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BACKGROUND: Enoyl-CoA hydratase short-chain 1 (ECHS1) is an enzyme involved in the metabolism of branched chain amino acids and fatty acids. Mutations in the ECHS1 gene lead to mitochondrial short-chain enoyl-CoA hydratase 1 deficiency, resulting in the accumulation of intermediates of valine. This is one of the most common causative genes in mitochondrial diseases. While genetic analysis studies have diagnosed numerous cases with ECHS1 variants, the increasing number of variants of uncertain significance (VUS) in genetic diagnosis is a major problem. METHODS: Here, we constructed an assay system to verify VUS function for ECHS1 gene. A high-throughput assay using ECHS1 knockout cells was performed to index these phenotypes by expressing cDNAs containing VUS. In parallel with the VUS validation system, a genetic analysis of samples from patients with mitochondrial disease was performed. The effect on gene expression in cases was verified by RNA-seq and proteome analysis. RESULTS: The functional validation of VUS identified novel variants causing loss of ECHS1 function. The VUS validation system also revealed the effect of the VUS in the compound heterozygous state and provided a new methodology for variant interpretation. Moreover, we performed multiomics analysis and identified a synonymous substitution p.P163= that results in splicing abnormality. The multiomics analysis complemented the diagnosis of some cases that could not be diagnosed by the VUS validation system. CONCLUSIONS: In summary, this study uncovered new ECHS1 cases based on VUS validation and omics analysis; these analyses are applicable to the functional evaluation of other genes associated with mitochondrial disease.
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Doenças Mitocondriais , Humanos , Fenótipo , Doenças Mitocondriais/diagnóstico , Doenças Mitocondriais/genética , Mutação/genética , Enoil-CoA Hidratase/genética , Enoil-CoA Hidratase/metabolismo , Testes GenéticosRESUMO
The identification of the m.12207G > A variant in MT-TS2, (NC_012920.1:m.12207G > A) was first reported in 2006. The affected individual presented with developmental delay, feeding difficulty, proximal muscle weakness, and lesions within her basal ganglia, with heteroplasmy levels of 92% in muscle and no evidence of maternal inheritance. Herein, we report a case involving a 16-year-old boy with the same pathogenic variation and different phenotype, including sensorineural deafness, epilepsy, and intellectual disability, without diabetes mellitus (DM). His mother and maternal grandmother had similar but milder symptoms with DM. Heteroplasmy levels of the proband in blood, saliva, and urinary sediments were 31.3%, 52.6%, and 73.9%, respectively, while those of his mother were 13.8%, 22.1%, and 29.4%, respectively. The differences in the symptoms might be explained by the different levels of heteroplasmy. To our knowledge, this is the first familial report of the m.12207G > A variant in MT-TS2 that causes DM. The present case showed milder neurological symptoms than did the former report, and suggests the presence of a good phenotype-genotype correlation within this family.
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Pathogenic mitochondrial DNA heteroplasmy has mainly been assessed with bulk sequencing in individuals with mitochondrial disease. However, the distribution of heteroplasmy at the single-cell level in skin fibroblasts obtained from individuals, together with detailed clinical and biochemical information, remains to be investigated. We used the mitochondrial DNA single-cell assay for the transposase-accessible chromatin sequencing method. Skin fibroblasts were obtained from six individuals with mitochondrial disease and pathogenic m.3243A>G variants of differing severity. Different distributions of heteroplasmy at the single-cell level were identified in skin fibroblasts from all six individuals. Four individuals with different outcomes showed similar averaged heteroplasmy rates with normal mitochondrial respiratory chain enzyme activity, while the distribution of single-cell heteroplasmy patterns differed among the individuals. This study showed different heteroplasmy distribution patterns at the single-cell level in individuals with the m.3243A>G variant, who had a similar averaged heteroplasmy rates with normal mitochondrial respiratory chain enzyme activity. Whether such different heteroplasmy distribution patterns explain the different clinical outcomes should be assessed further in future studies. Measuring heteroplasmy of pathogenic mitochondrial DNA variants at the single-cell level could be important in individuals with mitochondrial disease.
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DNA Mitocondrial , Doenças Mitocondriais , Humanos , DNA Mitocondrial/genética , Heteroplasmia , Doenças Mitocondriais/genética , Mitocôndrias/genéticaRESUMO
The m.14453G > A mutation in MT-ND6 has been described in a few patients with mitochondrial encephalomyopathy, lactic acidosis, and stroke-like episodes or Leigh syndrome.However, the clinical spectrum and molecular characteristics are unclear.Here, we present four infantile-onset patients with m.14453G > A-associated Leigh syndrome. All four patients had brainstem lesions with basal ganglia lesions, and two patients had cardiac manifestations. Decreased ND6 protein expression and immunoreactivity were observed in patient-derived samples. There was no clear correlation between heteroplasmy levels and onset age or between heteroplasmy levels and phenotype; however, infantile onset was associated with Leigh syndrome.
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Doença de Leigh , Encefalomiopatias Mitocondriais , DNA Mitocondrial/genética , Heteroplasmia , Humanos , Doença de Leigh/genética , Mutação , ProbabilidadeRESUMO
OBJECTIVE: Neonatal-onset mitochondrial disease has not been fully characterised owing to its heterogeneity. We analysed neonatal-onset mitochondrial disease in Japan to clarify its clinical features, molecular diagnosis and prognosis. DESIGN: Retrospective observational study from January 2004 to March 2020. SETTING: Population based. PATIENTS: Patients (281) with neonatal-onset mitochondrial disease diagnosed by biochemical and genetic approaches. INTERVENTIONS: None. MAIN OUTCOME MEASURES: Disease types, initial symptoms, biochemical findings, molecular diagnosis and prognosis. RESULTS: Of the 281 patients, multisystem mitochondrial disease was found in 194, Leigh syndrome in 26, cardiomyopathy in 38 and hepatopathy in 23 patients. Of the 321 initial symptoms, 236 occurred within 2 days of birth. Using biochemical approaches, 182 patients were diagnosed by mitochondrial respiratory chain enzyme activity rate and 89 by oxygen consumption rate. The remaining 10 patients were diagnosed using a genetic approach. Genetic analysis revealed 69 patients had nuclear DNA variants in 36 genes, 11 of 15 patients had mitochondrial DNA variants in five genes and four patients had single large deletion. The Cox proportional hazards regression analysis showed the effects of Leigh syndrome (HR=0.15, 95% CI 0.04 to 0.63, p=0.010) and molecular diagnosis (HR=1.87, 95% CI 1.18 to 2.96, p=0.008) on survival. CONCLUSIONS: Neonatal-onset mitochondrial disease has a heterogenous aetiology. The number of diagnoses can be increased, and clarity regarding prognosis can be achieved by comprehensive biochemical and molecular analyses using appropriate tissue samples.
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Doença de Leigh , Doenças Mitocondriais , DNA Mitocondrial/genética , Humanos , Recém-Nascido , Doença de Leigh/diagnóstico , Doença de Leigh/genética , Doenças Mitocondriais/diagnóstico , Doenças Mitocondriais/genética , Mutação , PrognósticoRESUMO
INTRODUCTION: Autosomal dominant mitochondrial DNA depletion syndrome (MTDPS-12A) is characterized by severe hypotonia from birth due to a mutation in the adenine nucleotide translocator 1 (ANT1). CASE REPORT: A 4-year-old female patient diagnosed with neonatal-onset mitochondrial disease, who had good cognitive function while receiving antiepileptic treatment, presented with sudden-onset status epilepticus with facial and limb myoclonus persisting for more than 30 min. Subsequently, she developed epileptic encephalopathy. Brain MRI showed progressive ventricular enlargement and marked white matter atrophy. She was unable to perform verbal communication or make eye contact and fingertip movements. She lacked any signs of cardiomyopathy. Sanger sequencing demonstrated a heterozygous de novo mutation of c.239G>A (p.Arg80His) in SLC25A4. Her right quadriceps muscle tissue showed lowered complexes I, III, and IV activities and mitochondria DNA depletion (mitochondria/nuclear DNA: 14.6 ± 2.2%) through the quantitative polymerase chain reaction. She was definitively diagnosed with MTDPS-12A. CONCLUSION: Status epilepticus causes encephalopathy in patients with MTDPS-12A. Reducing the energy requirement on the cardiac muscle and brain may be a treatment strategy for patients with MTDPS-12A. Therefore, seizure management and preventive treatment of status epilepticus are considered to be important for maintaining neurodevelopmental outcomes.
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Translocador 1 do Nucleotídeo Adenina/genética , Encefalopatias , DNA Mitocondrial/genética , Doenças Mitocondriais , Doenças Musculares , Estado Epiléptico , Encefalopatias/diagnóstico , Encefalopatias/etiologia , Pré-Escolar , Feminino , Humanos , Doenças Mitocondriais/complicações , Doenças Mitocondriais/diagnóstico , Doenças Mitocondriais/genética , Doenças Musculares/diagnóstico , Doenças Musculares/etiologia , Estado Epiléptico/diagnóstico , Estado Epiléptico/etiologia , SíndromeRESUMO
BACKGROUND: Cardiomyopathy is a risk factor for poor prognosis in pediatric patients with mitochondrial disease. However, other risk factors including genetic factors related to poor prognosis in mitochondrial disease has yet to be fully elucidated. METHODS AND RESULTS: Between January 2004 and September 2019, we enrolled 223 consecutive pediatric mitochondrial disease patients aged <18 years with a confirmed genetic diagnosis, including 114 with nuclear gene mutations, 89 patients with mitochondrial DNA (mtDNA) point mutations, 11 with mtDNA single large-scale deletions and 9 with chromosomal aberrations. Cardiomyopathy at baseline was observed in 46 patients (21%). Hazard ratios (HR) and 95% confidence intervals (CI) were calculated for all-cause mortality. Over a median follow-up of 36 months (12-77), there were 85 deaths (38%). The overall survival rate was significantly lower in patients with cardiomyopathy than in those without (p < 0.001, log-rank test). By multivariable analysis, left ventricular (LV) hypertrophy (HR = 4.6; 95% CI: 2.8-7.3), neonatal onset (HR = 2.9; 95% CI: 1.8-4.5) and chromosomal aberrations (HR = 2.9; 95% CI: 1.3-6.5) were independent predictors of all-cause mortality. Patients with LV hypertrophy with neonatal onset and/or chromosomal aberrations had higher mortality (100% in 21 patients) than those with LV hypertrophy alone (71% in 14 patients). CONCLUSION: In pediatric patients with mitochondrial disease, cardiomyopathy was common (21%) and was associated with increased mortality. LV hypertrophy, neonatal onset and chromosomal aberrations were independent predictors of all-cause mortality. Prognosis is particularly unfavorable if LV hypertrophy is combined with neonatal onset and/or chromosomal aberrations.
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Cardiomiopatias , Doenças Mitocondriais , Cardiomiopatias/diagnóstico , Cardiomiopatias/genética , Criança , Patrimônio Genético , Humanos , Hipertrofia Ventricular Esquerda , Recém-Nascido , Doenças Mitocondriais/diagnóstico , Doenças Mitocondriais/epidemiologia , Doenças Mitocondriais/genética , Prognóstico , Fatores de RiscoRESUMO
The identification of the m.4412Gâ¯>â¯A MT-TM (mt-tRNAMet) mutation was first reported in 2019. The affected individual presented with childhood-onset seizures and myopathy and bilateral basal ganglia changes, with heteroplasmy levels in muscle as high as 90%. Here, we describe another adult-onset patient with the same mutation and additional phenotypes, including hearing impairment, cerebellar ataxia, progressive dementia, and myopathy. The 10% heteroplasmy level observed in skin fibroblasts from this patient are lower than those in the previously reported patient. Our report suggests possible clinical heterogeneity in patients with mitochondrial tRNA mutations based on heteroplasmy levels.
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Heteroplasmia , Doenças Mitocondriais/genética , Polimorfismo de Nucleotídeo Único , RNA de Transferência de Metionina/genética , Idade de Início , Ataxia Cerebelar/genética , Demência/genética , Feminino , Perda Auditiva/genética , Humanos , Pessoa de Meia-Idade , Doenças Musculares/genética , FenótipoRESUMO
BACKGROUND: In about half of all patients with a suspected monogenic disease, genomic investigations fail to identify the diagnosis. A contributing factor is the difficulty with repetitive regions of the genome, such as those generated by segmental duplications. The ATAD3 locus is one such region, in which recessive deletions and dominant duplications have recently been reported to cause lethal perinatal mitochondrial diseases characterized by pontocerebellar hypoplasia or cardiomyopathy, respectively. METHODS: Whole exome, whole genome and long-read DNA sequencing techniques combined with studies of RNA and quantitative proteomics were used to investigate 17 subjects from 16 unrelated families with suspected mitochondrial disease. FINDINGS: We report six different de novo duplications in the ATAD3 gene locus causing a distinctive presentation including lethal perinatal cardiomyopathy, persistent hyperlactacidemia, and frequently corneal clouding or cataracts and encephalopathy. The recurrent 68 Kb ATAD3 duplications are identifiable from genome and exome sequencing but usually missed by microarrays. The ATAD3 duplications result in the formation of identical chimeric ATAD3A/ATAD3C proteins, altered ATAD3 complexes and a striking reduction in mitochondrial oxidative phosphorylation complex I and its activity in heart tissue. CONCLUSIONS: ATAD3 duplications appear to act in a dominant-negative manner and the de novo inheritance infers a low recurrence risk for families, unlike most pediatric mitochondrial diseases. More than 350 genes underlie mitochondrial diseases. In our experience the ATAD3 locus is now one of the five most common causes of nuclear-encoded pediatric mitochondrial disease but the repetitive nature of the locus means ATAD3 diagnoses may be frequently missed by current genomic strategies. FUNDING: Australian NHMRC, US Department of Defense, Japanese AMED and JSPS agencies, Australian Genomics Health Alliance and Australian Mito Foundation.
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Cardiomiopatias , Insuficiência Cardíaca , Doenças Mitocondriais , ATPases Associadas a Diversas Atividades Celulares/genética , Austrália , Criança , Humanos , Proteínas de Membrana/genética , Doenças Mitocondriais/genética , Proteínas Mitocondriais/genética , Estados UnidosRESUMO
AIMS: Mitochondrial cardiomyopathy (MCM) is difficult to make a definite diagnosis because of various cardiovascular phenotypes and no diagnostic criteria in the pathology examination. We aim to add myocardial pathology to the diagnostic criteria for mitochondrial respiratory chain disorders. METHODS: Quantitative analysis of mitochondria using electron microscopy and immunohistopathological analysis with respiratory chain enzyme antibodies were performed in 11 patients with hypertrophic or restrictive cardiomyopathy who underwent endomyocardial biopsy for possible MCM . Respiratory chain enzymatic assay in biopsied myocardium and genetic studies were also performed in all the subjects to define MCM. RESULTS: Four patients were diagnosed with MCM according to the recent criteria of mitochondrial respiratory chain disorders. Using electron microscopy with quantitative analysis, the volume density of mitochondria within cardiac muscle cells was significantly increased in the MCM group compared with the non-MCM group (p=0.007). Immunohistopathological results were compatible with the result of the respiratory chain enzymatic assay. CONCLUSIONS: Pathological diagnosis of MCM could be confirmed by a quantitative study of electron microscopy and immunohistopathological analysis using the mitochondrial respiratory chain enzyme subunit antibody.
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BACKGROUND: Mitochondrial DNA depletion syndrome (MTDPS) is part of a group of mitochondrial diseases characterized by a reduction in mitochondrial DNA copy number. Most MTDPS is caused by mutations in genes that disrupt deoxyribonucleotide metabolism. METHODS: We performed the whole-exome sequencing of a hepato-encephalopathy patient with MTDPS and functional analyses to determine the clinical significance of the identified variant. RESULTS: Here, whole-exome sequencing of a patient presenting with hepato-encephalopathy and MTDPS identified a novel homozygous frameshift variant, c.13_29del (p.Trp6Profs*71) in MICOS13. MICOS13 (also known as QIL1, MIC13, or C19orf70) is a component of the MICOS complex, which plays crucial roles in the maintenance of cristae junctions at the mitochondrial inner membrane. We found loss of MICOS13 protein and fewer cristae structures in the mitochondria of fibroblasts derived from the patient. Stable expression of a wild-type MICOS13 cDNA in the patients fibroblasts using a lentivirus system rescued mitochondrial respiratory chain complex deficiencies. CONCLUSION: Our findings suggest that the novel c.13_29del (p.Trp6Profs*71) MICOS13 variant causes hepato-encephalopathy with MTDPS. We propose that MICOS13 is classified as the cause of MTDPS.
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Proteínas de Membrana/genética , Miopatias Mitocondriais/genética , Proteínas Mitocondriais/genética , Células Cultivadas , Feminino , Fibroblastos/metabolismo , Fibroblastos/ultraestrutura , Mutação da Fase de Leitura , Homozigoto , Humanos , Lactente , Proteínas de Membrana/metabolismo , Mitocôndrias/metabolismo , Mitocôndrias/ultraestrutura , Miopatias Mitocondriais/patologia , Proteínas Mitocondriais/metabolismoRESUMO
Leigh syndrome is a major phenotype of mitochondrial diseases in children. With new therapeutic options being proposed, assessing the mortality and clinical condition of Leigh syndrome patients is crucial for evaluating therapeutics. As data are scarce in Japan, we analysed the mortality rate and clinical condition of Japanese Leigh syndrome patients that we diagnosed since 2007. Data from 166 Japanese patients diagnosed with Leigh syndrome from 2007 to 2017 were reviewed. Patients' present status, method of ventilation and feeding, and degree of disability as of April 2018 was analysed. Overall, 124 (74.7%) were living, 40 (24.1%) were deceased, and 2 (1.2%) were lost to follow-up. Median age of living patients was 8 years (1-39 years). Median length of disease course was 91 months for living patients and 23.5 months for deceased patients. Nearly 90% of deaths occurred by age 6. Mortality rate of patients with onset before 6 months of age was significantly higher than that of onset after 6 months. All patients with neonatal onset were either deceased or bedridden. MT-ATP6 deficiency caused by m.8993T>G mutation and MT-ND5 deficiency induced a severe form of Leigh syndrome. Patients with NDUFAF6, ECHS1, and SURF1 deficiency had relatively mild symptoms and better survival. The impact of onset age on prognosis varied across the genetic diagnoses. The clinical condition of many patients was poor; however, few did not require mechanical ventilation or tube-feeding and were not physically dependent. Early disease onset and genetic diagnosis may have prognostic value.
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Doença de Leigh/genética , Doença de Leigh/mortalidade , Adolescente , Adulto , Idade de Início , Criança , Pré-Escolar , DNA/genética , DNA Mitocondrial/genética , Feminino , Humanos , Lactente , Japão/epidemiologia , Estimativa de Kaplan-Meier , Doença de Leigh/diagnóstico , Imageamento por Ressonância Magnética , Masculino , Mutação/genética , Fenótipo , Taxa de Sobrevida , Adulto JovemRESUMO
Homozygosity mapping is a well-known technique to identify runs of homozygous variants that are likely to harbor genes responsible for autosomal recessive disease, but a comparable method for autosomal dominant traits has been lacking. We developed an approach to map dominant disease genes based on heterozygosity frequencies of sequence variants in the immediate vicinity of a dominant trait. We demonstrate through theoretical analysis that DNA variants surrounding an inherited dominant disease variant tend to have increased heterozygosity compared with variants elsewhere in the genome. We confirm existence of this phenomenon in sequence data with known dominant pathogenic variants obtained on family members and in unrelated population controls. A computer-based approach to estimating empirical significance levels associated with our test statistics shows genome-wide p-values smaller than 0.05 for many but not all of the individuals carrying a pathogenic variant.
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Mapeamento Cromossômico/métodos , Biologia Computacional/métodos , Heterozigoto , Predisposição Genética para Doença , Variação Genética , Humanos , Modelos GenéticosRESUMO
AIMS: Cardiac myosin light chain kinase (cMLCK) phosphorylates ventricular myosin regulatory light chain 2 (MLC2v) and regulates sarcomere and cardiomyocyte organization. However, few data exist regarding the relationship between cMLCK mutations and MLC2v phosphorylation, particularly in terms of developing familial dilated cardiomyopathy (DCM) in whom cMLCK gene mutations were identified. The purpose of the present study was to investigate functional consequences of cMLCK mutations in DCM patients. METHODS AND RESULTS: The diagnosis of DCM was based on the patients' history and on echocardiography. We screened cMLCK gene mutations in DCM probands with high resolution melting analysis. Known DCM-causing genes mutations were excluded by exome sequencing of family members. MLC2v phosphorylation was analysed by Phos-tag sodium dodecyl sulfate-polyacrylamide gel electrophoresis assays. We also performed ADP-Glo assays for determining the total amount of adenosine triphosphate used in the kinase reaction. Unrelated DCM probands (109 males and 40 females) were enrolled in this study, of which 16 were familial and 133 sporadic. By mutation screening, a truncation variant of c1915-1 g>t (p.Pro639Valfs*15) was identified, which was not detected in 400 chromosomes of 200 healthy volunteers; it is listed in the Human Genetic Variation Database with an allele frequency < 0.001. In the proband, the presence of mutations in known DCM-causing genes was excluded with exome analysis. Familial analysis identified a 19-year-old male carrier who manifested slight left ventricular dilation with preserved systolic function. Phosphorylation assays analysed by Phos-tag SDS-PAGE revealed that the identified p.Pro639Valfs*15 mutation results in a complete lack of kinase activity, although it did not affect wild-type cMLCK activity. ADP-Glo assays confirmed that the mutant cMLCK had no kinase activity, whereas wild-type cMLCK had a Km value of 5.93 ± 1.47 µM and a Vmax of 1.28 ± 0.03 mol/min/mol kinase. CONCLUSIONS: These results demonstrate that a truncation mutation in the cMLCK gene p.Pro639Valfs*15 can be associated with significant impairment of MLC2v phosphorylation and possibly with development of DCM, although a larger study of DCM patients is required to determine the prevalence of this mutation and further strengthen its association with disease development.
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Cardiomiopatia Dilatada/genética , DNA/genética , Ventrículos do Coração/metabolismo , Mutação , Miócitos Cardíacos/metabolismo , Quinase de Cadeia Leve de Miosina/genética , Adulto , Cardiomiopatia Dilatada/metabolismo , Cardiomiopatia Dilatada/fisiopatologia , Análise Mutacional de DNA , Ecocardiografia , Feminino , Ventrículos do Coração/diagnóstico por imagem , Humanos , Masculino , Pessoa de Meia-Idade , Contração Miocárdica/fisiologia , Miócitos Cardíacos/patologia , Quinase de Cadeia Leve de Miosina/metabolismo , Linhagem , Sarcômeros/metabolismo , Sarcômeros/patologia , Adulto JovemRESUMO
BACKGROUND: Cardiomyopathy is a reported indicator of poor prognosis in children with mitochondrial disease. However, the association between prognosis and the genetic background of cardiomyopathy in children with mitochondrial disease has yet to be fully elucidated. METHODS AND RESULTS: Of 137 children with mitochondrial disease whose genetic diagnosis was made between 2004 and 2018, 29 had mitochondrial cardiomyopathy (21%). After a median follow-up of 35â¯months, the overall survival rate was significantly lower in patients with cardiomyopathy than in those without (pâ¯<â¯0.001). Ten-year Kaplan-Meier estimates of overall survival were 18 and 67%, respectively. Among the 21 cardiomyopathy patients who died, two died within one month of birth (COQ4 in one patient, and COX10 in one patient), ten died within one year (BOLA3 in three patients, QRSL1 in two patients, large chromosomal deletions in two patients, MT-ATP6/8 in one patient, MT-TL1 in one patient, and TAZ gene in one patient), and nine died after one year (MT-ND5 in three patients, MT-TL1 in three patients, ACAD9 in one patient, KARS in one patient, and MT-TV in one patient). In the three patients with mitochondrial DNA mutations whose cardiac tissues were available, high heteroplasmy rates in the cardiac tissue were observed for m.8528T>C (90%, died at 2â¯months of age) and m.3243A>G (90 and 80%, died at 12 and 13â¯years of age, respectively). CONCLUSIONS: In children with mitochondrial disease, cardiomyopathy was common (21%) and was associated with increased mortality. Genetic analysis coupled with detailed phenotyping could be useful for prognosis.
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Cardiomiopatias/diagnóstico , Cardiomiopatias/genética , DNA Mitocondrial/genética , Patrimônio Genético , Doenças Mitocondriais/diagnóstico , Doenças Mitocondriais/genética , Cardiomiopatias/epidemiologia , Criança , Pré-Escolar , Feminino , Humanos , Lactente , Recém-Nascido , Masculino , Doenças Mitocondriais/epidemiologia , Mutação/genética , PrognósticoRESUMO
The diagnosis of Barth syndrome is challenging owing to the wide phenotypic spectrum with allelic heterogeneity. Here we report 3 cases of Barth syndrome with phenotypic and allelic heterogeneity that were diagnosed by different approaches, including whole exome sequencing and final confirmation by reverse-transcription polymease chain reaction.
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Síndrome de Barth/diagnóstico , Fatores de Transcrição/genética , Aciltransferases , Síndrome de Barth/genética , Humanos , Lactente , Recém-Nascido , Masculino , Mutação , Fenótipo , Reação em Cadeia da Polimerase Via Transcriptase Reversa/métodos , Sequenciamento do Exoma/métodosRESUMO
High-density oligonucleotide arrays have widely been used to detect pathogenic chromosomal deletions. In addition to high-density oligonucleotide arrays, programs using whole-exome sequencing have become available for estimating copy-number variations using depth of coverage. Here, we propose a new statistical method, HDR-del, to prioritize pathogenic chromosomal deletions based on Hamming distance in exome sequencing. In vcf (variant call format) files generated from exome sequencing, hemizygous chromosomal deletion regions lack heterozygous variants and lead to apparent long runs of homozygosity (ROH). In our Hamming distance ratio (HDR)-del approach, we calculate the "difference" in heterozygous status between an affected individual and control individuals using the HDR over all candidate chromosomal deletion regions defined as ROH longer than 1Mbp. Using a suitable test statistic, which is expected to be large for a true pathogenic deletion region, we prioritize candidate chromosomal deletion regions based on this statistic. In our approach, we were able to considerably narrow down true pathogenic chromosomal deletion regions, which were confirmed by high-density oligonucleotide arrays in four mitochondrial disease patients. Our HDR-del approach represents an easy method for detecting chromosomal deletions.