OBJECTIVE: To analyze the clinical and genetic characteristics of a child with restricted cardiomyopathy (RCM) and phenylketonuria (PKU), and summarize the clinical characteristics and genetic diversity of RCM in children through a literature review. METHODS: A child with RCM in conjunct with PKU who was admitted to the Children's Hospital Affiliated to Zhengzhou University in June 2020 due to edema of eyelids and lower limbs for 1 year and aggravation for over 1 month was selected as the study subject. Relevant clinical data were collected. Peripheral blood samples of the child and his parents were collected for whole exome sequencing (WES). Candidate variants were validated by Sanger sequencing and bioinformatic analysis. Childhood, TNNI3 gene and restricted cardiomyopathy were used as the keywords to search the Wanfang data knowledge service platform, Chinese Journal Full-text database and PubMed database, and the search period was limited to from the time of establishment till August 2022. Clinical manifestations and characteristics of the TNNI3 gene variants were summarized. RESULTS: The child, a 2-year-old-and-4-month-old male, had normal intelligence, facial features and normal hair and skin color, but his motor and physical development was delayed, in addition with edema of bilateral eyelids and lower limbs. The results of WES and Sanger sequencing revealed that he has harbored compound heterozygous variants of the PAH gene, namely c.331C>T (p.R111X) and c.940C>A (p.P341T), which were inherited from his father and mother, respectively. In addition, he has also harbored a de novo heterozygous variant of c.508C>T (p.R170W) of the TNNI3 gene. Based on the guidelines from the American College of Medical Genetics and Genomics (ACMG), the TNNI3: c.508C>T (p.R170W) was classified as a pathogenic variant (PS2+PS4+PM2_Supporting+PM5), PAH: c.331C>T (p.R111X) as a pathogenic variant (PVS1+PM2_Supporting+PM3+PP4), and c.940C>A (p.P341T) as a likely pathogenic variant (PM2_Supporting+PM3+PM5+PP4). In total 30 children with RCM caused by TNNI3 gene variants were retrieved, with a male-to-female ratio of 1 : 1.55 and manifestations including heart failure, sinus rhythm, bi-atrial enlargement, ST-T wave change, ventricular restricted filling, and decreased ventricular diastolic function. In total 16 variants of the TNNI3 gene were identified, among which c.575G>A was the most common, and all cases had conformed to an autosomal dominant inheritance. CONCLUSION Phenylalanine hydroxylase deficiency and RCM are rare diseases with complex clinical manifestations. The PAH: c.331C>T (p.R111X)/c.940C>A (p.P341T) and TNNI3: c.508C>T (p.R170W) variants probably underlay the RCM and PKU in this child.
Humans ; Male ; Cardiomyopathy, Restrictive ; Computational Biology ; Diastole ; Mutation ; Phenylketonurias ; Child, Preschool

OBJECTIVES: To analyze the results of neonatal screening for congenital hypothyroidism (CH) and hyperphenylalaninemia (HPA) in Zhejiang province from 1999 to 2022. METHODS: A total of 11 922 318 newborns were screened from September 1999 and December 2022 in Zhejiang province. The blood thyroid stimulating hormone (TSH) levels were measured by a fluorescence method and blood phenylalanine (Phe) levels were measured by fluorescence method or tandem mass spectrometry. TSH≥9 μIU/mL was considered positive for CH, while Phe>120 μmol/L and/or Phe/Tyr ratio>2.0 were considered positive for HPA. The positive newborns in screening were recalled, and the gene variations were detected by high-throughput sequencing and MassARRAY tests. RESULTS: The overall neonatal screening rate during 1999-2022 was 89.41% (11 922 318/13 333 929) and the screening rate was increased from 6.46% in 1999 to 100.0% in 2022. A total of 8924 cases of CH were diagnosed among screened newborns with an incidence rate of 1/1336. A total of 563 cases of HPA were diagnosed, including 508 cases of classic phenylketonuria (cPKU) and 55 cases of tetrahydrobiopterin deficiency (BH4D), with an incidence rate of 1/21 176. Ninety-seven out of 8924 cases of CH underwent genetic analysis. Gene mutations were detected in 9 CH related genes, the highest frequency mutations were found in DUOX2 gene (69.0%) with c.3329G>A (p.R1110Q) (18.2%) and c.1588A>T (p.K530X) (17.3%) as the hotspot mutations. There were 81 PAH gene variants detected in a total of 250 cases of cPKU, and c728G>A (p.R243Q) (24.4%), c.721C>T (p.R241C) (15.0%) were the hotspot mutations. Meanwhile 7 novel variants in PAH gene were detected: c.107C>A (p.S36*), c.137G>T (p.G46V), c.148A>G(p.K50E), c.285C>T (p.I95I), c.843-10delTTCC, exon4-7del and c.1066-2A>G. There were 12 PTS gene variants detected in 36 cases of BH4D, and c.259C>T (p.P87S) (31.9%) was the hotspot mutation. CONCLUSIONS The incident of CH has increased from 1999 to 2022 in Zhejiang province, and it is higher than that of national and global levels; while the incidence of HPA is similar to the national average. DUOX2 gene variation is the most common in CH patients; c.728G>A (p.R243Q) is the hotspot mutation in cPKU patients, while c.259C>T (p.P87S) is the hotspot mutation in BH4D patients.
Humans ; Infant, Newborn ; Neonatal Screening ; Dual Oxidases ; Congenital Hypothyroidism/genetics* ; Phenylketonurias/genetics* ; Thyrotropin

OBJECTIVES: To retrospectively analyze the variation and characteristics of phenylalanine hydroxylase (PAH) gene, and to observe the long-term treatment effect and follow-up of newborns with PAH deficiency. METHODS: Clinical data, treatment and follow-up results of 198 patients with PAH deficiency diagnosed by newborn screening in Jinan from 1996 to 2021 were collected. The genetic analysis of 55 patients with PAH deficiency diagnosed by newborn screening in Jinan and 213 patients referred from the surrounding areas of Jinan were summarized. Gene variations were checked by a customized Panel gene detection method. Blood phenylalanine-concentration and physical development indicators including height and weight were regularly monitored. Intellectual development was assessed using a neuropsychological development scale for patients aged 0-6 years and academic performance, and brain injury in patients was assessed using brain magnetic resonance imaging. RESULTS: c.728G>A, c.158G>A, c.721C>T, c.1068C>A, c.611A>G variations were common in PAH gene. The genotype of c.158G>A variation is compound heterozygous variation, with mainly a mild hyperpheny-lalaninemia. 168 patients with PAH deficiency who were followed-up regularly had normal physical development without dwarfism or malnutrition. Among the 33 preschool patients who underwent mental development assessment, 2 were mentally retarded and the initial treatment age was older than 6 months. Nine patients with an average age of (17.13±2.42) years completed brain magnetic resonance imaging, one case was normal, and 8 cases were abnormal. There were patchy or patchy hyperintense foci near the bilateral lateral ventricles on T₂WI, and the intellectual development was normal. Compared with the other eight patients, the blood phenylalanine concentration of the normal child was better and stably controlled within the ideal range. CONCLUSIONS c.728G>A, c.158G>A, c.721C>T, c.1068C>A, c.611A>G variations were common in PAH gene. After standardized treatment, most patients with PAH deficiency diagnosed by screening can obtain normal growth and intellectual development in adolescence, but there are different degrees of organic lesions in the cerebral white matter.
Child ; Child, Preschool ; Adolescent ; Humans ; Infant, Newborn ; Young Adult ; Adult ; Neonatal Screening ; Follow-Up Studies ; Retrospective Studies ; Phenylketonurias/genetics* ; Phenylalanine Hydroxylase/genetics* ; Phenylalanine/therapeutic use* ; Mutation

Newborn screening (NBS) plays a significant role in reducing the risk of birth defects. NBS in China began in the early 1980s. Under the protection of laws and regulations and the leadership of the national health administration, approved screening centers in public hospitals took the responsibility for publicity, screening, diagnosis, treatment, follow-up and management of birth defects. As of 2022, 31 provinces (autonomous regions and municipalities directly under the central government) have carried out NBS for phenylketonuria, congenital hypothyroidism, and hearing loss, 23 provinces have carried out screening for glucose-6-phosphate dehydrogenase (with a screening rate of 89.24%), and 24 provinces have carried out screening for congenital adrenal cortical hyperplasia (91.45% screening rate). Over the past four decades, screening techniques have evolved from bacterial inhibition, fluorescence analysis, and tandem mass spectrometry for the detection of biochemical markers to genetic testing, which has greatly contributed to the expansion of the types of diseases screened for. The combined use of metabolomics and genomics is currently being explored. Effective management and rigorous quality control of NBS are prerequisites for improving the quality and ensuring the accuracy of screening. The Quality Management System for Newborn Screening System Network (QMS-NBS), established by the National Center for Clinical Laboratories, covers all screening centers and related blood collection agencies. The operation of the QMS-NBS allows the quality and performance of screening to be transparent and measurable, ensuring the quality and efficiency of screening. This article provides an overview of the history of NBS, especially the evolution of policies for the NBS in China, the construction of screening institutions, the number of newborns screened, the incidence rates of screened diseases, the changes in screening technology, the expansion of new diseases screened for, and the quality control of NBS. Overall, the progress in NBS in China has not only benefited from the development and standardization at the technological level, but also benefited from the construction of policies, regulations and ethics.
Infant, Newborn ; Humans ; Neonatal Screening ; Phenylketonurias ; Genetic Testing ; Congenital Hypothyroidism ; China

Consensus ; Humans ; Nutritional Requirements ; Phenylalanine ; blood ; Phenylalanine Hydroxylase ; deficiency ; Phenylketonurias ; diagnosis ; diet therapy ; genetics ; Practice Guidelines as Topic

External quality assessment (EQA) trials of conventional newborn screening tests for phenylketonuria, galactosemia, congenital adrenal hyperplasia, maple syrup urine disease, homocystinuria, and congenital hypothyroidism, as well as extended newborn screening tests using tandem mass spectrometry, were performed twice in 2016 and 2017. A total of 44 specimens in the form of dried blood spots were distributed in each trial to 16 laboratories. The response rate of these laboratories was 100%. The mean, standard deviation, coefficient of variation, median, and cut-offs were evaluated for each analyte in the newborn screening tests. EQA trials for the analyses of methylmalonic acid, vanillylmandelic acid, catecholamines, metanephrines, organic acids, and amino acids were also performed. A well-designed EQA program and continuous education would improve the performance of metabolite testing.
Adrenal Hyperplasia, Congenital ; Amino Acids ; Catecholamines ; Congenital Hypothyroidism ; Education ; Galactosemias ; Homocystinuria ; Humans ; Infant, Newborn ; Korea* ; Maple Syrup Urine Disease ; Mass Screening ; Methylmalonic Acid ; Phenylketonurias ; Tandem Mass Spectrometry ; Vanilmandelic Acid

OBJECTIVE: To explore the characteristics of PAH gene variants among 113 phenylketonuria patients from Henan Province. METHODS: The 13 exons of the PAH gene were subjected to PCR amplification and direct sequencing. Large fragment deletion and duplication of the PAH gene were detected with a multiple ligation-dependent probe amplification (MLPA) assay. RESULTS: In total 195 point variants and 3 large fragment deletions were detected among the 226 alleles, with the detection rates being 86.28% and 1.33%, respectively. Variants of p.Arg243Gln (18.14%), p.Arg111X (6.19%), p.Arg53His (5.31%), EX6-96A>G (5.31%), p.Tyr356X (4.87%) and p.Val399Val (4.42%) were relatively common. Most of the variants were located in exons 7, 11, 3 and 6. Missense variations were most common. Four novel variations were detected, which included c.1016C>A (p.Ser339Tyr), c.1000T>C (p.Cys334Arg), c.1110G>T (p.Glu370Asp), and IVS6+1G>T. CONCLUSION The PAH gene variations in Henan Province have featured extensive allelic heterogeneity and variety.
China ; Exons ; Humans ; Mutation, Missense ; Phenylalanine Hydroxylase ; genetics ; Phenylketonurias ; genetics ; Point Mutation ; Sequence Deletion

OBJECTIVETo explore the characteristics of (PAH) gene mutations among patients with phenylketonuria (PKU) from Linyi area of Shandong Province.

METHODSFor 51 children affected with PKU and their parents, the 13 exons and their flanking intronic sequences of the PAH gene were directly sequenced with Sanger method.

RESULTSPAH gene mutations were detected in all of the 102 alleles of the patients, which included 31 types of mutations. Common mutations included R243Q (17/102, 16.67%), IVS4-1G to A (9/102, 8.82%), R241C (8/102, 7.84%), R111X (8/102, 7.84%), and V399V (8/102, 7.84%). In addition, two novel mutations, D101N, 345-347del, have been detected. The 31 types of mutations included missense, nonsense, deletion, and splicing mutations, which were mainly located in exons 7 (29, 28.43%), 11 (18, 17.65%), 3 (16, 15.69%) and 12 (13, 12.75%).

CONCLUSIONMutations of the PAH gene in Linyi region mainly distributed in exons 7, 11, and 3, and the most common mutation were R243Q. Two novel mutations, D101N and 345-347del, have been detected.

Base Sequence ; Child ; Child, Preschool ; China ; Exons ; Female ; Humans ; Infant ; Male ; Molecular Sequence Data ; Mutation ; Phenylalanine Hydroxylase ; genetics ; Phenylketonurias ; enzymology ; genetics

OBJECTIVETo delineate the mutation spectrum of phenylalanine hydroxylase (PAH) gene among patients affected with phenylalanine hydroxylase deficiency (PAHD) in Henan Province of China, and to explore the correlation between the genotype and the phenotype.

METHODSA total of 155 affected children were recruited. Potential mutation of the PAH gene were analyzed by direct sequencing. The genotype-phenotype correlation was analyzed by matching the expected and observed phenotypes.

RESULTSOver 72 mutations and 108 genotypes have been identified. There were 7 homozygous mutations, including 1 case with EX6-96A>G/EX6-96A>G, 1 with R241C/R241C, 1 with R413P/R413P, and 4 with R243Q/R243Q. Among these, 6 patients have presented classic PKU phenotypes, except for a R241C/R241C genotype which has led to mild PKU. In 104 patients carrying compound PAH mutations, 52 were classic, 34 were mild and 39 had mild HPA. Patients who were heterozygous for EX6-96A>G/R241C, R243Q/A434D, EX6-96A>G/R413P and EX6-96A>G/ R241C were found with both the classic PKU and mild PKU phenotypes. Common mutations associated with mild HPA have included R53H, R243Q, V399V and H107R. The common mutations associated with mild PKU included R243Q, R241C, EX6-96A>G, and IVS4-1G>A. The prevalent mutations in classic PKU were R243Q, EX6-96A>G and V399V. The consistency between prediction of the biochemical genotype and observed phenotype was 77.78%, especially in classic PKU, the consistency was up to 82.14%. Significant correlations were disclosed between pretreatment levels of phenylalanine and AV sum (r=-0.6729, P < 0.01).

CONCLUSIONThe mutation spectrum of PAH gene in Henan seems to differ from that of other regions. Independent assortment of mutant alleles may result in a complex genotype-phenotype correlation, but the genotypes of PAHD patients have correlated with the phenotype.

Female ; Genotype ; Humans ; Male ; Mutation ; Phenotype ; Phenylalanine Hydroxylase ; genetics ; Phenylketonurias ; genetics

Two external quality assessment (EQA) trials of conventional newborn screening tests for phenylketonuria, galactosemia, congenital adrenal hyperplasia, maple syrup urine disease, homocystinuria, and congenital hypothyroidism, as well as newborn screening tests using tandem mass spectrometry, were performed in 2015. A total of 44 specimens in the form of dried blood spots were distributed to 16 laboratories and the response rate of these laboratories was 100%. The mean, standard deviation, coefficient of variation, median, and cut-offs were evaluated for each analyte in the newborn screening tests. Two EQA trials for the analyses of methylmalonic acid, vanillylmandelic acid, catecholamines, metanephrines, organic acids, and amino acids were also performed. A well-designed EQA program and continuous education would improve the performance of biochemical genetics tests.
Adrenal Hyperplasia, Congenital ; Amino Acids ; Catecholamines ; Congenital Hypothyroidism ; Education ; Galactosemias ; Homocystinuria ; Humans ; Infant, Newborn ; Korea* ; Maple Syrup Urine Disease ; Mass Screening ; Methylmalonic Acid ; Molecular Biology* ; Phenylketonurias ; Tandem Mass Spectrometry ; Vanilmandelic Acid