Prenatal diagnosis and genetic analysis of 13 fetuses with 16p11.2 microdeletion/microduplication

Objective To investigate the value of standardized ultrasound screening in diagnosis of fetal congenital heart disease (CHD) during the first trimester. Methods This study retrospectively analyzed the clinical data of 8 383 fetuses who received ultrasound screening during the first trimester in the Dongguan Maternal and Child Health Hospital from September 2015 to December 2016. Standardized ultrasound was performed to observe fetal heart position, apical direction, apical four-chamber view, three vessels and trachea view and the thickness of nuchal translucency (NT). Fetuses with thickened NT or fetal CHD observed during the first and second trimester were followed up. Pregnancy outcomes and the growth of newborns within one year after birth were recorded and analyzed. Pathological results after the termination of pregnancy were compared with the results of routine karyotyping and chromosome microarray analysis (CMA). Results (1) A total of 27 cases of fetal CHD were identified during the first trimester giving a detection rate of 0.32% (27/8 383). These included ten (37.0%) of single atrium and/or single ventricle, seven (25.9%) of endocardial cushion defect (including two complicated by persistent arterial trunk), three (11.1%) of hypoplastic right heart syndrome, three (11.1%) of interventricular septal defect, two (7.4%) of hypoplastic left heart syndrome, one (3.7%) of mirror-image dextrocardia and one (3.7%) of right atrial enlargement and severe tricuspid regurgitation. Nineteen out of the 27 cases had NT thickening (NT≥3.0 mm) and 17 of them had a cystic hygroma (NT≥6.0 mm). Among the 27 cases, 22 were terminated in the first trimester which autopsy results were consistent with ultrasound and the other five were rescreened during the second trimester. Thirteen out of the 27 cases received chorionic villus sampling, and seven of them were found to have chromosomal abnormalities by karyotyping and CMA, among whom one was microdeletion of 22q11. (2) Twenty-one cases of CHD were detected in the second-trimester ultrasound screening, including five initially identified in the first trimester. These cases included four (19.0%) of complex cardiac malformations (with three or more malformations), four (19.0%) of interventricular septal defect, three (14.3%) of dextroaortic arch, left subclavian artery vagus and 'U' shaped vascular ring, three (14.3%) of hypoplastic right heart syndrome (including one complicated by coronary artery-right ventricular fistula and one by interventricular septal defect), two (9.5%) of transposition of the great arteries, two (9.5%) of tetralogy of Fallot, one (4.8%) of hypoplastic left heart syndrome, one (4.8%) of Taussig-Bing anomaly and one (4.8%) of coarctation of the aorta. Among the 16 cases first identified in the second trimester, eight had NT thickening, including one with cystic hygroma. Among the 21 cases, two were lost to follow-up after being transferred to another hospital; four with negative results in karyotype analysis and CMA were delivered vaginally at term (37-40 gestational weeks) with 1-min Apgar scores of ten points and postpartum ultrasound of the baby was consistent with the second-trimester ultrasound screening; 15 were terminated and the autopsy confirmed those findings in the second-trimester ultrasound screening. Eleven out of the 21 cases received amniocentesis and five of them were found to be abnormal according to karyotype analysis and CMA, including one of microdeletion of 22q11. Conclusions Standardized first-trimester ultrasound screening is important and of great clinical value in the diagnosis of fetal CHD. Increased NT thickness could be a key indicator of fetal CHD and chromosomal abnormalities in early pregnancy. CMA may facilitate detecting the abnormality of genetic material in fetuses with normal chromosome karyotype. Key words: Heart defects, congenital; Ultrasonography, prenatal; Pregnancy trimester, first; Microarray analysis

Regarding increased nuchal translucency (NT), the cutoff values used are heterogeneous in clinical practice, this study aims to assess the efficacy of prenatal detection for chromosomal abnormalities and pregnancy outcomes in fetuses with varying NT thicknesses, in order to provide data that supports informed prenatal diagnosis and genetic counseling for such cases. We included 2,272 pregnant women under 35 with singleton pregnancies who underwent invasive prenatal diagnosis between 2014 and 2022. The cohort comprised 2,010 fetuses with increased NT (≥2.5 mm) and 262 fetuses with normal NT but exhibiting a single soft marker. Prenatal diagnoses were supported by chromosomal microarray (CMA) and copy number variation sequencing (CNV-seq) analyses. The detection rates of numerical chromosomal abnormalities were 15.4% (309/2,010) and 17.3% (297/1,717) in the NT ≥2.5 and ≥ 3.0 groups, respectively. Pathogenic/likely pathogenic CNV incidence increased with NT thickness (χ2 = 8.60, p < 0.05), peaking at 8.7% (22/254) in the NT 4.5-5.4 mm group. Structural defects were found in 18.4% of fetuses with NT values between 2.5 mm and 2.9 mm. Chromosomal abnormality rates in the isolated increased NT groups of 2.5-2.9 mm and 3.0-3.4 mm were 6.7% (16/239) and 10.0% (47/470), respectively, with no statistical significance (χ2 = 2.14, p > 0.05). Fetuses with NT thickness between 2.5 and 2.9 mm combined with the presence of soft markers or non-lethal structural abnormalities exhibited a significantly higher chromosomal abnormality risk (19.0%) compared to fetuses with isolated increased NT ranging from 3.5 to 4.4 mm (13.0%). Pregnancy termination rates increased with NT thickness (χ2 = 435.18, p < 0.0001), ranging from 12.0% (30/249) in the NT 2.5-2.9 mm group to 87.0% (141/162) in the NT ≥ 6.5 mm group. CMA or CNV-seq exhibited good performance in identifying genomic aberrations in pregnancies with increased NT thickness. NT ranging from 2.5 mm to 2.9 mm elevated the risk of fetal chromosomal abnormalities, particularly when combined with other soft markers.

Chromosome microarray analysis (CMA) and whole exome sequencing (WES) are increasingly utilized in prenatal diagnosis of abnormal ultrasound findings, but studies on correlation between pathogenic copy number variations (pCNVs) and single-gene mutations in fetuses with nuchal translucency (NT) thickening/cystic hygroma (CH), and pregnancy outcomes, are rare. This study aimed to investigate clinical value of CMA and WES for NT thickening/CH in fetuses, explore genetic correlation between fetal NT thickening and CH, and analyze pregnancy outcomes. We retrospectively selected 215 pregnant women diagnosed with fetal NT thickening (NT > 95th)/CH who underwent invasive prenatal diagnosis at our hospital from January 2020 to June 2022. With negative chromosomal karyotype analysis (KA) and CMA results, patients voluntarily underwent WES. Patients were grouped by NT thickening/CH, and application value of KA, CMA, and WES examined. Ultrasound findings, pregnancy outcomes, and fetal growth post-birth were followed during mid/late pregnancy and post-delivery. Abnormalities in chromosomal number were detected in 28 of 215 samples, with a detection rate of 13.0%, and pCNVs were detected in 12 cases, with a detection rate of 5.6%. The most common abnormality in fetuses from both groups suggested by CMA was 22q11.21 microdeletion-microduplication syndrome. 35 patients with negative KA and CMA results underwent WES, and single gene variants were detected in 12 fetuses, with an abnormality rate of 34.3%. The incidence of adverse pregnancy outcomes was 28.2% in the NT thickening group and 82.9% in the CH group (P < 0.05). Overall, fetal NT thickening/CH was associated with genetic abnormalities, WES further improved the diagnosis of abnormal fetuses after negative KA and CMA results in both groups, and the incidence of adverse pregnancy outcomes was lower in the NT thickening group than in the CH group. The management of pregnancy outcomes could guide clinical genetic counselling.

Congenital talipes equinovarus (clubfoot)

ObjectiveTo assess prenatal diagnosis and pregnancy outcomes in twin pregnancies where one fetus has nuchal translucency (NT) above the 95th percentile.MethodIn this retrospective analysis, 130 twin pregnancies (260 fetuses) in which one twin had an NT measurement above the 95th percentile while that of the other twin was normal were analyzed. Prenatal diagnostic results such as G bands, chromosomal microarray analysis, ultrasound findings, and pregnancy outcomes were reviewed.ResultsKaryotype analysis and CMA results revealed that 15 (15.6 percent, 15/96) fetuses exhibited chromosomal abnormalities and that 13 fetuses were Variant of Uncertain Significance. Chromosome abnormalities were detected at a rate of 8.9% (5/56) in the DCT group and 25.0% (10/40) in the MCT group (p = 0.033, X2 = 4.571). 2 fetuses in DCT (3.9 percent, 2/51) and 4 fetuses in MCT (13.3 percent, 4/30) (p = 0.187) revealed structural abnormalities among the cases with normal prenatal diagnosis. Fetuses in the DCT group had an overall survival rate of 75.4 percent (95/126), whereas those in the MCT group had a survival rate of 60.4 percent (81/134) (p = 0.01, X2 = 6.636). According to the findings of Logistics regression analysis, NT thickening, maternal age and method of conception were all significant risk factors for chromosome abnormalities.ConclusionIn twin pregnancies with one fetus with NT above the 95th percentile, the prevalence of fetal structural abnormalities of the MCT group and the DCT group were comparable. Pregnant women’s age and mode of pregnancy are risk factors for chromosomal abnormalities.

ObjectiveThis study aimed to explore genetic etiologies of conotruncal defects (CTDs) in fetuses by analyzing the results of different genetic tests and to assess pregnancy outcomes of fetuses with CTD in a Chinese prenatal cohort.MethodsA total of 146 fetuses that underwent invasive prenatal genetic testing for CTD at the prenatal diagnosis center of the International Peace Maternity and Child Health Hospital between January 2018 and December 2022 were retrospectively analyzed.All of them underwent chromosomal microarray analysis (CMA) and karyotype analysis, but only 27 underwent whole‐exome sequencing (WES). The results of these different genetic testing methods were collected and compared. Data on pregnancy outcomes and neonate prognosis were obtained from the electronic medical records and postpartum telephone follow‐up.ResultsAmong the 146 fetuses with CTD, the chromosome abnormality rate was 22.6% (33/146), and the detection rate of abnormal chromosomes in CMA and karyotype was 15.8% (23/146). Among the 27 pregnant women who accepted WES, five had pathogenic variants. The detection rate of abnormal chromosomes by CMA was higher in infants with an interrupted aortic arch (IAA) than in the other infant groups (60.0%, χ2 = 11.661, P = 0.045, c2‐test or Fisher's exact test).ConclusionCongenital heart disease is a complex congenital heart disease with an etiology closely related to genetic abnormalities. Therefore, invasive genetic testing and CMA are recommended as first‐line tests for all fetuses with CTD detected by prenatal ultrasound, especially IAA. WES should be recommended when necessary.

A new comprehensive paradigm for prenatal diagnosis: seeing the forest through the trees.

Objective To investigate the genetic risk and pregnancy outcomes of fetuses who had the sonographic diagnosis of vertebral abnormalities (VA). Methods Fifty-two fetuses with sonographically detected VA (excluding neural tube defects) were included in the study. Data on prenatal ultrasound scan, prenatal genetic testing by amniocentesis, and pregnancy outcomes were collected and reviewed. Results Four types of VA were identified among 52 fetuses: butterfly vertebrae (26.9%, 14/52), hemivertebrae (59.6%, 31/52), hemivertebrae combined with butterfly vertebrae (9.6%, 5/52), and block vertebrae (3.9%, 2/52). Of the 52 fetuses, 33 presented VA as the sole sonographic anomaly, while the remaining 19 had associated anomalies. The positive rate of prenatal diagnosis for fetuses with VA was 19.2% (10/52). Chromosomal analysis, including karyotyping and chromosomal microarray analysis (CMA), detected one case of mosaic trisomy 9 and six cases of (likely) pathogenic copy number variants (CNVs). Whole exome sequencing (WES) identified four likely pathogenic variants in three cases with negative CMA results, specifically c.5110-1G > A in FLNB, c.8366G > A in KMT2D, and c.1275_1283dup as well as c.870 + 2T > C in DLL3. Among the 10 cases with diagnostic genetic testing results, seven fetuses exhibited isolated VA. There was no significant difference in the diagnostic rates between the isolated VA group (21.2%, 7/33) and the non-isolated VA group (15.8%, 3/19) (odds ratio [OR] 0.696, 95% confidence interval [CI] 0.157-3.087, p = 0.910). However, the live birth rate was significantly higher in the isolated VA group (71.9%, 23/32) compared to the non-isolated VA group (38.9%, 7/18) (OR 4.016, 95% CI 1.184-13.622, p = 0.022). Among the 30 live birth cases, two underwent spinal surgery and another two were identified with additional abnormalities. Following appropriate interventions, no apparent abnormalities were observed in the growth and development of 30 live birth cases. Conclusion Invasive prenatal diagnosis is recommended for all fetuses diagnosed with VA, regardless of whether associated anomalies are present. WES can enhance the diagnostic yield for fetuses with negative CMA results. Fetuses with isolated VA can have favorable pregnancy outcomes when genetic testing results are negative. However, long-term follow-up remains necessary for the assessment of the prognosis of these fetuses.

Background: There are few studies on the detection rate by chromosomal microarray analysis (CMA) of the prenatal diagnosis of talipes equinovarus (TE) compared to conventional karyotyping. We aimed to explore the molecular etiology of fetal TE and examine the detection rate by CMA, which provides more information for the clinical screening and genetic counseling of TE. Methods: In this retrospective study, pregnancies diagnosed with fetal TE were enrolled and clinical data for all cases were retrieved from our medical record database, including demographic data for pregnancies, ultrasound findings, karyotype/CMA results, and pregnant and perinatal outcomes. Results: Among the 164 patients, 17 (10.4%) clinically significant variants were detected by CMA. In 148 singleton pregnancies, the diagnostic rate of clinically significant variants was significantly higher in the non-isolated TE group than in the isolated TE group (10/37, 27.0% vs. 6/111, 5.4%, P < 0.001). In twin pregnancies, 1 (6.3%) pathogenic copy number variant was present in the other 16 twin pregnancies. Conclusions: This study demonstrates that CMA is useful for the prenatal genetic diagnosis of fetal TE. Fetal TE with the associated structural malformation correlates with a higher probability of clinically significant variants. This data may aid prenatal diagnosis and genetic counseling for fetal TE.

Genetic etiology plays a critical role in fetal ventriculomegaly (VM). However, the studies on chromosomal copy number variants (CNVs) in fetal VM are limited. This study aimed to investigate the chromosomal CNVs in fetuses with mild to moderate VM, and explore its genotype-phenotype correlation. A total of 242 fetuses with mild to moderate VM detected by prenatal ultrasound were enrolled in our study from October 2018 to October 2022. All cases underwent chromosomal microarray analysis (CMA) and G-banding simultaneously. All VM cases were classified different subgroups according to the maternal age, severity, VM distribution and presence/absence of other ultrasound abnormalities. The pregnancy outcomes and health conditions after birth were followed up. We also performed a pooled analysis regarding likely pathogenic and pathogenic CNVs (LP/P CNVs) for VM. The detection rate of chromosomal abnormalities by karyotyping was 9.1% (22/242), whereas it was 16.5% (40/242) when CMA was conducted (P < 0.05). The total detection rate of chromosomal abnormalities by karyotyping and CMA was 21.1% (51/242). A 12.0% incremental yield of CMA over karyotyping was observed. The detection rate of total genetic variants in fetuses with bilateral VM was significantly higher than in fetuses with unilateral VM (30.0% vs. 16.7%, P = 0.017). No significant differences were discovered between isolated VM and non-isolated VM, or between mild and moderate VM, or between advanced maternal age (AMA) and non-AMA (all P > 0.05). 28 fetuses with VM were terminated and 214 fetuses were delivered: one presented developmental delay and one presented congenital heart disease. The VM cases with both positive CMA and karyotypic results had a higher rate of termination of pregnancy than those with either a positive CMA or karyotypic result, or both negative testing results (P < 0.001). The combination of CMA and karyotyping should be adopted to improve the positive detection rate of chromosomal abnormalities for VM. The total genetic abnormalities detected using both techniques would affect the final pregnancy outcomes. LP/P CNVs at 16p11.2, 17p13, and 22q11.21 were identified as the top three chromosomal hotspots associated with VM, which would enable genetic counselors to provide more precise genetic counseling for VM pregnancies.

Objective To investigate the sonographic diagnosis and genetic counseling of fetal minorabnormalities. Methods Cases of fetal minorabnormalities detected by prenatal ultrasound in our hospital were Peking union Medical College Hospital between 2014 and 2017 enrolled in this retrospective study. The results of ultrasonography, genetic testing, and pregnancy outcomes were analyzed. Results Seventeen fetuses with minorabnormalities were detected by ultrasound, among which 10 cases showed the isolated finding, 2 cases showed two findings, and 5 cases showed more than three findings. Abnormal ultrasound findings were as follows: fetal biometry small for gestational age in 7 cases, choroid plexus cysts in 2, echogenic intracardiac focus in 1, echogenic bowel in 4, pyelestasis in 2, mild ventriculomegaly in 2, single umbilical artery in 2, persistent right umbilical vein in 2, persistent left superior vena cava in 1, absent nasal bone in 4, hand anomalies in 3, foot anomaly in 1, abnormal amniotic fluid in 2, and umbilical cord abnormalities in 1. All cases underwent the genetic analysis including karyotype, fluorescent in situ hybridization (FISH), and chromosomal microarray analysis (CMA). All of them showed normal results in karyotype and FISH, but CMA identified different copy number variants (CNVs). Seven cases had pathogenic CNVs, and 10 cases had variants of uncertain significance (VOUS), including 4 with VOUS-likely pathogenic, 3 with VOUS-likely benign, and 3 with VOUS-unknown. After counseling 9 cases chose to continue the pregnancy and the remaining 8 cases chose to terminate the pregnancy. Conclusions Careful examination and follow-up should be recommended in fetal minor abnormalitieson ultrasound. Prenatal genetic analysis could be recommended in such cases. Compared with traditional karyotyping, CMA could provide more information related with the prognosis of the fetuses, and prenatal counseling should be based on CMA results combined with ultrasound findings. Key words: Ultrasonography; Genetic analysis; Chromosomal microarray analysis; Copy number variants

Isodicentric Y chromosomes [idic(Y)] are frequently detected in patients with disorders of sex development (DSDs). However, prenatal cases are rarely reported. We performed comprehensive prenatal diagnostic evaluations in three fetuses with mosaic idic(Y) and followed up on pregnancy outcomes to explore the genotype-phenotype correlation. Amniotic fluid cells from three fetuses were subjected to karyotyping and chromosomal microarray analysis (CMA), with confirmation by fluorescence in situ hybridization (FISH) and real-time quantitative PCR (qPCR). Subsequently, parental karyotypes were analyzed using peripheral blood samples to assess inheritance (de novo vs. inherited). All three fetuses exhibited mosaic 45,X with idic(Y) karyotypes, which exhibited distinct breakpoints (Yp11.32, Yq11.221, and Yq11.223). Two fetuses had additional 46,XY cell lines. The SRY gene was retained in all cases. In Fetus 1, the AZF region was intact, whereas in Fetus 2 and Fetus 3, AZFb + c and AZFc were deleted. Fetus 1 was born alive following genetic counseling. Notably, a right adrenal neuroblastoma was detected during late gestation and successfully resected postnatally. The other two pregnant women elected for pregnancy termination. The integration of multiple technologies is essential for accurate prenatal diagnosis in cases of mosaic idic(Y). Our findings, including the rare association with neuroblastoma, contribute new insights into the phenotypic spectrum of this condition. Long-term follow-up and multidisciplinary clinical management are crucial for optimizing outcomes in live-born patients.

(1) Objective: To investigate the prenatal diagnosis and genetic counseling for 16p11.2 microdeletion syndrome and to evaluate its pregnancy outcome. (2) Methods: This study included 4968 pregnant women who selected invasive prenatal diagnoses from 1 January 2017 to 1 August 2022. These 4698 pregnancies underwent chromosomal microarray analysis (CMA), data on 81 fetuses diagnosed with 16p11.2 microdeletion syndrome based on prenatal ultrasound features and genetic test results were recorded, and their pregnancy outcome was evaluated. (3) Results: 1.63% of fetuses (81/4968) were diagnosed with 16p11.2 microdeletion syndrome. Among these, there were skeletal malformations in 48.15% of the 81 fetuses, cardiovascular malformations in 30.86%, central nervous system malformations (CNS) in 11.11%, digestive system structural abnormalities in 6.17%, and isolated ultrasonography markers in 3.70%. (4) Conclusions: 16p11.2 microdeletion syndrome can display various systemic ultrasound abnormalities in the perinatal period but vertebral malformations are the most common. Our study is the first to report that TBX1 and CJA5 are associated with 16p11.2 microdeletion syndrome, expanding the disease spectrum of 16p11.2 microdeletion syndrome. In our study, the ventricular septal defect is the main feature of cardiac structural abnormalities caused by 16p11.2 microdeletion syndrome. In addition, our study highlights the use of CMA in 16p11.2 microdeletion syndrome, analyzed their genetic results, and evaluated the follow-up prognosis, which can be useful for prenatal diagnosis and genetic counseling.

The aim of this study was to present prenatal ultrasound findings, molecular testing results and pregnancy outcomes of cases with 22q11.2 deletion (del22q11.2) diagnosed prenatally. A total of 76 foetuses were included. All cases were diagnosed by using chromosomal microarray analysis. Data on prenatal diagnosis, ultrasound findings, pregnancy outcomes and inheritance of del22q11.2 were reviewed. Congenital heart defects (CHDs) were the most common indications (47/76, 61.8%) for prenatal testing and were isolated in 52.6% (40/76). The constitution of CHDs comprised predominantly of conotruncal defects (61.7%; 29/47). Other cardiac anomalies were encountered in 38.3% (18/47) of cases. Extracardiac findings, including unilateral multicystic dysplastic kidney, clubfoot, increased nuchal translucency, intrauterine growth retardation and polyhydramnios, were found in 31.6% (24/76) of cases, and were combined with CHDs in 7 cases. Twelve cases had normal sonographic scans at the time of prenatal diagnosis. Foetal CHDs, especially conotruncal defects, are the most predictive association with del22q11.2. The information about del22q11.2 should also be part of the contents in comprehensive pre-test counselling even for those who are referred for diagnostic testing with foetal extracardiac findings. Impact statement What is already known on this subject? 22q11.2 deletion (del22q11.2) is the most common microdeletion syndrome in humans. At present, the main indications for prenatal testing for del22q11.2 are pregnancies of abnormal sonographic findings, especially foetal congenital heart defects. What do the results of this study add? Many extracardiac malformations, including some lethal or mildly non-specific ones, could be associated with foetal del22q11.2. There were also del22q11.2 foetuses had normal sonographic scans at the time of prenatal diagnosis. What are the implications of these findings for clinical practice and/or further research? The information about del22q11.2 should also be part of the contents in comprehensive pre-test counselling even for those who are referred for diagnostic testing with indications other than foetal cardiac anomalies.

To explore the clinical phenotype, pregnancy outcome and follow-up of fetuses with 15q11.2BP1-BP2 microdeletions in order to provide a basis for prenatal and reproductive consultation. From March 2019 to December 2023, 20 fetuses who were diagnosed with 15q11.2BP1-BP2 microdeletion syndrome at the Prenatal Diagnosis Center of General Hospital of Ningxia Medical University were selected as the study subjects. Results of genetic testing and ultrasound examination, outcome of pregnancy, and postnatal follow-up were retrospectively analyzed. This study has been approved by the Ethics Committee of General Hospital of Ningxia Medical University ([2020]0520B). None of the 20 fetuses was found to have chromosomal abnormality, whilst all were found to harbor a 15q11.2 BP1-BP2 microdeletion by low-depth whole genome sequencing (CNV-seq). The range of deletions was determined as 0.26 ~ 0.87 Mb, and all were rated as pathogenic CNVs. Three fetuses had abnormal ultrasound findings, including 1 with widened renal pelvis, 1 with agenesis of corpus callosum, and 1 with nuchal fold thickening. Parental verification in 10 couples verified that two fetal deletions were de novo, whilst the remaining eight were inherited from a phenotypically normal parent. Following genetic counseling, three couples had opted to terminate the pregnancy, whilst the remaining 17 had continued with the pregnancy until delivery. The 17 liveborns were followed up for 2 months to 5 years, with no obvious abnormality in growth and development noted. CNV-seq plays an important role in the prenatal diagnosis of 15q11.2 BP1-BP2 microdeletions. Such deletions may not always lead to disease phenotypes. Individualized consultation and long-term follow-up, in combination with intrauterine ultrasound and parental verification are necessary.