Acceptability of Prenatal Screening Tests Among Expectant Mothers in India: Insights and Implications for Public Health.

INTRODUCTION: Prenatal screening tests are essential for preventing common genetic disorders, yet their acceptability among pregnant women in India remains unexplored. This study aims to investigate the acceptability of prenatal screening tests and their correlation with demographic characteristics among pregnant women in India. METHODS: A cross-sectional study was conducted at a tertiary care, public hospital, involving 200 pregnant women. Data were collected through a self-administered questionnaire assessing demographic information and the acceptability of prenatal screening tests. Statistical analysis included chi-square tests and logistic regression. RESULTS: Most participants demonstrated adequate acceptability toward prenatal screening tests, with 73% scoring above the threshold. Factors associated with higher acceptability included younger maternal age, second-trimester gestational age, higher education, salaried employment, and urban residence. However, factors such as parity, consanguinity, mode of conception, and family history of genetic disease showed no significant associations. CONCLUSION: The study highlights positive attitudes toward prenatal screening tests among pregnant women in India, particularly among younger, more educated, and urban populations. These findings emphasize the need for targeted interventions to enhance awareness and accessibility of prenatal screening, ultimately contributing to the reduction of the genetic disorder burden in India.

Non-invasive prenatal test identifies circulating cell-free DNA chromosomal abnormalities derived from clonal hematopoiesis in aggressive hematological malignancies.

Liquid biopsy is a minimally invasive diagnostic tool for identification of tumor-related mutations in circulating cell-free DNA (cfDNA). The aim of this study was to investigate feasibility, sensitivity, and specificity of non-invasive prenatal test (NIPT) for identification of chromosomal abnormalities in cfDNA from a total of 77 consecutive patients with non-Hodgkin B-cell lymphomas, Hodgkin lymphoma (HL), or plasma cell dyscrasia. In this case series, half of patients had at least one alteration, more frequently in chromosome 6 (23.1%), chromosome 9 (20.5%), and chromosomes 3 and 18 (16.7%), with losses of chromosome 6 and gains of chromosome 7 negatively impacting on overall survival (OS), with a 5-year OS of 26.9% and a median OS of 14.6 months, respectively (P = 0.0009 and P = 0.0004). Moreover, B-cell lymphomas had the highest NIPT positivity, especially those with aggressive lymphomas, while patients with plasma cell dyscrasia with extramedullary disease had a higher NIPT positivity compared to conventional cytogenetics analysis and a worse outcome. Therefore, we proposed a NIPT-based liquid biopsy a complementary minimally invasive tool for chromosomal abnormality detection in hematological malignancies. However, prospective studies on larger cohorts are needed to validate clinical utility of NIPT-based liquid biopsy in routinely clinical practice.

Understanding the Awareness of Prenatal Genetic Screening Tests Among Pregnant Women in India: A Cross-Sectional Study.

Introduction Genetic disorders pose a significant health challenge in India, with chromosomal abnormalities ranking second only to congenital anomalies in terms of disease burden. Prenatal testing offers a crucial strategy for identifying and managing these disorders. However, the awareness and understanding of prenatal screening tests among pregnant women in India remain understudied. This study aims to fill this gap by investigating the awareness quotient of prenatal screening tests for genetic disorders among pregnant women in India. Methods A hospital-based cross-sectional study was conducted at the Genetics Unit, Department of Anatomy, and Department of Obstetrics and Gynecology, All India Institute of Medical Sciences, Mangalagiri. Ethical clearance was obtained, and data were collected using a self-administered questionnaire covering demographic characteristics and awareness assessment. Descriptive statistics, chi-square tests, and logistic regression analysis were employed for data analysis. Results Among the 200 pregnant women surveyed, the majority demonstrated inadequate awareness of prenatal screening tests for genetic disorders, with only 36.5% possessing adequate knowledge. Significant associations were found between awareness levels and factors such as age, trimester of pregnancy, and education level. Notably, awareness about non-invasive prenatal testing (NIPT) was notably low at 7%, indicating a need for targeted educational interventions. Comparison with international studies revealed varying levels of awareness across different populations, highlighting the influence of socio-cultural factors and healthcare systems. Conclusion This study underscores the need for improved awareness of prenatal screening tests among pregnant women in India. Addressing disparities in awareness, particularly among younger age groups and those with lower education levels, is crucial for informed decision-making in prenatal care. Targeted educational interventions can empower pregnant women to make informed choices, ultimately contributing to better maternal and child health outcomes. Further research should explore the effectiveness of such interventions in diverse settings to enhance prenatal care delivery.

Association between fetal fraction of cell-free DNA and adverse pregnancy outcomes.

PURPOSE: To determine the association between fetal fraction (FF) levels in cell-free fetal DNA (cffDNA) testing and adverse pregnancy outcomes. METHODS: This retrospective cohort study, conducted at a single center, involved 2063 pregnant women with normal 1st and 2nd trimester non-invasive prenatal test (NIPT) results between 2016 and 2021. Pregnancy outcomes were examined by determining the < 4% and < 5th percentile (3.6%) cut-off values for low fetal fraction (LFF). Pregnancy outcomes were also examined by dividing the FF into population-based quartiles. Adverse pregnancy outcomes were pregnancy-induced hypertensive diseases (PIHD), gestational diabetes mellitus (GDM), spontaneous preterm birth (PTB), intrahepatic cholestasis of pregnancy (ICP), small for gestational age (SGA), large for gestational age (LGA), low birth weight (LBW), macrosomia, and 1st and 5th minutes low APGAR scores (< 7). RESULTS: PIHD was significantly higher in LFF (< 4% and < 5th percentile) cases (p = 0.015 and p < 0.001, respectively). However, in population-based quartiles of FF, PIHD did not differ significantly between groups. Composite adverse maternal outcomes were significantly higher in the FF < 4% group (p = 0.042). When analyzes were adjusted for maternal age, BMI, and gestational age at NIPT, significance was maintained at < 4%, < 5th percentile LFF for PIHD, and < 4% LFF for composite adverse maternal outcomes. However, there was no significant relationship between LFF with GDM, ICP and PTB. Additionally, there was no significant association between low APGAR scores, SGA, LGA, LBW, macrosomia, and LFF concerning neonatal outcomes. CONCLUSION: Our study showed that LFF in pregnant women with normal NIPT results may be a predictor of subsequent PIHD.

Adverse pregnancy outcome in fetuses with early increased nuchal translucency: prospective cohort study.

OBJECTIVES: An increased nuchal translucency (NT) thickness of ≥ 3.5 mm is a well-established marker for congenital anomalies and adverse pregnancy outcome between 11 and 14 weeks' gestation, but little is known about its performance as a screening tool before 11 weeks. We aimed to investigate, in a prospective setting, whether fetuses with increased NT before 11 weeks are at risk for adverse pregnancy outcome. METHODS: This was a prospective cohort study including pregnant women with a viable fetus with NT ≥ 2.5 mm and a crown-rump length (CRL) < 45 mm. All included women were referred to our fetal medicine unit (FMU) and scheduled for a follow-up scan where the NT was remeasured after 1 week when the CRL was > 45 mm. Two groups were evaluated: cases with a normalized NT (< 3.5 mm) and cases with persistently increased NT (≥ 3.5 mm). The cases were monitored until 4 weeks after delivery. The main outcome was a composite adverse outcome of aneuploidy, other genetic disorders, structural anomalies and pregnancy loss. We performed subgroup analyses of NT thickness at inclusion and normalized or persistently increased NT at follow-up. RESULTS: The study included 109 cases, of which 39 (35.8%) had an adverse pregnancy outcome. Of these, 64.1% (25/39) were aneuploid, corresponding to 22.9% (25/109) of the total study population. In the subgroups of NT thickness at inclusion of 2.5-3.4 mm, 3.5-4.4 mm and ≥ 4.5 mm, an adverse outcome was reported in 22.0% (9/41), 40.0% (18/45) and 52.2% (12/23), respectively. In fetuses with a normalized NT and without ultrasound abnormalities at the follow-up scan, the incidence of adverse outcome was 8.5% (5/59), of which 5.1% (3/59) cases were aneuploid. CONCLUSIONS: Fetuses with an early increased NT thickness are at considerable risk of an adverse pregnancy outcome, even if the NT normalizes after 11 weeks. Not all congenital anomalies can be diagnosed with routine first-trimester screening, such as non-invasive prenatal testing and/or a first-trimester anomaly scan. Therefore, expectant parents should always be referred to a FMU for detailed ultrasonography. Invasive prenatal testing should be offered if an increased NT of ≥ 2.5 mm is observed before 11 weeks' gestation. © 2024 The Authors. Ultrasound in Obstetrics & Gynecology published by John Wiley & Sons Ltd on behalf of International Society of Ultrasound in Obstetrics and Gynecology.

Resultado adverso del embarazo en fetos con aumento precoz de la translucencia nucal: estudio prospectivo de cohortes OBJETIVOS: El aumento del grosor de la translucencia nucal (TN) de ≥3,5 mm es un marcador bien establecido de anomalías congénitas y resultados adversos del embarazo entre las semanas 11 y 14 de gestación, pero se sabe poco sobre su rendimiento como herramienta de cribado antes de las 11 semanas. El objetivo fue investigar, en un contexto prospectivo, si los fetos con aumento de la TN antes de las 11 semanas corren riesgo de presentar resultados adversos del embarazo. MÉTODOS: Se trató de un estudio prospectivo de cohortes que incluyó a embarazadas con un feto viable con una TN ≥2,5 mm y una longitud céfalo­caudal (LCC) <45 mm. Todas las mujeres incluidas fueron remitidas a una unidad de medicina fetal (UMF) y con cita para una prueba de seguimiento en la que se volvió a medir la TN al cabo de 1 semana cuando la LCC era >45 mm. Se evaluaron dos grupos: casos con una TN normalizada (<3.5 mm) y casos con una TN persistentemente aumentada (≥3,5 mm). A los casos se les dio seguimiento hasta 4 semanas después del parto. El resultado principal fue un resultado adverso compuesto de aneuploidía, otros trastornos genéticos, anomalías estructurales y pérdida del embarazo. Se realizaron análisis de subgrupos del grosor de la TN en el momento de la inclusión y de la TN normalizada o persistentemente aumentada en el seguimiento. RESULTADOS: El estudio incluyó 109 casos, de los cuales 39 (35,8%) tuvieron un resultado adverso del embarazo. De ellos, el 64,1% (25/39) eran aneuploides, lo que supone el 22,9% (25/109) de la población total del estudio. En los subgrupos de grosor de la TN en el momento de la inclusión de 2,5­3,4 mm, 3,5­4,4 mm y ≥4,5 mm, se notificó un resultado adverso en el 22,0% (9/41), el 40,0% (18/45) y el 52,2% (12/23), respectivamente. En los fetos con una TN normalizada y sin anomalías ecográficas en la ecografía de seguimiento, la incidencia de resultados adversos fue del 8,5% (5/59), de los cuales el 5,1% (3/59) de los casos eran aneuploides. CONCLUSIONES: Los fetos con un aumento precoz del grosor de la TN corren un riesgo considerable de sufrir un resultado adverso del embarazo, incluso si la TN se normaliza después de 11 semanas. No todas las anomalías congénitas pueden diagnosticarse con un cribado rutinario en el primer trimestre, como las pruebas prenatales no invasivas y/o una ecografía de anomalías en el primer trimestre. Por lo tanto, los futuros padres siempre deben ser remitidos a una UMF para una ecografía detallada. Se debería ofrecer una prueba prenatal invasiva si se observa un aumento de la TN de ≥2,5 mm antes de las 11 semanas de gestación.

Cell-Free Fetal DNA Screening Analysis in Korean Pregnant Women: Six Years of Experience and a Retrospective Study of 9327 Patients Analyzed from 2017 to 2022.

Cell-free DNA (cfDNA) screening for normal fetal aneuploidy has been widely adopted worldwide due to its convenience, non-invasiveness, and high positive predictive rate. We retrospectively evaluated 9327 Korean women with single pregnancies who underwent a non-invasive prenatal test (NIPT) to investigate how various factors such as maternal weight, age, and the method of conception affect the fetal fraction (FF). The average FF was 9.15 ± 3.31%, which decreased significantly as the maternal body mass index (BMI) increased (p < 0.001). The highly obese group showed a 'no-call' rate of 8.01%, which is higher than that of the normal weight group (0.33%). The FF was 8.74 ± 3.20% when mothers were in their 40s, and lower than that when in their 30s (9.23 ± 3.34, p < 0.001) and in the natural pregnancy group (9.31% ± 3.33). The FF of male fetuses was observed to be approximately 2.76% higher on average than that of female fetuses. As the gestational age increased, there was no significant increase in the fraction of fetuses up to 21 weeks compared to that at 10-12 weeks, and a significant increase was observed in the case of 21 weeks or more. The FFs in the NIPT high-risk result group compared to that in the low-risk group were not significantly different (p = 0.62). In conclusion, BMI was the factor most associated with the fetal fraction. Although the NIPT is a highly prevalent method in prenatal analysis, factors affecting the fetal fraction should be thoroughly analyzed to obtain more accurate results.

The effectiveness of non-invasive prenatal test technology and the prenatal screening algorithm based on various methods for determining foetal aneuploidy.

Objective: The purpose was to evaluate the effectiveness of a non-invasive prenatal test (NIPT) using mass parallel sequencing (MPS) to detect trisomy 13, 18, 21 and fetal sex chromosome abnormalities in maternal blood samples by isolating freely circulating foetal extracellular DNA (eDNA), and to develop an algorithm for prenatal screening. Material and Methods: The research methods used included blood sampling from patients, isolation of eDNA, determination of DNA concentration and quality, library preparation for sequencing, MPS using an Illumina HiSeq2000, positive and negative control samples, monitoring, and analysis of results using the distributed algorithms platform based on calculations of z-value and the average absolute deviation. Pregnant women were divided into two groups based on gestational age at sampling, group 1; 9-14 weeks and group 2; 15-27 weeks. Results: A total of 377 pregnant women were included with a mean (range) age of 33 (23-44) years. The mean gestational age at the time of blood sampling in group 1 was 11 (9-14) weeks, and in group 2 was 21 (15-27) weeks. In the first group, three cases of trisomy 18 chromosomes were detected in patients aged 43 years old, and female children were subsequently born with Edwards syndrome. In the second group, one case of trisomy 21 was detected in a patient aged 36 years and the pregnancy was terminated at 25 weeks. Conclusion: The analysis of freely circulating foetal eDNA was a sensitive method for detecting chromosomal abnormalities. The study has a practical significance, since the NIPT for frequent aneuploidy considerably exceeds the effectiveness of traditional screening methods and allows identifying chromosomal disorders starting from the 9th week of the gestation period.

Comparing Non-invasive Prenatal Testing With Invasive Testing for the Detection of Trisomy 21.

Background Non-invasive prenatal test (NIPT) is an intermediate step between serum screening and invasive diagnostic testing. It involves analysis of the cell-free fetal DNA (cffDNA) present in the maternal blood sample for determining the likelihood of fetal aneuploidy. Owing to its high sensitivity and specificity, NIPT has quickly gained popularity across the globe since its introduction to clinical practice, making it an attractive alternative to the available screening and diagnostic tests in use. Amniocentesis is currently the gold standard test for obtaining fetal DNA and diagnosing fetal trisomy prenatally, but it is invasive and has procedure-related adverse effects. This study aims to compare NIPT and amniocentesis in pregnancies screened positive for fetal trisomy. Material and methods This is an analytic cross-sectional prospective study conducted in the Department of Obstetrics & Gynecology, Patna Medical College and Hospital, for two and half years from December 2018 to June 2021. A total of 34 pregnant women screened positive for trisomy 21, attending the antenatal care outpatient department, in their second trimester, with their written consent, were enrolled in the study. Results Out of 34 pregnant patients, three refused NIPT and directly opted for amniocentesis. A total of 31 pregnant women have undergone NIPT. A total of 28 cases were positive for trisomy 21 on both NIPT and amniocentesis. The sensitivity of NIPT was 100% with the confidence interval being 87.66% to 100.00%. The specificity of NIPT was 100% with the confidence interval being 29.24% to 100.00%. Conclusion The high performance and effectiveness of NIPT are undeniable. Though the process by which this test has to be integrated into the clinical practice needs more study and should be determined with meticulous assessment.

A dPCR-NIPT assay for detections of trisomies 21, 18 and 13 in a single-tube reaction-could it replace serum biochemical tests as a primary maternal plasma screening tool?

BACKGROUND: The next generation sequencing (NGS) based non-invasive prenatal test (NIPT) has outplayed the traditional serum biochemical tests (SBT) in screen of fetal aneuploidies with a high sensitivity and specificity. However, it has not been widely used as a primary screen tool due to its high cost and the cheaper SBT is still the choice for primary screen even with well-known shortages in sensitivity and specificity. Here, we report a multiplex droplet digital PCR NIPT (dPCR-NIPT) assay that can detect trisomies 21, 18 and 13 (T21, T18 and T13) in a single tube reaction with a better sensitivity and specificity than the SBT and a much cheaper price than the NGS-NIPT. METHODS: In this study, the dPCR-NIPT assay's non-clinical characteristics were evaluated to verify the cell free fetal DNA (cffDNA) fraction enrichment efficiencies, the target cell free DNA (cfDNA) concentration enrichment, the analytical sensitivity, and the sample quality control on the minimum concentration of cfDNA required for the assay. We validated the clinical performance for this assay by blindly testing 283 clinical maternal plasma samples, including 36 trisomic positive samples, from high risk pregnancies to access its sensitivity and specificity. The cost effectiveness of using the dPCR-NIPT assay as the primary screen tool was also analyzed and compared to that of the existing contingent strategy (CS) using the SBT as the primary screen tool and the strategy of NGS-NIPT as the first-tier screen tool in a simulating situation. RESULTS: For the non-clinical characteristics, the sample processing reagents could enrich the cffDNA fraction by around 2 folds, and the analytical sensitivity showed that the assay was able to detect trisomies at a cffDNA fraction as low as 5% and the extracted cfDNA concentration as low as 0.2 ng/µL. By testing the 283 clinical samples, the dPCR-NIPT assay demonstrated a detection sensitivity of 100% and a specificity of 95.12%. Compared to the existing CS and the NGS-NIPT as the first-tier screen strategy, dPCR-NIPT assay used as a primary screen tool followed by the NGS-NIPT rescreen is the most economical approach to screen pregnant women for fetal aneuploidies without sacrificing the positive detection rate. CONCLUSION: This is the first report on a dPCR-NIPT assay, consisting of all the necessary reagents from sample processing to multiplex dPCR amplification, can detect T21, T18 and T13 in a single tube reaction. The study results reveal that this assay has a sensitivity and specificity superior to the SBT and a cost much lower than the NGS-NIPT. Thus, from both the test performance and the economic benefit points of views, using the dPCR-NIPT assay to replace the SBT as a primary screen tool followed by the NGS-NIPT rescreen would be a better approach than the existing CS for detection of fetal aneuploidies in maternal plasma.

A study on non-invasive prenatal screening for the detection of aneuploidy.

OBJECTIVES: To explore the feasibility of clinical application of non-invasive prenatal screening to detect aneuploidy diseases. MATERIAL AND METHODS: A total of 14,574 singleton pregnant women who underwent Non-invasive prenatal testing (NIPT) in the Southern Hospital from 2015 to June 2017 were selected, and 6471 pregnant women with twin pregnancy who underwent NIPT in the laboratory of Bei Rui He Kang Southern Hospital from June 2016 to October 2017 were included in this study. We analyzed NIPT screening efficiency (sensitivity, specificity) in twin pregnancies and singleton pregnancies, compared the positive detection rate of NIPT in patients with or without clinical symptoms. All NIPT high-risk results were validated by karyotyping, which were further verified by the follow-up physical examination of the neonatal. RESULTS: A total of 68 cases of twin pregnancy abnormalities were detected by NIPT, including 18 cases of trisomy 21, 6 cases of trisomy 18, 1 case of trisomy 13, 39 cases of Spinocerebellar ataxias (SCAs), and 4 cases of other chromosomal abnormalities. The sensitivity for trisomy 21, 18, and 13 and sex chromosome abnormality was 100%; the specificity for trisomy 21, 18, and 13 and sex chromosome abnormality was 99.97%, 99.95%, 99.97%, and 99.91% respectively. The screening efficiency was similar to that of singleton pregnancy, indicating that the NIPT technology in our laboratory for screening for aneuploidy diseases in twin pregnancy has reached the accuracy level of singleton pregnancy screening. There was a statistical difference between the risk group and the non-risk group in pregnant women with singleton pregnancy. The screening efficiency of NIPT was higher in pregnant women in the risk group, which implies that the clinical application of NIPT is inclined to detect high-risk group. CONCLUSIONS: Non-invasive prenatal testing (NIPT) is a rapid and safe screening method with high efficiency. Non-invasive prenatal testing (NIPT) is used for the screening of aneuploidy in twin pregnancy. The efficiency is similar to that of singleton pregnancy, indicating the feasibility of clinical application. However, the efficiency of NIPT screening tends to favor the detection in high-risk groups.

The impact of prenatal screening tests on prenatal diagnosis in Taiwan from 2006 to 2019: a regional cohort study.

BACKGROUND: The purpose of this study is to evaluate the impact of prenatal screening tests on prenatal diagnosis in Taiwan's 14 years from 2006 to 2019. METHODS: The prenatal screening methods evolved from the second-trimester serum screening to combined first-trimester screening (cFTS) and then followed by the non-invasive cell-free DNA prenatal test (NIPT). The data used by the Department of Statistics, the Ministry of Health and Welfare and Department of Household Registration, Ministry of the Interior public website. RESULTS: This regional registry-based cohort retrospective study examined a total of 2,775,792 births from January 2006 to December 2019. The proportion of advanced maternal age (AMA) pregnancies increased from 11.63% in 2006 to 30.94% in 2019. Overall, invasive diagnostic testing was used in 87.22% of AMA pregnancies. The prenatal detection rate of trisomy 21 and 18 increased from 74.1% and 83.3% in 2006 to 96.9% and 98.8% in 2019, respectively. CONCLUSION: During the second-trimester and cFTS periods, the percentage of AMA pregnancies increased every year and the number of invasive procedures also accompany with increased percentage of AMA. However, during the period that NIPT were implemented, the percentage of invasive procedures decreased.

Droplet-based digital PCR for non-invasive prenatal genetic diagnosis of α and ß-thalassemia.

Non-invasive prenatal diagnosis (NIPD) of isolated cell-free DNA from maternal plasma has been applied to detect monogenic diseases in the fetus. Droplet digital PCR (ddPCR) is a sensitive and quantitative technique for NIPD. In the present study, the development and evaluation of ddPCR-based assays for common α and ß-thalassemia variants amongst the Asian population was described; specifically, Southeast Asian (SEA) deletion, HbE, and 41/42 (-CTTT). SEA is caused by deletion of a 20 kb region surrounding the α-globin gene, whilst HbE and 41/42 (-CTTT) are caused by point mutations on the ß-globin gene. Cell-free DNA samples from 46 singleton pregnant women who were carriers of these mutations were isolated and quantified using ddPCR with specially designed probes for each target allele. Allelic copy number calculation and likelihood ratio tests were used to classify fetal genotypes. Classification performances were evaluated against ground truth fetal genotypes obtained from conventional amniocentesis. Copy number variation analysis of SEA deletion accurately classified fetal genotypes in 20 out of 22 cases with an area under the receiver operating characteristic curve of 0.98 for detecting Hb Bart's hydrops fetalis. For HbE cases, 10 out of 16 samples were correctly classified, and three were inconclusive. For 41/42 (-CTTT) cases, 2 out of 8 were correctly classified, and four were inconclusive. The correct genotype was not rejected in any inconclusive case and may be resolved with additional ddPCR experiments. These results indicate that ddPCR-based analysis of maternal plasma can become an accurate and effective NIPD for SEA deletion α-(0) thalassemia. Although the performance of ddPCR on HbE and 41/42 (-CTTT) mutations were not sufficient for clinical application, these results may serve as a foundation for future works in this field.

Novel method of real-time PCR-based screening for common fetal trisomies.

BACKGROUND: The non-invasive prenatal test (NIPT) is based on next generation sequencing (NGS) and is used for screening for fetal trisomy. However, it is time-consuming and technically difficult. Recently, peptide nucleic acid (PNA) probe-based real-time polymerase chain reaction (RT-PCR) was developed. This study aimed to examine the performance of the RT-PCR-based NIPT for screening of common fetal trisomies METHODS: From stored maternal plasma, RT-PCR was performed using Patio™ NIPT Detection Kit. In melting curve analysis, the height of melting peaks of target chromosome and reference chromosome was calculated as a peak ratio. The adjusted peak ratio of 8 markers with correction factors in each target chromosome was summated and calculated to z-score. The cut-off value for each target chromosome was established for classification (low risk vs. high risk for trisomy) whose performance was obtained in the validation phase. RESULTS: 330 plasma samples from pregnant women with normal fetus and 22 trisomy cell-line samples were used to establish the optimal cut-off values for z-score of each target chromosome. In the validation phase, 1023 samples from pregnant women including 22 cases with fetal trisomy and 1001 cases of normal control were used. The RT-PCR-based NIPT showed 95.45% sensitivity [95% confidence interval (CI) 77.16-99.88%], 98.60% specificity (95% CI 97.66-99.23%), and 98.53% accuracy (95% CI 97.59-99.18%) for the identification of trisomy 21, 18, or 13. Of 1023 samples, fifteen cases were mismatched for classification [one case as a false negative (false negative rate: 4.5%) and 14 cases as false positives (false positive rate: 1.4%)]. CONCLUSION: The RT-PCR-based NIPT showed high sensitivity and specificity for the detection of common fetal trisomies and it could be a feasible alternative to NGS-based NIPT.

First Case Report of Maternal Mosaic Tetrasomy 9p Incidentally Detected on Non-Invasive Prenatal Testing.

Tetrasomy 9p (ORPHA:3390) is a rare syndrome, hallmarked by growth retardation; psychomotor delay; mild to moderate intellectual disability; and a spectrum of skeletal, cardiac, renal and urogenital defects. Here we present a Chinese female with good past health who conceived her pregnancy naturally. Non-invasive prenatal testing (NIPT) showed multiple chromosomal aberrations were consistently detected in two sampling times, which included elevation in DNA from chromosome 9p. Amniocentesis was performed and sent for chromosomal microarray, which was normal. Maternal karyotype revealed that mos 47,XX,+dic(9;9)(q21.1;q21.1)(24)/46,XX(9) presents mosaic tetrasomy for the short arm of chromosome 9p and is related to the NIPT results showing elevation in DNA from chromosome 9p. The pregnancy was uneventful, and the patient was delivered at term. Maternal samples were obtained at two different time points after delivery showed the same multiple chromosomal aberrations detected during pregnancy. This is a first report on an unusual case of mosaic isodicentric tetrasomy 9p in a healthy adult with normal intellect. With widespread adoption of NIPT for screening fetal aneuploidy and genome-wide copy number changes, a rise in incidental detection of maternal rare genetic syndrome will bring challenges in our current approach to genetic counselling and prenatal diagnosis.

A placental trisomy 2 detected by NIPT evolved in a fetal small Supernumerary Marker Chromosome (sSMC).

BACKGROUND: Non-invasive prenatal testing (NIPT) is a rapidly developing and widely used method in the prenatal screening. Recently, the widespread use of the NIPT caused a neglecting of the limitations of this technology. CASE PRESENTATION: The 38-year-old woman underwent amniocentesis because of a high risk of trisomy 2 revealed by the genome-wide Non-Invasive Prenatal Test (NIPT). The invasive prenatal diagnosis revealed the mosaicism for a small supernumerary marker chromosome sSMC derived from chromosome 2. Interphase fluorescence in situ hybridization (FISH) on uncultured amniocytes revealed three signals of centromere 2 in 30% of the cells. GTG-banded metaphases revealed abnormal karyotype (47,XX,+mar[21]/46,XX[19]) and was confirmed by array comparative genomic hybridization (aCGH). Cytogenetic analyses (FISH, aCGH, karyotype) on fetal skin biopsies were performed and confirmed the genomic gain of the centromeric region of chromosome 2. In the placenta, three cell lines were detected: a normal cell line, a cell line with trisomy 2 and a third one with only the sSMC. CONCLUSION: Whole-genome Non-Invasive Prenatal Testing allows not only the identification of common fetal trisomies but also diagnosis of rare chromosomal abnormalities. Especially in such cases, it is extremely important to perform not only NIPT verification on a sample of material other than trophoblast, but also to apply appropriate research methods. Such conduct allows detailed analysis of the detected aberration, thus appropriate clinical validity.

Uptake of fetal aneuploidy screening after the introduction of the non-invasive prenatal test: A national population-based register study.

INTRODUCTION: The introduction of the non-invasive prenatal test (NIPT) has shifted the prenatal screening landscape. Countries are exploring ways to integrate NIPT in their national prenatal screening programs, either as a first- or second-tier test. This study aimed to describe how the uptake of fetal aneuploidy screening changed after the introduction of NIPT as a second-tier and as a first-tier test within the national prenatal screening program of the Netherlands. MATERIAL AND METHODS: A population-based register study in the Netherlands, recording uptake of fetal aneuploidy screening. Data from all pregnant women choosing to have the first-trimester combined test (FCT) or first-tier NIPT between January 2007 and March 2019 were retrospectively collected using national registration systems. Uptake percentages for fetal aneuploidy screening (FCT and NIPT) were calculated and stratified by region and maternal age. Statistical significance was determined using trend analysis and chi-squared tests. RESULTS: Between 2007 and 2013 FCT uptake increased from 14.8% to 29.5% (P = .004). In April 2014 NIPT was introduced as a second-tier test for high-risk women after FCT (TRIDENT-1 study). FCT uptake rose from 29.5% in 2013 to 34.2% in 2015 (P < .0001). After the introduction of NIPT as a first-tier test for all women in April 2017 (TRIDENT-2 study), FCT uptake declined significantly from 35.8% in 2016 to 2.6% in 2018 (P < .0001). NIPT uptake increased to 43.4% in 2018. Regionally, NIPT uptake ranged from 31.8% to 67.9%. Total uptake (FCT and NIPT) between 2007 and 2018 increased significantly from 14.8% to 45.9% (P < .0001). However, total uptake stabilized at 46% for both years of TRIDENT-2 (April 2017-March 2019). CONCLUSIONS: An increase in total fetal aneuploidy screening uptake up to 45.9% was observed after the introduction of NIPT. Uptake appears to have stabilized within a year after introducing first-tier NIPT.

Clinical management of pregnancies with positive screening results for rare autosomal aneuploidies at a single center.

OBJECTIVE: To review our experiences on clinical management of pregnancies with positive noninvasive prenatal testing (NIPT) results for rare autosomal aneuploidies (RAAs) at a single center. METHODS: We performed a retrospective study and reviewed data from 18,016 pregnancies undergoing NIPT at a single center in China from March 2017 to February 2020. Depending on the patient's choice, women with positive screening results for RAAs underwent chromosomal microarray analysis for invasive prenatal diagnosis. RESULTS: Thirty-three positive cases for RAAs were identified, with a positive screening rate of 0.18%. The most common RAA was trisomy 7 (33.3%), while trisomies for other chromosomes were less frequent. Monosomies involving chromosomes 16, 14, and 22 were observed. Twenty-eight cases of RAAs underwent invasive diagnosis. Abnormal pregnancy outcomes were observed in four cases, including true fetal mosaicism (n=1), partial uniparental disomy (n=1), miscarriage (n=1), and structural anomalies on ultrasound (n=1). CONCLUSIONS: RAAs at NIPT might be associated with fetal uniparental disomy, mosaic aneuploidy, and poor pregnancy outcomes, but most positive cases have normal pregnancy outcomes. For RAAs, genetic counseling on the potential risks of abnormal NIPT results, as well as on benefits and limitations of invasive prenatal diagnosis, might help guide clinical management.

Association between low fetal fraction of cell free DNA at the early second-trimester and adverse pregnancy outcomes.

OBJECTIVE: The purpose of this study was to examine whether low fetal fraction (FF) of cell free DNA is associated with risks of adverse pregnancy outcomes. METHODS: This was a historical cohort study of 2191 women with singleton pregnancies who had non-invasive prenatal test (NIPT) at 13 to 26 weeks of gestation. Data were collected from prenatal screening system and hospital records. Main outcome was the subsequent diagnosis of gestational diabetes mellitus (GDM), intrahepatic cholestasis of pregnancy (ICP), preeclampsia (PE), pregnancy induced hypertension (PIH) and preterm birth (PTB). Logistic regression analysis was performed to evaluate the association between LFF and adverse pregnancy outcomes. RESULTS: The prevalence of GDM, ICP, PE, PIH and PTB was 23.87% (523), 4.02% (88), 2.92% (64), 2.83% (62) and 6.85% (150), respectively. Low FF, defined as less than the 10th percentile, was associated with an increased risk of PE (adjusted OR = 2.06, 95% CI: 1.07-3.98) and early PTB (<34 weeks' gestation: adjusted OR = 3.09, 95% CI: 1.21-7.92). In addition, low FF, defined as less than the 5th percentile, was associated with an increased risk of low birth weight babies (<2500 g: adjusted OR = 2.50, 95% CI: 1.01-6.17). However, there was no significant association between low FF and GDM, as well as ICP and PIH. CONCLUSION: Our study provides evidence that low FF is associated with PE and early PTB. Further exploration of the clinical significance of low FF is warranted.

Strategy for Use of Genome-Wide Non-Invasive Prenatal Testing for Rare Autosomal Aneuploidies and Unbalanced Structural Chromosomal Anomalies.

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.

Rethinking counselling in prenatal screening: An ethical analysis of informed consent in the context of non-invasive prenatal testing (NIPT).

Informed consent is a key condition for prenatal screening programmes to reach their aim of promoting reproductive autonomy. Reaching this aim is currently being challenged with the introduction of non-invasive prenatal testing (NIPT) in first-trimester prenatal screening programmes: amongst others its procedural ease-it only requires a blood draw and reaches high levels of reliability-might hinder women's understanding that they should make a personal, informed decision about screening. We offer arguments for a renewed recognition and use of informed consent compared to informed choice, and for a focus on value-consistent choices and personalized informational preferences. We argue for a three-step counselling model in which three decision moments are distinguished and differently addressed: (1) professionals explore women's values concerning whether and why they wish to know whether their baby has a genetic disorder; (2) women receive layered medical-technical information and are asked to make a decision about screening; (3) during post-test counselling, women are supported in decision-making about the continuation or termination of their pregnancy. This model might also be applicable in other fields of genetic (pre-test) counselling, where techniques for expanding genome analysis and burdensome test-outcomes challenge counselling of patients.