Progress of research on clinical use of non-invasive prenatal screening for special groups of pregnant women / 中华医学遗传学杂志

As a prenatal testing for chromosomal abnormalities, non-invasive prenatal testing (NIPT) has been integrated into prenatal healthcare service. NIPT has shown a high sensitivity and specificity for screening fetal trisomies 13, 18 and 21, and has attained excellent clinical results. With the propagation of the NIPT screening, international organizations have issued guidelines and comments for its clinical utility with regular updating. China has also developed guidelines for NIPT in 2016. NIPT guidelines in various countries have provided valuable guidance for its target diseases and suitable patient groups, but there has been few research data on its clinical application for special groups of patients. Based on the guidelines and comments of various professional bodies and published data on the clinical utility of NIPT, in addition with consideration of the conditions in China, clinical utility of NIPT for particular groups of pregnant women, including those with advanced maternal age, obesity, twin pregnancy and fetal ultrasonographic anomalies, are reviewed. The value of genetic counseling for NIPT is also emphasized, which is critical for the clinical application of NIPT.

Identification of a novel c.1A>G variant of GDAP1 gene in a pedigree affected with autosomal recessive fibula atrophy / 中华医学遗传学杂志

OBJECTIVE@#To explore the genetic basis for a pedigree affected with Charcot-Marie-Tooth (CMT) disease through high-throughput sequencing.@*METHODS@#Potential variants of the genes associated with CMT were screened by next-generation sequencing (NGS) of the members of the pedigree.@*RESULTS@#NGS has revealed that the two affected sisters both harbored homozygous c.1A>G variant of the GDAP1 gene, which caused replacement of the first amino acid Methionine by Valine (p.Met1Val). Their parents were both carriers of the heterozygous c.1A>G variant. The variant was unreported previously and has an extremely low frequency in the population. Meanwhile, one of the sisters and the mother also carried heterozygous c.710A>T variant of the BAG3 gene.@*CONCLUSION@#The homozygous c.1A>G variant of the GDAP1 gene probably underlay the CMT in both children. Above result has enabled clinical diagnosis and genetic counseling for this pedigree.

Identification of a novel PAX6 mutation in a sporadic case with congenital aniridia / 中华医学遗传学杂志

OBJECTIVE@#To identify mutation of the PAX6 gene in a patient with congenital aniridia.@*METHODS@#DNA was extracted from peripheral blood sample of the patient and analyzed by direct PCR-Sanger sequencing.@*RESULTS@#The proband was found to harbor a heterozygous c.239T>A (p.Ile80Asn) mutation of the PAX6 gene. The same mutation was not found in his parents and 150 healthy controls.@*CONCLUSION@#A novel mutation of the PAX6 gene has been identified in a sporadic case with congenital aniridia.

Prenatal diagnosis and PAH gene mutations in 55 pedigrees with phenylketonuria / 中华围产医学杂志

Objective To investigate phenylalanine hydroxylase ( PA H ) gene mutations and to perform prenatal diagnosis in 55 pedigrees with classical phenylketonuria (PKU). Methods Subjects of this study were 55 probands diagnosed with PKU in the Gansu Provincial Maternal and Child Health Care Hospital from 2013 to 2017 and their pedigrees. Sanger sequencing/Multiplex ligation-dependent probe amplification (MLPA) was used to investigate PA H gene mutations in these probands and their parents. Sanger sequencing/MLPA, linkage analysis of three common short tandem repeats (STR) including PAH-26, PAH-STR and PAH-32 in the PA H gene and paternity testing were used in combination for prenatal diagnosis of 60 fetuses in the 55 pedigrees. Results Among the 110 alleles in the 55 probands, 108 mutant alleles (98.2%) were found by Sanger sequencing. The 108 mutant alleles located in 38 regions resulting in 22 missense mutations, nine splice site mutations, five nonsense mutations and two microdeletion. The most common mutations were c.728G>A (22.2%, 24/108), c.442-1G>A (5.6%, 6/108), c.611A>G (5.6%, 6/108), c.764T>C (5.6%, 6/108), c.1068C>A (5.6%, 6/108) and c.331C>T (4.6%, 5/108). Loss of heterozygosity in 4-5 and 4-7 exons were detected by MLPA in two probands, in which only one mutation was unidentified. Prenatal diagnosis for the 60 fetuses were successfully performed. Among them, 17 fetuses (28.3%) were affected, 29 fetuses (48.3%) were heterozygous carriers and fetuses 14(23.4%) were unaffected ones. Conclusions Combination of Sanger sequencing/MLPA, linkage analysis and paternity testing could provide accurate prenatal diagnosis in pedigrees with PKU.

Analysis of SOX10 gene mutation in a family affected with Waardenburg syndrome type II / 中华医学遗传学杂志

OBJECTIVE To detect potential mutation of SOX10 gene in a pedigree affected with Warrdenburg syndrome type II. METHODS Genomic DNA was extracted from peripheral blood samples of the proband and his family members. Exons and flanking sequences of MITF, PAX3, SOX10, SNAI2, END3 and ENDRB genes were analyzed by chip capturing and high throughput sequencing. Suspected mutations were verified with Sanger sequencing. RESULTS A c.127C>T (p.R43X) mutation of the SOX10 gene was detected in the proband, for which both parents showed a wild-type genotype. CONCLUSION The c.127C>T (p.R43X) mutation of SOX10 gene probably underlies the ocular symptoms and hearing loss of the proband.

Prenatal diagnosis and mutation analysis of fibroblast growth factor receptor 3 gene in achondroplasia / 中华围产医学杂志

Objective To explore the value of prenatal genetical diagnosis by mutation analysis of achondroplasia (ACH) fibroblast growth factor receptor 3 (FGFR3) gene.Methods Genomic DNA from nine ACH patients and their parents in Gansu Maternal and Child Health Hospital from July,2010 to December,2014 was prepared for polymerase chain reaction.Direct sequencing revealed the samples were performed after amplification of exon 10 of FGFR3 containing the potential mutation.Fetal DNA was extracted from cells in both amniotic fluid and umbilical cord,and then exon 10 of FGFR3 was also tested.Three fetuses with short-limb dysplasia were also included and prenatal diagnosis was offered to them through amniocentesis or cordocentesis.Results Prenatal ultrasonography test showed shorter femoral length,which was less than 2-3 standard deviation of normal reference dysplasia fetal performance for femoral short.Femur length is lower than 2-3 standard deviation minus normal value,and discrepancy in biparietal diameter compared with fetuses at the same gestational age.In the four families with one ACH parent,c.1138G ＞ A heterozygous mutation was detected in all of the four mothers,while two fetuses among them showed c.1138G ＞ A heterozygous mutation mutation and the other two were normal.There were other two fetuses with c.1138G ＞ A heterozygous mutation from other two families,one's father had c.1138G ＞ A heterozygous mutations,but not the mother,the other had c.1138G ＞ A heterozygous mutations in both the mother and father.Among the three families with unaffected parents but each had a de novo c.1138G ＞ A mutation child,no mutation of c.1138G ＞ A genotype was detected in their fetuses,neither in the three fetus with short limb dysplasia.Four fetuses with a c.1138G ＞ A mutation and three with short-limb dysplasia were terminated.The other five fetuses whose genotype was normal were born and healthy with normal phenotype at one-year-old follow-up.Conclusion FGFR3 genetic analysis could provide information for genetic counseling and prenatal diagnosis for ACH parents or parents who had an ACH baby to prevent birth defect.

Mutational spectrum of phenylalanine hydroxylase gene in Ningxia patients with phenylketonuria / 中华实用儿科临床杂志

Objective To analyze the mutation distribution of phenylalanine hydroxylase(PAH)gene in pa-tients with phenylketonuria(PKU)in Ningxia,and understand the hot spots and regions of PAH gene. Methods First-ly,6 hot mutation exons including 3,5,6,7,11,12 and their surrounding introns of the PAH gene in 30 patients with PKU in Ningxia were directly sequenced. And then the last 7 exons of 1,2,4,8,9,10,13 were sequenced for the patients in which 2 mutant alleles were not characterized. Multiplex ligation - dependent probe amplification (MLPA)was performed for the identification of uncharacterized mutant alleles after PAH sequence analysis of patients with PKU. Results Among 60 alleles,there were 58 mutant alleles(96. 7% ). Forty - six(81. 6% )mutant alleles were found in the exons 3,5,6,7,11,12. A total of 23 various mutations were detected,including missense(n = 9), splicing(n = 9),nonsense(n = 2),small deletion(n = 2)and large deletion(n = 1). The most common mutations were R243Q(18. 3% ),IVS4 - 1G ﹥ A(11. 7% )and R111X(11. 7% ). Among them,a novel mutation N393del was detec-ted in exon 11. MLPA identified a large deletion(c. - 1932 + 3402del)in 3 patients,1 of them was homoallelic muta-tion,and the others were heteroallelic mutation. Conclusions There are obvious hotspots and hot spot areas of PAH gene in PKU patients in Ningxia. There is a large deletion mutation in PKU patients in Ningxia. The MLPA is an effec-tive assay to detect large deletion in PAH gene.

Congenital central hypoventilation syndrome: analysis of PHOX2B gene mutation in a case / 中华医学遗传学杂志

OBJECTIVE To report on the phenotype of an infant with central hypoventilation syndrome (CCHS) and result of PHOX2B gene mutation analysis for the purpose of genetic counseling and prenatal diagnosis. METHODS Clinical data of an infant with CCHS was collected and analyzed. Potential mutation of PHOX2B gene was analyzed by amplified fragment length polymorphism (amp-FLP) and DNA sequencing. RESULTS The patient had typical clinical features of CCHS including frequent hypoventilation during sleeping, hypoxemia and hypercapnia which could be corrected by continuous ventilatory support. She also had repeated bruising and was difficult-to-wean, but without any cardiac, pulmonary, neuromuscular or brainstem lesions. DNA sequencing and amp-FLP of the PHOX2B gene showed that the patient has carried a polyalanine expansion repeat mutation (PARM) in exon 3. A 27 bp duplication was confirmed in the repeat sequence of 20 alanines by cloned and sequenced. This has led to an expansion of the repeat tract to 29 alanines. The genotype was therefore 20/29. CONCLUSION A patient with CCHS has been described. Mutation screening of PHOX2B gene can be used as an important support for diagnosis and genetic counseling for such patients.

Associations between androgen receptor CAG & GGN repeat polymorphism & recurrent spontaneous abortions in Chinese women.

Background & objectives: Recurrent spontaneous abortion (RSA) is a reproductive problem that occurs in women in reproductive age with a frequency of 1-3 per cent. Previous studies have reported high levels of serum androgens to be associated with RSAs. At the molecular level, the effect of androgens is mediated through the activation of the androgen receptor (AR). The CAG and GGN repeat polymorphisms of the AR gene are associated with the AR activity. We hypothesize that the AR CAG/GGN repeat polymorphism may be associated with levels of serum androgens. Thus, this study as undertaken to evaluate the relationship between CAG/GGN repeats in exon 1 of the AR gene in women with RSAs. Methods: This case-control study was performed in Ningxia, PR China, including 149 women with RSAs and 210 controls. The CAG and GGN repeats of the AR gene were genotyped using a PCR-based assay and were analyzed using Peak Scanner Software v1.0 to determine the CAG/GGN repeat length. Results: CAG repeats ranged from 15 to 29 in the RSA patients, compared to 14 to 35 in the control group. The median value of CAG repeats was 22 for the RSA group and 24 for control group. The total AR CAG alleles (≤22 repeats), shorter AR CAG alleles (≤22 repeats), and biallelic means (≤22.5 repeats) were significantly different in the RSA group in comparison to the control group (P<0.001, P<0.01). The median value of the GGN repeats was 23 for the cases and 22 for controls. The total number of AR GGN alleles (≤23 repeats) was significantly different in the RSA group compared to the control group (pP<0.5). There was no difference between the RSA group and the control groups in regards to shorter alleles, longer alleles, and biallelic means. Interpretation & conclusions: Our observation suggests that the CAG and GGN repeat length is shorter in women with RSAs as compared with controls and that shorter CAG and GGN repeats may be pathogenic for RSAs in Chinese women. Further studies need to be done in different ethnic populations.

The mutation analysis of PAH gene and prenatal diagnosis in classical phenylketonuria family / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To characterize the mutation spectrum of phenylalanine hydroxylase (PAH) gene and perform prenatal diagnosis for families with classical phenylketonuria.</p><p><b>METHODS</b>By stratified sequencing, mutations were detected in the exons and flaking introns of PAH gene of 44 families with classical phenylketonuria. 47 fetuses were diagnosed by combined sequencing with linkage analysis of three common short tandem repeats (STR) (PAH-STR, PAH-26 and PAH-32) in the PAH gene.</p><p><b>RESULTS</b>Thirty-one types of mutations were identified. A total of 84 mutations were identified in 88 alleles (95.45%), in which the most common mutation have been R243Q (21.59%), EX6-96A>G (6.82%), IVS4-1G>A (5.86%) and IVS7+2T>A (5.86%). Most mutations were found in exons 3, 5, 6, 7, 11 and 12. The polymorphism information content (PIC) of these three STR markers was 0.71 (PAH-STR), 0.48 (PAH-26) and 0.40 (PAH-32), respectively. Prenatal diagnosis was performed successfully with the combined method in 47 fetuses of 44 classical phenylketonuria families. Among them, 11 (23.4%) were diagnosed as affected, 24 (51.1%) as carriers, and 12 (25.5%) as unaffected.</p><p><b>CONCLUSION</b>Prenatal diagnosis can be achieved efficiently and accurately by stratified sequencing of PAH gene and linkage analysis of STR for classical phenylketonuria families.</p>

Clinical characteristics and genetic analysis in one case of incontinentia pigmenti / 临床儿科杂志

Objective To explore the clinical manifestations and the deletion mutation in NEMO gene in incontinentia pigmenti. Methods The clinical manifestations of one neonatal infant were analyzed. By long PCR ampliifcation, the deletion mutations in NEMO gene and pseudogene ΔNEMO were detected. Results The clinical manifestations were typical skin lesions. Histopathological examination showed focal edema sponge and gathered or scattered eosinophilic granulocytes. The deletion of exons 4-10 in both NEMO andΔNEMO genes were detected in the patient. Conclusions Incontinentia pigmenti is a rare X chromosome linked dominant genetic disease. It has typical clinical manifestations and pathological changes, and deletion mutation in NEMO gene.