Investigation of the mitochondrial energy metabolism from intact mitochondria by radiochemical substrate oxidation reveals defects of the pyruvate oxidation (PO) which is seen in an abnormal ratio of the oxidation of pyruvate+malate versus acetylcarnitine+malate. In approximately 3/4 of these patients defects of either one of the subunits of pyruvate dehydrogenase or of the regulatory phosphatases could be identified. Evaluation of the cofactor status in the remaining patients revealed an at least 4-fold decreased amount of thiamine pyrophosphate in the muscle of patients from 3 unrelated families. The clinical course of the 5 patients was characterised by metabolic crises after the first year of life with mild lactic acidosis. MRI showed a Leigh-like picture and lactate elevation in spectroscopy. Two of the patients died in infancy. Investigation of the known disease causing genes of the thiamine metabolism did not reveal pathogenic mutations. In another patient with reduced PO a severe deficiency of the lipoic acid prosthetic group was found. This patient presented with neonatal seizures. Brain sonography showed the development of a severe generalised multicystic encephalopathy. Echocardiography revealed an enlargement of the ventricular septum. He died at the age of 4 years. Pyruvate carboxylase (PC) deficiency is an autosomal recessive disease with three subtypes. Patients homozygous for the c.1828 G>A mutation in the PC gene belong to Type A, which typically has infantile-onset of developmental delay, hypotonia, and moderate lactic acidemia. We report the early brain MRI abnormalities of three patients homozygous for the c.1828 G>A mutation at 3 days, 4 days and 2 months of life respectively. The patients had bilateral cystic abnormalities in differing locations of the brain. One patient also developed diffuse brain edema. Neuropathology is available from two additional patients who died at 11 and 32 months. One patient had mild ventriculomegaly, and focal cavitation of the right caudate and right cerebellar white matter. The other patient had scattered multifocal demyelination and gliosis of the cerebral white matter. Although our patients have identical genotype and phenotype, the areas of neurologic injury in PC deficiency are diverse and varied. Our patients experience repeated episodes of severe metabolic decompensation, leading to profound developmental delay, independent of time of treatment implementation, frequency of metabolic decompensation and presence or absence of seizures. Despite the later clinical disease onset of the c.1828 G>A mutation, neurological injury can be observed on neuroimaging in the neonatal period. Elevated urinary excretion of 3-methylglutaconic acid (3-MGA) is considered rare in patients suspected of a metabolic disorder. In 3-MGAuria I, 3-MGA stems from leucine degradation. In all other types mitochondrial dysfunction is thought to be the common denominator. We investigated the clinical, metabolic and genetic data of 386 patients with 3-MGA-uria. 3-MGA-uria was often found in correlation with disorders not reported earlier in association with 3-MGA-uria (organic acidurias, urea cycle disorders, haematological/neuromuscular disorders). Mitochondrial dysfunction was indeed the common denominator for most of the patients. But, also a disturbed cholesterol biosynthesis can lead to 3-MGA-uria (Smith-Lemli-Opitz syndrome, glycogen storage disorder). Furthermore, we investigated 597 patients with mutations in 48 mitochondria-associated genes, thus having genetically proven mitochondrial disorders. 8.3% of these patients presented 3-MGA-uria. We show, that it was frequently seen in Complex V related disorders, in patients with mitochondrial DNA depletion or Pearson syndrome. Deficiencies of complex I and mitochondrial translational defects were less frequently associated with 3-MGAuria, which was also never reported in association with deficiencies of the complexes II-IV. With these data we will improve the diagnostic approach to the patient with 3-MGA-uria in general and type IV in particular. The paper further reclassifies the 3-MGA-urias. Background: Nuclear genetic defects in TMEM70 gene have been described in a subgroup of patients with 3-methylglutaconic aciduria type IV showing ATP synthase deficiency. Clinical presentation includes: cardiomyopathy, nervous system involvement and lactic acidosis. We report the clinical findings of two Spanish patients with TMEM70 mutations. Case reports: Patient 1 is the first son of healthy consanguineous parents diagnosed of intrauterine growth retardation and hypertrophic cardiomyopathy in neonatal period. He showed lactic acidemia with increased L/P ratio and 3-methylglutaconic aciduria, normal mitochondrial respiratory chain activities and normal brain MRI. He has developed: special facial phenotype, mild growth and psychomotor retardation and multiple decompensations. He is homozygous for c.317-2A>G mutation. Patient 2 is the second daughter of healthy consanguineous parents born premature after intrauterine growth retardation. She presented acute deterioration with lactic acidosis, increased L/P ratio and 3methylglutaconic aciduria. Mitochondrial respiratory chain activities showed deficiency of complex II+III. Brain MRI showed corpus callosum dysgenesis and cerebellar atrophy. She has developed: special facial phenotype, mild growth and psychomotor retardation, hypertrophic cardiomyopathy and multiple decompensations. She is compound heterozygous for the common c.317-2A>G and a new c.211-450_317-568del mutations. Conclusion: Patients with 3-methylglutaconic aciduria and cardiomyopathy should be screened for TMEM70 mutations. Today new diagnostic approaches are being developing for such "young" forms of mito diseases as mitochondrial hepatoencephalomyopathies. The algorithm for AS includes test for 4 frequent mutations (W748S, G268A, A467T, G848S) and afterwards whole POLG gene sequencing. For patients with clinical features different from AS, but still with strongly suggestion of hepatoencephalomyopathy we performed test for frequent POLG mutations and then sequencing of DGUOK and MPV17 genes. We revealed molecular defect in 9 AS patients and 5 MDS patients. The main pitfalls were: genetic heterogeneity-although AS has one gene, patients with other forms of hepatoencephalomyopathy may have mutation either in DGUOK, MPV17, POLG genes. Clinical data couldn't help us to foresee what gene is mutated; tissue specific accumulation of mtDNA depletionwe revealed only 2 patients with significant depletion of mtDNA (with DGUOK and MPV17 mutations consequently); search for 2nd allele-In our group of patients we didn't detect the second mutant allele in 4 cases (3 patients with hepatoencephalopathy and 1 patient with AS). Large POLG gene rearrangements are known being analyzed with CGH-method. 4) clinical polymorphism-differential diagnosis could be provide among several other forms if IMDs.