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Identification of a novel TTC19 mutation in a Portuguese family with complex III deficiency
584
Abstracts
a
Center for Developmental Therapeutics, University of Washington, Seattle, WA, United States b Department of Pediatrics, Children's Hospital of Philadelphia, Philadelphia, PA, United States c Departments of Pharmacology and Cancer Biology, Biochemistry and Internal Medicine, Duke University, Durham, NC, United States
Center for Molecular and Mitochondrial Medicine and Genetics, University of California, Irvine, Irvine, CA 92697-3940, USA c lnstitute of Human Respiratory Disease, Xinqiao Hospital, Third Military Medical University, Chongqing 400037, China d Graduate School of Arts & Sciences, Wake Forest University, Winston-Salem, NC 27157-1001, USA
Mitochondrial function is emerging as the keystone of an evergrowing and fairly extraordinary ensemble of clinical conditions. However, mitochondrial diseases remain difficult to diagnose. Although many other valuable approaches are being used to understand how mitochondria function, the use of molecular genetics in C. elegans possesses powerful and unique advantages. The excellent genetics, superb catalogue of mutations, well-defined behaviors, and invariant cell lineage led to our choice of C. elegans for study. Studies in C. elegans have shown that mitochondrial function in the nematode is extremely similar to that seen in mammals. This has led us to study mitochondrial function in disease by analyzing C. elegans mutants with defects in different steps of the electron transport chain (ETC). Methods: We combined the use of gene set expression analysis (GSEA), protein set enrichment analysis (PSEA; proteomics) and metabolomics to study the global changes in three mutants affecting the ETC in comparison to wildtype. The mutants studied were a complex I mutant (gas-1), a complex II mutant (mev-1) and a mutant in coenzyme Q synthesis (clk-1). For GSEA and metabolomics, we isolated RNA and metabolites from whole worms as first day adults. For proteomics, we isolated protein from mitochondria of first day adults. Pathways identified as changed in GSEA and PSEA platforms were studied further by introducing mutations directly affecting those pathways. The expression platforms were then compared to the metabolomic results. Results: Comparison of gene expression and proteomic arrays were revealed multiple concordant trends. For example pathways of 1) glycolosis/gluconeogenesis, 2) valine, isoleucine and leucine metabolism (both synthesis and degradation), 3) ubiquinone metabolism, 4) arginine/proline metabolism, 5) TCA cycle, 6) fatty acid metabolism and 7) glyoxylate metabolism were found upregulated in gas-1 and mev-1. The expression results reported earlier by Falk et al. corroborate the proteomic results reported here. Inhibition of ubiquinone metabolism by the clk-1 mutation, which by itself does not cause a strong phenotype, caused a sterile phenotype when combined with gas-1 or mev-1. Inhibition of the glyoxylate pathway (gei-7), upregulated in both the GSEA and PSEA in gas-1, was lethal when combined with gas-1. Studies combining gei-7 with mev-1 are in progress. Discussion: A systems-based approach to characterizing the phenotypes of mitochondrial mutants may aid both in diagnosis of the diseases and understanding the interactions of multiple defects. Such understanding is likely to be more useful than any one platform used alone.
The distinction between mild pathogenic mtDNA mutations and population polymorphisms can be ambiguous since both are homoplasmic, alter conserved functions, and correlate with disease. One possible explanation for this ambiguity is that the same variant may have different consequences in different contexts. Although not one of the original mtDNA disease causing mutations, 3394 has been found in significantly increased frequencies in LHON pedigrees (Brown et al., 1995). A case in point is the mtDNA ND1 T3394C (Y30H) missense mutation. This mutation has been associated with LHON when occurring on macrohaplogroup N mtDNAs (Liang et al., 2009). However, in Asia the 3394C variant is most commonly associated with the M9 haplogroup which is rare at low elevations but increases in frequency with elevation, to an average of 25% in residents of the Tibetan Plateau (OR = 23.7). In high-altitude Tibetan and Indian populations, the 3394C variant occurs on five different macrohaplogroup M haplogroup backgrounds and it is enriched on the M9 background in Tibet and the C4a4 background on the Indian Deccan Plateau (OR = 21.9). When occurring on macrohaplogroup N mtDNAs including B4c and F1 this variant causes reduced complex I and reduced cellular respiration on macrohaplogroups N haplogroups B4c and F1 mtDNAs but not in the M9 haplogroup. On the M9 background the 3394C variant is associated with a complex I activity that is equal to or higher than that of the 3394T variant on the B4c and F1 backgrounds. Hence, the 3394C variant can either be deleterious or beneficial depending on its haplogroup and environmental context. Thus this mtDNA variant fulfills the criteria for a common variant that predisposes to a “complex” disease. 1 Brown MD, Torroni A, Reckord CL, & Wallace DC (1995) Phylogenetic analysis of Leber's hereditary optic neuropathy mitochondrial DNA's indicates multiple independent occurrences of the common mutations. Human Mutation 6(4):311–325. 2 Liang M, et al. (2009) Leber's hereditary optic neuropathy is associated with mitochondrial ND1 T3394C mutation. Biochemical and Biophysical Research Communications 383(3):286–292.
doi:10.1016/j.mito.2012.07.085
95 Leber Hereditary Optic Neuropathy (LHON) associated mutation 3394 is also a high-altitude adaptive polymorphism Presenter: Leonardo S. Alves Leonardo S. Alvesa,b,d, Fuyun Jic, Mark S. Sharpleya,b, Olga Derbenevaa,b, Dimitra Chalkiaa,b, Maria Lvovaa,b, Guisheng Qianc, Lorna G. Moorec, Douglas C. Wallacea,b a Center for Mitochondrial and Epigenomic Medicine, Children's Hospital of Philadelphia, University of Pennsylvania, 3501 Civic Center Boulevard Philadelphia, PA 19104-4302, USA
b
doi:10.1016/j.mito.2012.07.086
96 Identification of a novel TTC19 mutation in a Portuguese family with complex III deficiency Presenter: Célia Nogueira Célia Nogueiraa, José Barrosb, Maria José Sác, Luísa Azevedod, Filippo M. Santorellie, Laura Vilarinhoa a National Institute of Health, Genetics Department, INSA, Oporto, Portugal b Oporto Hospital Center, Neurology Unit, Hospital S. António, Oporto, Portugal c Neurology Unit, Hospital S. João, Oporto, Portugal d IPATIMUP, Population Genetics, Oporto University, Oporto, Portugal e Fundazione Stella Maris, Molecular Medicine & Neurogenetics, IRCCS, Pisa, Italy Defects of mitochondrial complex III (CIII) are a relatively rare cause of mitochondrial dysfunction. CIII or ubiquinol-cytochrome c reductase is the third component of the mitochondrial respiratory chain and catalyzes the electron transfer from reduced coenzyme Q to cytochrome c and is composed of 11 subunits; one encoded by
Abstracts
mitochondrial DNA (MT-CYB) and the remaining by nuclear genes. BCS1L gene is a CIII assembly factor. Mutations in MT-CYB and BCS1L genes account for the vast majority of mutations leading to CIII deficiency, and are associated with a wide range of neuromuscular disorders. The human tetratricopeptide 19 (TTC19), encodes a poorly understood member of tetratricopeptide repeat domain 19 located on chromosome 17 and appears to be involved in the correct assembly of CIII. Recently, mutations in TTC19 have been described in three unrelated Italian kindred in association with a severe neurodegenerative disease. Here we present a consanguineous Portuguese family where a severe biochemical deficiency of complex III enzyme activity occurred in four siblings in association with neurological manifestations suggestive of cerebellar ataxia combined with relentless psychiatric manifestations. Variability in age at onset and disease course was associated with a novel homozygous mutation in TTC19. We had first detected a biochemically deficient enzyme activity in the family, we had analyzed all structural genes part of CIII as well as BCS1L. Only the recent description of mutations in TTC19 raised high the suspect of a similar condition in the present family. The novel TTC19 mutation identified in this family, was homozygous in the four patients, heterozygous in their parents and in two healthy relatives, and it was absent in ethnically-matched controls. The mutation predicts a frameshift, resulting in a truncated protein by the insertion of a premature stop codon. In summary, we are describing the 4th family identified in the world carrying a novel TTC19 mutation. Our data corroborate the genotype and phenotype variability presented by the affected family members and hopefully will contribute to a deeper understanding of the CIII-related disorders.
doi:10.1016/j.mito.2012.07.087
97 “Double-trouble” or digenic disorder in complex I deficiency Presenter: Ligia S. Almeida Lígia S. Almeidaa, Mariana Ferreiraa, Célia Nogueiraa, Fátima Furtadob, Teresinha Evangelistac, Filippo M. Santorellid, Laura Vilarinhoa a Research Unit, Dept. of Genetics, Medical Genetics Center, INSA, Porto, Portugal b Serviço de Pediatria, Hospital Distrital de Beja, Portugal c Hospital Sta. Maria, Neurologia, Lisboa, Portugal d Molecular Medicine & Neurodegenerative Diseases, IRCCS Fondazione Stella Maris, Pisa, Italy Complex I (CI) deficiency is a defect of OXPHOS caused by mutations in the mitochondrial or nuclear genomes. To date diseasecausing mutations have been reported in all mitochondrial-encoded subunits and 22 nuclear genes. In about 50% of the patients no mutations are found, suggesting that undiscovered factors are an important cause of disease. In this study we report a consanguineous family from Southern Portugal with three affected children where CI deficiency could not be clarified. The affected children presented similar clinical findings with early onset of the disease having the three of them a fatal outcome. A reduced activity of CI was detected in two of the patients that led us to investigate, at the molecular level, some CI associated genes. After studying some CI associated genes no mutations were detected. Interestingly, all patients presented 3-methylglutaconic acid in the urinary organic acids and it is known that POLG gene is involved in the etiology 3-methylglutaconic acidurias. In only one of the patients, presenting also with mtDNA depletion, the p.G848S/p.Q1236H mutations were found in POLG.
585
Besides the efforts done it remains unsolved whether this family due to the high consanguinity could have two different disorders or if a yet unknown gene, leading to CI deficiency, could be involved. doi:10.1016/j.mito.2012.07.088
101 Metabolic adaptations in neurons with complex IV deficiency Presenter: Francisca Diaz Francisca Diaza, Sofia Garciaa, Carlos T. Moraesa,b a Department of Neurology, University of Miami, Miller School of Medicine, Miami, FL, 33136, USA b Department of Cell Biology and Anatomy, University of Miami, Miller School of Medicine, Miami, FL, 33136, USA We created a mitochondrial encephalopathy mouse model with complex IV (cytochrome c oxidase or COX) deficiency by the ablation of the COX10 gene in neurons. COX10 encodes a heme a farnesyl transferase indispensable for COX assembly and function. The COX10 KO had a reduced life span and the majority of the mice died between 8 and 14 months of age. The COX10 KO did not display an overt phenotype until 4 months of age, long before their death, when distinctive behavioral abnormalities developed (cycles of hyper and hypo activities). The onset of the mitochondrial defect was not accompanied by neuronal death in the COX10 KO mice. A progressive COX deficiency was observed in both cortex and hippocampus homogenates showing about 70% of control values at 2 months of age and rapidly declining to 24% by 4 months. The first signs of neuronal degeneration were observed only at about 4 months of age with tunel positive neurons found in cingulate cortex, hippocampus/dentate gyrus and piriform cortex. This early neurodegeneration developed into a severe cortical atrophy by 8 to 10 months of age. We investigated early metabolic adaptations responsible for neuronal survival to maintain the energetic crisis triggered by the mitochondrial defect. We observed an increased glucose uptake in the COX10 KO brain when compared to control mice. Metabolomic analysis revealed an increase in the levels of several glycolyticpentose phosphate pathway intermediates and certain amino acids. These results were in agreement with an increase in the activity of one of the regulatory enzymes of glycolysis (hexokinase) at 3 months of age. Interestingly, western blot analysis revealed that glycolysis was modulated by enhanced activity and not by overexpression of the enzymes. We are currently investigating the molecular mechanisms responsible for the upregulation of the glycolytic pathway. The COX10 conditional brain KO constitutes an excellent model of mitochondrial encephalopathy to study the molecular bases of metabolic adaptations leading to neuronal survival to compensate for an energetic crisis caused by a mitochondrial defect. doi:10.1016/j.mito.2012.07.089
102 NextGen Sequencing of the complete mtDNA genome: 20% estimated positive cases among recent 117 patients Presenter: Carolyn H. Buzin Carolyn H. Buzin, Christian Neckelmann, Melissa Gasper, Juliana O. Barba, William A. Scaringe, Steve S. Sommer MEDomics, 426N. San Gabriel Ave., Azusa, CA 91702, United States The complete mitochondrial genome (mtDNA), including all 37 genes and the control region, was sequenced 10,000+ times by Next
Abstracts
a
Center for Developmental Therapeutics, University of Washington, Seattle, WA, United States b Department of Pediatrics, Children's Hospital of Philadelphia, Philadelphia, PA, United States c Departments of Pharmacology and Cancer Biology, Biochemistry and Internal Medicine, Duke University, Durham, NC, United States
Center for Molecular and Mitochondrial Medicine and Genetics, University of California, Irvine, Irvine, CA 92697-3940, USA c lnstitute of Human Respiratory Disease, Xinqiao Hospital, Third Military Medical University, Chongqing 400037, China d Graduate School of Arts & Sciences, Wake Forest University, Winston-Salem, NC 27157-1001, USA
Mitochondrial function is emerging as the keystone of an evergrowing and fairly extraordinary ensemble of clinical conditions. However, mitochondrial diseases remain difficult to diagnose. Although many other valuable approaches are being used to understand how mitochondria function, the use of molecular genetics in C. elegans possesses powerful and unique advantages. The excellent genetics, superb catalogue of mutations, well-defined behaviors, and invariant cell lineage led to our choice of C. elegans for study. Studies in C. elegans have shown that mitochondrial function in the nematode is extremely similar to that seen in mammals. This has led us to study mitochondrial function in disease by analyzing C. elegans mutants with defects in different steps of the electron transport chain (ETC). Methods: We combined the use of gene set expression analysis (GSEA), protein set enrichment analysis (PSEA; proteomics) and metabolomics to study the global changes in three mutants affecting the ETC in comparison to wildtype. The mutants studied were a complex I mutant (gas-1), a complex II mutant (mev-1) and a mutant in coenzyme Q synthesis (clk-1). For GSEA and metabolomics, we isolated RNA and metabolites from whole worms as first day adults. For proteomics, we isolated protein from mitochondria of first day adults. Pathways identified as changed in GSEA and PSEA platforms were studied further by introducing mutations directly affecting those pathways. The expression platforms were then compared to the metabolomic results. Results: Comparison of gene expression and proteomic arrays were revealed multiple concordant trends. For example pathways of 1) glycolosis/gluconeogenesis, 2) valine, isoleucine and leucine metabolism (both synthesis and degradation), 3) ubiquinone metabolism, 4) arginine/proline metabolism, 5) TCA cycle, 6) fatty acid metabolism and 7) glyoxylate metabolism were found upregulated in gas-1 and mev-1. The expression results reported earlier by Falk et al. corroborate the proteomic results reported here. Inhibition of ubiquinone metabolism by the clk-1 mutation, which by itself does not cause a strong phenotype, caused a sterile phenotype when combined with gas-1 or mev-1. Inhibition of the glyoxylate pathway (gei-7), upregulated in both the GSEA and PSEA in gas-1, was lethal when combined with gas-1. Studies combining gei-7 with mev-1 are in progress. Discussion: A systems-based approach to characterizing the phenotypes of mitochondrial mutants may aid both in diagnosis of the diseases and understanding the interactions of multiple defects. Such understanding is likely to be more useful than any one platform used alone.
The distinction between mild pathogenic mtDNA mutations and population polymorphisms can be ambiguous since both are homoplasmic, alter conserved functions, and correlate with disease. One possible explanation for this ambiguity is that the same variant may have different consequences in different contexts. Although not one of the original mtDNA disease causing mutations, 3394 has been found in significantly increased frequencies in LHON pedigrees (Brown et al., 1995). A case in point is the mtDNA ND1 T3394C (Y30H) missense mutation. This mutation has been associated with LHON when occurring on macrohaplogroup N mtDNAs (Liang et al., 2009). However, in Asia the 3394C variant is most commonly associated with the M9 haplogroup which is rare at low elevations but increases in frequency with elevation, to an average of 25% in residents of the Tibetan Plateau (OR = 23.7). In high-altitude Tibetan and Indian populations, the 3394C variant occurs on five different macrohaplogroup M haplogroup backgrounds and it is enriched on the M9 background in Tibet and the C4a4 background on the Indian Deccan Plateau (OR = 21.9). When occurring on macrohaplogroup N mtDNAs including B4c and F1 this variant causes reduced complex I and reduced cellular respiration on macrohaplogroups N haplogroups B4c and F1 mtDNAs but not in the M9 haplogroup. On the M9 background the 3394C variant is associated with a complex I activity that is equal to or higher than that of the 3394T variant on the B4c and F1 backgrounds. Hence, the 3394C variant can either be deleterious or beneficial depending on its haplogroup and environmental context. Thus this mtDNA variant fulfills the criteria for a common variant that predisposes to a “complex” disease. 1 Brown MD, Torroni A, Reckord CL, & Wallace DC (1995) Phylogenetic analysis of Leber's hereditary optic neuropathy mitochondrial DNA's indicates multiple independent occurrences of the common mutations. Human Mutation 6(4):311–325. 2 Liang M, et al. (2009) Leber's hereditary optic neuropathy is associated with mitochondrial ND1 T3394C mutation. Biochemical and Biophysical Research Communications 383(3):286–292.
doi:10.1016/j.mito.2012.07.085
95 Leber Hereditary Optic Neuropathy (LHON) associated mutation 3394 is also a high-altitude adaptive polymorphism Presenter: Leonardo S. Alves Leonardo S. Alvesa,b,d, Fuyun Jic, Mark S. Sharpleya,b, Olga Derbenevaa,b, Dimitra Chalkiaa,b, Maria Lvovaa,b, Guisheng Qianc, Lorna G. Moorec, Douglas C. Wallacea,b a Center for Mitochondrial and Epigenomic Medicine, Children's Hospital of Philadelphia, University of Pennsylvania, 3501 Civic Center Boulevard Philadelphia, PA 19104-4302, USA
b
doi:10.1016/j.mito.2012.07.086
96 Identification of a novel TTC19 mutation in a Portuguese family with complex III deficiency Presenter: Célia Nogueira Célia Nogueiraa, José Barrosb, Maria José Sác, Luísa Azevedod, Filippo M. Santorellie, Laura Vilarinhoa a National Institute of Health, Genetics Department, INSA, Oporto, Portugal b Oporto Hospital Center, Neurology Unit, Hospital S. António, Oporto, Portugal c Neurology Unit, Hospital S. João, Oporto, Portugal d IPATIMUP, Population Genetics, Oporto University, Oporto, Portugal e Fundazione Stella Maris, Molecular Medicine & Neurogenetics, IRCCS, Pisa, Italy Defects of mitochondrial complex III (CIII) are a relatively rare cause of mitochondrial dysfunction. CIII or ubiquinol-cytochrome c reductase is the third component of the mitochondrial respiratory chain and catalyzes the electron transfer from reduced coenzyme Q to cytochrome c and is composed of 11 subunits; one encoded by
Abstracts
mitochondrial DNA (MT-CYB) and the remaining by nuclear genes. BCS1L gene is a CIII assembly factor. Mutations in MT-CYB and BCS1L genes account for the vast majority of mutations leading to CIII deficiency, and are associated with a wide range of neuromuscular disorders. The human tetratricopeptide 19 (TTC19), encodes a poorly understood member of tetratricopeptide repeat domain 19 located on chromosome 17 and appears to be involved in the correct assembly of CIII. Recently, mutations in TTC19 have been described in three unrelated Italian kindred in association with a severe neurodegenerative disease. Here we present a consanguineous Portuguese family where a severe biochemical deficiency of complex III enzyme activity occurred in four siblings in association with neurological manifestations suggestive of cerebellar ataxia combined with relentless psychiatric manifestations. Variability in age at onset and disease course was associated with a novel homozygous mutation in TTC19. We had first detected a biochemically deficient enzyme activity in the family, we had analyzed all structural genes part of CIII as well as BCS1L. Only the recent description of mutations in TTC19 raised high the suspect of a similar condition in the present family. The novel TTC19 mutation identified in this family, was homozygous in the four patients, heterozygous in their parents and in two healthy relatives, and it was absent in ethnically-matched controls. The mutation predicts a frameshift, resulting in a truncated protein by the insertion of a premature stop codon. In summary, we are describing the 4th family identified in the world carrying a novel TTC19 mutation. Our data corroborate the genotype and phenotype variability presented by the affected family members and hopefully will contribute to a deeper understanding of the CIII-related disorders.
doi:10.1016/j.mito.2012.07.087
97 “Double-trouble” or digenic disorder in complex I deficiency Presenter: Ligia S. Almeida Lígia S. Almeidaa, Mariana Ferreiraa, Célia Nogueiraa, Fátima Furtadob, Teresinha Evangelistac, Filippo M. Santorellid, Laura Vilarinhoa a Research Unit, Dept. of Genetics, Medical Genetics Center, INSA, Porto, Portugal b Serviço de Pediatria, Hospital Distrital de Beja, Portugal c Hospital Sta. Maria, Neurologia, Lisboa, Portugal d Molecular Medicine & Neurodegenerative Diseases, IRCCS Fondazione Stella Maris, Pisa, Italy Complex I (CI) deficiency is a defect of OXPHOS caused by mutations in the mitochondrial or nuclear genomes. To date diseasecausing mutations have been reported in all mitochondrial-encoded subunits and 22 nuclear genes. In about 50% of the patients no mutations are found, suggesting that undiscovered factors are an important cause of disease. In this study we report a consanguineous family from Southern Portugal with three affected children where CI deficiency could not be clarified. The affected children presented similar clinical findings with early onset of the disease having the three of them a fatal outcome. A reduced activity of CI was detected in two of the patients that led us to investigate, at the molecular level, some CI associated genes. After studying some CI associated genes no mutations were detected. Interestingly, all patients presented 3-methylglutaconic acid in the urinary organic acids and it is known that POLG gene is involved in the etiology 3-methylglutaconic acidurias. In only one of the patients, presenting also with mtDNA depletion, the p.G848S/p.Q1236H mutations were found in POLG.
585
Besides the efforts done it remains unsolved whether this family due to the high consanguinity could have two different disorders or if a yet unknown gene, leading to CI deficiency, could be involved. doi:10.1016/j.mito.2012.07.088
101 Metabolic adaptations in neurons with complex IV deficiency Presenter: Francisca Diaz Francisca Diaza, Sofia Garciaa, Carlos T. Moraesa,b a Department of Neurology, University of Miami, Miller School of Medicine, Miami, FL, 33136, USA b Department of Cell Biology and Anatomy, University of Miami, Miller School of Medicine, Miami, FL, 33136, USA We created a mitochondrial encephalopathy mouse model with complex IV (cytochrome c oxidase or COX) deficiency by the ablation of the COX10 gene in neurons. COX10 encodes a heme a farnesyl transferase indispensable for COX assembly and function. The COX10 KO had a reduced life span and the majority of the mice died between 8 and 14 months of age. The COX10 KO did not display an overt phenotype until 4 months of age, long before their death, when distinctive behavioral abnormalities developed (cycles of hyper and hypo activities). The onset of the mitochondrial defect was not accompanied by neuronal death in the COX10 KO mice. A progressive COX deficiency was observed in both cortex and hippocampus homogenates showing about 70% of control values at 2 months of age and rapidly declining to 24% by 4 months. The first signs of neuronal degeneration were observed only at about 4 months of age with tunel positive neurons found in cingulate cortex, hippocampus/dentate gyrus and piriform cortex. This early neurodegeneration developed into a severe cortical atrophy by 8 to 10 months of age. We investigated early metabolic adaptations responsible for neuronal survival to maintain the energetic crisis triggered by the mitochondrial defect. We observed an increased glucose uptake in the COX10 KO brain when compared to control mice. Metabolomic analysis revealed an increase in the levels of several glycolyticpentose phosphate pathway intermediates and certain amino acids. These results were in agreement with an increase in the activity of one of the regulatory enzymes of glycolysis (hexokinase) at 3 months of age. Interestingly, western blot analysis revealed that glycolysis was modulated by enhanced activity and not by overexpression of the enzymes. We are currently investigating the molecular mechanisms responsible for the upregulation of the glycolytic pathway. The COX10 conditional brain KO constitutes an excellent model of mitochondrial encephalopathy to study the molecular bases of metabolic adaptations leading to neuronal survival to compensate for an energetic crisis caused by a mitochondrial defect. doi:10.1016/j.mito.2012.07.089
102 NextGen Sequencing of the complete mtDNA genome: 20% estimated positive cases among recent 117 patients Presenter: Carolyn H. Buzin Carolyn H. Buzin, Christian Neckelmann, Melissa Gasper, Juliana O. Barba, William A. Scaringe, Steve S. Sommer MEDomics, 426N. San Gabriel Ave., Azusa, CA 91702, United States The complete mitochondrial genome (mtDNA), including all 37 genes and the control region, was sequenced 10,000+ times by Next