Fetal Chromosome Research Articles

Trends of fetal chromosome analysis by amniocentesis before and after beginning of noninvasive prenatal testing: A single-center experience in Japan.

This study is to investigate the role of amniocentesis for prenatal diagnosis before and after the beginning of noninvasive prenatal testing (NIPT) in Japan. We performed a retrospective analysis of genetic amniocentesis at mid-trimester (15-20 gestational weeks) for fetal karyotype analysis at Nagoya City University between April 2006 and March 2020. The indications, test results, and the detection rate of fetal abnormal karyotype were compared before (phase 1, P1) and after (phase 2, P2) beginning of NIPT at April 2013. A total of 2458 (P1: 1132, P2: 1326) amniocentesis were enrolled in this study. The most frequent indication was advanced maternal age in both phases (P1: 78.2% %, P2: 81.1%). In P2, 110 patients (8.3%) received amniocentesis after positive or nonreportable NIPT results. Other indications were fetal abnormal findings by ultrasounds (P1: 15.4%, P2: 17.7%), abnormal maternal serum screening results (P1: 8.0%, P2: 10%), previous child with fetal chromosome aberration (P1: 6.5%, P2: 3.5%), and translocation of either partner (P1:1.5%, P2: 2.1%). The detection rate for fetal chromosomal aberrations including all indications was significantly increased in P2 (15.9%, 95% CI 14.0-18.0) as compared to P1 (9.0%, 7.4-10.8). However, if the indication was only advanced maternal age, the positive detection rate kept low in both phases (P1: 5.2%, 3.7-7.1, P2: 4.2%, 2.9-5.9). Since the initiation of NIPT, the detection rate of fetal chromosomal abnormalities was higher in this study, suggesting that amniocentesis cannot be strongly recommended for advanced maternal age alone.

P–515 Clinical outcome of mosaic-blastocyst transfer versus euploid-blastocyst transfer in single frozen blastocyst transfer cycles

Abstract Study question What is the clinical outcome of transferring a mosaic blastocyst versus a euploid blastocyst in single frozen blastocyst transfer (sFBT) cycles? Summary answer Single mosaic blastocyst transfer has similar clinical outcome to single euploid blastocyst transfer. What is known already Embryonic mosaicism occurs when there are two or more distinct cell lines found in preimplantation embryos derived from IVF. Data from recent studies show that mosaic blastocysts have the potential to implant and can result in healthy live births. As a result, patients now have the option of transferring mosaic blastocyst when they do not have any euploid blastocyst available for transfer. However, the clinical outcome of transferring mosaic blastocyst has not been definitively reported. Thus, a retrospective study was conducted to compare the clinical outcome of mosaic sFBT and euploid sFBT. Study design, size, duration A total of 602 patients underwent frozen blastocyst transfer in Alpha IVF from January to October 2019 and had their blastocysts screened for aneuploidy. These patients were divided into 2 groups: 26 patients with mosaic blastocysts transferred (Group A, age ranged 19–44), and 576 patients with euploid blastocysts transferred (Group B, age ranged 21–44). The mean age of patients from Group A and B were 34.0 and 32.8 respectively (p > 0.05). Participants/materials, setting, methods All samples had their DNA libraries constructed for sequencing using Next Generation Sequencing according to manufacturer’s specification (IonTorrent, USA). All blastocysts were frozen for subsequent sFBT cycle (Cryotech, Japan). All thawed blastocysts for sFBT survived with morphologically intact inner cell mass and trophectoderm cells. The importance of antenatal confirmation of the fetal chromosome status was emphasized in patients from Group A. The clinical outcomes of both groups were analysed and compared. Main results and the role of chance No significant differences were seen in the clinical pregnancy and implantation rate of Group A and B (65.4% vs 63.0%; p > 0.05). The miscarriage rate of Group A and B were 23.5% and 14.0% respectively. Albeit the higher miscarriage rate in Group A, there was no statistical significance between these two groups (p > 0.05). Group A was further divided into two subgroups, Subgroup A1: low risk mosaic blastocyst transfer; Subgroup A2: high risk mosaic blastocyst transfer. In the comparison of Group A subgroups, the clinical pregnancy and implantation of Group A1 is higher than Group A2 (76.9% vs 44.4%). In addition, the miscarriage rate of Group A1 and A2 were 23.1% and 0.0% respectively. Interestingly, there was no statistical significance in clinical pregnancy rate, implantation rate and miscarriage rate between these two subgroups. Limitations, reasons for caution This is a retrospective study and the sample size was comparatively smaller in the mosaic blastocyst transfer group than the euploid blastocyst transfer group. Further studies with a larger sample size should be carried out to ascertain the clinical outcome. Wider implications of the findings: Single mosaic blastocyst transfer has similar clinical outcome to single euploid blastocyst transfer. Thus, mosaic blastocyst can be considered for transfer when no euploid blastocyst are available. Nevertheless, stringent antenatal surveillance for chromosomal abnormalities to confirm the chromosomal status of the fetus must be followed. Trial registration number Not applicable

Evaluation of the practical applications of fluorescence in situ hybridization in the prenatal diagnosis of positive noninvasive prenatal screenings

Objective To investigate the application value and limitations of fluorescence in situ hybridization (FISH) in prenatal diagnosis of positive results for trisomies 13, 18, 21 (T13, T18, T21) and sex chromosome aneuploidies (SCAs) indicated by noninvasive prenatal screening (NIPS). Methods Samples from women who underwent prenatal diagnosis for the indication of positive NIPS of T13, T18, T21, and SCAs were collected. Each sample was split into two for both karyotype analysis and FISH analysis. The efficiency and consistency of FISH were assessed for the detection of chromosome abnormalities in the indications of positive NIPS results compared with karyotyping. Results A total of 649 pregnant women who scored positive for clinical significance of fetal chromosome abnormalities by NIPS were enrolled in our study, including T 13 (6%), T18 (14.3%), T21 (44.7%), SCAs (35.0%). From the following diagnostic test, the positive predictive value (PPV) of NIPS for T13, T18, T21, and SCAs was 17.9, 60.2, 89.3, and 43.6% respectively. FISH analysis was successful in all samples. Compared with karyotyping, the sensitivity and specificity were 98.3 and 100%, respectively. 95.7% (621/649) were fully concordant with karyotyping. 3.2% (21/649) cases were incompletely concordant with the karyotyping, among these cases, the FISH analysis identified all the aneuploidies, but karyotyping analysis provided more information about the chromosomal structure. There were 7 cases (1.1%, 7/649) of anomalies diagnosed by karyotype but missed out by FISH, all of which occurred in cases with the indication of SCAs. If the indications were confined to cases with a positive NIPS of T13, T18, T21, the diagnostic consistency of the two methods almost perfectly agree, and all the aneuploidies were detected by the FISH assay. FISH analysis was highly consistent in determining whether the fetus was euploid or not in the prenatal diagnosis for the patients with positive NIPS results compared with karyotyping (kappa= 0.976, p < .01). Conclusion For the prenatal diagnostic indications of positive NIPS of T13, T18, T21, and SCAs, FISH was equally efficacious in identifying aneuploidies and provided a quick diagnosis to alleviate anxiety. However, the missed risk of FISH analysis for structural chromosomal abnormalities should be taken seriously and fully informed during genetic counseling.

The Optimal Cutoff Value of Z-scores Enhances the Judgment Accuracy of Noninvasive Prenatal Screening.

ObjectiveTo evaluate the accuracy of Z-scores of noninvasive prenatal screening (NIPS) in predicting 21, 18 trisomy, and X chromosome aneuploidy.MethodsA total of 39,310 prenatal women were recruited for NIPS from September 2015 to September 2020. Interventional prenatal diagnosis was applied to verify the diagnosis of NIPS-positive results. Logistic regression analysis was employed to relate the Z-scores to the positive predictive value (PPV) of NIPS-positive results. Using receiver operating characteristic (ROC) curves, we calculated the optimal cutoff value of Z-scores to predict fetal chromosome aneuploidy. According to the cutoff value, NIPS-positive results were divided into the medium Z-value (MZ) and high Z-value (HZ) groups, and PPV was calculated to access the accuracy of Z-scores.ResultsA total of 288 effective values of Z-scores were used as the final data set. The logistics regression analysis revealed that Z-scores were significantly associated with true-positive results for 21 trisomy (T21) and 18 trisomy (T18) (P < 0.05), whereas the same was not observed for X chromosome aneuploids (P > 0.05). The optimal cutoff value of the Z-score for T21, T18, XO, XXX, and XXY indicated by ROC curve analysis were 5.79, 6.05, −9.56, 5.89, and 4.47, and the area under the curve (AUC) were 0.89, 0.80, 0.48, 0.42, and 0.45, respectively. PPV in the HZ group was higher than that in the MZ group, and the application of the cutoff value reduced the false discovery rate (FDR), which was only 2.9% in the HZ group compared with 61.1% in the MZ group for T21 and T18. The difference in total PPV between the MZ and HZ groups for X chromosome aneuploids was statistically significant. Moreover, the PPV for XXX and XXY seemed to increase with Z-scores but not for XO.ConclusionThe Z-score is helpful for the accurate judgment of NIPS results and for clinical prenatal counseling. Especially for T21 and T18, Z-scores have an excellent clinical association, which is superior to that seen with X chromosome aneuploids. In addition, using Z-scores to judge NIPS results offers a certain reference value for XXX and XXY but not for XO.

Investigation on combined copy number variation sequencing and cytogenetic karyotyping for prenatal diagnosis

BackgroundWe aimed to evaluate the clinical value of copy number variation-sequencing (CNV-Seq) in combination with cytogenetic karyotyping in prenatal diagnosis.MethodsCNV-Seq and cytogenetic karyotyping were performed in parallel for 9452 prenatal samples for comparison of the diagnostic performance of the two methods, and to evaluate the screening performance of maternal age, maternal serum screening, fetal ultrasound scanning and noninvasive prenatal testing (NIPT) for fetal pathogenic copy number variation (CNV).ResultsAmong the 9452 prenatal samples, traditional karyotyping detected 704 cases (7.5%) of abnormal cytogenetic karyotypes, 171 (1.8%) chromosome polymorphism, 20 (0.2%) subtle structural variations, 74 (0.7%) mutual translocation (possibly balanced), 52 (0.6%) without karyotyping results, and 8431 (89.2%) normal cytogenetic karyotypes. Among the 8705 cases with normal karyotype, polymorphism, mutual translocation, or marker chromosome, CNV-Seq detected 63 cases (0.7%) of pathogenic chromosome microdeletion/duplication. Retrospectively, noninvasive prenatal testing (NIPT) had high sensitivity and specificity for the screening of fetal pathogenic CNV, and NIPT combining with maternal age, maternal serum screening or fetal ultrasound scanning, which improved the screening performance.ConclusionThe combined application of cytogenetic karyotyping and CNV-Seq significantly improved the detection rate of fetal pathogenic chromosome microdeletion/duplication. NIPT was recommended for the screening of pathogenic chromosome microdeletion/duplication, and NIPT combining with other screening methods further improved the screening performance for pathogenic fetal CNV.

Molecular diagnosis of sex chromosome mosaics in fetal amniotic cells.

Mosaicism can be observed in karyotype analyses of amniotic fluid cells. Differentiating between true mosaicism and pseudomosaicism and determining mosaic proportions can help avoid misjudgments by doctors and effectively reduce mental and physical harm to pregnant women. However, the detection of mosaicism and mosaic proportions via karyotype analysis and fluorescence in situ hybridization (FISH) is extremely complex. We have developed a novel approach, “segmental duplication quantitative fluorescent PCR” (SD-QF-PCR), to detect mosaicism and mosaic proportions.In this study, twenty control samples and fourteen mosaic samples were tested by first-line karyotype analysis; by second-line karyotype analysis, SD-QF-PCR and FISH were used to reassess fetal sex chromosome mosaicism and mosaic proportions.Detection of the 20 control samples by second-line karyotype analysis via FISH and SD-QF-PCR showed normal and consistent results. Among the 14 mosaic samples, the numbers of samples showing true mosaicism and pseudomosaicism detected by the three methods were 6 and 8, respectively.Our study demonstrates that SD-QF-PCR can be used as a complementary method to traditional cytogenetic analysis of amniotic fluid mosaics and has clinical application value.

An assessment of the analytical performance of non-invasive prenatal testing (NIPT) in detecting sex chromosome aneuploidies: 34,717-patient sample in a single prenatal diagnosis Centre in China.

The present study aimed to evaluate the efficacy of a non-invasive prenatal test (NIPT) in the detection of the sex chromosome aneuploidies (SCAs) at our prenatal diagnosis centre. Among a cohort of 34,717 pregnancies, maternal plasma samples from our prenatal diagnosis centre were subject to analysis of SCAs using NIPT detection. Pregnant women with NIPT positive results of SCAs were recommended to undergo an invasive prenatal diagnosis (i.e. karyotyping and fluorescence in situ hybridization) to validate the prediction value of NIPT. From 34,717 clinical pregnancies, 229 (0.66%) pregnancies were identified with SCAs. Of these, 78 (34.1%) cases were positive for 45,X and 151 (65.9%) cases comprised a sex chromosome trisomy. Of the 229 positive NIPT results, 193 (84.3%) cases had accepted an invasive diagnosis involving karyotyping analysis of the amniotic fluid, which confirmed 67 cases (34.7%) as true positive, as well as 126 cases (65.3%) as false positive. The positive predictive values were 23.07%, 50%, 36% and 27.27% respectively. The remaining 36 (15.7%) cases declined a prenatal diagnosis. The termination rates of 45,X, 47,XXY, 47,XXX and 47,XYY were 20.5%,46%,12.9% and 11.5% respectively. NIPT demonstrated a lower accuracy in predicting monosomy X than sex chromosome trisomies. After invasive testing, the fetal chromosome with 45,X and 47,XXY were terminated more often than those with 47,XXX, 47,XYY. Because NIPT is a screening test, false positive/negative cases exist, and pre- and post-test counselling is essential for informing patients about the benefits and limitations of the test. Confirmatory testing of abnormal results is recommended prenatally or after birth, and the importance of confirmatory testing and benefits of early diagnosis should be addressed.

Genomic and structural abnormalities of the chromosome set of the fetus, depending on the age of pregnant women

To study the frequency and spectrum of fetal chromosomal abnormalities in pregnant women, depending on age, a cytogenetic study of 2248 biopsies of the chorion and placenta, cord blood lymphocytes and amniocytes was carried out in 2016-2020. The work used the generally accepted methods of direct preparation of chromosome preparation and methods of cell cultivation. The analysis of the preparations was performed based on GTG, CBG staining method and FISH method, using DNA probes for centromeric regions 13, 18, 21 and X, Y chromosomes. Numerical and structural abnormalities of chromosomes were considered according to the ISCN 2013 system. As a result of the study, 272 (12.1%) cases of genomic and structural abnormalities of the fetus chromosomes of the fetus of pregnant women were registered. Genomic abnormalities were 69.1%, and chromosome structural rearrangements - 30.9%. Among genomic disorders, the frequency of the Down syndrome in the karyotype of the fetus was 58.5%. Frequency of occurrence of fetal chromosomal abnormalities varies significantly depending on the age group of pregnant women. The frequency of fetal chromosomal pathology in pregnant women under 20 years old was 4.4%, at the age from 20 to 34 years old - 7.6%, from 35 to 39 years old - 16.3%, from 40 to 44 years old - 19.8% , after 45 years figure increased significantly - up to 29.6%. In older pregnant women, when compared with pregnant women of the reproductive period from 20 to 34 years, when the highest level of pregnancy rate (54.6%) in the population is observed, there is an increase in the frequency of fetal chromosomal pathology by 2.1; 2.6 and 3.9 times, respectively.

Copy number variation sequencing combined with quantitative fluorescence polymerase chain reaction in clinical application of pregnancy loss.

In this study, we evaluated the feasibility of the combining CNV-seq and quantitative fluorescence polymerase chain reaction (QF-PCR) for miscarriage analysis in clinical practice. Over a 35-month period, a total of 389 fetal specimens including 356 chorionic villi and 33 fetal muscle tissues were analyzed by CNV-seq and QF-PCR. Relationships between the risk factors (e.g., advanced maternal age, abnormal pregnancy history, and gestational age) and incidence of these chromosomal abnormalities were further analyzed by subgroup. Clinically significant chromosomal abnormalities were identified in 58.95% cases. Aneuploidy was the most common abnormality (46.84%), followed by polyploidy (8.16%) and structural chromosome anomalies (3.95%). In sub-group analysis, significant differences were found in the total frequency of chromosomal abnormalities between the early abortion and the late abortion group, as well as in the distribution of chromosomal abnormalities between the advanced and the younger maternal age group. Meanwhile, the results of the logistic regression analysis identified a trend suggesting that the percentage of fetal chromosomal abnormalities is significantly higher in advanced maternal age, lesser gestational age, and lesser number of prior miscarriages. Our study suggests that CNV-seq and QF-PCR are efficient and reliable technologies in the fetal chromosome analysis of miscarriages and could be used as a routine selection method for the genetic analysis of spontaneous abortion.

Economic impact of using maternal plasma cell-free DNA testing to guide further workup in recurrent pregnancy loss.

ObjectiveWe have previously demonstrated that maternal‐plasma cell‐free DNA (cfDNA)‐testing can detect chromosomal anomalies in recurrent pregnancy loss (RPL) with 81.8% sensitivity and 90.3% specificity. Here we assess whether this is cost effective in guiding further workup in RPLs.MethodA decision‐analytic model was developed to compare the cost of various RPL management pathways: (1) current American Society for Reproductive Medicine (ASRM) RPL workup; (2) microarray or karyotyping analysis of products of conception (POCs) and RPL workup only for euploid cases; and (3) cfDNA testing and RPL workup only for euploid cases. Sample accessibility, failure rates, and sensitivity were specified for each test. Costs of sample collection, genetic tests, and RPL workup were considered. Analysis outcomes included detection rate of chromosomal anomaly and cost per patient tested.ResultsIn comparison to existing cytogenetic testing on POCs, cfDNA testing pathway allowed for better sample accessibility with a lower cost per patient. In addition, using cfDNA to guide further workup significantly increases the number of causative fetal chromosome anomalies detected, reducing the number of patients undergoing unnecessary workup resulting in an overall cost savings.ConclusionOur study showed that inclusion of cfDNA testing is a cost‐effective approach to guide RPL workup.

Expanding the Scope of Non-invasive Prenatal Testing to Detect Fetal Chromosomal Copy Number Variations.

Non-invasive prenatal testing (NIPT) for common fetal trisomies is effective. However, the usefulness of cell-free DNA testing to detect other chromosomal abnormalities is poorly understood. We analyzed the positive rate at different read depths in next-generation sequencing (NGS) and identified a strategy for fetal copy number variant (CNV) detection in NIPT. Pregnant women who underwent NIPT by NGS at read depths of 4–6 M and fetuses with suspected CNVs were analyzed by amniocentesis and chromosomal microarray analysis (CMA). These fetus samples were re-sequenced at a read depth of 25 M and the positive detection rate was determined. With the increase in read depth, the positive CNV detection rate increased. The positive CNV detection rates at 25 M with small fragments were higher by NGS than by karyotype analysis. Increasing read depth in NGS improves the positive CNV detection rate while lowering the false positive detection rate. NIPT by NGS may be an accurate method of fetal chromosome analysis and reduce the rate of birth defects.

The correlation between maternal age and fetal sex chromosome aneuploidies: a 8-year single institution experience in China

BackgroundAlthough a few studies have investigated a possible association between maternal age and fetal sex chromosome aneuploidies (SCAs), most of these studies were limited to advanced maternal age (AMA) women and the results were conflicting. This study aimed to investigate the correlation between maternal age and common fetal SCAs (including 45,X, 47,XXY, 47,XXX and 47,XYY) in pregnant women of different ages that not only limited to AMA women. We retrospectively investigated a 8-year experience of prenatal diagnosis for fetal chromosome aberrations by second-trimester amniocentesis at a university teaching hospital in China. 20,409 amniotic fluid specimens collected at 19–22+6 gestational weeks were included in this study. The women were categorized into five age groups (≤ 23, 24–28, 29–33, 34–38, 39+ years) based on maternal age at the time of amniocentesis and entered as a categorical variable in all samples. The correlation between fetal SCAs and maternal age was determined using the logistic regression analysis. A chi-square test was performed to compare the incidence of fetal SCAs among age groups.ResultsA total of 179 cases of fetal SCAs were detected, and the incidence was 8.77‰ (about 1/114). The incidence of fetal SCAs increased significantly with advancing maternal age (SE, 0.014; odds ratio, 1.044; P = 0.002). Specifically, the incidence of 45,X (SE, 0.037; odds ratio, 0.916; P = 0.017) and 47,XXY (SE, 0.024; odds ratio, 1.127; P = 0.000) had significant correlation with maternal age, while the incidence of 47,XXX and 47,XYY had no correlation with maternal age (P = 0.473; P = 0.272, respectively). The incidence of fetal SCAs was also significantly different among age groups (χ2 = 10.197, P = 0.037 < 0.05), from 5.81 per 1000 fetuses at the 24–28 years to 10.92 per 1000 at the 39+ years.ConclusionsMaternal age was ascertained to be a strong risk factor for fetal SCAs.

Amniocentesis and Next Generation Sequencing (NGS)-Based Noninvasive Prenatal DNA Testing (NIPT) for Prenatal Diagnosis of Fetal Chromosomal Disorders.

ObjectiveThe present study aimed to evaluate and analyze the results of karyotyping by amniocentesis and next generation sequencing (NGS)-based noninvasive prenatal DNA testing (NIPT) for the prenatal diagnosis of fetal chromosomal disorders.MethodsA total of 2267 high-risk pregnant females with the indications for prenatal diagnosis who met the enrollment criteria between January 2015 and May 2019 at the Affiliated Hospital of Inner Mongolia Medical University were included and underwent NGS-based NIPT in the present study. Amniocentesis, chromosome karyotyping by cell culture, and follow-up of the pregnancy outcomes were also conducted in the NIPT-positive pregnant females to assess the consistency between NIPT and results of karyotyping by amniocentesis.ResultsAmong the 2267 cases, 29 cases were positive for NIPT, including 10 cases with a high risk of trisomy 21, 2 cases with a high risk of trisomy 18, 2 cases with a high risk of chromosome 13, and 20 cases with sex chromosome abnormalities. All the above NIPT-positive cases underwent amniocentesis, and 20 cases were eventually diagnosed. The sensitivity and specificity of NIPT for the diagnosis of trisomy 21, trisomy 13, and trisomy 18 were 100%, 99.96%, 100%, and 99.96%, 100%, 100%, respectively, and the positive predictive values were 91.67%, 66.67%, and 100%, respectively.ConclusionNGS of the fetal free DNA from the peripheral blood of pregnant females was an important complement to the prenatal diagnosis of chromosomal disorders represented by fetal chromosome aneuploidy with high sensitivity and specificity. In combination with the traditional karyotyping by amniocentesis, it could improve the diagnostic efficacy for fetal chromosomal disorders.

Application analysis of noninvasive prenatal testing for fetal chromosome copy number variations in Chinese laboratories

Objective: To investigate the general situation, detection range, testing reagents, and clinical performance of non-invasive prenatal testing (NIPT) for fetal chromosomal copy number variations (CNVs) in Chinese laboratories. Methods: The National Center for Clinical Laboratories of the National Health Commission designed a questionnaire for the detection of CNVs by NIPT, which included the investigation of whether the laboratory has carried out NIPT to detect CNVs and its testing scope, reagents/platforms, intended uses, screening populations and clinical performance. The questionnaires were distributed to 355 laboratories in 31 provinces, autonomous regions, and municipalities across the country on October, 2020. Further, the feedbacks were statistical analyzed. Results: Two hundred and twenty-eight laboratories had performed NIPT to detect CNVs, including 116 types of CNVs, and more than 95% of laboratories chose to detect the CNVs of 5p15 deletion, 22q11.2 deletion, 1p36 deletion, and 15q11.2 deletion. All testing reagents used were laboratory-developed tests and were based on massive parallel sequencing, the minimum amount of sequencing data was 3-15 M reads, the detection limit of fetal fraction was 3%-5%, and the minimum size of variants that can be detected was 1-5 Mb. The proportion of laboratories that apply CNVs testing for daily project, voluntary requirements of patients, and scientific research were 58.8% (134/228), 57.5% (131/228), and 20.6% (47/228), respectively. One hundred and thirty-four laboratories were fully or partially aware of the clinical performance of NIPT to detect microdeletion/microduplication syndromes, and the laboratories' declared sensitivity of NIPT for Cri du Chat syndrome, 22q11.2 deletion syndrome, 1p36 deletion syndrome, and Angelman syndrome were 50.0%-100%, 60.0%-100%, 50.0%-100%, and 33.3%-100%, and the positive predictive values were 9.0%-50.0%, 18.0%-100%, 20.0%-30.0%, and 20.0%. Conclusion: The detection of CNVs by NIPT in Chinese laboratories need to be standardized. Laboratories should detect CNVs with clear clinical significance in accordance with the guidelines, conduct performance validation of the reagents, then perform NIPT test and provide adequate interpretation after mastering the clinical performance sufficiently.

Effectiveness of non-invasive prenatal screening for the detection of fetal sex chromosome anomalies

To evaluate the efficacy of non-invasive prenatal screening (NIPS) for fetal sex chromosome anomalies. A retrospective analysis was carried out for 20 802 women undergoing NIPS screening. For 165 cases suspected for fetal sex chromosomal anomalies, the results of invasive prenatal diagnosis were obtained. Among the 165 cases suspected for fetal sex chromosome anomalies, 129 have accepted invasive prenatal diagnosis, and 45 were confirmed, which yielded a positive predictive value of 34.88%. These included 16 cases of 47,XYY, 10 cases of 47,XXY, 6 cases of 45,X/46,XX, 5 cases of 47,XXX, 3 cases of 45,X, 1 case of 45,X/46,X,i(X)(q10), 1 case of 45,X/46,X,del(X)(q22), 1 case of 46,X,del(X)(q22), 1 case of 46,X,del(X)(p11) and 1 case of Xp22.31 1.2 Mb deletion. NIPS has limited value for detecting fetal sex chromosome anomalies. Karyotyping analysis combined with other diagnostic techniques can offer effective prenatal diagnosis for suspected cases.

Value of non-invasive prenatal testing for the detection of fetal chromosomal copy number variations

To explore the value of non-invasive prenatal testing (NIPT) for the detection of fetal chromosome copy number variations (CNVs). Clinical data of 18 661 pregnant women who underwent NIPT were collected. For fetuses suspected for carrying CNVs, amniotic fluid samples were collected for chromosomal karyotyping and/or chromosomal microarray analysis (CMA). Among all samples, NIPT suggested that 58 fetuses carried trisomy 21, 18 carried trisomy 18, 19 carried trisomy 13, 1 carried trisomies 18 and 21. Eighty eight women accepted invasive prenatal diagnosis. The results of CMA in 59 cases were consistent with those of NIPT, which yielded a consistency rate of 67.05%. In addition, 37 cases of fetal CNVs were detected by NIPT, of which 19 (15 microdeletions and 4 microduplications) have accepted invasive prenatal diagnosis. In 14 cases, the results were consistency with those of NIPT, with a consistent rate of 73.68%. NIPT features high sensitivity and accuracy. Invasive prenatal diagnosis should be considered for CNVs detected by NIPT, and by tracing its parental origin, it can provide guidance for clinical practice.

Adoption of a non-invasive prenatal test (NIPT) in prenatal screening in Moscow: first results

The objective — To assess the effectiveness of including NIPT in the structure of prenatal diagnostics in Moscow. Material and Methods — Totally 5,181 pregnancies undergoing screening for fetal trisomy using NIPT during the period from 01.04.2020 to 30.09.2020 in Russia. According to the results of biochemical blood test, the patients were divided into two groups: group of high risk (cut-off ≥1:100) (n=208) and group of intermediate risk (cut-off 1:101 – 1:2500) (n=4,973). Patients at high-risk cell-free DNA (cfDNA) were offered an invasive procedure, followed by genetic analysis (cytogenetic or molecular karyotyping). Results — Among the analysed samples, 117 (2.3%) had a high risk of the following common fetal chromosome abnormalities by NIPT: trisomy 21 in 50 cases, trisomy 18 in 17 cases, trisomy 13 in 5 cases, and sex chromosome aneuploidy (SCA) in 22 cases. Additionally, rare autosomal trisomies and/or subchromosomal arrangements were revealed in 23 cases. We found associations between cfDNA concentration and high risk of aneuploidies (particularly trisomy 21) and fetal sex and between low fetal fraction (FF) and body mass index (BMI) as well as maternal weight. Additionally, a high risk of trisomy 21 was associated with the term gestation. Conclusion — The effectiveness of technological resources that are based on cfDNA testing for detecting abnormal fetal chromosome numbers and other chromosomal anomalies is high and reduce rates of false positive results. Therefore, NIPT should be more widely used as a first-line screening method.

Non-invasive prenatal testing for the prenatal screening of sex chromosome aneuploidies: A systematic review and meta-analysis of diagnostic test accuracy studies.

BackgroundThere is little evidence on the performance of non‐invasive prenatal testing (NIPT) for the detection of fetal sex chromosomal imbalances. In this review, we aimed to appraise and synthesize the literature on the performance of NIPT for the prenatal detection of fetal sex chromosome aneuploidies.MethodsWe performed our literature search in PubMed, Embase, Cochrane Library, Web of Science, and CADTH. Study selection and data extraction were performed by two reviewers independently. There were no restrictions on the study population. Meta‐analyses were performed with “R” software. Pooled sensitivities and specificities with their 95% CI were estimated using a random‐effects model. Heterogeneity between studies was assessed by a Q test.ResultsBased on 11 studies in high prior risk pregnancies, including 116 affected fetuses in aggregate, Massively Parallel Shotgun Sequencing (MPSS) had a sensitivity of 93.9% (95% CI 84.1%, 97.8%) and a specificity of 99.6% (95% CI 98.7%, 99.9%) for the detection of 45,X. Based on four studies in high‐risk pregnancies, with 83 affected fetuses in aggregate, Targeted Massively Parallel Sequencing (TMPS) had a sensitivity of 83.2% (95% CI 49.6%, 96.2%) and specificity was 99.8% (95% CI 98.3%, 100%) for the detection of 45,X. In mixed‐risk pregnancies, the sensitivity of TMPS for the detection of 45,X was 90.9% (2 studies; 95% CI 70%, 97.7%) and specificity 99.9% (2 studies; 95% CI 99.4%, 100%); MPSS data were not available in such pregnancies. Based on smaller numbers of studies, and small numbers of affected fetuses in either high‐risk or mixed‐risk pregnancies (using either MPSS or TMPS), the sensitivities and specificities were equal to or greater than 76.2% for 47,XXX, 47,XXY and 47, XYY. The test failures for SCAs were 0.2% (95% CI 0%, 13.6%) for MPSS and 5.6% (95% CI 3.7%, 8.4%) for TMPS.ConclusionHigh‐quality studies are still desirable in order to estimate the performance of NIPT for the detection of sex chromosome imbalances.

Prenatal diagnosis of 913 fetuses samples using copy number variation sequencing.

BackgroundThe present study aimed to explore the etiological relationship between fetal abnormalities and copy number variations (CNVs) with the aim of intervening and preventing the birth of children with birth defects in time.MethodsSamples of 913 fetuses with puncture indications were collected from January 2017 to December 2019. Karyotype analysis and CNV sequencing (CNV‐seq) testing was performed for fetuses with ultrasonic abnormalities, a high risk of Down’s syndrome and an adverse birth history. All cases were followed up.ResultsIn total, 123 cases (13.47%) had abnormal karyotypes, including 109 cases with chromosome number abnormalities and 14 cases of chromosomal structural abnormalities. Thirty‐seven (4.05%) cases with pathogenic CNVs were detected. The detection rate of pathogenicity CNVs was 12.82% for mixed indications, followed by 7.5% for an adverse birth history, 5.88% at high risk of non‐invasive prenatal testing, 5.00% with an abnormal ultrasonic marker, 1.89% at high risk of screening for Down's syndrome and 1.45% with advanced maternal age. There were 12 (1.31%) cases with microduplications and 25 (2.74%) cases with microdeletions. Trisomy 21 (39.02%), trisomy 18 (13.82%) and Turner syndrome (9.76%) were the top three chromosome abnormalities. There were 104, 746 and 63 cases in the 11–13 weeks, 14–27 weeks 28–38 weeks gestational ages, respectively. The abnormal rates of fetal chromosome aneuploidy and the rate of pathogenic CNVs were decreased and increased with the increase of gestational age (p < 0.05), respectively.ConclusionsCompared with karyotype analysis, CNV‐seq can improve the detection rate of chromosomal abnormalities. CNV‐seq combined karyotype analysis should be performed simultaneously in fetuses with puncture indications.

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

BackgroundNon-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 presentationThe 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.ConclusionWhole-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.