Mitochondrial 3-hydroxy-3-methylglutaryl-CoA synthase 2 (HMGCS2) deficiency is a rare, potentially life-threatening autosomal recessive disorder resulting from mutations in the HMGCS2 gene, leading to impaired ketogenesis. We systematically reviewed the clinical presentations, biochemical and genetic abnormalities in 93 reported cases and 2 new patients diagnosed based on biochemical findings. Reported onset ages ranged from 3 months to 6 years, mostly before the age of 3. Children younger than one year old are more prone to a severe clinical course. In most patients, the initial metabolic decompensation occurs after an episode of gastroenteritis or gastroenteritis-like symptoms. Other commonly observed symptoms during the first clinical episode included poor intake, altered consciousness, dyspnea, seizures and hepatomegaly. Severity was correlated with the number of truncating mutations. Most patients presented with acute metabolic decompensation with hypoglycemia, dicarboxyluria and inadequate ketonuria. Dicarboxylic acid levels were elevated in 54/56 cases. The organic acid 4-hydroxy-6-methyl-2-pyrone (4HMP) was detected in 33/35 urine samples taken during the acute episodes, but typically only retrospectively. The plasma C2/C0 acylcarnitine ratio was abnormal in 16/18 (88.9 %) of acute plasma samples, but only in 2/6 (33 %) of DBS samples. Other metabolites that have been reported are hydroxyhexenoic acid, 3,5-dihydroxyhexanoic (1,5 lactone), glutaric acidand 3-OH-isovaleric acid. Laboratories should look for 4HMP in urinary organic acid analysis and an increased plasma C2/C0 acylcarnitine ratio to facilitate the diagnosis of HMGCS2 deficiency, especially in cases of metabolic decompensation with dicarboxyluria without adequate ketonuria.