Mitochondrial acetoacetyl-CoA thiolase deficiency: neonatal onset

We present the case of a 4-day-old newborn with serious dehydration, polypnea, hypertonus and lethargy. Blood analysis showed severe metabolic acidosis with ketonemia, ketonuria and elevation of the GAP anion. Urine analysis revealed increased excretion of 2-methyl-3-hydroxybutyrate acid, tiglycine, and 2-methylacetoacetate acid. Neonatal onset of mitochondrial acetoacetyl-CoA thiolase (T2) deficiency is exceptional. Most patients have no clinical symptoms in the neonatal period. This entity should be considered in patients with acute metabolic acidosis and ketosis with normal glycemia and aciduria. The urine contains large amounts of 2-methylacetoacetate and its decarboxylation products. In the neonatal period, this inherited disorder of metabolism can produce severe hydroelectrolyte disorders in the form of a gradual process or acute episodes, which can occasionally be fatal.     

Heterogeneity of defects in mitochondrial acetoacetyl-CoA thiolase biosynthesis in fibroblasts from four patients with 3-ketothiolase deficiency

Deficient mitochondrial acetoacetyl-CoA thiolase in fibroblasts from four patients with 3-ketothiolase deficiency was studied using immunochemical methods. We also examined fibroblasts from two heterozygotes, the mother and the brother of the case 1 patient, identified on the basis of the results of the enzyme activity measurements, using 2-methylacetoacetyl-CoA as substrate. The results were as follows: 1) in fibroblasts from all four patients, the thiolase activity using acetoacetyl-CoA was not activated by K+, although that of the controls and the heterozygotes was activated about 2-fold. 2) by immunoblot analyses, mitochondrial acetoacetyl-CoA thiolase was not detectable in fibroblasts from cases 2 and 3, although a very faint band was seen in tissues from cases 1 and 4. However, the band of mitochondrial 3-ketoacyl-CoA thiolase was clearly detected in all patients to the same extent as in the controls. 3) mitochondrial acetoacetyl-CoA thiolase was observed after pulse labeling for 1-h and a 72-h chase period in three cell lines (cases 1, 2, and 4), but was fainter compared to the controls. In another cell line (case 3), a fluorographic band at the same position was detected following a 1-h pulse, but disappeared following a 6-h chase. These results demonstrate heterogeneity in the enzyme defect resulting in a deficiency of mitochondrial acetoacetyl-CoA thiolase in fibroblasts from patients with 3-ketothiolase deficiency.     

Sickle-cell disease not identified by newborn screening because of prior transfusion

Erythrocyte transfusion can impair detection of sickle-cell disease, galactosemia, or biotinidase deficiency with newborn screening. We report on 4 infants with SCD in whom delayed diagnosis was associated with neonatal transfusion. In 2 cases, the initial newborn screening showed no hemoglobin S. In no case was the recommended screening >/=120 days from the last transfusion obtained. Two children had significant SCD-related morbidity before diagnosis.     

Decline of acute encephalopathic crises in children with glutaryl-CoA dehydrogenase deficiency identified by newborn screening in Germany

Glutaryl-CoA dehydrogenase (GCDH) deficiency is a rare neurometabolic disorder that is considered treatable if patients are identified before the onset of acute encephalopathic crises. To allow early identification of affected individuals, tandem mass spectrometry-based newborn screening for GCDH deficiency has been started in Germany in 1999. We prospectively followed neonatally screened patients (n=38) and compared the neurologic outcome with patients from a historical cohort (n=62). In the majority of neonatally screened children, the onset of encephalopathic crises has been prevented (89%), whereas acute encephalopathic crises or progressive neurologic impairment was common in the historical cohort. Neonatal screening in combination with intensive management is effective--even assuming ascertainment bias in the historical cohort. Similar proportions of commonest mutations and biochemical phenotypes (high and low excretors) were found in neonatally screened and historical patients. However, potential predictor variables for mild clinical phenotypes are not yet known and thus a selection of these patients by newborn screening is not excluded. No patient was known to be missed by newborn screening from 1999 to 2005. In conclusion, this study confirms that newborn screening for GCDH deficiency in combination with intensive management is beneficial.     

线粒体乙酰乙酰辅酶A硫解酶缺乏症一家系报告并文献复习

目的探讨线粒体乙酰乙酰辅酶A硫解酶缺乏症(简称T2缺乏症)的诊断及预后。方法回顾性分析一家系中3例T2缺乏症患儿的临床资料,并复习相关文献。结果例1和例2为先证者,男孩,同卵双胞胎,因发热、呕吐、气促入院,血气分析均显示严重的代谢性酸中毒,尿气相色谱质谱均显示3-羟基丁酸、2-甲基-3-羟基丁酸、甲基巴豆酰甘氨酸-1、3-甲基巴豆酰甘氨酸-1明显增高,二羧酸轻微升高;血串联质谱仪分析结果均显示,C5∶1、C5-OH、C4-OH升高。例3,为例1和例2同胞姐姐,5月龄时因严重代谢性酸中毒住院治疗。3例患儿行ACAT1基因检测均为复合杂合突变,c.622 CT(p R208X)和c.653 CT(p S218F)。结论对于以酸中毒为突出表现的患儿,需警惕T2缺乏症,及时行尿气相色谱质谱和血串联质谱分析,以早期诊断和治疗。     

Epidemiology of congenital toxoplasmosis identified by population-based newborn screening in Massachusetts.

BACKGROUND: Fourteen years of newborn screening in Massachusetts for congenital toxoplasmosis infection identified subpopulations that appeared to have higher rates of infection. Elaborating an epidemiologic profile and risk correlates might aid implementing targeted prenatal education and newborn screening strategies with the goal of early postnatal treatment to prevent morbidity. OBJECTIVE: To describe the epidemiology of congenital toxoplasmosis in Massachusetts and risk correlates of infection using birth certificate data. METHODS: A case-control study was conducted based on Massachusetts birth certificate data. Cases were all infants with congenital toxoplasmosis identified by statewide universal newborn screening from 1988 to 1999. Controls were all children born on the same day as those infants in Massachusetts. RESULTS: Factors that strongly predicted congenital toxoplasmosis infection were mother's country of birth outside the US (especially the southeast Asian refugee origin countries of Cambodia and Laos), mother's educational level and higher gravidity. CONCLUSIONS: More extensive, culturally and linguistically appropriate, prenatal education is needed for pregnant women, regardless of a mother's educational level, especially for non-US-born mothers, and not focused only on primiparous women. Other states may be able to use their state-specific birth certificate data to compare risk profiles with those of Massachusetts to guide a toxoplasmosis screening policy on the basis of population similarities and differences.     

Detection of neonatal carnitine palmitoyltransferase II deficiency by expanded newborn screening with tandem mass spectrometry

The introduction of tandem mass spectrometry to newborn screening has substantially expanded our ability to diagnose metabolic diseases in the newborn period. We report the first case of neonatal carnitine palmitoyltransferase deficiency II detected by expanded newborn screening with tandem mass spectrometry. The neonate presented with dysmorphic facial features, structural malformations, renal failure, seizures, and cardiac arrythmias and died on the third day of life. This experience illustrates the importance of expanded newborn screening to avoid missing a metabolic diagnosis in early infantile death.     

Mitochondrial acetoacetyl-CoA thiolase (T2) deficiency: T2-deficient patients with "mild" mutation(s) were previously misinterpreted as normal by the coupled assay with tiglyl-CoA

Mitochondrial acetoacetyl-CoA thiolase (T2) deficiency is an inborn error of metabolism that affects the catabolism of isoleucine and ketone bodies. This disorder is characterized by intermittent ketoacidotic episodes. Recently, we diagnosed T2 deficiency in two patients (GK45 and GK47) by the absence of potassium ion-activated acetoacetyl-CoA thiolase activity, whereas these patients were previously misinterpreted as normal by a coupled assay with tiglyl-CoA as a substrate. This method has been widely used for the enzymatic diagnosis of the T2 deficiency in the United States and Europe. We hypothesized that some residual T2 activity showed normal results in the assay. To prove this hypothesis, we analyzed these two patients together with three typical T2-deficient patients (GK46, GK49, and GK50) at the DNA level. Expression analysis of mutant cDNAs clearly showed that GK45 and GK47 had "mild" mutations (A132G, D339-V340insD) that retained some residual T2 activity, at least one of two mutant alleles, whereas the other three patients had null mutations (c.52-53insC, G152A, H397D, and IVS8+1g>t) in either allele. These results raise the possibility that T2-deficient patients with mild mutations have been misinterpreted as normal by the coupled assay with tiglyl-CoA.     

Diagnosis of mitochondrial trifunctional protein deficiency in a blood spot from the newborn screening card by tandem mass spectrometry and DNA analysis.

Trifunctional protein (TFP) plays a significant role in the mitochondrial beta-oxidation of long-chain fatty acids. Its deficiency impairs the energy generating function of this pathway and causes hypoketotic hypoglycemia once hepatic glycogen stores are depleted. A Reye-like syndrome, cardiomyopathy, and sudden death may follow. The diagnosis is based on demonstration of significantly decreased enzyme activity of at least two of the three involved enzymes in fibroblasts. The possibility of prospective diagnosis of TFP deficiency by newborn screening using tandem mass spectrometry (MS/MS) has not been evaluated. We report the postmortem diagnosis of a male newborn, who suffered sudden death at 2 wk of age, and his younger sister, who died of cardiomyopathy complicated by acute heart failure at the age of 6 mo, after she had acquired a common viral infection. Blood spots from the original newborn screening cards were the only remaining material from the patients. Analysis by MS/MS revealed acylcarnitine profiles consistent with either TFP or long-chain 3-hydroxyacyl coenzyme A dehydrogenase (LCHAD) deficiency. To prove the diagnosis, the alpha- and beta-subunit genes coding for TFP were examined. The patients were compound heterozygous for a 4-bp-deletion and an a-->g missense mutation, both in the same exon 3 donor consensus splice site. This is the first report of the diagnosis of TFP deficiency using blood spots obtained for newborn screening and suggests that TFP deficiency may be detectable by prospective newborn screening using MS/MS.     

Nutritional vitamin B12 deficiency: Two cases detected by routine newborn urinary screening

We describe two asymptomatic newborns with nutritional vitamin B₁₂ deficiency in whom increased urinary methylmalonic acid was detected by routine neonatal screening at 3 weeks of age. Both infants were exclusively breast-fed. One mother suffered from pernicious anaemia, and the other was a strict vegetarian. Both mothers had no clinical or haematological abnormality, aside from a borderline mean corpuscular volume for the vegetarian mother. This report illustrates the early appearance of functional vitamin B₁₂ deficiency in breastfed infants of vitamin B₁₂-depleted mothers. It also demonstrates that urinary methylmalonic acid measurement is a sensitive indicator of tissue vitamin B₁₂ deficiency.     

The first three years of screening for medium chain acyl-CoA dehydrogenase deficiency (MCADD) by newborn screening ontario

Background  

Medium chain acyl-CoA dehydrogenase deficiency (MCADD) is a disorder of mitochondrial fatty acid oxidation and is one of the most common inborn errors of metabolism. Identification of MCADD via newborn screening permits the introduction of interventions that can significantly reduce associated morbidity and mortality. This study reports on the first three years of newborn screening for MCADD in Ontario, Canada.     

MS/MS-based newborn and family screening detects asymptomatic patients with very-long-chain acyl-CoA dehydrogenase deficiency

OBJECTIVES: To determine whether asymptomatic persons with biochemical evidence of very long-chain acyl-CoA dehydrogenase (VLCAD) deficiency identified through expanded newborn screening with tandem mass spectometry have confirmed disease. STUDY DESIGN: We characterized 8 asymptomatic VLCAD-deficient individuals by enzyme and/or mutational analysis and compared them with clinically diagnosed, symptomatic patients with regard to mutations, enzyme activity, phenotype, and age of disease onset. RESULTS: VLCAD molecular analyses in 6 unrelated patients revealed the previously reported V243A mutation, associated with hepatic or myopathic phenotypes, on 7/12 alleles. All other mutations were also missense mutations. Residual VLCAD activities of 6% to 11% of normal were consistent with milder phenotypes. In these identified individuals treated prospectively with dietary modification as preventive measures, clinical symptoms did not develop during follow-up. CONCLUSIONS: MS/MS-based newborn screening correctly identifies VLCAD-deficient individuals. Based on mutational and enzymatic findings, these infants probably are at risk of future disease. Because life-threatening metabolic derangement can occur even in otherwise mild phenotypes, we advocate universal newborn screening programs for VLCAD deficiency to detect affected patients and prevent development of metabolic crises. Longer-term follow-up is essential to define outcomes, the definite risk of future disease, and appropriate treatment recommendations.     

An asymptomatic infant with isolated 3-methylcrotonyl-coenzyme A carboxylase deficiency detected by newborn screening for maple syrup urine disease

We describe an asymptomatic male infant with isolated 3-methylcrotonyl-coenzyme A carboxylase deficiency who came to medical attention by newborn mass screening due to elevated blood leucine. The diagnosis was made by abnormal urinary organic acids at 20 days of age and was confirmed by assay of the carboxylase activities in cultured skin fibroblasts. Conclusion More attention should be paid to slight elevations of leucine levels in newborn mass screening. Urinary organic acid analysis should be performed in conspicuous cases. Received: 9 October 1996 / Received in revised form and accepted: 18 February 1997     

Molecular characterisation and neuropsychological outcome of 21 patients with profound biotinidase deficiency detected by newborn screening and family studies

Early recognition by newborn screening and oral biotin supplementation may prevent clinical and neurological deficits in profound biotinidase deficiency (residual plasma biotinidase activity <10%). In order to evaluate possible correlations of molecular characteristics, onset and continuation of treatment and clinical outcome, we investigated 21 patients detected by newborn screening and consecutive family investigations. In 18 patients found by newborn screening, the range of biotinidase activities was 0%-9% residual activity. Application of a sensitive HPLC assay enabled us to discriminate five patients with residual biotinidase activities <1%. Two patients with zero activities were homozygous for the G98:d7i3 mutation and three patients with activities <1% carried mutations G98:d7i3, R157H, and Q456H. The mutation spectrum of the remaining patients included T532M, A171T+D444H, V62M,C432W, and D444H. Evaluation of clinical and neuropsychological outcome showed that only patients with biotinidase activities <1% exhibited characteristic clinical symptoms within the first weeks of life whereas five patients with residual activities of 1.2%-4.6% did not develop clinical symptoms even when not treated until 3.5-21 years. In all patients treated with biotin within the first weeks of life, neuropsychological outcome was normal whereas abnormal in three out of five patients tested for IQ and treated after the age of 3.5 years. CONCLUSION: The clinical and molecular spectrum of profound biotinidase deficiency is heterogeneous. Early onset of symptoms is predicted by the presence of zero residual activity as measured by sensitive assays and by homozygosity for the G98:d7i3 mutation. In patients with higher residual activities and variable mutational spectrum, correlation with the onset and severity of symptoms cannot be made.     

Pancreatic phenotype in infants with cystic fibrosis identified by mutation screening

Objective

To determine the pancreatic phenotype of infants with cystic fibrosis (CF) diagnosed in the first week of life by a combined immunoreactive trypsin/mutation screening program.

Design

A prospective evaluation of pancreatic function in infants with CF at the time of neonatal diagnosis and up to the age of 12.

Setting

Two different centres (Verona, Italy and Westmead, Australia) to enable comparison of results between two regions where <60% or ⩾90% of patients, respectively, have at least one single ΔF508 a mutation.

Patients

315 children with CF including 149 at Verona and 166 at Westmead.

Interventions

Fat balance studies over 3–5 days and pancreatic stimulation tests with main outcome measures being faecal fat or pancreatic colipase secretion. Patients with malabsorption are pancreatic insufficient (PI) or with normal absorption and pancreatic sufficient (PS).

Results

34 infants (23%) at Verona and 46 (28%) at Westmead were PS at diagnosis. 15% of those with two class I, II or III “severe” mutations and 26/28 (93%) of those with class IV or V mutations were PS at this early age. Of the 80 infants with PS, 20 became PI before the age of 12. All 20 had two severe mutations.

Conclusion

Neonatal mutational screening programs for CF are less likely to detect PS patients with non‐ΔF508 mutations. Of PS patients who are detected, those with two severe class I, II or III mutations are at particularly high risk of becoming PI during early childhood.Neonatal screening for cystic fibrosis (CF) began in the early 1980s with measurement of immunoreactive trypsin (IRT) assays on dried blood spots obtained from infants in the first 4–6 weeks of life.^1,2 Questions were raised about the reliability of these programs as they had not been validated for the 10%–15% of patients with normal fat absorption (pancreatic sufficient or PS).³ Subsequently however, 37% of screened infants were found to be PS,^4,5 that is 2–3 times the 10%–15% observed in older patients.³ Furthermore, follow‐up studies demonstrated that nearly 50% of the infants with PS near birth developed fat malabsorption (pancreatic insufficient or PI) in later childhood, thus accounting for the lower proportion of PS patients at an older age. It is significant that all those who transitioned from being PS to PI had two “severe” class I, II or III cystic fibrosis transmembrane conductance regulator (CFTR) mutations,⁶ as defined previously,^7,8 whereas those with persistent PS had at least one class IV or V “mild” mutation.Following the discovery of the CFTR gene,⁹ most screening programs have modified their screening strategy.¹⁰ In regions where more than 90% of patients with CF have at least one ΔF508 mutation, analysis for ΔF508 is performed on week 1 neonatal blood spots with an elevated IRT level.¹¹ This strategy has been further modified in regions (particularly southern Europe) where ΔF508 is only present in 40%–60% of patients, by screening for an extended panel of non‐ΔF508 mutations (usually severe class I, II or III) common to that region.^12,13 These IRT/DNA screening strategies are, however, not without problems. For instance, in regions where ΔF508 is responsible for CF in over 90% of cases, up to 10% of patients have non‐ΔF508 mutations with a preponderance of PS patients¹⁴ and therefore screening for ΔF508 alone would not detect such patients.Currently, only limited data are available on the pancreatic phenotype of infants with CF diagnosed by IRT/DNA screening from a study¹⁵ restricted to 27 infants with only class I or II mutations. The present study was thus undertaken, firstly to determine the pancreatic phenotype of infants in either a ΔF508 predominant or non‐dominant region, and secondly, to determine what proportion of infants initially with PS later transitioned to being PI and whether this was determined by the presence of two severe mutations.