Abstract
Maple syrup urine disease (MSUD) is a rare autosomal recessive disorder that affects the degradation of branched chain amino acids (BCAAs). Only a few cases of MSUD have been documented in Mainland China. In this report, 8 patients (4 females and 4 males) with MSUD from 8 unrelated Chinese Han families were diagnosed at the age of 6 days to 4 months. All the coding regions and exon/intron boundaries of BCKDHA, BCDKHB, DBT and DLD genes were analyzed by targeted NGS in the 8 MSUD pedigrees. Targeted NGS revealed 2 pedigrees with MSUD Ia, 5 pedigrees with Ib, 1 pedigree with MSUD II. Totally, 13 variants were detected, including 2 variants (p.Ala216Val and p.Gly281Arg) in BCKDHA gene, 10 variants (p.Gly95Ala, p.Ser171Pro, p.Phe175Leu, p.Arg183Trp, p.Lys222Thr, p.Arg285Ter, p.Arg111Ter, p.S184Pfs*46, p.Arg170Cys, p.I160Ffs*25) in BCKDHB gene, 1 variant (p.Arg431Ter) in DBT gene. In addition, 4 previously unidentified variants (p.Gly281Arg in BCKDHA gene, p.Ser171Pro, p.Gly95Ala and p.Lys222Thr in BCKDHB gene) were identified. NGS plus Sanger sequencing detection is effective and accurate for gene diagnosis. Computational structural modeling indicated that these novel variations probably affect structural stability and considered as likely pathogenic variants.
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Introduction
Maple syrup urine disease (MSUD, OMIM # 248600) is a hereditary branched-chain amino acid metabolism disorder caused by branched chain α-ketoacid dehydrogenase multi-enzyme complex (BCKDC). Common clinical manifestations of MSUD are feeding difficulties, epilepsy, mental retardation, ketonuria and maple-like body odor. Without timely intervention, the disease progresses rapidly, and the mortality and disability rate are very high. According to the phenotype, MSUD can be divided into 5 types1: classic, intermediate, intermittent, thiamine-responsive and dihydrolipoylamide dehydrogenase (E3) deficiency. The classic type is the most common and severe type in the neonatal period, accounting for 75% affected infants. It usually occurs 4–9 days after birth. The liver BCKDC activity in classic type children is often lower than 2% in healthy children, manifested as ketoacidosis, neurologic damage and mental retardation2. 20% are intermediate or intermittent type, the intermediate type usually shows a continuously increased concentration of branched-chain amino acids (BCAAs), accompanied by nervous system damage3. Intermittent type usually occurs from 5 months to 2 years with mild symptoms4, prognosis of the thiamine-responsive type is better than classic type, with BCKDA activity of 2% -40%, and some children can survive for a long time5. The activity of dihydrolipoamide acyl dehydrogenase (E3) -deficient BCKDA is 25% lower than that of normal children, and it is characterized by low tension, stunting, and lactic acidosis6.
According to the Guidelines for the Diagnosis and Treatment of Maple syrup urine disease (2019 Edition, China), MSUD is diagnosed based on the typical clinical manifestations such as nervous system injury and urine maple sugar odor, elevated plasma level of BCAAs (leucine, isoleucine, and valine) and allo-isoleucine and elevated urine level of branched-chain hydroxyacids and ketoacids(BCKAs). Isoleucine > 5 μmol/L detected by plasma amino acid analyzer can be clinically diagnosed as MSUD. Increased isoleucine and alloisoleucine are the gold standard for diagnosis, and 50% of patients have leucine levels exceeding 1500 μmol/L at the time of diagnosis. Determination of BCKAD complex enzyme activity and variation analysis of four genes (BCKDHA, BCKDHB, DBT and DLD) are helpful for definite diagnosis. Tandem mass spectrometry (MSMS) was used to analyze the blood amino acid profile, with leucine, isoleucine and valine levels as the main indicators and leucine/phenylalanine and valine/phenylalanine ratios as the secondary indicators. Those who screened positive were further diagnosed by urine organic acid, plasma amino acid and gene analysis.
MSUD is inherited in autosomal recessive pattern, and it is very rare in most populations, with an incidence of 1:185,0007. BCKDC is located in the mitochondrial inner membrane and consists of 4 subunits: Elα, E1β, E2, E3, which are encoded by BCKDHA, BCKDHB, DBT and DLD genes, respectively8. According to the involved subunit, MSUD is divided into the following types: (1) type Ia (OMIM 608348), caused by biallelic pathogenic variants in BCKDHA gene encoding the Elα subunit (2) type I b (OMIM 248611), caused by biallelic pathogenic variants in BCKDHB gene encoding the E1β subunit (3) type II (OMlM 248610), caused by biallelic pathogenic variants in DBT gene encoding E2 subunit (4) type III (OMIM 238331), caused by biallelic pathogenic variants in DLD gene encoding E3 subunit7. Another two subtypes Type IV and type V are specific kinase and phosphatase gene mutation types, respectively.
MSUD is a genetically heterogeneous disease, and the traditional sequencing technology is time-consuming and costly. High-throughput sequencing technology based on target gene capture for sequencing of the four genes can simultaneously detect gene mutations in the causative genes, not only providing accurate genetic diagnosis results for patients, but also providing clinicians with the basis for differential diagnosis, drug treatment, subsequent genetic counseling, and prenatal diagnosis. In this study, we applied targeted high-throughput sequencing to sequence the target regions of BCKDHA, BCKDHB, DBT and DLD genes in peripheral blood samples of patients or parents in 8 families with MSUD, and Sanger sequencing validation was subsequently performed for confirmation of suspected pathogenic variants.
Methods
Subjects
This is a retrospective study of clinical cases from the First Affiliated Hospital of Zhengzhou University between 2015 and 2020. Eight unrelated families of Chinese Han nationality that had given birth to children affected with MSUD were collected from a single center (Fig. 1). Written informed consent was obtained from the legal guardians. All of the procedures and informed consent were approved by the Medical Ethics Committee of the First Affiliated Hospital of Zhengzhou University (KS-2018-KY-36), and were performed according to the principles of the Declaration of Helsinki.
Blood amino acid and ester acylcarnitine spectra analysis
Urine and venous blood was collected from the children on an empty stomach for more than 4 h. Urinary organic acid analysis was performed using gas chromatography mass spectrometry (GC–MS) and blood amino acid and ester-acylcarnitine profiling was performed using liquid chromatography-tandem mass spectrometry (LC–MS/MS).
DNA extraction
Blood samples (2 ml) were collected from each patient and their parents in families 3 and 7 by venipuncture in EDTA tubes. For the remaining six families, parental blood samples were collected. Genomic DNA was extracted from peripheral blood leukocytes using a DNA Blood Mini Kit (Qiagen, Cat.No.51106, Germany) according to the recommended protocol.
Targeted next-generation sequencing
Targeted genes were chosen according to OMIM database (https://omim.org/) and were designed by the MyGenostics company (Beijing, China). Metabolic disease gene panel was specifically captured and enriched using array-based hybridization chip (NimbleGen, Madison, USA) followed by HiSeq2000 (Illumina, San Diego, USA) sequencing to generate paired read 100 bp according to the manufacturer’s protocol. Then, the final products were amplified by PCR and validated using the Agilent Bioanalyzer. Fastq-format reads were aligned to the human reference genome (GRCh37/hg19, https://genome.ucsc.edu/) using BWA9 software (Burrows Wheeler Aligner). Base quality score recalibration together with SNP and short Indel calling was conducted with GATK 3.810. Quality metrics were evaluated the average depth was 80 × per sample, with at least 97% of the target region covered by 10 × reads or more. The VCF files were then annotated using SnpEff11. Variants with > 1% frequency in the population variant databases-1000 Genomes Project12, Exome Variant Server (EVS, http://evs.gs.washington.edu/EVS/) and Exome Aggregation Consortium (ExAC, http://exac.broadinstitute.org) or > 5% frequency in the local database with 150 exome datasets were filtered, and subsequently intergenic, intronic, and synonymous variants were filtered, except those located at canonical splice sites. By searching the Human Gene Mutation Database (HGMD, http://www.hgmd.org/) to clarify whether the variant is a known pathogenic variant. The nomenclature of new variants was based on the international gene variant nomenclature system (http://www.hgvs.org/mutnomen).
Validation tests of Sanger sequencing
Gene tool software was used for designing primers for suspected variants. Routine PCR reactions were performed. PCR products were purified and directly sequenced on ABl3130-xl gene sequencing instrument using the ABIBigDye3.1 sequencing kit (Thermo Fisher Scientific, USA), and the sequencing data were compared and analyzed using ABI Sequencing Analysis 5.1.1 software.
In silico webservers and structure prediction
Multiple sequence alignments were performed using HomoloGene database (http://www.ncbi.nlm.nih.gov/homologene) to verify the degree of conservation. The pathogenicity of the variants was then evaluated using three in silico webservers, PolyPhen2 (http://genetics.bwh.harvard.edu/pph2/), SIFT (http://provean.jcvi.org/index.php) and Mutation Taster (http://www.mutationtaster.org). The American College of Medical Genetics and Genomics (ACMG) guideline was applied to assess novel variants’ pathogenicity. Computational modeling was carried out to observe the effect of new missense variants on protein structure. Three-dimensional structure of the target protein sequence was constructed using PyMOL protein model structure simulation software to determine the effect of amino acid substitution on protein structure.
Summary of published data
A literature search by using the PubMed and WanFang databases were conducted to identify reported variants in Chinese MSUD patients.The searches were using the Keywords “Maple syrup urine disease (MSUD)” or “BCKDHA” or “BCKDHB” or “DBT” or “DLD” and “Chinese”.
Results
Characteristics of recruited subjects
Between 2015 and 2020, a total of 8 families were collected in our study. The study design is shown in Fig. 1. Characteristics of these cases are shown in Table 1. Only children in family 3 and 7 accepted timely diagnosis and treatment after neonatal screening. All children in the 8 families were screened by tandem mass spectrometry and received positive screening results. As is shown in Table 1, the remaining 6 children developed the disease from 3 days to 4 months, and died at 16 days, 2 months, 20 days, 1 month, 10 days and 1 month, respectively.
Molecular analysis in BCKDHA, BCKDHB and DBT genes
NGS was performed in children of families 3 and 7 and couples of the other 6 MSUD families to detect the sequence variation in each exon of the 4 causative genes (BCKDHA, BCDKHB, DBT, DLD) associated with MSUD. After alignment with the hg19 sequence, the variants were filtered by excluding the SNPs (normal frequency > 0.05) reported in the dbSNP137 database, Hapmap database and 1000 Genome database. Suspected variants in the causative genes associated with the 8 families were shown in Table 2.
Sanger sequencing results
The suspected variants found by NGS were confirmed by Sanger sequencing. The patient in family 3 carried BCKDHB gene c.511T>C(p.Ser171Pro) and c. c.508C>T(p.Arg170Cys) compound heterozygous variants, and the child in pedigree 7 carried c.523T>C(p.Phe175Leu) and c.478-552del(p.I160Ffs*25) compound heterozygous variants. Their parents were heterozygous carriers of the respective variant. Heterozygous variants in the same causative gene of MSUD were detected in both couples in the remaining six families. Gene sequences of four novel variant in BCKDHA and BCKDHB genes were shown in Fig. 2.
In silico prediction of novel gene variant sequences
Mutation Taster and PolyPhen-2 analysis showed that four novel missense variants: p.Gly95Ala, p.Gly281Arg, p.Lys222Thr and p.Ser171Pro were highly likely to be pathogenic/deleterious variants. We use The American College of Medical Genetics and Genomics (ACMG)13 guideline to assess these novel variants’ pathogenicity in Table 2.
Three-dimensional structure of proteins
The predicted three-dimensional structures of 4 novel variants in BCKDHA and BCKDHB genes were shown in Fig. 3. In BCKDHA gene, Glycine 281 is located in the random coil structure of protein secondary structure. Glycine lacks side chain (only one H-bond). After variation to arginine, arginine is a basic amino acid with ions, affecting the stability of E1α tertiary structure, thus affecting protein function.
In BCKDHB gene, As is shown in Fig. 3d, Gly95 is located in the β-turn region, and due to the lack of side chains (only one H atom) by Glycine, there is no steric hindrance, allowing a U-shaped turn of the peptide chain by 180° in the β-turn. Variation of Glycine to Alanine, which forms some steric hindrance, is likely to change the turning of the β-turn, causing change in local steric conformation.
In Fig. 3e, Serine at position 171 is located in the α-helix of protein secondary structure, forming hydrogen bonds with Leucine at position 174, Phenylalanine at position 175 and Tyrosine at position 167. After variation to Proline, the hydrogen bonds with Leucine at position 167 and Phenylalanine at position 175 disappear, affecting the stability of protein secondary structure. Therefore, it is speculated that p.Ser171Pro variant has a greater impact on protein function.
In Fig. 3f, Amino acid 222 is located in the α-helix of the secondary structure of the protein, amino acid 222 forms hydrogen bonds with amino acids 79, 83, 218, 225, 252, and 254. After variation to Threonine, it reforms hydrogen bonds with amino acids 76, 218, and 225. The secondary structure of the protein is changed, which disrupts the stability of the protein and may affect the cleavage and activation function of the protein.
Published data analysis
Collectively, 15 studies were included for analysis of genotype and phenotype (Supplementary Table S1). In the combined data with ours, there were 61 cases in reported Chinese patients, of which 17 (27.9%) cases had BCKDHA gene variations, 35 (57.4%) cases had BCKDHB gene variations, 9 (14.7%) cases carried DBT gene variations. And no patients carried DLD gene variations. According to the clinical manifestations and laboratory tests, Forty-five (73.4%) were diagnosed as classic type (13/BCKDHA, 25/BCKDHB, 7/DBT), 1 the intermittent type (1/BCKDHA), 9 the intermediate type (2/BCKDHA, 7/BCKDHB), 4 the thiamine-responsive type (1/BCKDHA, 1/BCKDHB, 2/DBT). We counted variants with frequency number > 2 in Chinese patients (Table 3).
Discussion
Maple syrup urine diabetes is a branched-chain amino acid metabolism disease caused by mitochondrial branched-chain α-keto acid dehydrogenase (BCKDC) deficiency. Scaini et al.14 suggested that cognitive impairment after accumulation of branched-chain amino acids is mainly due to oxidative damage to the brain. The clinical manifestations of MSUD are lack of specificity with rapid onset. The detection of amino acid levels and the ratio between related amino acids in hemofilter paper by tandem mass spectrometry15 allow for early screening of MSUD and provide an important basis for further diagnosis and treatment. In this study, the results of blood tandem mass spectrometry in all families showed that both leucine and valine were significantly higher, accompanied by amino acid ratio changes, consistent with MSUD biochemical findings.
In our study, a total of 13 variants (15.4% located in the BCKDHA gene, 76.9% in the BCKDHB gene, and 7.7% in the DBT gene, no variants in DLD gene) were identified in 16 alleles in 8 families. In the systematic literature review of MSUD reported in Chinese population, 81 mutations have been detected in 61 patients in China, including the 8 patients in our study. There are 26 (32.1%) gene variants located in the BCKDHA gene, 45 (55.6%) gene variants in the BCKDHB gene, 10 (12.3%) gene variants in the DBT gene, no variants in the DLD gene. The BCKDHB gene may be a major variant type of MSUD in the Chinese population. Gene variations of MSUD patients are mainly concentrated in the BCKDHB gene, followed by BCKDHA and DBT genes16. Current study suggested that DLD gene variants account for 13%17. While our data are inconsistent with this, DLD gene variants may be very rare in Chinese population. MSUD gene has high allelic heterogeneity, with the exception of gene mutation hotspots found in minority of ethnic groups, such as the most common mutation in the Mennonite community being the BCKDHA gene c.1312T>A (p.Tyr393Asn)18, Portuguese gypsy mutation hotspot c.117delC19. The BCKDHB gene c.538G>C was a common mutation found in Ashkenazi Jews20, and exon 5 of the BCKDHB gene may be a region of genetic variation and a hotspot region21,22. Hotspot mutations are not found in the remaining population23,24,25. There were no significant hotspot mutations have been identified in the Chinese population26,27,28,29,30,31,32. Variants c.331C>T and c.853C>T in BCKDHB gene may be relatively common in Chinese patients.
Four variants were novel variants, one located in BCKDHA gene (p. Gly281Arg) and three located in BCKDHB gene (p. Gly95Ala, p.Lys222Thr, p.Ser171Pro) illustrates that the disease has high allelic heterogeneity. Protein structure prediction was carried out for BCKDHA, BCKDHB in order to analyze the variants in a visual way. In BCKDHA, variation p. Gly281Arg is present in coil and minimal change has been discovered in the protein structure. In BCKDHB, p. Gly95Ala is present in the β-turn region, which may change the turning of the β-turn and causing change in local steric conformation. Two variants (p.Lys222Thr and p.Ser171Pro) are present in the α-helix of the secondary structure of the protein and thus have great impact on the protein function. According to the American College of Medical Genetics and Genomics (ACMG) standard and guidelines, the three variants (p.Gly281Arg of BCKDHA, p.Gly95Ala and p.Ser171Pro of BCKDHB) are variants of uncertain significance (PM3 + PP3 + PP4), and p.Lys222Thr of BCKDHB is likely pathogenic (PM3 + PM5 + PP3 + PP4). All the novel variants are predicted to be disease causing by prediction software (PolyPhen2, SIFT and Mutation Taster). In summary, these four novel variant may be causative variants.
The relationship between MSUD genotype and phenotype has not yet been established. The incidence of the disease is low, and fewer cases are included in each study, making it difficult to obtain an exact genotype–phenotype relationship. Current studies suggest that patients with BCKDHA and BCKDHB gene variants mostly present with classical type, BCKDH activity is less than 2%. DBT gene variants accounts for about 24%, and most of them are thiamine responsive type6. The clinical manifestations of patients are relatively mild, including developmental retardation and hypotonia. Patients with nonsense variations presented the severe classic phenotype. Variations in p.Arg111Ter and p.Arg285Ter in BCKDHB gene generate premature termination codons and the encoded protein has serious effects on the activity of the complex33. Our cases in family 5 and 6 carry the nonsense variants p.Arg111Ter and p.Arg285Ter, respectively, and they have classic phenotype. However, the same type of genetic variation also leads to different clinical phenotypes. For example, In BCKDHA gene, p.Glu327Lys has been reported to be associated with intermediate phenotype8 while the same variation results in classic phenotype in patients in another study22. In our study, patient 4 carries the missense variation p.Arg183Trp in the BCKDHB gene and shows intermediate phenotype, while the patient who had the same variation showed classic phenotype in a previous report34. Therefore, we could not establish any genotype–phenotype correlation in our patients with MSUD. Half of our cases are classical phenotypes and half are intermediate phenotypes. Majority of patients with intermediate phenotype had variants in the BCKDHB gene. All the three genes are implicated in classic phenotype. Most of Chinese patients carried DBT gene variations were diagnosed as classic type, while in Norway35, patients had DBT gene variants were intermittent phenotype. All patients with classic phenotype have worst clinical outcome.
MSUD is a fatal and disabling inherited metabolic disease which is difficult to treat, and has a poor prognosis. Untreated classical children mostly die shortly after birth. The principle of treatment is to remove the inducement, reduce the toxic effect of blood leucine, correct acute metabolic disorders, maintain plasma branched-chain amino acids in the ideal range, and ensure good nutrition and growth and development36. MSUD treatment mainly includes acute phase management, dietary management and vitamin B1 treatment. In recent years, liver transplantation for MSUD has been reported37,38. However, shortage of liver sources, high cost, and the need to take immunosuppressive agents for a long time after surgery are disadvantages of this treatment. Currently, the best preventive strategy for the disease is to avoid the birth of affected children through prenatal diagnosis. When MUSD is clinically suspected, capture-based high throughput sequencing followed by Sanger sequencing confirmation allows for accurate detection of gene mutations in the causative genes in an effective manner.
In conclusion, we present the clinical characteristics and 16 variants in 8 patients with MSUD and explore the genotype–phenotype relationship. We identified four pathogenic variants in the BCKDHA and BCKDHB gene by applying high throughput sequencing technology based on target gene capture for sequencing, which have not been previously reported in the Chinese population. This article will contribute to a better understanding of the MSUD variation spectrum identified so far. NGS combined with Sanger sequencing can detect gene variants in the causative genes in an effective way, providing clinicians with the basis for differential diagnosis, drug treatment, subsequent genetic counseling and prenatal diagnosis.
Change history
20 October 2021
A Correction to this paper has been published: https://doi.org/10.1038/s41598-021-00718-4
References
Miryounesi, M., Ghafouri-Fard, S., Goodarzi, H. & Fardaei, M. A New missense mutation in the BCKDHB gene causes the classic form of maple syrup urine disease (MSUD). J. Pediatr. Endocrinol. Metab. 28, 673–675 (2015).
Couce, M. L. et al. Evolution of maple syrup urine disease in patients diagnosed by newborn screening versus late diagnosis. Eur. J. Paediatr. Neurol. 19, 652–659 (2015).
Pode-Shakked, N. et al. Clues and challenges in the diagnosis of intermittent maple syrup urine disease. Eur. J. Med. Genet. 63, 103901 (2020).
Axler, O. & Holmquist, P. Intermittent maple syrup urine disease: Two case reports. Pediatrics 133, e458–e460 (2014).
Scriver, C. R., Clow, C. L. & George, H. So-called thiamin-responsive maple syrup urine disease: 15-year follow-up of the original patient. J. Pediatr. 107, 763 (1985).
Mitsubuchi, H., Owada, M. & Endo, F. Markers associated with inborn errors of metabolism of branched-chain amino acids and their relevance to upper levels of intake in healthy people: An implication from clinical and molecular investigations on maple syrup urine disease. J. Nutr. 135, 1565S-1570S (2005).
Nellis, M. M. et al. Relationship of causative genetic mutations in maple syrup urine disease with their clinical expression. Mol. Genet. Metab. 80, 189–195 (2003).
Rodriguez-Pombo, P., Navarrete, R., Merinero, B., Gomez-Puertas, P. & Ugarte, M. Mutational spectrum of maple syrup urine disease in Spain. Hum. Mutat. 27, 715 (2006).
Li, H. & Durbin, R. Fast and accurate short read alignment with Burrows-Wheeler transform. Bioinformatics 25, 1754 (2009).
McKenna, A. et al. The genome analysis toolkit: A MapReduce framework for analyzing next-generation DNA sequencing data. Genome Res. 20, 1297 (2010).
Cingolani, P. et al. A program for annotating and predicting the effects of single nucleotide polymorphisms, SnpEff: SNPs in the genome of Drosophila melanogaster strain w1118; iso-2; iso-3. Fly 6, 80 (2012).
Abecasis, G. R. et al. A map of human genome variation from population-scale sequencing. Nature 467, 1061 (2010).
Richards, S. et al. Standards and guidelines for the interpretation of sequence variants: A joint consensus recommendation of the American College of Medical Genetics and Genomics and the Association for Molecular Pathology. Genet. Med. 17, 405–424 (2015).
Scaini, G. et al. Antioxidant administration prevents memory impairment in an animal model of maple syrup urine disease. Behav. Brain Res. 231, 92–96 (2012).
Chace, D. H. et al. Rapid diagnosis of maple syrup urine disease in blood spots from newborns by tandem mass spectrometry. Clin. Chem. 41, 62–68 (1995).
Ali, E. Z. & Ngu, L. Fourteen new mutations of BCKDHA, BCKDHB and DBT genes associated with maple syrup urine disease (MSUD) in Malaysian population. Mol. Genet. Metab. Rep. 17, 22–30 (2018).
Georgiou, T. et al. Maple syrup urine disease in cypriot families: Identification of three novel mutations and biochemical characterization of the PThr211Met mutation in the E1alpha subunit. Genet. Test Mol. Biomarkers. 13, 657–664 (2009).
Love-Gregory, L. D., Grasela, J., Hillman, R. E. & Phillips, C. L. Evidence of common ancestry for the maple syrup urine disease (MSUD) Y438N allele in non-mennonite MSUD patients. Mol. Genet. Metab. 75, 79–90 (2002).
Quental, S. et al. Revisiting MSUD in portuguese gypsies: Evidence for a founder mutation and for a mutational hotspot within the BCKDHA gene. Ann. Hum. Genet. 73, 298–303 (2009).
Edelmann, L. et al. Maple syrup urine disease: Identification and carrier-frequency determination of a novel founder mutation in the Ashkenazi Jewish population. Am. J. Hum. Genet. 69, 863–868 (2001).
Su, L. et al. Two homozygous mutations in the exon 5 of BCKDHB gene that may cause the classic form of maple syrup urine disease. Metab. Brain Dis. 32, 765–772 (2017).
Gupta, D. et al. Identification of mutations, genotype-phenotype correlation and prenatal diagnosis of maple syrup urine disease in Indian patients. Eur. J. Med. Genet. 58, 471–478 (2015).
Abiri, M. et al. Identification of six novel mutations in Iranian patients with maple syrup urine disease and their in silico analysis. Mutat. Res. 786, 34–40 (2016).
Imtiaz, F. et al. Twenty novel mutations in BCKDHA, BCKDHB and DBT genes in a cohort of 52 Saudi Arabian patients with maple syrup urine disease. Mol. Genet. Metab. Rep. 11, 17–23 (2017).
Li, X. et al. Clinical, biochemical and genetic features of 13 children with maple syrup urine disease. Chin. J. Appl. Clin. Pediatr. 31, 569–572 (2016).
Yang, N. et al. Analysis of gene mutations in chinese patients with maple syrup urine disease. Mol. Genet. Metab. 106, 412–418 (2012).
Li, X. et al. Eleven novel mutations of the BCKDHA, BCKDHB and DBT genes associated with maple syrup urine disease in the Chinese population: Report on eight cases. Eur. J. Med. Genet. 58, 617–623 (2015).
Li, W. et al. Silico analysis of a novel mutation C550delT in a Chinese patient with maple syrup urine disease. Clin. Case Rep. 6, 1989–1993 (2018).
Liu, G. et al. A novel whole gene deletion of BCKDHB by alu-mediated non-allelic recombination in a chinese patient with maple syrup urine disease. Front. Genet. 9, 145 (2018).
Li, X. et al. Clinical characteristics and mutation analysis of five Chinese patients with maple syrup urine disease. Metab. Brain Dis. 33, 741–751 (2018).
Sun, W. et al. Identification of eight novel mutations in 11 Chinese patients with maple syrup urine disease. World J. Pediatr. 16, 401–410 (2020).
Yang, C., Linpeng, S., Cao, Y. & Wu, L. Identification of six novel mutations in five infants with suspected maple syrup urine disease based on blood and urine metabolism screening. Gene 710, 9–16 (2019).
Flaschker, N. et al. Description of the mutations in 15 subjects with variant forms of maple syrup urine disease. J. Inherit. Metab. Dis. 30, 903–909 (2007).
Abiri, M. et al. Maple syrup urine disease mutation spectrum in a cohort of 40 consanguineous patients and insilico analysis of novel mutations. Metab. Brain Dis. 34, 1145–1156 (2019).
Brodtkorb, E. et al. Four novel mutations identified in norwegian patients result in intermittent maple syrup urine disease when combined with the R301C mutation. Mol. Genet. Metab. 100, 324–332 (2010).
Yang, C. et al. Progress of research on maple syrup disease. Chin. J. Med. Genet. 737, 1–10 (2019).
Chin, H. L. et al. Two consecutive partial liver transplants in a patient with classic maple syrup urine disease. Mol. Genet. Metab. Rep. 4, 49–52 (2015).
Basturk, A. et al. Liver transplantation from a live donor to a patient with maple syrup urine disease: Two case reports. Turk. Pediatr. Ars. 53, 113–116 (2018).
Acknowledgements
We would like to thank our colleagues. Without their referrals this would be incomplete. Last but not least we would like to thank the patients and their families for their sacrifices and giving us their precious blood and urine samples and clinical information. All authors reviewed the manuscript.
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This work was supported financially by National Natural Science Foundation of China (81701533, 81672110) for which we are grateful.
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F.X. and Z.X. wrote the main manuscript text. B.Y. and F.Y. prepared figures. Z.X. prepared table. All authors analyzed data, wrote and approved the manuscript.
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The original online version of this Article was revised: The original version of this Article contained an error in the order of the author names, which was incorrectly given as Xiaohua Fang, Xiaofan Zhu, Yin Feng, Ying Bai, Xuechao Zhao, Xiangdong Kong & Ning Liu.
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Fang, X., Zhu, X., Feng, Y. et al. Genetic analysis by targeted next-generation sequencing and novel variation identification of maple syrup urine disease in Chinese Han population. Sci Rep 11, 18939 (2021). https://doi.org/10.1038/s41598-021-98357-2
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DOI: https://doi.org/10.1038/s41598-021-98357-2
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