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Background: Patients with atypical values of HCG and/or PAPP-A are at higher risk of chromosomal abnormality and vascular complications of pregnancy. The performance of cfDNA in this particular population has not yet been evaluated., Objectives: The primary objective was to evaluate the usefulness and reliability of cfDNA in screening for trisomy 21, 18 and 13 for patients with HCG < 0.25 multiple of median (MoM), HCG > 5.0 MoM and/or PAPP-A < 0.25 MoM, PAPP-A > 2.5 MoM. The secondary objective was to evaluate the contribution of cfDNA assay for the prediction of pregnancy's vascular complications., Method: Between June 2016 and July 2017, we analysed a women cohort from all over France who had at least one first trimester serum biomarker outside of normal range, in a retrospective, observational and multicentre study. Patients were included if they had a single pregnancy, normal first trimester ultrasound examination, whatever the result of the combined first trimester screening test was. The cfDNA was analysed by massive parallel sequencing technique. The accuracy of cfDNA assay was evaluated by calculation of sensitivity and specificity, and multivariate regression analysis was used to search for predictive factors for pregnancy's vascular complications., Results: Among the 498 patients who underwent a cfDNA assay in this context, twenty-one (4.2%) were excluded because of loss to follow-up. Out of 477, test failure occurred for four patients initially, reduced to two patients (0.4%) after redrawn. CfDNA was positive for Trisomy 21 (n = 19), Trisomy 18 (n = 6) and Trisomy 13 (n = 1) and negative in 449. The sensitivity of cfDNA assay for trisomy 21 screening was 100% (19/19) (IC 95% 82.4-100) and specificity 100% (458/458) (IC 95% 99.2-100). Among the 447 patients included for prediction of vascular complications, there were four cases of pregnancy induced hypertension and 10 cases of preeclampsia, for which no predictive factor was identified. Intra Uterine growth restriction under 5th percentile (n = 44, 9.8%) was significantly associated with a low fetal fraction (OR = 0.87, IC 95% 0.79-0.96, p = 0.006)., Conclusion: cfDNA assay is an effective and reliable tool for women with atypical profile of first trimester serum biomarkers.

Objectives: To evaluate the failure rate and performance of cell-free DNA (cfDNA) testing, mainly in terms of detection rates for trisomy 21, performed by 2 laboratories using different analytical methods., Methods: cfDNA testing was performed on 2,870 pregnancies with the HarmonyTM Prenatal Test using the targeted digital analysis of selected regions (DANSR) method, and on 2,635 pregnancies with the "Cerba test" using the genome-wide massively parallel sequencing (GW-MPS) method, with available outcomes. Propensity score analysis was used to match patients between the 2 groups. A comparison of the detection rates for trisomy 21 between the 2 laboratories was made., Results: In all, 2,811 patients in the Harmony group and 2,530 patients in the Cerba group had no trisomy 21, 18, or 13. Postmatched comparisons of the patient characteristics indicated a higher no-result rate in the Harmony group (1.30%) than in the Cerba group (0.75%; p = 0.039). All 41 cases of trisomy 21 in the Harmony group and 93 cases in the Cerba group were detected., Conclusions: Both methods of cfDNA testing showed low no-result rates and a comparable performance in detecting trisomy 21; yet GW-MPS had a slightly lower no-result rate than the DANSR method., (© 2018 S. Karger AG, Basel.)

Purpose: Cell-free DNA (cfDNA) as a primary screening test has been available for years but few studies have addressed this option in a prospective manner. The question is of interest after reports that maternal serum screening (MSS) is less accurate for pregnancies resulting from assisted reproduction technologies (ART) than for spontaneous pregnancies (SP)., Methods: A prospective interventional study was designed to address the performances of cfDNA compared with MSS in pregnancies with or without ART. Each patient was offered both MSS and cfDNA testing. The primary analysis cohort ultimately included 794 patients with a spontaneous pregnancy (SP) (n = 472) or pregnancy obtained after ART (n = 322)., Results: Overall, the false-positive rate and positive predictive value were 6.6% and 8.8% for MSS but 0% and 100% for cfDNA. MSS false-positive rate and positive predictive values were clearly poorer in the ART group (11.7% and 2.6%) than in the SP group (3.2% and 21.1%). The global rates of invasive procedures were 1.9% (15/794) with cfDNA but 8.4% (65/794) if MSS alone was proposed., Conclusion: cfDNA achieved better performance than MSS in both spontaneous and ART pregnancies, thus decreasing the number of invasive procedures. Our findings suggest that cfDNA should be considered for primary screening, especially in pregnancies obtained after ART.

Context: Prenatal dexamethasone (DEX) treatment has been proposed since 1984 to prevent genital virilization in girls with congenital adrenal hyperplasia (CAH). DEX is effective in CAH females if initiated before the sixth week of gestation, but its safety in children treated in utero remains controversial regarding cognitive functions., Objective: To avoid prenatal DEX in males and initiate DEX in due time in CAH females, we proposed in 2002 a protocol for fetal sex determination in the maternal serum (SRY test)., Design and Setting: We conducted a retrospective study of the management of 258 fetuses in the period 2002 through 2011 in pregnancies managed in referent medical centers with an institutional practice., Patients: A total of 258 fetuses at risk of CAH (134 males and 124 females) were included., Intervention: DEX was offered after informed consent to pregnant women., Main Outcome Measure: The sensitivity of an early SRY test was evaluated after data collection., Results: The SRY test is sensitive from 4 weeks and 5 days of gestation. It avoided prenatal DEX in 68% of males, and this percentage increased over the years. DEX was maintained until prenatal diagnosis in non-CAH females. Virilization was prevented in 12 CAH girls treated at the latest at 6 weeks gestation and minimized in 3 girls treated between 6 and 7 weeks gestation. Maternal tolerance was correct. No fetal malformations were noted in the 154 children treated in utero., Conclusions: The SRY test is reliable to avoid prenatal DEX in males, but its application must be improved. Prenatal DEX should be maintained to prevent virilization and traumatic surgery in CAH girls after informed consent and information provided to families about the benefit to risk ratio in limiting hyperandrogenism during fetal life. Our large multicentric French cohort has helped to better assess the risks previously reported.

Coelocentesis may represent the ideal technique for early prenatal diagnosis. This study aimed to quantify the number of cells in coelomic fluid and to investigate the feasibility of interphase fluorescence in-situ hybridization (FISH) in uncultured coelomic cells for chromosomes X and Y in 12 samples of 0.4-0.8 ml of coelomic fluid obtained by transvaginal puncture at 8-9 weeks of gestation. It was found that the density of cells in the coelomic fluid was low and variable ranging from 0 to 10,600 cells/ml. The FISH analysis failed in three cases because of the absence or remarkably low number of cells. Among the remaining nine cases, FISH analysis led to an unambiguous result in all the samples except two in whom the FISH analysis clearly demonstrated a high count of maternal cells whereas the fluid was apparently not blood stained. The presence of such maternal cells, while their source and nature remaining unexplained, stressed the question of the absolute necessity to prove the fetal origin of the cells analysed. Whatever the cytogenetic analysis performed on coelomic fluid, combining a systematic exclusion of significant maternal contamination is recommended, using multiplex polymerase chain reaction for short tandem repeat analysis to cytogenetic analyses.

Background: Coelocentesis may represent the ideal technique for very early prenatal diagnosis. Although cell density in coelomic fluid (CF) is very low, the results of analyses on the cellular compartment have been proposed for prenatal diagnosis., Methods and Results: We aimed to evaluate the amount of total DNA (i.e. cellular and cell-free) in 14 samples (0.4-0.8 ml) of CF, taken from women at 8- to 9-week gestation, who are about to undergo termination of pregnancy, and to assess the feasibility of multiple single-gene analyses using multiplex real-time PCR. We found that the amount of total DNA in the CF was very low and varied widely. Genetic testing using multiplex real-time PCR was successfully achieved in 10 of 14 samples (71%). However, when considering samples that could provide a reliable prenatal diagnosis (i.e. successful PCR analysis and no marked maternal contamination), reliable CF-DNA-based prenatal diagnoses were obtained in only 8 of the 14 (58%) samples., Conclusion: The development of highly reliable procedures adapted to pauci-cellular CF is crucially needed before coelocentesis could be proposed for early prenatal diagnosis of genetic diseases before 10 weeks.

Rhesus (Rh) D blood group incompatibility between the pregnant woman and her fetus is a significant problem due to the possibility of maternal alloimmunization and consequent hemolytic disease of the newborn. The RhD-negative blood group is found in 15% of whites, 3-5% of black Africans, and is rare in Asians. Advances in both our understanding of the RHD locus and its variants, as well as technical improvements in the extraction and amplification of cell-free fetal DNA in maternal plasma, have led to incorporation of noninvasive diagnosis of RHD genotype into routine prenatal care in the United Kingdom, France, and the Netherlands. In this commentary we examine the experience to date with large-scale clinical trials performed in the European Union, describe approaches to reduce false-positive and false-negative results, and review ongoing research to standardize assays and reduce costs using automated assays. False-negative cases are mainly due to either a lack of fetal DNA in the maternal sample due to early gestation or insensitive methods. False-positive cases are due to genotypic variants observed in individuals of African ancestry. Noninvasive prenatal diagnosis of fetal Rhesus D genotype is sensitive and accurate and has been widely validated in Europe. The United States should begin to undertake clinical trials to bring this technology to patient care as soon as possible.

Objectives: Trisomy 21 mosaicism associated with a structural rearrangement of chromosome 21 is uncommon. We report on two prenatal diagnoses in which karyotypes showed mosaicism with an aberrant cell line, including a structural rearrangement of chromosome 21., Methods: Both these cases were associated with increased nuchal translucency. Conventional and molecular cytogenetic analyses were performed on uncultured and cultured trophoblast and amniotic fluid cells., Results: In Case 1, analysis of trophoblast cells revealed an abnormal karyotype of 47,XX,+mar.ish der(13/21)(D13Z1/D21Z1+)/46,XX. The amniocentesis showed a free non-mosaic trisomy 21. In Case 2, the trophoblast direct analysis showed a normal male karyotype whereas the long-term culture revealed a mosaicism for a dicentric long-arm isochromosome 21: 46,XY,idic(21)(p11)/45,XY,-21/46,XY. Amniocentesis showed an unbalanced non-mosaic karyotype 46,XY,idic(21)(p11) resulting therefore in trisomy for the long arm of chromosome 21., Conclusion: Our cases underline the importance of combining the direct analysis and long-term culture of trophoblast and emphasise the need for confirmatory studies in other tissues when mosaicism of structural rearrangement is encountered in chorionic villi. The meiotic and mitotic mechanisms of formation of these structural rearrangements of chromosome 21 are discussed., (Copyright 2003 John Wiley & Sons, Ltd.)

Fetal RHD genotype determination is useful in the management of sensitized RhD-negative pregnant women. It can be ascertained early during pregnancy by chorionic villus sampling (CVS) or amniocentesis. However, these procedures are invasive, resulting both in an increased risk of fetal loss and in an increased severity of immunization due to fetomaternal haemorrhage. A reliable determination of RHD genotype by fetal DNA analysis in maternal serum during the first trimester of pregnancy is reported in this study. One hundred and six sera from RhD-negative pregnant women were obtained during the first trimester of pregnancy. These sera were tested for the presence of RHD gene using a new real-time polymerase chain reaction assay and the results compared with those obtained later in pregnancy on amniotic fluid cells and by RHD serology of the new-born. All sera from women carrying a RhD-positive fetus (n = 62) gave positive results for RHD gene detection and sera from women carrying a RhD-negative fetus (n = 40) were negative. The high level of accuracy of fetal RHD genotyping obtained in this study could enable this technique to be offered on a routine basis for the management of RhD-negative patients during the first trimester of pregnancy.

The purpose of this study was to determine the accuracy of the none-invasive prenatal determination of polymerase chain reaction (PCR)-based fetal RhD genotyping. Study design: A prospective case series was undertaken on all RhD-negative pregnant women presenting for genetic counseling in our prenatal diagnosis center from January 2001 until December 2002. Results were compared with serologic RhD typing of the newborns. Results: Among the 285 pregnant women who participated in the study, fetal RhD status could be determined for 283 patients. In 2 cases, the RhD-negative phenotype of the mother was not the result of a complete RHD gene deletion, and therefore, the status of the fetus could not be determined. Neither false-negative nor false-positive results were observed. Conclusion: The present report demonstrates that a reliable fetal RHD genotype determination can be achieved with 100% accuracy. It is therefore possible to consider that such an assay could be systematically proposed to all RhD-negative pregnant women in order to more effectively utilize RhD prophylaxis. [ABSTRACT FROM AUTHOR]

Fetal DNA and cells present in maternal blood have previously been used for non-invasive prenatal diagnosis. However, some fetal cells can persist in maternal blood after a previous pregnancy. Fetal rhesus status and sex determination have been performed by using amplification by real-time polymerase chain reaction (PCR) of fetal DNA sequences present in maternal circulation; no false-positive results related to persistent fetal DNA from a previous pregnancy have been reported. This idea has recently been challenged. An SRY real-time PCR assay was performed on the serum of 67 pregnant women carrying a female fetus but having previously given birth to at least one boy and on the serum of 30 healthy non-pregnant women with a past male pregnancy. In all cases, serum was negative for the SRY gene. These data suggest that fetal DNA from a previous pregnancy cannot be detected in maternal serum, even by using a highly sensitive technique. Therefore, non-invasive prenatal diagnosis by fetal sex determination for women at risk of producing children with X-linked disorders, and fetal RHD genotyping is reliable and secure as previously demonstrated. [ABSTRACT FROM AUTHOR]

The article focuses on a new concept for non-invasive prenatal diagnosis of aneuploidies by analysis of free fetal DNA circulating in maternal plasma. Free fetal DNA has been successfully used for the determination of fetal sex and rhesus D status on a routine basis by some specialized laboratories. Ravinder Dhallan and colleagues proposed the idea that single-nucleotide polymorphisms (SNPs) could be used to determine the number of fetal chromosomes in maternal blood. Though the approach is not new, the method that Dhallan proposes is. This approach can be used to detect trisomy 21, an aneuploidy that causes Down's syndrome. Details of clinical studies are presented.

Atypical fetal chromosomal anomalies are more frequent than previously recognized and can affect fetal development. We propose a screening strategy for a genome-wide non-invasive prenatal test (NIPT) to detect these atypical chromosomal anomalies (ACAs). Two sample cohorts were tested. Assay performances were determined using Cohort A, which consisted of 192 biobanked plasma samples—42 with ACAs, and 150 without. The rate of additional invasive diagnostic procedures was determined using Cohort B, which consisted of 3097 pregnant women referred for routine NIPT. Of the 192 samples in Cohort A, there were four initial test failures and six discordant calls; overall sensitivity was 88.1% (37/42; CI 75.00–94.81) and specificity was 99.3% (145/146; CI 96.22–99.88). In Cohort B, there were 90 first-pass failures (2.9%). The rate of positive results indicating an anomaly was 1.2% (36/3007) and 0.57% (17/3007) when limited to significant unbalanced chromosomal anomalies and trisomies 8, 9, 12, 14, 15, 16, and 22. These results show that genome-wide NIPT can screen for ACAs with an acceptable sensitivity and a small increase in invasive testing, particularly for women with increased risk following maternal serum screening and by limiting screening to structural anomalies and the most clinically meaningful trisomies. [ABSTRACT FROM AUTHOR]

We report a case of fetal hyperechogenic bowel diagnosed at midgestation that was associated with fetal parvovirus B19 infection. Isolated hyperechogenic bowel was detected at 25 weeks. Cystic fibrosis, chromosomal abnormalities and cytomegalovirus infection were excluded, whereas polymerase chain reaction DNA for parvovirus B 19 was found positive on amniotic fluid. The hyperechogenic bowel decreased with complete resolution by 32 weeks of gestation. No other signs of fetal B19 infection were detected prenatally and the baby had normal postnatal outcome. This case provides additional arguments in favor of a possible intestinal tropism of parvovirus B19 during fetal life. Fetal B19 infection should be systematically incorporated in the prenatal evaluation of isolated fetal hyperechogenic bowel. [ABSTRACT FROM AUTHOR]

OBJECTIVE: To evaluate the utility of noninvasive prenatal testing using cell-free circulating fetal DNA for detection of the three main autosomal fetal trisomies in the setting of ultrasonographically identified fetal anomalies. METHODS: Nine hundred patients at risk for fetal aneuploidy with or without ultrasonography anomalies and who underwent invasive procedures were included in the study. Cell-free DNA analysis was performed by massive parallel sequencing during a multicenter, noninterventional, prospective study and the results were compared with a fetal karyotype. RESULTS: Among all 900 pregnancies, cell-free DNA identified 76 of 76 (100%) fetal Down syndrome, 22 of 25 (88%) trisomy 18, and 12 of 12 (100%) trisomy 13. In those with a normal ultrasonogram and normal cell-free DNA analysis, karyotype identified 2 of 483 (0.4%) additional aneuploidies other than trisomies 13, 18, and 21. In those with an abnormal ultrasonogram and a normal cell-free DNA analysis, there were 23 of 290 (7.9%) additional pathogenic karyotypes. These additional aneuploidies included sex chromosome abnormalities and triploidy. The rates of additional aneuploidies not identifiable by standard cell-free DNA screening in the two groups is significantly different at P,.01. CONCLUSION: In women with fetal abnormalities by ultrasonography, the rate of pathogenic chromosome abnormalities missed by cell-free DNA was 8%. Noninvasive prenatal testing should not be offered to women with fetal abnormalities because a negative result is falsely reassuring. [ABSTRACT FROM AUTHOR]

Background: Congenital infection with Toxoplasma gondii can produce serious sequelae. However, there is little consensus about screening during pregnancy, and the tests used to establish a prenatal diagnosis of toxoplasmosis are complex and slow. We evaluated a simpler approach that is based on a polymerase-chain-reaction (PCR) test. Methods: Prenatal diagnostic tests, including ultrasonography, amniocentesis, and fetal-blood sampling, were performed in 2632 women with T. gondii infection acquired during pregnancy. In 339 consecutive women, a competitive PCR test for T. gondii was performed on amniotic fluid, and its results were compared with those of conventional diagnostic tests. The PCR test targets the B1 gene of T. gondii, uses an internal control, and can be completed in a day. Positive tests were confirmed by serologic testing of newborns or by autopsy in terminated pregnancies. Results: Overall, the risk of fetal infection was 7.4 percent, but it increased sharply with gestational age. Congenital infection was demonstrated in 34 of 339 fetuses by conventional methods, and the PCR test was positive in all 34. In three other fetuses, only the PCR test gave positive results, and follow-up testing confirmed the presence of congenital toxoplasmosis. The PCR test gave one false negative result but no false positive results. The PCR test performed better than conventional parasitologic methods (sensitivity, 97.4 percent vs. 89.5 percent; negative predictive value, 99.7 percent vs. 98.7 percent). Conclusions: For the prenatal diagnosis of congenital T. gondii infection, an approach based on a PCR test performed on amniotic fluid is rapid, safe, and accurate. (N Engl J Med 1994;331:695-9.) [ABSTRACT FROM AUTHOR]