Fetal phenotype of CHARGE syndrome with a molecular confirmation: a series of 13 cases.

Introduction CHARGE syndrome is an autosomal dominant genetic disorder with known pattern of features. The aim of the study was to present the fetal features of CHARGE syndrome to gain awareness that the antenatal characteristics can be very nonspecific. Case Presentation This was a retrospective study of 13 cases with CHARGE syndrome diagnosed by prenatal or postnatal genetic testing and physical examination. Two (15.4%; 2/13) had normal ultrasound scans during pregnancy. One (7.7%; 1/13) with first-trimester cystic hygroma presented intrauterine fetal demise (IUFD) at 16 weeks gestation. The remaining 10 (76.9%; 10/13) cases had abnormal ultrasound features in utero; among these, 1 had an increased nuchal translucency in the first trimester, 5 had second-trimester abnormal ultrasounds including micrognathia, cardiac defects and facial defects, and 4 third-trimester abnormal ultrasounds including micrognathia, isolated fetal growth restriction and polyhydramnios. Among the 11 cases with abnormal prenatal ultrasound scans, no fetus could reach the diagnostic criteria of CHARGE syndrome if only based on the results of ultrasound. However, the diagnosis was made in all cases when CHD7 defects were detected. Discussion/Conclusion The CHARGE syndrome presents non-specific abnormal ultrasound markers in utero. Exome sequencing in the genetic work up will aid in prenatal diagnosis of this syndrome.

Single-cell RNA sequencing reveals cellular and molecular landscape of fetal cystic hygroma.

BACKGROUND: The molecular mechanism of fetal cystic hygroma (CH) is still unclear, and no study has previously reported the transcriptome changes of single cells in CH. In this study, single-cell transcriptome sequencing (scRNA-seq) was used to investigate the characteristics of cell subsets in the lesion tissues of CH patients. METHODS: Lymphoid tissue collected from CH patients and control donors for scRNA-seq analysis. Differentially expressed gene enrichment in major cell subpopulations as well as cell-cell communication were analyzed. At the same time, the expression and interactions of important VEGF signaling pathway molecules were analyzed, and potential transcription factors that could bind to KDR (VEGFR2) were predicted. RESULTS: The results of scRNA-seq showed that fibroblasts accounted for the largest proportion in the lymphatic lesions of CH patients. There was a significant increase in the proportion of lymphatic endothelial cell subsets between the cases and controls. The VEGF signaling pathway is enriched in lymphatic endothelial cells and participates in the regulation of cell-cell communication between lymphatic endothelial cells and other cells. The key regulatory gene KDR in the VEGF signaling pathway is highly expressed in CH patients and interacts with other differentially expressed EDN1, TAGLN, and CLDN5 Finally, we found that STAT1 could bind to the KDR promoter region, which may play an important role in promoting KDR up-regulation. CONCLUSION: Our comprehensive delineation of the cellular composition in tumor tissues of CH patients using single-cell RNA-sequencing identified the enrichment of lymphatic endothelial cells in CH and highlighted the activation of the VEGF signaling pathway in lymphoid endothelial cells as a potential modulator. The molecular and cellular pathogenesis of fetal cystic hygroma (CH) remains largely unknown. This study examined the distribution and gene expression signature of each cell subpopulation and the possible role of VEGF signaling in lymphatic endothelial cells in regulating the progression of CH by single-cell transcriptome sequencing. The enrichment of lymphatic endothelial cells in CH and the activation of the VEGF signaling pathway in lymphatic endothelial cells provide some clues to the pathogenesis of CH from the perspective of cell subpopulations.

Early Onset of Severe Anemia Caused by Hb Calgary (HBB: C.194G > T): Another Case Report.

Unstable hemoglobin (Hb) variants are a rare cause of congenital hemolytic anemia. We describe a Chinese girl who presented with transfusion-dependent anemia in early infancy. Her diagnosis of Hb Calgary [ß64(E8)Gly > Val; HBB:c.194G > T] was not made until molecular testing was performed at the age of 5 years. Our case highlights the importance of early genetic testing in order to make the diagnosis, which may not only be useful for patient management and family counseling, but also for avoiding further unnecessary investigative attempts.

A 6-Year Follow-up of a Chinese Child with Homozygous ß0-Thalaasemia and a Heterozygous KLF1 Mutation.

Patients with the genotype of ß0/ß0 for ß-thalassemia (ß-thal) usually behave as ß-thal major (ß-TM) phenotype which is transfusion-dependent. The pathophysiology of ß-thal is the imbalance between α/ß-globin chains. The degree of α/ß-globin imbalance can be reduced by the more effective synthesis of γ-globin chains, and increased Hb F levels, modifying clinical severity of ß-TM. We report a Chinese child who had homozygous ß0-thal and a heterozygous KLF1 mutation. The patient had a moderate anemia since 6 months old, keeping a baseline Hb value of 8.0-9.0 g/dL. She had normal development except for a short stature (3rd percentile) until 6 years old, when splenomegaly and facial bone deformities occurred. Although genetic alteration of KLF1 expression in ß0/ß0 patients can result in some degree of disease alleviation, our case shows that it is insufficient to ameliorate satisfactorily the presentation. This point should be borne in mind for physicians who provide the genetic counseling and prenatal diagnosis to at-risk families.

Prenatal diagnosis and outcomes in fetuses with duplex kidney.

OBJECTIVE: Duplex kidney is a relatively frequent form of urinary system abnormality. This study aimed to elucidate the value of chromosomal microarray analysis (CMA) and whole exome sequencing (WES) for duplex kidney and the perinatal outcomes of duplex kidney fetuses. METHODS: This retrospective cohort study included 63 patients with duplex kidney diagnosed using antenatal ultrasound between August 2013 and January 2023. We reviewed the clinical characteristics, genetic test results, and pregnancy outcomes of the patients. RESULTS: Among the 63 cases based on the inclusion criteria, the CMA detected seven (11.1%) clinically significant variants and nine variants of uncertain significance (VUS), and the pathogenic/likely pathogenic (P/LP) copy number variations (CNVs) in the recurrent region that were associated with prenatal duplex kidney included 17q12, 17p13.3, and 22q11.2. No significant disparity was observed in the CMA detection rate between the unilateral and bilateral groups, or between the isolated and non-isolated groups. WES identified three (50%) P/LP single-gene variants in six fetuses with duplex kidney. We detected the following pathogenic genes in the duplex kidney fetuses: KMT2D, SMPD4, and FANCI. Pregnancy termination in cases where clinically significant variants were detected by genetic testing was different in statistical significance from that in cases with negative results (9/10, 90.0% vs 8/48, 16.7%, P < 0.001). CONCLUSION: This study elucidated the value of CMA and WES for fetal duplex kidney, proving that CMA and WES may be useful tools in prenatal diagnosis and genetic counseling.

Prenatal diagnosis of polycystic renal diseases: diagnostic yield, novel disease-causing variants, and genotype-phenotype correlations.

BACKGROUND: Polycystic renal disease is a frequent congenital anomaly of the kidneys, but research using chromosomal microarray analysis and exome sequencing in fetuses with polycystic renal disease remains sparse, with most studies focusing on the multisystem or genitourinary system. OBJECTIVE: This study aimed to assess the detection rate of detectable genetic causes of fetal polycystic renal disease at different levels, novel disease-causing variants, and genotype-phenotype correlations. STUDY DESIGN: This study included 220 fetal polycystic renal disease cases from January 2014 to June 2022. Cases were divided into the following 3 groups: isolated multicystic dysplastic kidneys, nonisolated multicystic dysplastic kidneys, and suspected polycystic kidney disease group. We reviewed data on maternal demographics, ultrasonographic results, chromosomal microarray analysis/exome sequencing results, and pregnancy outcomes. RESULTS: In our cohort, chromosomal microarray analysis identified 19 (8.6%) fetuses carrying chromosomal abnormalities, and the most common copy number variation was 17q12 microdeletion (7/220; 3.2%). Furthermore, 94 families chose to perform trio-exome sequencing testing, and 21 fetuses (22.3%) were found to harbor pathogenic/likely pathogenic variants. There was a significant difference in the live birth rate among the 3 groups (91/130 vs 46/80 vs 1/10; P<.001). Among 138 live birth cases, 106 (78.5%) underwent postnatal ultrasound review, of which 95 (89.6%) had a consistent prenatal-postnatal ultrasound diagnosis. CONCLUSION: For both isolated and nonisolated polycystic renal disease, our data showed high detection efficiency with both testing tools. The detection of novel pathogenic variants expands the known disease spectrum of polycystic renal disease-associated genes while enriching our understanding of the genotype-phenotype correlation. Therefore, we consider it feasible to perform chromosomal microarray analysis+exome sequencing testing in fetal polycystic renal disease. Moreover, prenatal-postnatal ultrasound concordance was greater, the live birth rate was higher, and prognosis was better when known genetic disorders were excluded, indicating that genetic testing results significantly influenced pregnancy decisions.

Association of prenatal thoracic ultrasound abnormalities with copy number variants at a single Chinese tertiary center.

OBJECTIVE: To systematically evaluate the association of prenatal thoracic ultrasound abnormalities with copy number variants (CNVs). METHODS: Chromosomal microarray (CMA) data and clinical characteristics from fetuses with thoracic ultrasound abnormalities were retrieved and analyzed. RESULTS: Thoracic ultrasound findings were mainly isolated except for fetal pleural effusion (FPE) and pulmonary hypoplasia. The diagnostic yield of CMA for thoracic anomaly was 9.66%, and FPE (17/68, 25%), pulmonary hypoplasia (1/8, 12.5%), and congenital diaphragmatic hernia (CDH) (6/79, 7.59%) indicated relatively high pathogenic/likely pathogenic (P/LP) CNV findings. The detection rate for P/LP CNVs was obviously increased in non-isolated thoracic anomalies (27.91% vs. 1.96%, P < 0.0001), non-isolated FPE (37.78% vs. 0%, P = 0.0007) and non-isolated congenital pulmonary airway malformation (CPAM) (27.27% vs. 0%, P < 0.0001), and significantly different among thoracic anomalies. Additionally, the rate of termination of pregnancy in cases with non-isolated thoracic anomalies (58.49% vs. 12.34%, P < 0.0001) and P/LP CNVs (85.71% vs. 24.15%, P < 0.0001) was obviously increased. CONCLUSION: The present study expanded phenotype spectrums for particular recurrent CNVs. FPE, CDH, and pulmonary hypoplasia indicated relatively high P/LP CNV findings among common thoracic ultrasound abnormalities, CPAM associated with other ultrasound abnormalities increased the incidence of diagnostic CNVs, while bronchopulmonary sequestration might not be associated with positive CNVs. The present data recommended CMA application for cases with prenatal thoracic ultrasound abnormalities, especially non-isolated FPE, non-isolated CPAM, CDH, and pulmonary hypoplasia.

Clinical and molecular analysis of nine fetal cases with clinically significant variants causing nemaline myopathy.

OBJECTIVE: To present the prenatal features and postnatal outcomes of pregnancies with fetal nemaline myopathy (NM). STUDY DESIGN: This was a retrospective study of nine cases with NM diagnosed by prenatal or postnatal clinical features and confirmed by genetic testing. Clinical and laboratory data were collected and reviewed for these cases, including maternal demographics, prenatal sonographic findings, exome sequencing (ES) results, and pregnancy outcomes. RESULTS: All of the nine cases were detected to have NM-causing variants, involving NEB gene in 2 cases, ACTA1 in 3 cases, KLHL40 in 3 cases, and TPM2 in 1 case. Almost all (8/9) had normal first-trimester ultrasound scans except one who had an increased nuchal translucency. Seven (7/9) cases had second-trimester abnormal ultrasounds with fetal akinesia and/or extremity anomalies. Two (2/9) had only third-trimester abnormal ultrasounds with fetal akinesia and polyhydramnios, with one combined with fetal growth restriction. Four pregnancies with a positive prenatal ES were terminated, while five having not receiving prenatal ES continued to term. Only one infant survived 1 year old, and four passed away within 12 months. CONCLUSION: Prenatal ultrasound can detect clues that lead to the diagnosis of NM, such as reduced or absent fetal movements, polyhydramnios and extremity anomalies.

Isolated polyhydramnios: Is a genetic evaluation of value?

OBJECTIVE: To analyze the risk for genetic aberrations and pregnancy outcomes in pregnancies with isolated polyhydramnios. STUDY DESIGN: This was a retrospective study of singleton pregnancies complicated by isolated polyhydramnios that underwent genetic amniocentesis between 2016 and 2021. Clinical and laboratory data were collected and reviewed for these cases, including maternal demographics, prenatal sonographic findings, chromosomal microarray results, and pregnancy outcomes. RESULTS: A total of 94 singleton pregnancies were included. Three (3.2%) cases with chromosomal abnormalities were detected, including 2 case of trisomy 21 and 1 of 22q21.1 microdeletion. One case was diagnosed as Prader-Willi syndrome caused by maternal uniparental disomy of chromosome 15. Perinatal death occurred in 1 case with severe polyhydramnios, and was retrospectively diagnosed as Bartter syndrome. Of the 90 infants survived, two were identified to have single gene disorders after birth by whole exome sequencing. CONCLUSION: We first attempted to determine the value of exome sequencing in pregnancies with isolated polyhydramnios. Our results warrant more studies to evaluate advanced genetic testing technologies used in such pregnancies.

Prenatal genetic diagnosis associated with fetal ventricular septal defect: an assessment based on chromosomal microarray analysis and exome sequencing.

Objective: In the study, we investigated the genetic etiology of the ventricular septal defect (VSD) and comprehensively evaluated the diagnosis rate of prenatal chromosomal microarray analysis (CMA) and exome sequencing (ES) for VSD to provide evidence for genetic counseling. Methods: We carried out chromosomal microarray analysis (CMA) on 468 fetuses with VSD and exome sequencing (ES) on 51 fetuses. Results: In our cohort, 68 (14.5%) VSD fetuses received a genetic diagnosis, including 61 (13.03%, 61/468) cases with chromosomal abnormalities and seven (13.7%, 7/51) cases with gene sequence variants. The detection rate of total pathogenic and likely pathogenic gene variations in the non-isolated VSD group (61/335, 18.2%, 55 by QF-PCR/karyotype/CMA + 6 by ES) was significantly higher than that in the isolated VSD group (7/133, 5.3%, 6 by QF-PCR/karyotype/CMA + 1 by ES, p = 0.000). The most common copy number variation (CNV) was 22q11.2 microdeletion syndrome. Additionally, we found six previously unreported variants, which expanded the variation spectrum of VSD-related genes. Conclusion: In this study, CNVs and sequence variants were found in 13.03% and 13.7% of cases, respectively. ES can be recommended for fetuses with VSD without chromosome abnormalities and pathogenic CNVs, especially those that are combined with other ultrasound abnormalities.

Prenatal diagnosis of autosomal recessive Noonan syndrome associated with biallelic LZTR1 variants presented with thick nuchal translucency and cardiac abnormalities.

Noonan syndrome (NS) is a common clinical variable disease characterized by a number of features, mainly including congenital heart defects, short stature, and a variable degree of developmental delay. This disorder is transmitted mostly in an autosomal dominant manner and is genetically heterogeneous. We report three prenatal cases of LZTR1-related recessive NS. One case had a recurrent cystic hygroma at 13 weeks gestation and the pregnancy was terminated. Two cases had an increased nuchal translucency at 12 weeks' gestation, but a normal second trimester ultrasound; both presented with hypertrophic cardiomyopathy in the third trimester. The two infants were diagnosed with NS after birth. All of the three cases had invasive genetic investigations during pregnancy, and trio exome sequencing revealed biallelic likely pathogenic or pathogenic LZTR1 variants in the fetuses. All parents were LZTR1 variant carriers. Our report further strengthens the association of LZTR1 with an autosomal recessive form of NS. The affected fetuses are more likely to have cardiac anomalies. Clarification of molecular diagnosis has important implications in these families because they carry a 25% recurrence risk.

Prenatal diagnosis and perinatal outcomes of twin pregnancies disharmonious for one fetus with nuchal translucency above the 95th percentile.

OBJECTIVE: To assess prenatal diagnosis and pregnancy outcomes in twin pregnancies where one fetus has nuchal translucency (NT) above the 95th percentile. METHOD: In 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. RESULTS: Karyotype 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. CONCLUSION: In 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.

Molecular spectrum and prevalence of thalassemia investigated by third-generation sequencing in the Dongguan region of Guangdong Province, Southern China.

BACKGROUND: PCR, Sanger sequencing and NGS are often employed for carrier screening of thalassemia but all of these methods have limitations. In this study, we evaluated a new third-generation sequencing-based approach termed comprehensive analysis of thalassemia alleles (CATSA) to explore the prevalence of thalassemia in the Dongguan region of southern China. METHODS: 19,932 subjects were recruited for thalassemia screening and hemoglobin testing was performed for each of them. Routine PCR was performed for all the hemoglobin testing-positive subjects and CATSA was conducted for randomly selected subjects from hemoglobin testing-positive and negative subjects. RESULTS: In the 2716 subjects tested both by PCR and CATSA, 2569 had the same results and 147 had discordant results between the two methods. Sanger sequencing, specially designed PCR and MLPA confirmed the results of CATSA were all correct. In total, CATSA correctly detected 787 subjects with variants while routine PCR correctly detected 640 subjects with variants. CATSA yielded a 5.42% (147 of 2716) increment compared with routine PCR. In the 447 hemoglobin testing-negative subjects, CATSA identified pathogenic variants in 12 subjects. Moreover, CATSA identified a novel deletion (chr16:171262-202032) in the α-globin gene cluster. As a result, the deduced carrier frequency of α-thalassemia,ß-thalassemia and α-/ß-thalassemia was 5.62%, 3.85% and 0.93%, respectively. CONCLUSIONS: Our study demonstrated CATSA was a more comprehensive and precise approach than the routine PCR in a large scale of samples, which is highly beneficial for carrier screening of thalassemia. It provided a broader molecular spectrum of hemoglobinopathies and a better basis for a control program in Dongguan region.