Search

Purpose: To evaluate the utilization of single-embryo transfer (SET) and preimplantation genetic testing (PGT) in gestational carrier IVF cycles in the USA with donor oocyte and examine the impact on live birth and multiple gestation., Methods: Retrospective cohort study using the Society of Assisted Reproductive Technology (SART) clinic database of 4776 donor oocyte-recipient IVF cycles in which a GC was used. The cycles were separated into 4 groups by use of PGT and number of embryos transferred as follows: (1) PGT and single-embryo transfer (PGT-SET); (2) PGT and multiple embryo transfer (PGT-MET); (3) no PGT and SET (NoPGT-SET); (4) no PGT and MET (NoPGT-MET). Primary outcomes were live birth rate (LBR) and multiple pregnancy rate (MPR)., Results: More than one blastocyst was transferred in 48.7% (2323/4774) of the cycles. When ≥1 blastocyst was transferred, with or without the use of PGT, the MPR was 45.5% and 42.0%, respectively. In comparison, in the PGT-SET and NoPGT-SET groups, the MPR was 1.4% (8/579) and 3.3% (29/883), respectively. Live birth rates increased with the use of PGT-A and with MET., Conclusion: This study shows that SET, with or without PGT, is associated with a significantly reduced MPR in donor oocyte-recipient GC IVF cycles while maintaining high LBR. It also demonstrates that many infertility centers in the USA are not adhering to ASRM embryo transfer guidelines. Our findings highlight an opportunity to increase GC safety, which ultimately may lead to widened access to this increasingly restricted service outside the USA.

The success of a pregnancy and the birth of a heathy baby following embryo transfer are conditioned by many factors, including embryo quality and the uterine environment. While we keep looking for more indicators of embryo quality, it also is critical to understand what constitutes a favorable uterine environment leading to a successful pregnancy and birth. This issue of JARG offers new insights on both components-so called by some "the seed and the soil"-and also highlights the critical interactions between the two. Collectively, these publications are contributing to a better understanding of basic embryology and reproductive biology. They could lead to multiple applications to mitigate infertility issues; however, our knowledge base remains rudimentary when it comes to sorting out the 'soil or seed' dilemma. The call from all authors for more research in their respective areas resonates within the ART community. Recognizing the practical and ethical limitations of studies in human patients also reemphasizes the need for solid research in multiple animal models to better understand what constitutes the best recipe for successful embryo transfer outcomes.

Purpose: Excess embryos transferred (ET) (> plurality at birth) and fetal heartbeats (FHB) at 6 weeks' gestation are associated with reductions in birthweight and gestation, but prior studies have been limited by small sample sizes and limited IVF data. This analysis evaluated associations between excess ET, excess FHB, and adverse perinatal outcomes, including the risk of nonchromosomal birth defects., Methods: Live births conceived via IVF from Massachusetts, New York, North Carolina, and Texas included 138,435 children born 2004-2013 (Texas), 2004-2016 (Massachusetts and North Carolina), and 2004-2017 (New York) were classified by ET and FHB. Major birth defects were reported by statewide registries within the first year of life. Logistic regression was used to estimate adjusted odds ratios (AORs) and 95% CIs of the risks of a major nonchromosomal birth defect, small-for-gestational age birthweight (SGA), low birthweight (LBW), and preterm birth (≤36 weeks), by excess ET, and excess ET + excess FHB, by plurality at birth (singletons and twins)., Results: In singletons with [2 ET, FHB =1] and [≥3 ET, FHB=1], risks [AOR (95% CI)] were increased, respectively, for major nonchromosomal birth defects [1.13 (1.00-1.27) and 1.18 (1.00-1.38)], SGA [1.10 (1.03-1.17) and 1.15 (1.05-1.26)], LBW [1.09 (1.02-1.13) and 1.17 (1.07-1.27)], and preterm birth [1.06 (1.00-1.12) and 1.14 (1.06-1.23)]. With excess ET + excess FHB, risks of all adverse outcomes except major nonchromosomal birth defects increased further for both singletons and twins., Conclusion: Excess embryos transferred are associated with increased risks for nonchromosomal birth defects, reduced birthweight, and prematurity in IVF-conceived births.

Background: This paper describes the clinical practice and performance of cell-free DNA sequencing-based non-invasive prenatal testing (NIPT) as a screening method for fetal trisomy 21, 18, and 13 (T21, T18, and T13) and sex chromosome aneuploidies (SCA) in a general Italian pregnancy population., Methods: The AMES-accredited laboratory offers NIPT in maternal blood as a screening test for fetal T21, T18, T13 and SCA. Samples were sequenced on a NextSeq 550 (Illumina) using the VeriSeq NIPT Solution v1 assay., Results: A retrospective analysis was performed on 36,456 consecutive maternal blood samples, including 35,650 singleton pregnancies, 800 twin pregnancies, and 6 triplet pregnancies. Samples were tested between April 2017 and September 2019. The cohort included 46% elevated-risk and 54% low-risk patients. A result indicative of a classic trisomy was found in 356 (1%) of singleton or twin samples: 254 T21, 69 T18, and 33 T13. In addition, 145 results (0.4%) were indicative of a SCA. Of the combined 501 screen-positive cases, 484 had confirmatory diagnostic testing. NIPT results were confirmed in 99.2% (247/249) of T21 cases, 91.2% (62/68) of T18 cases, 84.4% (27/32) of T13 cases, and 86.7% (117/135) of SCA cases. In the 35,955 cases reported as unaffected by a classic trisomy or SCA, no false negative cases were reported. Assuming that false negative results would be reported, the sensitivity of NIPT was 100.00% for T21 (95% Cl 98.47-100.0), T18 (95% Cl 94.17-100.0), and T13 (95% Cl 87.54-100.0). The specificities were 99.99% (95% Cl 99.98-100.0), 99.98% (95% Cl 99.96-100.0), 99.99% (95% Cl 99.97-100.0), and 99.95% (95% Cl 99.92-99.97) for T21, T18, T13, and SCA, respectively., Conclusion: This retrospective analysis of a large cohort of consecutive patients who had whole-genome sequencing-based NIPT for classic trisomies and SCA shows excellent detection rates and low false positive rates.

Purpose: To establish blastocyst freezing criteria for day 7 blastocyst (day 7 BL) for single vitrified-warmed blastocyst transfer (SVBT) by examining the diameter of blastocysts., Methods: Patients who underwent day 7 BL transfer cycles (1143 cycles, mean age: 38.5 ± 3.5) and randomly selected patients after 1:1 matching who underwent day 6 BL transfer cycles and day 2-single-embryo transfer (SET) cycles were used for analysis. Comparison of the miscarriage (per clinical pregnancy) and live birth rates were made among day 2-SET, day 7 BL, and day 6 BL. These blastocyst groups were stratified into six groups based on blastocyst diameter, namely, 180 μm, 190 μm, 200 μm, 210 μm, over 220 μm, and hatched, for making the freezing criteria., Results: For each diameter, 180 μm, 190 μm, 200 μm, 210 μm, over 220 μm, and hatched, the live birth rates of day 7 BL after SVBT were 9.0%, 11.9%, 11.5%, 15.6%, 20.0%, and 19.9%, respectively. Compared with the 14.6% live birth rate of the day 2-SET group, the live birth rate of 220 μm day 7 BL was significantly higher (P < 0.05) and was around the same in other diameter groups., Conclusion: Our study demonstrates that sufficient live birth rates can be obtained after SVBT even from blastocysts on day 7 when blastocysts were vitrified at expanded blastocyst stage of over 180 μm of diameter or at hatched blastocyst stage and were transferred at the optimal time. This is the first study to establish a day 7 blastocyst freezing criteria using blastocyst diameter, which is an objective assessment way.

Purpose: To evaluate the association of objectively measured physical activity (PA) and sedentary behaviour before and during in vitro fertilization (IVF) with controlled ovarian stimulation (COS) and pregnancy outcomes., Methods: This longitudinal study involved 107 infertile women undergoing IVF treatment. PA and sedentary behaviour were measured for 14 consecutive days using accelerometry as follows: (1) before IVF treatment, (2) during IVF at the implantation time, immediately after embryo transfer, and (3) after positive pregnancy test. Total screen time was assessed by questionnaires. COS results were measured as the number of oocytes and embryos obtained, and the study outcomes included positive hCG, clinical pregnancy, and live birth., Results: Compared with baseline activity levels, women significantly reduced their PA and increased sedentary behaviour during IVF (p ≤ 0.001). Higher average PA, light PA, and ratio between breaks in every ≥ 30-min blocks of sedentary time showed positive associations, while sedentary time, number, and time accumulated in blocks of ≥ 30 min of sedentary time associated negatively with oocyte and embryo counts (all p < 0.05). Women with high total screen time during non-work days (≥ 7 h) obtained 4.7 oocytes (p = 0.005) and 2.8 embryos (p = 0.008) less in COS. PA and sedentary behaviour before and during IVF did not affect the positive hCG, clinical pregnancy, and live birth outcomes., Conclusion: Our study results suggest that higher time spent in PA and lower time spent in sedentary behaviour before entering assisted reproduction is associated with better COS outcomes, while activity levels before and during IVF do not affect the implantation, pregnancy, and live birth outcomes.

The incidence of multiple pregnancy has significantly increased over the past decades, reaching different statistics to double, triple, or even overcome these numerical orders globally. Zygosity and chorionicity are the key elements in the multiple pregnancy but the placentation issue should be correlated primarily with zygosity, unlike chorionicity that should be correlated with the outcome and complications of multifetal gestation. Multiple pregnancy is by itself a special maternal-fetal condition, and the monochorionic one, moreover, due to specific complications. These aspects make early assessment of chorionicity and amnionicity a priority. Ultrasound is essential in pregnancy but pathological placental examination after delivery is complementary, in order to have a complete overview of potential mechanisms and pathogenesis affecting twin gestation. In this review, we highlight both ultrasound aspects specific to multifetal placentation, complemented by macro and microscopic morphological aspects, which underpin the obstetric imaging.

The aim of the report was to present a method of zygosity determination in multiple pregnancy. The study wascarried out on same-sex neonates born as a result of spontaneous quadruplet pregnancy. Zygosity was determinedby DNA profiling. The pregnancy was confirmed to be polyzygotic. DNA profiling may be used as a method ofzygosity determination in multiple pregnancy.

In the present paper, we highlight the need to study very early maternal-embryo interactions and discuss how these interactions can be addressed. Bovine species normally carry one or, less frequently, two embryos to term; there are very rare cases of triplets or higher-order multiple pregnancies in which all the offspring are born alive. Multiple-embryo transfer (MET) in cattle allows for the detection of endometrial responses in scenarios where single-embryo transfer would not. Although MET is non-physiological, the present study shows that at the very early embryonic stages, a uterus carrying zona-enclosed embryos does not exhibit non-physiological reactions. On the contrary, MET should be considered the sum of multiple individual effects triggered by developing embryos. We provide arguments to support our hypothesis that describe a rationale for current work with MET, and we discuss alternative hypotheses. Using cattle as a model, we describe how technical approaches to analyzing zona-enclosed early embryo-maternal interactions (i.e., transcriptomics, proteomics, and endometrial cell culture) can help identify molecular changes that may be difficult to observe when only a single embryo is present. We conclude that MET can be used for studying very early maternal-embryo interactions in vivo in monotocous species. Free Spanish abstract: A Spanish translation of this abstract is freely available at http://www.reproduction-online.org/content/150/2/R35/suppl/DC1., (© 2015 Society for Reproduction and Fertility.)

Background: A multiple pregnancy is now considered to be the most common adverse outcome associated with in vitro fertilization (IVF). As a consequence, the identification of women with the best chances of embryo implantation is a challenge in IVF program, in which the objective is to offer elective single-embryo transfer (eSET) without decreasing the pregnancy rate. To date, a range of hormonal and clinical parameters have been used to optimize eSET but none have significant predictive value. This variability could be due to genetic predispositions related to single-nucleotide polymorphisms (SNPs). Here, we assessed the individual and combined impacts of thirteen SNPs that reportedly influence the outcome of in vitro fertilisation (IVF) on the embryo implantation rate for patients undergoing intracytoplasmic sperm injection program (ICSI)., Materials and Methods: A 13 gene polymorphisms: FSHR(Asn680Ser), p53(Arg72Pro), AMH(Ile49Ser), ESR2(+1730G>A), ESR1(-397T>C), BMP15(-9C>G), MTHFR1(677C>T), MTHFR2(1298A>C), HLA-G(-725C>G), VEGF(+405G>C), TNFα(-308A>G), AMHR(-482A>G), PAI-1(4G/5G), multiplex PCR assay was designed to genotype women undergoing ICSI program. We analyzed the total patients population (n = 428) and a subgroup with homogeneous characteristics (n = 112)., Results: Only the VEGF(+405G>C) and TNFα(-308A>G) polymorphisms impacted fertilization, embryo implantation and pregnancy rates. Moreover, the combined VEGF+405.GG and TNFα-308.AG or AA genotype occurred significantly more frequently in women with high implantation potential. In contrast, the VEGF+405.CC and TNFα-308.GG combination was associated with a low implantation rate., Conclusion: We identified associations between VEGF(+405G>C) and TNFα(-308A>G) polymorphisms (when considered singly or as combinations) and the embryo implantation rate. These associations may be predictive of embryo implantation and could help to define populations in which elective single-embryo transfer should be recommended (or, conversely, ruled out). However, the mechanism underlying the function of these polymorphisms in embryo implantation remains to be determined and the associations observed here must be confirmed in a larger, more heterogeneous cohort.

Exposure to poor maternal nutrition around the time of conception results in an early prepartum activation of the fetal pituitary-adrenal axis and in increased adrenal growth and stress response after birth associated with epigenetic changes in a differentially methylated region (DMR) of adrenal IGF2/H19. We have determined the effects of maternal undernutrition during the periconceptional period (PCUN: 70% of control intake from 60 days before until 6 days after conception) and early preimplantation period (PIUN: 70% of control intake for 6 days after conception) on fetal plasma ACTH and cortisol concentrations and fetal adrenal ACTHR, StAR, 3βHSD, CYP11B, CYP17, TGFβ1, IGF1, IGF1R, IGF2, and IGF2R mRNA expression and the methylation level of sites within the DMRs of IGF2/H19 and IGF2R in the adrenal of twin and singleton fetuses at 136-138 days gestation. Being a twin resulted in a delayed prepartum increase in fetal ACTH and in a lower cortisol response to CRH in the control but not PCUN and PIUN groups. PCUN, but not PIUN, resulted in an increase in adrenal weight and CYP17 expression in singletons, a decrease in adrenal IGF2 expression in singletons, and an increase in adrenal IGF2R expression in both twins and singletons. IGF2/H19 and IGF2R DMR methylation levels and ACTHR expression were lower in the twin adrenal. Thus, exposure of the oocyte and embryo to maternal undernutrition or to the environment of a twin pregnancy have differential effects on epigenetic and other factors that regulate fetal adrenal growth and IGF2 and IGF2R expression., (Copyright © 2014 the American Physiological Society.)

For the past three decades, assisted reproductive technologies (ART) have revolutionized infertility treatments. The use of ART is thought to be safe. However, early investigations suggested that children born as a result of ART had higher risk of diseases with epigenetic etiologies, including imprinting disorders caused by a lack of maternal methylation at imprinting control elements. In addition, large epidemiology studies have highlighted an increased risk of obstetric complications, including severe intrauterine growth restriction (IUGR) in babies conceived using ART. It is plausible that the increased frequency of IUGR may be due to abnormal imprinting because these transcripts are key for normal fetal growth and development. To address this, we have collected a large cohort of placenta and cord blood samples from ART conceptions and compared the imprinting status with appropriate non-ART population. Using a custom DNA methylation array that simultaneously quantifies 25 imprinted differentially methylated regions, we observed similar epigenetic profiles between groups. A multiplex Sequenom iPLEX allelic expression assay revealed monoallelic expression for 11 imprinted transcripts in our placenta cohort. We also observe appropriate gestational age-dependent methylation dynamics at retrotransposable elements and promoters associated with growth genes in ART placental biopsies. This study confirms that children conceived by ART do not show variability in imprinted regulation and that loss-of-imprinting is not commonly associated with nonsyndromic IUGR or prematurity.

Background: Currently very few noninvasive molecular genetic approaches are available to determine zygosity for twin pregnancies in clinical laboratories. This study aimed to develop a novel method to determine zygosity by using maternal plasma target region sequencing., Methods: We constructed a statistic model to calculate the possibility of each zygosity type using likelihood ratios ( Li ) and empirical dynamic thresholds targeting at 4,524 single nucleotide polymorphisms (SNPs) loci on 22 autosomes. Then two dizygotic (DZ) twin pregnancies,two monozygotic (MZ) twin pregnancies and two singletons were recruited to evaluate the performance of our novel method. Finally we estimated the sensitivity and specificity of the model in silico under different cell-free fetal DNA (cff-DNA) concentration and sequence depth., Results/conclusions: We obtained 8.90 Gbp sequencing data on average for six clinical samples. Two samples were classified as DZ with L values of 1.891 and 1.554, higher than the dynamic DZ cut-off values of 1.162 and 1.172, respectively. Another two samples were judged as MZ with 0.763 and 0.784 of L values, lower than the MZ cut-off values of 0.903 and 0.918. And the rest two singleton samples were regarded as MZ twins, with L values of 0.639 and 0.757, lower than the MZ cut-off values of 0.921 and 0.799. In silico, the estimated sensitivity of our noninvasive zygosity determination was 99.90% under 10% total cff-DNA concentration with 2 Gbp sequence data. As the cff-DNA concentration increased to 15%, the specificity was as high as 97% with 3.50 Gbp sequence data, much higher than 80% with 10% cff-DNA concentration., Significance: This study presents the feasibility to noninvasively determine zygosity of twin pregnancy using target region sequencing, and illustrates the sensitivity and specificity under various detecting condition. Our method can act as an alternative approach for zygosity determination of twin pregnancies in clinical practice.

Hedgehog signaling is involved in regulation of ovarian function in Drosophila, but its role in regulating mammalian ovarian folliculogenesis is less clear. Therefore, gene expression of Indian hedgehog (IHH) and its type 1 receptor, patched 1 (PTCH1), were quantified in bovine granulosa (GC) or theca (TC) cells of small (1-5 mm) antral follicles by in situ hybridization and of larger (5-17 mm) antral follicles by real-time RT-PCR from ovaries of cyclic cows genetically selected (Twinner) or not selected (control) for twin ovulations. Expression of IHH mRNA was localized to GC and cumulus cells, whereas PTCH1 mRNA was greater in TC than in GC. Estrogen-active (E-A; follicular fluid concentration of estradiol > progesterone) versus estrogen-inactive follicles had a greater abundance of mRNA for IHH in GC and PTCH1 in TC. Abundance of IHH mRNA in GC was not affected by cow genotype, whereas TC PTCH1 mRNA was less in large E-A follicles of Twinners than in controls. In vitro, estradiol and wingless-type (WNT) 3A increased IHH mRNA in IGF1-treated GC. IGF1 and BMP4 treatments decreased PTCH1 mRNA in small TC. Estradiol and LH increased PTCH1 mRNA in IGF1-treated TC from large and small follicles, respectively. In summary, functional status of ovarian follicles was associated with differences in hedgehog signaling in GC and TC. We hypothesize that as follicles grow and develop, increased free IGF1 may suppress expression of IHH mRNA by GC and PTCH1 mRNA by TC, and these effects are regulated in a paracrine way by estradiol and other intra- and extragonadal factors.

Objective: To perform a cost-benefit analysis of preimplantation genetic diagnosis (PGD) for carrier couples of cystic fibrosis (CF) compared with the alternative of natural conception (NC) followed by prenatal testing and termination of affected pregnancies., Design: Cost-benefit analysis using a decision analytic model., Setting: Outpatient reproductive health practices., Patient(s): A simulated cohort of 1,000 female patients., Intervention(s): We calculated the net benefit of giving birth to a child as the present value of lifetime earnings minus lifetime medical costs., Main Outcome Measure(s): Net benefits in dollars., Result(s): When used for women younger than 35 years of age, the net benefit of PGD over NC was $182,000 ($715,000 vs. $532,000, respectively). For women aged 35-40 years, the net benefit of PGD over NC was $114,000 ($634,000 vs. $520,000, respectively). For women older than 40 years, however, the net benefit of PGD over NC was -$148,000 ($302,000 vs. $450,000, respectively)., Conclusion(s): Preimplantation genetic diagnosis provides net economic benefits when used by carrier couples of CF. Although there is an upper limit of maternal age at which economic benefit can be demonstrated, carrier couples of CF should be offered PGD for prevention of an affected child., (Copyright 2010 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.)

Background: In studies of twin and triplet pregnancies, molecular genetic techniques have rarely been used to confirm zygosity yet this is the most accurate approach. The aim of this study was to present the cross-distribution of chorionicity and zygosity in triplet pregnancies as a function of their mode of conception using such techniques. Rates of monozygosity were studied simultaneously., Methods: Forty-nine consecutive sets of triplets were observed in the study, including 18 sets of spontaneously conceived (SC) triplet pregnancies and 31 sets resulting from assisted reproduction technologies (ARTs). Zygosity was determined through PCR-amplified microsatellite analysis. Chorionicity was determined by placental analysis in our department of fetopathology. Sets of triplets were considered as twin pairs in order to determine the rate of monozygosity., Results: For SC triplet pregnancies, the rate of monozygotic (MZ) twin pairs was 48%; 30% of dichorionic (DC) triplet pregnancies were MZ and 70% dizygotic (DZ); 20% of trichorionic (TC) triplet pregnancies were DZ and 80% trizygotic (TZ). For triplet pregnancies conceived using ART, the rate of MZ twin pairs was 6.5%; 100% of DC triplet pregnancies were DZ; 4% of TC triplet pregnancies were DZ and 96% TZ., Conclusions: This study is the first report to present the cross-distribution of chorionicity and zygosity in triplet pregnancies as a function of their mode of conception. In triplet pregnancies conceived using ART, DC triplets are always DZ, and TC triplets are almost always TZ.

Objective: To test for chromosomal abnormalities in a missed abortion involving a twin gestation., Design: Case report., Setting: University-based infertility practice., Patient(s): A 37-year-old G3P0030 with recurrent abortions presented with a fourth missed abortion involving a twin gestation in the first trimester., Intervention(s): Under anesthesia, hysteroscopy was performed and specimens obtained for karyotyping from each twin., Main Outcome Measure(s): Karyotype of each twin., Result(s): The karyotype of the conceptus on the left was 46,XY and that on the right was 46,XX., Conclusion(s): Direct hysteroscopic biopsies can successfully differentiate the karyotypes of each twin of a missed abortion.

Objective: To present serum androgen levels during pregnancy in a twin gestation complicated by maternal late onset 21-hydroxylase deficiency., Design: Case report., Setting: University teaching hospital reproductive endocrinology and infertility practice., Patient(s): A 27-year-old with nonclassic 21-hydroxylase deficiency and infertility, twin female fetuses, and elevated androgens., Intervention(s): Steroid replacement., Main Outcome Measure(s): Serum T and 17-hydroxyprogesterone (17-OHP) levels., Result(s): Elevated androgen levels persisted throughout pregnancy in spite of aggressive steroid replacement. However, twin girls were born without any evidence of virilization., Conclusion(s): The changes associated with a twin gestation may result in excessive stimulation of androgens in mothers with nonclassic 21-hydroxylase deficiency. However, the increased placental aromatase provides protection.

Objective: To determine the genotype of a triplet resulted from an IVF procedure, with discordant cystic fibrosis (CF) phenotype., Design: Molecular diagnosis of CF., Setting: Affected triplet followed at the CF Clinic Center of Children's Hospital Los Angeles was referred to Molecular Genetics Laboratory at Georgetown University Medical Center for comprehensive DNA analysis of the cystic fibrosis transmembrane regulator (CFTR) gene., Patient(s): Three affected children and the healthy parents with negative family history of CF., Main Outcome Measure(s): Temporal temperature gradient gel electrophoresis and direct DNA sequencing were used to detect and to identify the mutations., Result(s): The child with classic CF had DeltaF508 and R553X mutations. Two children with mild CF symptoms had DeltaF508 and R117C. The father carried two mutations, R553X and R117C. The mother is a carrier for DeltaF508., Conclusion(s): Mutational analysis of the CFTR gene should always be recommended to the infertile couples seeking for IVF. The CFTR mutation screening would be essential if the man has congenital bilateral absence of vas deferens (CBAVD) despite the negative family history of CF. Option of complete CFTR gene analysis at a cost of about 1,500-2,000 dollars should be made available if one mutation is found in the male partner with congenital bilateral absence of vas deferens.

Objective: To evaluate the value of short tandem repeat polymerase chain reaction (STR-PCR) in identification of zygosity in multiple pregnancy and detection of the common chromosomal trisomies., Methods: Amniotic fluid or fetal blood samples were collected from 38 fetuses in 17 multiple pregnant women who had indications for prenatal diagnosis or planed to be performed feticide. Of the 17 cases there were 13 sets of twins (26 fetuses) and 4 sets of triplets (12 fetuses). Parental blood samples were collected. STR-PCR technique was employed to determine the zygosity and detect trisomy 21, trisomy 18, trisomy 13, and trisomy sex chromosomes. Amniotic fluid or blood samples were collected from 18 singleton pregnant fetuses as controls for detecting chromosomal trisomies., Results: All 7 cases with gestation by assisted reproductive techniques were multizygotic. In 7 cases associated with one fetal malformation, only one set of twin was dizygotic. There were no above mentioned trisomies found in the multiple pregnancies while 2 trisomy 21, one trisomy 18 and 13 respectively were detected in the singleton pregnant group. Feticide was performed on 4 multizygotic cases in which 3 were triplets and 1 was twin associated with an abnormal fetus. Following-up showed that there was no any bad effect after procedure., Conclusion: STR-PCR can be used in identifying the zygosity correctly and quickly while being used in detecting chromosomal trisomies. It facilitates monitoring multiple pregnancy as well as selecting appropriate methods of feticide.

Background: The aim of this study was to establish the frequency of fetal cells in the maternal blood of multifetal pregnancies and compare this figure with singleton pregnancies., Methods: We obtained maternal blood from 31 pregnancies with 2-6 fetuses at 11-16 weeks gestation and from 50 normal singleton controls (11-14 weeks gestation). Fetal erythroblasts were isolated from maternal blood using triple density gradient separation and anti-CD71 magnetic cell-sorting techniques. The enriched erythroblasts were stained with Kleihauer-Giemsa and with fluorescent antibodies for the zeta (zeta), epsilon (epsilon) and gamma (gamma) globin chains. The percentage of fetal cells positive for each stain was calculated. Fluorescence in-situ hybridization (FISH) for X and Y chromosomes was also performed., Results: The percentage of erythroblasts enriched from maternal blood that stained positive for zeta, epsilon and gamma globin chains and with Kleihauer-Giemsa was significantly higher in the multifetal compared with singleton pregnancies. The median enriched percentage of positively stained erythroblasts was about three times higher in the twin than in singleton pregnancies (P < 0.0001), nearly twice as high in the triplet than in twin pregnancies (P < 0.01) and five times higher in the triplet than singleton pregnancies (P < 0.0001). FISH for Y chromosome confirmed the increase in fetal cell proportion in the multifetal pregnancies., Conclusions: These findings suggest that there is an increase in the physiological feto-maternal cell trafficking in multifetal pregnancies compared with singleton pregnancies, which is likely to be due to the increased placental surface area and vasculature.

Objective: To determine the influence of the A307/S680 and T307/N680 isoforms of the follicle-stimulating hormone receptor (FSHR) gene on the incidence of spontaneous human twinning., Design: Case-control study., Setting: Departments of Obstetrics and Gynecology and of Clinical Chemistry-Grosshadern, University Hospital Munich, Germany., Patient(s): Fifty-four mothers with dichorionic twin pregnancies and 92 singleton mothers as controls, who had conceived without assisted reproduction., Intervention(s): Exon 10 of the FSHR gene was screened for the G2105A/S680N mutation., Main Outcome Measure(s): Amplification of genomic DNA by the polymerase chain reaction followed by restriction fragment length polymorphism analysis., Result(s): Allele frequencies for the G2105A/ S680N substitution of the FSHR in twin mothers were not different from those of controls (genotype (isoform) [twins vs. controls]: G/G (S/S) [24.1% vs. 22.3%]; A/G (N/S) [57.4% vs. 55.4%]; A/A (N/N) [18.5% vs. 22.3%]). Subgroup analysis of women with three or more successful pregnancies gave a similar result (G/G (S/S) [17.7% vs. 13.7%]; A/G (N/S) [64.7% vs. 63.6%]; A/A (N/N) [17.6% vs. 22.7%]). There was no correlation between FSHR isoform and twinning., Conclusion(s): The S680N substitution of the follicle-stimulating hormone receptor is a common polymorphism not associated with spontaneous human twinning.

In multiple pregnancies, demands for folic acid are considerably increased. The most common inborn error of folate metabolism is mild methylenetetrahydrofolate reductase (MTHFR) deficiency due to the synthesis of a thermolabile variant of the enzyme with impaired catalytic activity which leads to reduced 5-methyltetrahydrofolate (5-methyl-THF) and mildly elevated homocysteine plasma concentrations when folate status is inadequate. To investigate whether the number of offspring is influenced by this mutation, we determined the frequency of the 677C-->T substitution in 156 singleton and 40 twin mothers with dichorionic placentation. The T allele frequency in singleton (0.30) and twin mothers (0.16) was significantly different (P = 0.011). Mothers with the 677C-->T mutation had a 2.28 times lower risk of having a twin pregnancy than those without (95% confidence interval = 1.18-4.66; P = 0.008). Our observation would explain, at least in part, the hereditary trait of multiple gestations and is in agreement with the ethnic distribution pattern of the T allele which has been found to be inversely correlated with the incidence of dichorionic twins. Our findings suggest that the MTHFR 677C-->T mutation interferes with human brood size, probably by influencing the proliferation of rapidly dividing embryonic and maternal cells.

On the basis of MEDLINE and manual searches, we examined the main papers in the English literature regarding risk factors for spontaneous (i.e. not related to fertility drug use) multiple births. The constant frequency of monozygotic (MZ) pregnancies over time and in different geographical areas suggests that the determination of MZ twins is largely unchanged over time, and that a genetic mechanism may have a role. In contrast, temporal and geographical trends observed in dizygotic (DZ) pregnancies suggest that environmental factors play a role in determining this condition. At present, maternal age and hereditary components are the best-defined determinants for spontaneous multiple births.

In 176 diamniotic twin pregnancies at 10-20 weeks of gestation, amniotic fluid for cytogenetic studies was successfully obtained from both sacs by the use of a single uterine entry. There were no cases of discordancy between sex at amniocentesis and birth. There were six pregnancies with fetal unbalanced chromosomal defects; in one pregnancy both fetuses were abnormal and in five pregnancies only one fetus was abnormal. The total fetal loss rate was 5.7% (20 of 352 fetuses), including six (1.7%) terminations or selective fetocides and 14 (4.0%) spontaneous deaths. In the 176 pregnancies there were five (2.8%) with no survivors, including one termination and four (2.3%) spontaneous miscarriages or intrauterine deaths. There are only two (1.1%) pregnancies in which amniocentesis could have contributed directly to the losses and therefore the procedure-related rate of fetal loss may be similar to that in singleton pregnancies. The median gestation at delivery was 37 (range 16-40) weeks and delivery before 32 weeks occurred in 9% of the pregnancies. The birth weight distribution was similar to that reported in singleton pregnancies. This study demonstrates that in twin pregnancies amniotic fluid for cytogenetic studies can be obtained successfully from both sacs by use of a single uterine entry. The risk of fetal loss from this procedure appears to be similar to that in singleton pregnancies.

On the basis of a family reconstitution of the rural Krummhörn population (Ostfriesland, Germany) of the eighteenth and nineteenth centuries, we pursued the question of to what extent the birth of twins contributed to the reproductive fitness of their mothers. The twinning rate was 16.2/1000; the secondary sex ratio among twins was 0.93, and it was 1.16 among their singleton siblings. Mothers of twins were older, had a longer generative life phase, and achieved higher age-specific fertility rates with shorter birth intervals. Parity effects on twinning tendency could not be detected. Twin maternities caused reproductive costs, namely, increased maternal, infant, and child mortality and obviously higher intrafamilial competition, because adult twins had fewer local marriage chances and to a higher degree were forced to emigrate. These reproductive disadvantages mean that the productivity of a male pair of twins, as measured by the number of live-born grandchildren, is clearly less than the productivity of a single boy. On the other hand, the birth of a female pair of twins led to more live-born grandchildren than the birth of a single girl. In sum, mothers of twins achieved greater reproductive success, with 13.5% more live-born grandchildren, than mothers of singletons only. The results are discussed against the background of Anderson's (1990) error hypothesis of twinning.

This paper analyzes from the mid 18th century to 1987 the birth records of the "Dariusleut," one of the three subgroups of the Hutterite population. The aim of this study is to describe several aspects of the twinning rate in a fertile population. The overall rate of twinning was 0.90%: 103 twins among all 11,492 maternities. The rate peaked at the 7th birth order and at the maternal age of 40 years and over. Until the mid 19th century when the Hutterites lived in Russia, the twinning rate was higher (1.5%), and it decreased during the migration period in the second half of the 19th century (0.7%). After the group had settled in the USA and Canada, the population maintained a twinning rate of 1.0% until 1965. After 1965 the rate decreased to 0.7%, partly due to a decline in fertility among women aged 30 years and over. There was a significant seasonal variation: the twinning rate decreased to 0.5% in May-July compared to 1.2% for the other three seasons during the years up to 1965 (P < 0.01), while more recent mothers did not show such a seasonal variation. The incidence of twin births in this population seems to have been influenced by environmental factors, which would change their effect seasonally and secularly.

In a sample of 160 Dutch twin pairs and their parents, we found that mothers of dizygotic twins had frequencies of the S and Z alleles at the protease inhibitor (Pi) locus that were 3 times higher than a control sample. Mothers of identical twins also had a higher frequency of S than controls. The S allele may thus both increase ovulation rate and enhance the success of multiple pregnancies. There was also an increased frequency of the S allele in fathers of dizygotic twins; however, this may be a secondary effect of assortative mating for family size (indicating by the number of siblings of the parents), for which a correlation of 0.2 was observed. Parents of dizygotic twins came from larger families than parents of monozygotic twins, but no effect of Pi type on family size was seen.