Genome-wide non-invasive prenatal testing in single- and multiple-pregnancies at any risk: Identification of maternal polymorphisms to reduce the number of unnecessary invasive confirmation testing.

OBJECTIVE: Non-invasive prenatal testing by targeted or genome-wide copy number profiling (cnNIPT) has the potential to outperform standard NIPT targeting the common trisomies 13, 18, and 21, only. Nevertheless, prospective results and outcome data on cnNIPT are still scarce and there is increasing evidence for maternal copy number variants (CNVs) interfering with results of both, standard and cnNIPT. STUDY DESIGN: We assessed the performance of cnNIPT in 3053 prospective and 116 retrospective cases with special consideration of maternal CNVs in singleton and multiple gestational pregnancies at any risk, as well as comprehensive follow-up. RESULTS: A result was achieved in 2998 (98.2%) of total prospective cases (89.2% analyzed genome-wide). Confirmed fetal chromosomal abnormalities were detected in 45 (1.5%) cases, of which five (11%) would have remained undetected in standard NIPTs. Additionally, we observed 4 likely fetal trisomies without follow-up and a likely phenotype associated placental partial trisomy 16. Moreover, we observed clinically relevant confirmed maternal CNVs in 9 (0.3%) cases and likely maternal clonal hematopoiesis in 3 (0.1%). For common fetal trisomies we prospectively observed a very high sensitivity (100% [95% CI: 91.96-100%]) and specificity (>99.9% [95% CI: 99.8-100%]), and positive predictive value (PPV) (97.8% [95% CI: 86.1-99.7%]), but our retrospective control cases demonstrated that due to cases of fetal restricted mosaicism the true sensitivity of NIPT is lower. After showing that 97.3% of small CNVs prospectively observed in 8.3% of genome-wide tests were mostly benign maternal variants, sensitivity (75.0% [95% CI: 19.4%-99.4%]), specificity (99.7% [99.5%-99.9%]) and PPV (30.0% [14.5%-52.1%]) for relevant fetal CNVs were relatively high, too. Maternal autoimmune disorders and medication, such as dalteparin, seem to impair assay quality. CONCLUSION: When maternal CNVs are recognized as such, cnNIPT showed a very high sensitivity, specificity and PPV for common trisomies in single and multiple pregnancies at any risk and very good values genome-wide. We found that the resolution for segmental aberrations is generally comparable to standard karyotyping, and exceeds the latter if the fetal fraction is above 10%, which allows detection of the 2.5 Mb 22q11.2 microdeletion associated with the velocardiofacial syndrome, even if the mother is not a carrier.

Accuracy of clinical diagnosis of anal sphincter defect: clinical evaluation versus 3D-transperineal ultrasound.

BACKGROUND: Obstetric anal sphincter injuries (OASIS) are common and an important factor in the etiology of anal incontinence. The objective of this study was to evaluate, classify and compare the agreement of clinically diagnosed third-degree sphincter tears with 3D-transperineal ultrasound (3D-TPUS) realized within 3-7 days post-delivery. METHODS: This is a retrospective observational study were 119 patients with third-degree obstetric anal sphincter tears were diagnosed and treated, 85 of those underwent a 3D-TPUS examination 3-7 days postpartum. We compared the proportion of third-degree perineal tears, classified with the clinical examination as grade 3a+b and grade 3c, with the 3D-TPUS. RESULTS: In 16 patients with clinically diagnosed third-degree perineal tears grade a and b, the ultrasound examination confirmed the lesion of the external anal sphincter (EAS) muscle, but in nine patients (56% of the cases) we found a lesion of the internal anal sphincter (IAS) muscle, missed by clinical examination. In the remaining 69 patients with the third-degree perineal tears grade c, the ultrasound examination confirmed both lesions (EAS and IAS muscles) in 56 women, but in 13 patients (19% of the cases) defects of the IAS muscle could not be confirmed by the ultrasound. CONCLUSIONS: There was moderate agreement regarding diagnosis of grade 3a+b and grade c perineal tears between ultrasound and clinical examination, so a combined use of clinical and ultrasound knowledge can improve the possibility to find a gold standard in the diagnosis of OASIS.

Comparison of Current Swiss Fetal Biometry Reference Charts with Reference Charts from 1999. Are Fetuses Getting Bigger?

PURPOSE: To create current fetal biometry reference ranges and to compare them with references published in 1999, from the same local area in order to generate data for secular trend in fetal size. MATERIALS AND METHODS: Applying the same methodology as previously published, we calculated reference ranges for biparietal diameter (BPD), occipitofrontal diameter (OFD), head circumference (HC), abdominal circumference (AC) and femur length (FL) in 7863 patients examined at the obstetric clinics in a cross-sectional, prospective study in a university setting from January 2008 to December 2014. In order to compare the new reference ranges with our previously published data, we used Z-Scores and displayed the pick-up of fetal biometry data below the 5th and above the 95th percentile using the previously published reference charts. RESULTS: The comparison of the charts showed a minimal but clinically relevant increase in mean fetal body measures (BPD, HC, AC). Applying the 1999 charts to the new dataset, we would classify only 162 of 339 fetuses (47.8 %) to be correctly below the 5th percentile for AC and only 134 of 349 (38.4 %) fetuses were correctly below the 5th percentile for HC. On the other hand, the 1999 charts classified 426 instead of 332 fetuses to be above the 95th percentile for AC, which means an overestimation of 28.3 %. CONCLUSION: Applying a similar methodology, study collective and clinical setting, our new charts showed clinically relevant differences compared to the 1999 charts. The data suggest that within one generation fetuses are getting bigger and regular updates of fetal reference charts are needed.

Prognostic Features and Long-Term Outcome in Patients with Isolated Fetal Ventriculomegaly.

OBJECTIVE: In order to provide aid for prenatal counseling in fetal isolated ventriculomegaly (IVM) on ultrasound, we recorded the latest long-term clinical and imaging outcomes of children with a mean age of 7.2 years (range 2.1-14.6). METHODS: In 72 fetuses with IVM, diagnosed between 1999 and 2011, the measurement quality of atrial diameter was reviewed in the axial plane. We assessed the association of characteristics of IVM with outcome parameters in the cohort and in subgroups. Prognostic values of significant associations were reported by receiver operating characteristic curve analysis. RESULTS: Cerebral anomalies were diagnosed postnatally in 42% and genetic disorders in 12% of 45 live births. Significant associations of outcome parameters were found between the degree of IVM and genetic disorders (p = 0.017) with an area under the curve (AUC) of 0.866, and between progression of IVM and motor impairment (p = 0.024) with an AUC of 0.789. No significant correlation was found with the other assessed outcome parameters. Furthermore, our subgroup analysis clearly showed that, if cerebral or genetic anomalies are not found postnatally, a favorable outcome may be expected. DISCUSSION: Diameter and progression in IVM are not significantly associated with most outcome parameters. Cerebral anomalies and genetic disorders may contribute to an unfavorable outcome.

Prenatal Megacystis - Is Prediction of Outcome and Renal Function Possible?

OBJECTIVE: To assess prenatal prognostic criteria for fetuses with megacystis in order to counsel parents. METHODS: In a retrospective observational study at a single tertiary referral center, we assessed the clinical course of 53 fetuses with megacystis cared for at the Department of Obstetrics of the University Hospital Zurich between 1995 and 2008 and followed them up for 2 to 12 years. We determined fetal karyotype, amniotic fluid volume and fetal urinary biochemistry as prenatal prognostic factors. The renal function of survivors was grouped according to age-related creatinine values. Using logistic regression analysis, gestational age-dependent discrimination curves and corresponding ROC curves for fetal urine, beta-2 microglobulin, osmolarity and chloride were calculated. RESULTS: 43 out of 53 fetuses underwent vesicocentesis, and spontaneous remission occurred in 3 fetuses. 15 fetuses survived, termination of pregnancy was requested in 23 cases, and 12 neonatal and 3 intrauterine deaths were observed. Reduced amniotic fluid volume showed a significant (p = 0.0027) increase of impaired renal function or perinatal death. Discrimination between survivors and non-survivors was complete for fetal urine beta-2 microglobulin with an area under the curve (AUC) of 1.0. For fetal urine osmolarity and fetal urinary chloride, the AUC was 0.81 and 0.76, respectively. CONCLUSION: The assessment of prognosis for fetal megacystis should include fetal k aryotyping, determination of amniotic fluid and assessment of fetal urine biochemistry. Gestational age-dependent regression lines disclose clinically relevant discrimination and can be used as selection criteria for fetal interventions and parental counselling.

Pregnancy outcomes regarding maternal serum AFP value in second trimester screening.

AIM: The aim of this study was to evaluate the predictive value of α-fetoprotein in maternal serum (MS-AFP) as a marker for diverse pregnancy outcomes. METHODS: The study was based on pregnancy and delivery data from 5520 women between 1999 and 2014 at University Hospital of Zurich (UHZ). INCLUSION CRITERIA: both MS-AFP and pregnancy outcome were known for the same pregnancy. Pregnancy outcomes and characteristics such as fetal malformation, intrauterine fetal death (IUFD) and intrauterine growth retardation as well as maternal age, weight before pregnancy, gestational age (GA) at delivery, newborn weight, length and head circumference were analyzed with respect to the MS-AFP value. MS-AFP value was categorized into three groups: elevated MS-AFP>2.5 multiples of the median (MoM), normal 0.5-2.49 MoM and decreased <0.5 MoM. RESULTS: Newborn weight (g) and length (cm) were significantly lower in the elevated MS-AFP (P<0.001) group, and infants had 1 week lower GA at delivery (P<0.05). In the group of elevated MS-AFP (n=46), 26.1% of pregnancies were significantly related to adverse pregnancy outcomes, such as fetal malformations, fetuses small for gestational age (SGA) and IUFD. Adverse pregnancy outcomes of 5.6% were registered in the group of normal MS-AFP and 7.3% in the group of low MS-AFP (P<0.05). CONCLUSION: MS-AFP level in the second trimester is still an important indicator of fetal surface malformations; however, ultrasound still outweighs as a screening method. Nevertheless, pregnant women with elevated MS-AFP values and with no sonographically detected fetal malformations should additionally receive the third trimester ultrasound examination to exclude other possible complications of pregnancy.

Noninvasive prenatal testing: more caution in counseling is needed in high risk pregnancies with ultrasound abnormalities.

OBJECTIVE: Non-invasive prenatal testing (NIPT) is increasingly being used in prenatal aneuploidy screening. The objective of this study was to assess the positive predictive value in our cohort of 68 cases with positive NIPT result. In addition, we wondered if the use of NIPT in cases with ultrasound abnormalities is appropriate, given the limited number of chromosomes investigated. DESIGN: We performed confirmative invasive testing using karyotyping, fluorescence in situ hybridization (FISH) and/or high-resolution chromosomal microarray analysis. RESULTS: In line with the published data, the positive NIPT result was confirmed in 64.7% of cases. Inconclusive and negative NIPT results followed by cytogenetically pathologic findings were encountered in three and in five cases, respectively. Four of the five fetuses with negative NIPT but pathologic cytogenetic findings were born with several malformations and diagnosed right after birth with severe genetic conditions. Of note, in all of those four cases, NIPT was offered despite the finding of major fetal ultrasound abnormalities and despite the fact that the family would not have opposed invasive testing or pregnancy termination. CONCLUSION: More education of health care providers and caution in counseling and interpretation of test results are needed in order to meet the challenges that this new test, which enriches our diagnostic options in prenatal testing, poses.

Outcome after prenatal diagnosis of congenital anomalies of the kidney and urinary tract.

UNLABELLED: Congenital anomalies of the kidney and urinary tract are common findings on fetal ultrasound. The aim of this prospective observational study was to describe outcome and risk factors in 115 patients born 1995-2001. All prenatally diagnosed children were stratified into low- and high-risk group and followed postnatally clinically and by imaging at defined endpoints. Risk factors were evaluated using odds ratios. Neonatal diagnosis included pelvi-ureteric junction obstruction (n = 33), vesicoureteral reflux (n = 27), solitary mild pelvic dilatation (postnatal anteroposterior diameter 5-10 mm; n = 25), and further diagnosis as primary obstructive megaureter, unilateral multicystic dysplastic kidney, renal dysplasia and posterior urethral valves. In 38 children with prenatal isolated hydronephrosis, ultrasound normalized at median age of 1.2 years (range 0.1-9). Surgery was performed in 34 children at median age of 0.4 years (0.1-10.8). Persistent renal anomalies without surgery were present in 43 children and followed in 36 for median time of 16 years (12.2-18). Oligohydramnios and postnatal bilateral anomalies were significantly associated with surgery and impaired renal function. CONCLUSION: The majority of children had a favourable postnatal outcome, in particular children with prenatally low risk, i.e. isolated uni- or bilateral hydronephrosis. Oligohydramnios and postnatal bilateral anomalies were risk factors for non-favourable outcome. WHAT IS KNOWN: • In congenital anomalies of the kidney and urinary tract significantly poorer outcome is known in patients with bilateral renal hypoplasia or solitary kidney associated with posterior urethral valves. • Other factors as proteinuria and vesicoureteral reflux were associated with a higher risk of progression to chronic renal failure in these patients. What is New: • Unlike other studies giving us above-mentioned information, we included all patients with any kind of prenatally diagnosed congenital anomalies of the kidney and urinary tract. Our study shows long-term follow up (median 16 years, range 12.2-18 years), especially in patients not needing surgery, but with persistent anomalies. • During postnatal long-term follow up (median 2.2 years, range 0.1-18 years) one third each showed normalization, need of surgery or persistence of anomalies without need of surgery. Our study revealed a good prognosis in the majority of these children, in particular with prenatally low risk, i.e. isolated uni- or bilateral hydronephrosis, and revealed oligohydramnios and postnatal bilateral anomalies as risk factors for a non-favourable outcome, defined as need of surgery, persistent anomalies with impaired renal function, end stage renal failure or death.

In utero and postnatal imaging findings of parasitic conjoined twins (ischiopagus parasiticus tetrapus).

Conjoined twins are a rare developmental anomaly with a reported prevalence of 1.47 per 100,000 births. We present an uncommon case of a parasitic ischiopagus tetrapus with a parasitic ischiopagus partial twin joined to the complete fetus at the level of the ischium diagnosed in utero by fetal MRI. The correct prenatal diagnosis led to birth by caesarean section. Prenatal MRI findings are presented and corroborated by postnatal imaging delineating the full extent and associated anomalies of this rare malformation. Differential diagnosis of duplicated lower extremities is discussed.

High-resolution chromosomal microarrays in prenatal diagnosis significantly increase diagnostic power.

OBJECTIVE: The objective of this study was to determine for the first time the reliability and the diagnostic power of high-resolution microarray testing in routine prenatal diagnostics. METHODS: We applied high-resolution chromosomal microarray testing in 464 cytogenetically normal prenatal samples with any indication for invasive testing. RESULTS: High-resolution testing revealed a diagnostic yield of 6.9% and 1.6% in cases of fetal ultrasound anomalies and cases of advanced maternal age (AMA), respectively, which is similar to previous studies using low-resolution microarrays. In three (0.6%) additional cases with an indication of AMA, an aberration in susceptibility risk loci was detected. Moreover, one case (0.2%) showed an X-linked aberration in a female fetus, a finding relevant for future family planning. We found the rate of cases, in which the parents had to be tested for interpretation of unreported copy number variants (3.7%), and the rate of remaining variants of unknown significance (0.4%) acceptably low. Of note, these findings did not cause termination of pregnancy after expert genetic counseling. The 0.4% rate of confined placental mosaicism was similar to that observed by conventional karyotyping and notably involved a case of placental microdeletion. CONCLUSION: High-resolution prenatal microarray testing is a reliable technique that increases diagnostic yield by at least 17.3% when compared with conventional karyotyping, without an increase in the frequency of variants of uncertain significance.

Risks of pregnancy and birth in obese primiparous women: an analysis of German perinatal statistics.

PURPOSE: To compare risks of pregnancy and birth in obese (body mass index, BMI ≥ 30) and normal weight women (BMI 18.5-24.99) giving birth to their first child. METHODS: We analysed data of 243,571 pregnancies in primiparous women from the German perinatal statistics of 1998-2000. We calculated odds ratios (ORs) with 95% confidence intervals (CIs) for selected pregnancy and birth risks. ORs were adjusted for the confounding factors age, smoking status, single mother status, and maternal education. RESULTS: Obesity during pregnancy is common in primiparous women (n = 19,130; 7.9% of all cases) and it is significantly associated with a number of risks of pregnancy and birth, including diabetes [OR 3.71 (95% CI 2.93; 4.71); p < 0.001], hypertension [OR 8.44 (7.91; 9.00); p < 0.001], preecalmpsia/eclampsia [OR 6.72 (6.30; 7.17); p < 0.001], intraamniotic infection [OR 2.33 (2.05; 2.64); p < 0.001], birth weight ≥ 4,000 g [OR 2.16 (2.05; 2.28); p < 0.001], and an increased rate of Caesarean section [OR 2.23 (2.15; 2.30); p < 0.001]. Some risks were less frequent in the obese such as cervical incompetence [OR 0.55 (0.48; 0.63); p < 0.001] and preterm labour [OR 0.47 (0.43; 0.51); p < 0.001]. CONCLUSIONS: Obesity during pregnancy is an important clinical problem in primiparous women because it is common and it is associated with a number of risks of pregnancy and birth. Because of these increased risks, obese women need special attention clinically during the course of their first pregnancy. Weight reduction before the first pregnancy is generally indicated in obese women to prevent the above-mentioned complications of pregnancy and birth.

Vitamin D receptor genotype BB is associated with higher serum osteocalcin in first pregnancy.

AIM: Serum osteocalcin was shown in a previous study on first trimester pregnant women to correlate with bone density and to distinguish between fast and slow bone losers. The objective of the present study is to examine whether serum osteocalcin is related to vitamin D receptor (VDR) BsmI polymorphism in pregnant women. STUDY DESIGN: We determined osteocalcin serum levels and VDR BsmI genotype in 97 healthy first trimester pregnant women consecutively recruited during six months. RESULTS: BB (21%), Bb (38%) and bb (41%) genotypes showed similar osteocalcin serum levels. However, in primigravidas (n=38) the BB genotype was significantly associated with higher mean osteocalcin level (9.67 ng/mL) than the Bb (8.07 ng/mL) and the bb genotype (8.14 ng/mL), respectively (P<0.05). The VDR genotype was the only independent parameter to correlate with serum osteocalcin (P<0.05). CONCLUSION: Only primigravidas show in the first trimester a relation between the bone formation parameter serum osteocalcin and the VDR genotype BB which indicates a higher risk of fractures. For further clinical applications serum osteocalcin and VDR genotype should be tested on a cohort of primigravidas including measurements of bone density.

Osteogenesis imperfecta type II: fetal magnetic resonance imaging findings.

We describe the prenatal magnetic resonance imaging (MRI) findings in a 25-week-old fetus with proven osteogenesis imperfecta type II. Fetal MRI findings are correlated with prenatal ultrasonography, conventional x-ray fetography and postpartal findings. Fetal MRI proves to be superior to conventional fetography in the evaluation of the skeletal findings. The high soft tissue resolution, the large field of view and the multiplanar imaging make it possible to study the non-ossified fetal skeleton in detail. Compared to prenatal ultrasonography, fetal MRI gives important additional information about the expected lung functionality by estimating fetal lung volume and signal intensity and rules out possible additional abnormalities of major fetal organs. These parameters may serve as valuable, additional prognostic markers in the prenatal diagnostic work-up.

Monochorionic-diamniotic twins discordant in gender from a naturally conceived pregnancy through postzygotic sex chromosome loss in a 47,XXY zygote.

OBJECTIVE: It is generally believed that monochorionic-diamniotic twin pregnancies result from one fertilized oocyte with both siblings having the same genotype and phenotype. In rare instances, due to somatic mutations or chromosome aberrations, the karyotypes and phenotypes of the two twins can differ. METHOD: We report cytogenetic, molecular genetic and clinical examinations in monochorionic-diamniotic twins discordant in gender. RESULTS: The monochorionic-diamniotic status of the twins was diagnosed by ultrasound and histologic examination of the placenta. Prenatal chromosome examination performed on amniocytes revealed a normal female karyotype in one and a 46,XX(26)/46,XY(3) karyotype in the other twin. Molecular examinations confirmed monozygosity despite discordant sex. Based on the cytogenetic and molecular results of lymphocytes and placental cells, the only explanation for gender discordance was that the conceptus originally had a 47,XXY chromosome complement. CONCLUSION: A 47,XXY zygote appears to have undergone a twinning process. A postzygotic loss of the X chromosome in some cells and the Y chromosome in other cells, either before or after twinning, resulted in 46,XX/46,XY mosaicism in both monozygotic (MZ) twins. The sex discordance of the MZ twins can be explained by different proportions of the 46,XX and 46,XY cell lines in the gonads and other tissues.

Pre- and postnatal findings in trisomy 17 mosaicism.

Trisomy 17 mosaicism is one of the rarest autosomal trisomies in humans. Thus far, only 23 cases have been described, most of them detected prenatally. In only five instances has mosaicism been demonstrated in lymphocytes and/or fibroblasts postnatally, and only in these have multiple congenital anomalies (MCA), facial dysmorphisms, and mental retardation been reported. Patients with trisomy 17 mosaicism at amniocentesis and a normal karyotype in blood and fibroblasts (n = 17) were always healthy. Here, we report on pre- and postnatal clinical, cytogenetic, molecular-cytogenetic, and molecular findings in four patients with trisomy 17 mosaicism. The first case was detected in cultured but not in short-term chorionic villi and amniocytes. Due to MCA on prenatal ultrasound examination the pregnancy was terminated. The second patient is a 13-month-old healthy boy, in whom low level trisomy 17 mosaicism was detected in cultured chorionic villi only. The third patient is a 2-year-old girl with growth retardation, developmental delay, MCA, and trisomy 17 mosaicism in amniocytes, fibroblasts, and placenta, but not in blood and buccal smear. The fourth patient is a 9-year-old boy with growth and mental retardation, sensoneurinal hearing loss, and MCA. Cytogenetic analyses showed trisomy 17 mosaicism in amniocytes, skin fibroblasts, and urinary sediment cells, whereas in blood and buccal smear a 46,XY karyotype was found. Molecular investigations in all four cases indicated biparental inheritance of chromosome 17. Formation of trisomy was most likely due to a maternal meiosis I error in Patient 1 and a postzygotic non-disjunction of the paternal chromosome 17 in Patient 4. Cerebellar malformations, reported in two cases from the literature and in two reported here may be a specific feature of trisomy 17 mosaicism. Since the aberration has rarely been reported in lymphocytes, chordocentesis is not indicated in prenatal diagnosis. Prenatal genetic counseling for trisomy 17 mosaicism in chorionic villi or amniocytes should consider that the clinical significance remains uncertain.

Postzygotic isochromosome formation as a cause for false-negative results from chorionic villus chromosome examinations.

OBJECTIVE: To investigate the origin and mechanisms of formation of isochromosomes 13q and 21q in instances where prenatal chromosome examination revealed a normal karyotype while postnatal chromosome examination from blood showed translocation trisomy 13 and 21. METHODS: G and/or Q-banded chromosome examinations from CVS cultures and lymphocyte chromosome examinations from two newborns. Microsatellite marker analysis of DNA from the probands and their parents. Prenatal ultrasonic examinations of the fetuses and postnatal clinical examinations of the probands. RESULTS: Short and long-term CVS examinations from two fetuses revealed normal karyotypes. Lymphocyte karyotypes of the newborns showed the karyotype 46,XY,i(21)(q10) in the first case and 46,XY,i(13)(q10) in the second. The isochromosomes 21q and 13q were shown, by microsatellite marker analysis of the patients and their parents, to be of maternal and paternal origin, respectively. CONCLUSION: Postzygotic isochromosome formation is one of the possible mechanisms that may lead to false-negative results of chorionic villus chromosome examinations, even if both short-term and long-term cultures are performed and give normal results.

Rigid overlay of volume sonography and MR image data of the female pelvic floor using a fiducial based alignment--feasibility due to a case series.

The visual combination of different medical image acquisition techniques (modalities) can lead to new modalities with enhanced informative content. In this paper, we present an overlay technique of magnetic resonance (MR) and 3D US image data sets of the female anal canal (internal and external sphincter) as a base for a new diagnostic modality. It is a new field of the application of the overlay technique. Three corresponding MR and US volume data sets from the female pelvic floor region were filtered using adaptive filtering techniques and overlayed (=registered rigidly) with a landmark based alignment method.