplet). In the transvaginal group, four pregnancies are currently ongoing (all beyond 28 weeks of gestation), 14 pregnancies resulted in a delivery of at least one live newborn (13 twins and one singleton), one patient had a late abortion at 24 weeks' gestation and another was delivered at 27 weeks' gestation due to severe pre-eclampsia. Transvaginal ultrasound-guided needle procedures are commonly practised in most in-vitro fertilization units. The employment of this route for embryo reduction, performed at an earlier gestational age and with the use of a non-toxic substance such as 0.9% saline solution, is advocated.     

Preimplantation genetic diagnosis of a large pericentric inversion of chromosome 5

We report the first established pregnancy using preimplantation genetic diagnosis in order to avoid chromosomal imbalance in the progeny of a woman carrying a large inversion of chromosome 5. This is also the first time where it has been possible to study the distribution of balanced and unbalanced gametes in a female inversion carrier. In total, 23 embryos were biopsied in two separate treatments and analysed by fluorescent in-situ hybridization. Of these, 10 were unbalanced, nine were balanced and for four the analysis was inconclusive. The diagnostic procedure was performed within 3.5 h. This allowed the biopsied embryos to be transferred the same day as the biopsy was taken (day 3). Two embryos were transferred each time, and in the second treatment a twin pregnancy with two chromosomally balanced fetuses was established. Healthy twins were delivered at 34 weeks of gestation.      

Twin pregnancy with a complete hydatidiform mole and co-existing fetus following in-vitro fertilization: case report.

Hydatidiform mole with a co-existing live fetus is a rare event. We report the case of a 41 year old Caucasian woman who attended for in-vitro fertilization. Three embryos, containing two apparently normal pronuclei, were transferred into the uterus. A twin pregnancy with a live fetus and a complete mole ensued. The pregnancy was managed conservatively until 28 weeks gestation when, following an episode of major antepartum haemorrhage, a live female infant was delivered by Caesarean section. The mole, weighing over 1.7 kg, was successfully evacuated. Following delivery, serum human chorionic gonadotrophin concentrations returned to baseline and remain within the normal range at 24 months. Both mother and daughter are well on assessment 24 months later.     

Multiple gestation pregnancy. The ESHRE Capri Workshop Group

Multiple gestation pregnancy rates are high in assisted reproductivetreatment cycles because of the perceived need to stimulateexcess follicles and transfer excess embryos in order to achievereasonable pregnancy rates. Perinatal mortality rates are, however,4-fold higher for twins and 6-fold higher for triplets thanfor singletons. Since the goal of infertility therapy is a healthychild, and multiple gestation puts that goal at risk, multiplepregnancy must be regarded as a serious complication of assistedreproductive treatment cycles. The 1999 ESHRE Capri Workshopaddressed the psychological, medical, social and financial implicationsof multiple pregnancy and discussed how it might be prevented.Multiple gestations are high risk pregnancies which may be complicatedby prematurity, low birthweight, pre-eclampsia, anaemia, postpartumhaemorrhage, intrauterine growth restriction, neonatal morbidityand high neonatal and infant mortality. Multiple gestation childrenmay suffer long-term consequences of perinatal complications,including cerebral palsy and learning disabilities. Even whenthe babies are healthy they must share their parents' attentionand may experience slow language development and behaviouralproblems. Current data indicate that the average hospital costper multiple gestation delivery is greater than the averagecost of in-vitro fertilization (IVF) and intracytoplasmic sperminjection (ICSI) cycles. Prevention is the most important meansof decreasing multiple gestation rates. Multiple gestation ratesin ovulation induction and superovulation cycles can be reducedby using lower dosage gonadotrophin regimens. If there are morethan three mature follicles, the cycle should be converted toan IVF cycle, or it should be cancelled and intercourse shouldbe avoided. In IVF cycles two embryos can be transferred withoutreducing birth rates in most circumstances. Embryo reductioninvolves extremely difficult decisions for infertile couplesand should be used only as a last resort. Assisted reproductivetreatment centres and registries should express cycle resultsas the proportion of singleton live births; twin and tripletrates should be reported separately as complications of theprocedures. Reducing the multiple gestation pregnancy rate shouldbe a high priority for assisted reproductive treatment programmes,despite the pressure from some patients to transfer more embryosin order to improve success. If nothing is done, public concernmay lead to legislation in many countries, a step that wouldbe unnecessary if assisted reproductive treatment programmesand registries took suitable steps to reduce multiple pregnancyrates.     

Comparison of alterations in fetal regional arterial vascular resistance in appropriate-for-gestational-age singleton, twin and triplet pregnancies.

The objective of this longitudinal study was to evaluate alterations in fetal vascular resistance of fetal peripheral arteries with advancing gestation in singleton appropriate-for-gestational-age (S-AGA), twin appropriate-for-gestational-age (Tw-AGA) and triplet appropriate-for-gestational-age (Tri-AGA) infants. Colour Doppler flow imaging and pulsed Doppler ultrasonographic examinations were performed on 35 S-AGA, 52 Tw-AGA and 12 Tri-AGA fetuses. The pulsatility index for middle cerebral artery (MCAPI), umbilical artery (UAPI), descending aorta (DAPI), splenic artery (SAPI), renal artery (RAPI) and femoral artery (FAPI) was measured as vascular resistance every 2 weeks after 15 weeks of menstrual age until delivery. Optimal models and normal ranges for pulsatility index for each artery in each group were generated. The alterations in various fetal regional arterial pulsatility indices with advancing gestational age showed no significant differences in S-AGA, Tw-AGA and Tri-AGA infants, respectively. These results suggest that there is no significant difference for regional arterial vascular resistance in AGA fetuses among singleton, twin, and triplet pregnancies, whereas there was a slight difference in fetal growth pattern among singleton, twin, and triplet pregnancies described in our previous investigation.     

Determination of twin zygosity using a commercially available STR analysis of 15 unlinked loci and the gender-determining marker amelogenin--a preliminary report

BACKGROUND: The aim of this preliminary study was to estimate the accuracy of the zygosity determination in twin pregnancies. METHODS: Seventy-three sets of twin pregnancies were enrolled in this study, including 48 sets of twins resulting from assisted reproductive techniques (ART) and 25 sets of spontaneously conceived twins. Determination of zygosity was made by PCR-amplified short tandem repeat (STR) analysis with a commercially available panel, comprising 15 autosomal, codominant, unlinked loci and the gender-determining marker, amelogenin. Monozygotic (MZ) twins were determined when all these unlinked loci and the gender-determining marker were identical. Chorionicity and placenta were examined after delivery of the newborns to check their relationships to the twin zygosities. RESULTS: Three of 48 (6.25%) sets of twins produced by ART and 18 of 25 (72%) sets of spontaneously conceived twins were MZ. Monozygosity could be evaluated based on 'like sex' in spontaneously conceived twins, because they had a greater incidence of MZs than those produced by artificial reproductive techniques. The MZ prediction rate was 78.6%, and the overestimated rate was 21.4% if the monochorionic like-sexed twins (LST) had a grossly single placenta. The underestimated rate of MZs was 26.7% when the dichorionic LST had apparently separate placentas. CONCLUSION: With the DNA-based 15 STR analysis amplified in a multiplex PCR, the determination of the zygosity in multifetal pregnancies is not only cost and time saving but also yields greater sensitivity and precision than conventional methods.     

Obstetrical results after embryonic reductions performed on 34 multiple pregnancies

Thirty-four women with multiple pregnancies (three or more fetuses) underwent embryonic reduction in order to reduce abortions, premature births or fetal growth-retardation by obtention of twins. Four early abortions occurred. Thirty pregnancies reached term and out of 60 fetuses, 58 infants were born alive. Fetal death in utero of one twin occurred in two pregnancies. The mean term until delivery was 36 +/- 2.8 weeks gestation and the prematurity rate was 51.7%. Of 55 neonates, 25 were underweight within the 10th percentile and 10 out of 55 neonates were underweight below the 3rd percentile. There were three deaths in the early neonatal period. The rate of perinatal mortality was 8.3%. Fifty-four children are currently healthy and one child has a mild axial hypotonia. A reduction in prematurity was observed with a gain of 2 weeks on reported data concerning triplet pregnancies. The rate of low-birth-weight infants was high, 63.5% being underweight at birth.     

Maternal serum concentrations of pregnancy associated placental protein A and pregnancy specific {beta}-1-glycoprotein in multifetal pregnancies before and after fetal reduction

Placental function in multifetal pregnancies before and afterembryo reduction was investigated by measuring maternal serumconcentrations of pregnancy associated placental protein-A (PAPP-A)and pregnancy specific -1-glycoprotein (SP-1). Three groupsof pregnant women were studied following assisted reproduction;groups 1 and 2, were 12 singleton and 12 twin pregnancies respectively,and group 3 comprised 12 women with multifetal pregnancies undergoingembryo reduction. PAPP-A and SP-1 were measured serially at8–21 weeks gestation. In all pregnancies, maternal serumPAPP-A and SP-1 increased with gestation. In twin pregnanciesthe mean concentrations of SP-1 were significantly higher thanin singletons at all gestations, whereas for PAPP-A, concentrationswere similar between these groups. In multifetal pregnanciesbefore embryo reduction, the serum concentrations of both proteinswere significantly higher than in twin pregnancies. Followingreduction, the concentrations of PAPP-A remained significantlyhigher than for twins throughout, whereas the concentrationsof SP-1 gradually converged towards those of twins; by 19 weeksthere was no difference between the means of the two groups.These findings suggest that circulating concentrations of SP-1reflect total placenta mass, which is proportional to the numberof live fetuses, whereas the pattern of PAPP-A changes suggeststhat this protein is produced by the placenta, decidua and othertissues.     

Human zona pellucida micromanipulation and monozygotic twinning frequency after IVF

To assess the association of zona pellucida micromanipulation and subsequent development of monozygotic twins, cases of assisted embryo hatching (AH) and intracytoplasmic sperm injection (ICSI) were identified and related to treatment type, implantation and zygosity data. Embryology records from all patients undergoing in-vitro fertilization (IVF) at this centre from January 1995 to March 1998 were reviewed. In this study, 3546 transfer cycles were completed, with clinical pregnancy established in 1911 (54% per transfer) patients undergoing a single IVF cycle. These pregnancies occurred in 1674 (88%) IVF cycles, 120 (6%) donor oocyte cycles (DER), and 117 (6%) frozen embryo transfer (FET) cycles. During the study period, 23 cases of monozygotic (MZ) twins were identified, representing an overall frequency of 1.2%. Chorionicity was determined by transvaginal ultrasound at 7 weeks when the number of embryos transferred was less than the number of fetal heart-beats, or when >1 fetal heartbeat per gestational sac was seen. Zygosity was confirmed by placental evaluation at delivery, and corroborated the antenatal diagnosis in all cases. Among IVF study patients the frequency of MZ twinning was not statistically different between zona manipulated and zona intact subgroups. While this investigation is the largest to date describing the relationship between MZ twins and zona procedures, studies with even greater statistical power are needed to clarify it more precisely, particularly in DER and FET settings. A greater overall frequency of MZ twinning for IVF patients may be a function of the higher number of embryos transferred in IVF, rather than discrete zona manipulations.     

Assisted reproductive technology in Europe, 1997. Results generated from European registers by ESHRE. European IVF-Monitoring Programme (EIM), for the European Society of Human Reproduction and Embryology (ESHRE)

European results of assisted reproductive techniques from treatments initiated during 1997 are presented in this first ESHRE report. Data were collected from 18 European countries, usually from already-existing national registers. A total of 482 clinics from these 18 countries reported 203 893 cycles. In 10 countries with complete registration, 133215 cycles were performed in a population of 174 million, corresponding to 765 cycles per million inhabitants. After IVF and intracytoplasmic sperm injection (ICSI), the distribution of transfer of one, two, three and four or more embryos was 11.5, 35.9, 38.4 and 14.3% respectively. Huge differences existed between countries. For IVF, the clinical pregnancy rate per transfer was 26.1%, and the delivery rate per embryo transfer 20.9%. For ICSI, the corresponding rates were 26.4% and 21.5%. Singleton, twin, triplet and quadruplet delivery rates for IVF were 70.4, 25.8, 3.6 and 0.2% respectively, giving a total multiple delivery rate for IVF of 29.6%. After ICSI, the corresponding rates were 71.7, 25.2, 2.9 and 0.1%, amounting to a total multiple delivery rate of 28.2%. The range of triplet delivery rates after IVF range from 0.4% to 11.9% among countries.     

Birth weight and age at menopause in Australian female twin pairs: exploration of the fetal origin hypothesis

In a twin sample where duration of gestation can be controlled, a specific example of the fetal origins hypothesis concerning association between low birth weight and early age at menopause is explored. The hypothesis is based on the physiologically plausible path from intrauterine growth retardation and reduced numbers of primary follicles to an earlier menopause. The sample comprised 323 Australian female twin pairs where both co-twins had reached menopause naturally and reported on their weight at birth. Regression analysis showed no linear association between the two variables (P = 0.371, r(2) = 0.0009). Intra-pair differences in age at menopause were investigated in the context of relative birth weight of co-twins. In 265 pairs an intra-pair birth weight difference was reported. In monozygotic (MZ) pairs (n = 168) this allowed for control of genetic effects as well as gestation duration. No significant differences dependent on birth weight relative to co-twin were found for age at natural menopause in either MZ or dizygotic (DZ) twin pairs, even in pairs whose birth weights differed markedly. There was some indication that twins with premature ovarian failure were heavier at birth than twins with normal or later menopausal age. We conclude that the hypothesis that lower birth weight is associated with earlier menopause is not supported by our data.     

Quadruplet pregnancy following transfer of two embryos: Case report

Following a long period of secondary unexplained infertility, a couple had assisted conception treatment with IVF and transfer of two embryos 2 days later. The woman conceived and was found to have a quadruplet pregnancy. Following spontaneous labour onset at 34 weeks gestation, she was delivered by immediate Caesarean section. All four babies (three boys, one girl) were eventually discharged home with the mother. All the boys were found to be of differing zygosity. The quadruplet pregnancy was attributed to natural conception following intercourse and fertilization of oocytes not collected at the time of IVF. The wisdom of intercourse during a cycle of IVF is discussed.     

What is the most relevant standard of success in assisted reproduction? The singleton, term gestation, live birth rate per cycle initiated: the BESST endpoint for assisted reproduction

Assisted reproduction programmes do not report success consistently. Rates vary with the definition used. Success must reflect delivery of healthy babies, and the burden of treatment to couples. We report the singleton, term gestation, live birth rate of a baby per assisted reproductive technology cycle initiated for a large IVF programme. We defined assisted reproductive technology cycles as those initiated with the intention of oocyte collection. We examined cycles conducted through Monash IVF in 2001. All women with positive pregnancy tests had first trimester ultrasonography. Obstetric outcomes were recorded. All babies had neonatal examinations conducted by paediatricians. A total of 644 positive pregnancy tests were recorded in 2600 cycles; 509 showed fetal heart motion. Of 448 deliveries, 328 were singleton and 120 were multiple. There were 290 singleton deliveries at term gestation. In 2001, a couple had an 11.1% chance of delivering a singleton, term gestation, live baby per assisted reproductive technology cycle begun. We suggest that delivery of a single, term gestation, live baby per cycle initiated is the most relevant standard of success. This statistic was 11.1% at Monash IVF. We encourage programmes to report this BESST (Birth Emphasizing a Successful Singleton at Term) outcome.     

Hydatidiform mole with coexisting twin pregnancy after gamete intra-fallopian transfer.

The pregnancy of a 31-year-old infertility patient is described. After gamete intra-Fallopian transfer, her pregnancy evolved uneventfully until the 18th week of gestation, when vaginal bleeding occurred. Ultrasonographic findings suggested a molar pregnancy with two live fetuses. At 24 weeks gestation, two male infants were spontaneously delivered. Fetal (46 XY) and molar (46 XX) karyotypes and post-mortem findings were consistent with a bizygotic twin pregnancy associated with a complete hydatidiform mole. The pathogenesis and obstetrical management are discussed.     

Achievement of pregnancy in globozoospermia with Y chromosome microdeletion after ICSI

Pregnancy achieved with sperm from a patient with globozoospermia is rare, even after ICSI, since the activation of the oocyte may not occur in this disorder. Therefore, activation of the oocytes by piezoelectricity or calcium ionophores has been suggested, although spontaneous activation of the oocyte after ICSI has been reported in some cases. We report a successful pregnancy in a couple in which the male partner had globozoospermia with microdeletions in the Y chromosome with no further assisted activation after ICSI. During the diagnostic study of the husband, increased numerical chromosome abnormalities after fluorescent in-situ hybridization (FISH) and microdeletions in AZFa; sY86 and AZFb; sY 131 were detected. Out of the 13 oocytes injected, four fertilized and a twin pregnancy was obtained after replacement of four embryos. Healthy twin girls were delivered after a term pregnancy. Some patients with globozoospermia may also have Y chromosome microdeletions, which subsequently may be inherited by the male offspring in cases of achievement of pregnancy.