A novel mitochondrial DNA variant in MT-ND6: m.14430A>C p.(Trp82Gly) identified in a patient with Leigh syndrome and complex I deficiency.

Leigh syndrome is a severe progressive mitochondrial disorder mainly affecting children under the age of 5 years. It is caused by pathogenic variants in any one of more than 75 known genes in the nuclear or mitochondrial genomes. A 19-week-old male infant presented with lactic acidosis and encephalopathy following a 2-week history of irritability, neuroregression and poor weight gain. He was hypotonic with pathological reflexes, impaired vision, and nystagmus. Brain MRI showed extensive bilateral symmetrical T2 hyperintense lesions in basal ganglia, thalami, and brainstem. Metabolic workup showed elevated serum alanine, and heavy lactic aciduria with increased ketones, fumarate, malate, and alpha-ketoglutarate as well as reduced succinate on urine organic acid analysis. Lactic acidemia persisted, with only a marginally elevated lactate:pyruvate ratio (16.46, ref. 0-10). He demised at age 7 months due to respiratory failure. Exome sequencing followed by virtual gene panel analysis for pyruvate metabolism and mitochondrial defects could not identify any nuclear cause for Leigh syndrome. Mitochondrial DNA (mtDNA) genome sequencing revealed 88% heteroplasmy for a novel variant, NC_012920.1(MT-ND6):m.14430A>C p.(Trp82Gly), in blood DNA. This variant was absent from the unaffected mother's blood, fibroblast, and urine DNA, and detected at a level of 5% in her muscle DNA. Mitochondrial respiratory chain analysis revealed markedly reduced mitochondrial complex I activity in patient fibroblasts (34% of parent and control cells), and reduced NADH-linked respirometry (less than half of parental and control cells), while complex II driven respirometry remained intact. The combined clinical, genetic, and biochemical findings suggest that the novel MT-ND6 variant is the likely cause of Leigh syndrome in this patient. The mitochondrial ND6 protein is a subunit of complex I. An interesting finding was the absence of a significantly elevated lactate:pyruvate ratio in the presence of severe lactatemia, which directed initial diagnostic efforts towards excluding a pyruvate metabolism defect. This case highlights the value of a multidisciplinary approach and complete genetic workup to diagnosing mitochondrial disorders in South African patients.

KCTD7-related progressive myoclonic epilepsy: Report of 42 cases and review of literature.

OBJECTIVE: KCTD7-related progressive myoclonic epilepsy (PME) is a rare autosomal-recessive disorder. This study aimed to describe the clinical details and genetic variants in a large international cohort. METHODS: Families with molecularly confirmed diagnoses of KCTD7-related PME were identified through international collaboration. Furthermore, a systematic review was done to identify previously reported cases. Salient demographic, epilepsy, treatment, genetic testing, electroencephalographic (EEG), and imaging-related variables were collected and summarized. RESULTS: Forty-two patients (36 families) were included. The median age at first seizure was 14 months (interquartile range = 11.75-22.5). Myoclonic seizures were frequently the first seizure type noted (n = 18, 43.9%). EEG and brain magnetic resonance imaging findings were variable. Many patients exhibited delayed development with subsequent progressive regression (n = 16, 38.1%). Twenty-one cases with genetic testing available (55%) had previously reported variants in KCTD7, and 17 cases (45%) had novel variants in KCTD7 gene. Six patients died in the cohort (age range = 1.5-21 years). The systematic review identified 23 eligible studies and further identified 59 previously reported cases of KCTD7-related disorders from the literature. The phenotype for the majority of the reported cases was consistent with a PME (n = 52, 88%). Other reported phenotypes in the literature included opsoclonus myoclonus ataxia syndrome (n = 2), myoclonus dystonia (n = 2), and neuronal ceroid lipofuscinosis (n = 3). Eight published cases died over time (14%, age range = 3-18 years). SIGNIFICANCE: This study cohort and systematic review consolidated the phenotypic spectrum and natural history of KCTD7-related disorders. Early onset drug-resistant epilepsy, relentless neuroregression, and severe neurological sequalae were common. Better understanding of the natural history may help future clinical trials.

Precision medicine for developmental and epileptic encephalopathies in Africa-strategies for a resource-limited setting.

PURPOSE: Sub-Saharan Africa bears the highest burden of epilepsy worldwide. A presumed proportion is genetic, but this etiology is buried under the burden of infections and perinatal insults in a setting of limited awareness and few options for testing. Children with developmental and epileptic encephalopathies (DEEs) are most severely affected by this diagnostic gap in Africa, because the rate of actionable findings is highest in DEE-associated genes. METHODS: We tested 234 genetically naive South African children diagnosed with/possible DEE using gene panels, exome sequencing, and chromosomal microarray. Statistical comparison of electroclinical features in children with and children without candidate variants was performed to identify characteristics most likely predictive of a positive genetic finding. RESULTS: Of the 41 (of 234) children with likely/pathogenic variants, 26 had variants supporting precision therapy. Multivariate regression modeling highlighted neonatal or infantile-onset seizures and movement abnormalities as predictive of a positive genetic finding. We used this, coupled with an emphasis on precision medicine outcomes, to propose the pragmatic "Think-Genetics" strategy for early recognition of a possible genetic etiology. CONCLUSION: Our findings emphasize the importance of an early genetic diagnosis in DEE. We designed the Think-Genetics strategy for early recognition, appropriate interim management, and genetic testing for DEE in resource-constrained settings.

Mitochondrial molecular genetic results in a South African cohort: divergent mitochondrial and nuclear DNA findings.

AIMS: Mitochondrial diseases form one of the largest groups of inborn errors of metabolism. The birth prevalence is approximately 1/5000 in well-studied populations, but little has been reported from Sub-Saharan Africa. The aim of this study was to describe the genetics underlying mitochondrial disease in South Africa. METHODS: An audit was performed on all mitochondrial disease genetic testing performed in Cape Town, South Africa. RESULTS: Of 1614 samples tested for mitochondrial DNA (mtDNA) or nuclear DNA (nDNA) variants in South Africa between 1994 and 2019, there were 155 (9.6 %) positive results. Pathogenic mtDNA variants accounted for 113 (73%)/155, from 96 families. Mitochondrial encephalopathy with lactic acidosis and stroke-like episodes, 37 (33%)/113, Leber's hereditary optic neuropathy, 26 (23%)/113, and single large mtDNA deletions, 22 (20%)/113, accounted for 76%. Thirty eight of 42 nDNA-positive results were homozygous for the MPV17 pathogenic variant c.106C>T (p.[Gln36Ter, Ser25Profs*49]) causing infantile neurohepatopathy, one of the largest homozygous groups reported in the literature. The other nDNA variants were in TAZ1, CPT2, BOLA3 and SERAC1. None were identified in SURF1, POLG or PDHA1. CONCLUSIONS: Finding a large group with a homozygous nuclear pathogenic variant emphasises the importance of looking for possible founder effects. The absence of other widely described pathogenic nDNA variants in this cohort may be due to reduced prevalence or insufficient testing. As advances in therapeutics develop, it is critical to develop diagnostic platforms on the African subcontinent so that population-specific genetic variations can be identified.

Pitfalls of relying on genetic testing only to diagnose inherited metabolic disorders in non-western populations - 5 cases of pyruvate dehydrogenase deficiency from South Africa.

Pyruvate dehydrogenase complex (PDHC) deficiencies are a group of mainly infantile onset disorders stemming from defects in pyruvate catabolism. They are characterised by severe lactic acidosis and progressive neurodegeneration.Although the PDHA1 gene is implicated in most cases of PDHC deficiency worldwide, no pathogenic variants have been reported in South African patients to date, despite availability of PDHA1 sequencing in the state diagnostic setting. METHODS: DNA from five patients with low to absent PDHC activity in fibroblasts were subjected to PDHC deficiency gene panel analysis. Included in the panel were: PDHA1, PDHB, DLAT, DLD, PDHX, BOLA3, GLRX5, IBA57, LIAS, LIPT1, LIPT2, NFU1, PDP1, PDP2, SLC19A2, SLC19A3, SLC25A19, SLC25A26, TPK1 and FBXL4. RESULTS: No pathogenic variants were identified in 4 out of 5 cases investigated. A homozygous frame-shift mutation was detected in the BOLA3 gene in one patient, supporting a diagnosis of multiple mitochondrial dysfunction syndrome type 2. DISCUSSION: A single, novel, homozygous BOLA3 frame-shift mutation was detected in a black South African child with severe neurodegenerative disease and very low to absent PDHC enzyme activity. This finding of a homozygous mutation in a patient from a non-consanguineous background may indicate a need for further investigation in clinically similar cases as well as heterozygous carrier rates in unaffected individuals from the same ethnic background.The paucity of identifiable mutations in 4 out of 5 South African patients with confirmed PDHC deficiency highlights the dangers in relying on Western population based genetic panels for diagnosing rare metabolic disease in genetically understudied populations.

The First Case of Riboflavin Transporter Deficiency in sub-Saharan Africa.

This report describes the first case of a child with genetically confirmed Brown-Vialetto-van Laere syndrome in sub-Saharan Africa. This is an extremely rare clinical condition that presents with an auditory neuropathy, bulbar palsy, stridor, muscle weakness, and respiratory compromise that manifests with diaphragmatic and vocal cord paralysis. It is an autosomal recessive condition for which the genetic mutation has only recently been linked to a riboflavin transporter deficiency. We describe an 11-month-old affected male infant. He has required long-term respiratory support and a gastrostomy tube to support feeding. With high-dose riboflavin supplementation, he had limited recovery of motor function. His respiratory chain enzyme studies were abnormal suggestive of mitochondrial (mt) dysfunction. In the setting of limited resources, recognition of this striking clinical phenotype is important to highlight, specifically regarding the genetic implications of the condition and the potentially remedial response to vitamin supplementation.

MT-ND5 Mutation Exhibits Highly Variable Neurological Manifestations at Low Mutant Load.

Mutations in the m.13094T>C MT-ND5 gene have been previously described in three cases of Leigh Syndrome (LS). In this retrospective, international cohort study we identified 20 clinically affected individuals (13 families) and four asymptomatic carriers. Ten patients were deceased at the time of analysis (median age of death was 10years (range: 5·4months-37years, IQR=17·9years). Nine patients manifested with LS, one with mitochondrial encephalomyopathy, lactic acidosis and stroke-like episodes (MELAS), and one with Leber hereditary optic neuropathy. The remaining nine patients presented with either overlapping syndromes or isolated neurological symptoms. Mitochondrial respiratory chain activity analysis was normal in five out of ten muscle biopsies. We confirmed maternal inheritance in six families, and demonstrated marked variability in tissue segregation, and phenotypic expression at relatively low blood mutant loads. Neuropathological studies of two patients manifesting with LS/MELAS showed prominent capillary proliferation, microvacuolation and severe neuronal cell loss in the brainstem and cerebellum, with conspicuous absence of basal ganglia involvement. These findings suggest that whole mtDNA genome sequencing should be considered in patients with suspected mitochondrial disease presenting with complex neurological manifestations, which would identify over 300 known pathogenic variants including the m.13094T>C.

Progressive deafness-dystonia due to SERAC1 mutations: A study of 67 cases.

OBJECTIVE: 3-Methylglutaconic aciduria, dystonia-deafness, hepatopathy, encephalopathy, Leigh-like syndrome (MEGDHEL) syndrome is caused by biallelic variants in SERAC1. METHODS: This multicenter study addressed the course of disease for each organ system. Metabolic, neuroradiological, and genetic findings are reported. RESULTS: Sixty-seven individuals (39 previously unreported) from 59 families were included (age range = 5 days-33.4 years, median age = 9 years). A total of 41 different SERAC1 variants were identified, including 20 that have not been reported before. With the exception of 2 families with a milder phenotype, all affected individuals showed a strikingly homogeneous phenotype and time course. Severe, reversible neonatal liver dysfunction and hypoglycemia were seen in >40% of all cases. Starting at a median age of 6 months, muscular hypotonia (91%) was seen, followed by progressive spasticity (82%, median onset = 15 months) and dystonia (82%, 18 months). The majority of affected individuals never learned to walk (68%). Seventy-nine percent suffered hearing loss, 58% never learned to speak, and nearly all had significant intellectual disability (88%). Magnetic resonance imaging features were accordingly homogenous, with bilateral basal ganglia involvement (98%); the characteristic "putaminal eye" was seen in 53%. The urinary marker 3-methylglutaconic aciduria was present in virtually all patients (98%). Supportive treatment focused on spasticity and drooling, and was effective in the individuals treated; hearing aids or cochlear implants did not improve communication skills. INTERPRETATION: MEGDHEL syndrome is a progressive deafness-dystonia syndrome with frequent and reversible neonatal liver involvement and a strikingly homogenous course of disease. Ann Neurol 2017;82:1004-1015.

Could we offer mitochondrial donation or similar assisted reproductive technology to South African patients with mitochondrial DNA disease?

The decision of the UK House of Commons in 2015 to endorse the use of pioneering in vitro fertilisation techniques to protect future generations from the risk of mitochondrial DNA (mtDNA) disease has sparked worldwide controversy and debate. The availability of such technologies could benefit women at risk of transmitting deleterious mutations. MtDNA disease certainly occurs in South Africa (SA) in all population groups. However, diagnostic strategies and practices for identifying individuals who would benefit from technologies such as IVF have in the past been suboptimal in this country. New developments in the molecular diagnostic services available to SA patients, as well as better education of referring clinicians and the implementation of more structured, population-appropriate diagnostic strategies, may open the floor to this debate in SA.

The usefulness of electroencephalograms in a survey of children from the Western cape of South Africa.

There are few dedicated pediatric neurophysiology services in Africa. Optimizing the efficiency of these units is essential for the large populations they serve. Two hundred eighty-seven electroencephalogram (EEG) studies, performed in a sub-Saharan neurophysiology unit, were analyzed for referral sources, appropriateness of referral, reporting consistency, and use in confirming epilepsy or a neurologic process. Pediatric neurologists requested 24% of the electroencephalogram studies and pediatricians 45%. Pediatric neurologists requested more appropriate referrals than pediatricians or nonspecialists. The electroencephalogram studies assisted management in 40% of cases. Electroencephalogram studies for nonepileptic indications, such as syncope, were highly predictable and typically normal. Education and training in the management of children with epilepsy could improve requesting practice and the subsequent usefulness of electroencephalograms in the diagnosis of epilepsy syndromes. This is especially important in the context of a resource-poor country.