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Newborn screening (NBS) for Krabbe disease (KD) is performed by measurement of galactocerebrosidase (GALC) activity as the primary test. This revealed that GALC activity has poor specificity for KD. Psychosine (PSY) was proposed as a disease marker useful to reduce the false positive rate for NBS and for disease monitoring. We report a highly sensitive PSY assay that allows identification of KD patients with minimal PSY elevations. PSY was extracted from dried blood spots or erythrocytes with methanol containing d5-PSY as internal standard, and measured by liquid chromatography–tandem mass spectrometry. Analysis of PSY in samples from controls (N = 209), GALC pseudodeficiency carriers (N = 55), GALC pathogenic variant carriers (N = 27), patients with infantile KD (N = 26), and patients with late-onset KD (N = 11) allowed for the development of an effective laboratory screening and diagnostic algorithm. Additional longitudinal measurements were used to track therapeutic efficacy of hematopoietic stem cell transplantion (HSCT). This study supports PSY quantitation as a critical component of NBS for KD. It helps to differentiate infantile from later onset KD variants, as well as from GALC variant and pseudodeficiency carriers. Additionally, this study provides further data that PSY measurement can be useful to monitor KD progression before and after treatment.

The prevalence of Fabry disease (FD) in adult patients with suspected hypertrophic cardiomyopathy (HCM) has been reported between 0.3% and 4%. Fabry disease-specific therapy necessitates early diagnosis; however, the optimal screening strategy and cost efficacy of routine α-galactosidase A (α-gal A) vs comprehensive galactosidase alpha gene (GLA) testing remain poorly understood. We identified 1192 patients who underwent routine α-gal A screening between January 1, 2011, and December 31, 2017, for suspected HCM. Cost efficacy was explored using prevalence and cost estimates. Ten patients had reduced α-gal A enzyme activity, and 5 (3 women) were ultimately diagnosed with FD (prevalence estimate, 0.42%). An alternative cardiac diagnosis was made in 3 patients with mildly reduced enzyme activity. Two women with reduced borderline enzyme levels did not undergo confirmatory testing, but FD was not suspected. The number needed to screen to diagnose 1 patient with FD in a similar cohort is estimated at 238 (5 new cases per 1192 at-risk individuals) at a cost of approximately US $24,000 per diagnosis. We identified a 0.42% prevalence of FD using routine α-gal A screening in adult patients referred to a dedicated HCM center in the United States. Compared with more comprehensive genetic testing strategies, we identified a similar prevalence of FD at a lower cost per diagnosis.

Enzyme-based newborn screening for Mucopolysaccharidosis type I (MPS I) has a high false-positive rate due to the prevalence of pseudodeficiency alleles, often resulting in unnecessary and costly follow up. The glycosaminoglycans (GAGs), dermatan sulfate (DS) and heparan sulfate (HS) are both substrates for &alpha, l-iduronidase (IDUA). These GAGs are elevated in patients with MPS I and have been shown to be promising biomarkers for both primary and second-tier testing. Since February 2016, we have measured DS and HS in 1213 specimens submitted on infants at risk for MPS I based on newborn screening. Molecular correlation was available for 157 of the tested cases. Samples from infants with MPS I confirmed by IDUA molecular analysis all had significantly elevated levels of DS and HS compared to those with confirmed pseudodeficiency and/or heterozygosity. Analysis of our testing population and correlation with molecular results identified few discrepant outcomes and uncovered no evidence of false-negative cases. We have demonstrated that blood spot GAGs analysis accurately discriminates between patients with confirmed MPS I and false-positive cases due to pseudodeficiency or heterozygosity and increases the specificity of newborn screening for MPS I.

Patients with Anderson-Fabry Disease (AFD) and severe left ventricular hypertrophy complicated by left ventricular outflow tract (LVOT) obstruction may benefit from surgical septal myectomy (SSM). Mid- and late outcomes following surgery have not been established, and we sought to better characterize postoperative outcomes following septal myectomy. Between January 2011 and June 2017, 7 patients (6 females) with AFD underwent SSM. The median (range) age at the time of surgery was 53 (37–72) years; 4 patients had a positive family history of AFD and a preoperative diagnosis of AFD. Extracardiac features suggestive of AFD were present in 3 patients and all but 1 (female) had reduced α-galactosidase A activity. All patients had severe left ventricular hypertrophy and LVOT obstruction on transthoracic echocardiography. Preoperatively, all patients were severely symptomatic with New York Heart Association (NYHA) class III symptoms. There was no early mortality following surgery. The median in-hospital length of stay was 5 (4–7) days with 6 patients reporting NYHA class II or less symptoms at 3 month follow-up. Long-term outcomes were favorable with 4 patients reporting sustained NYHA class II or less symptoms, but 2 patients had recurrence of NYHA class III symptoms without evidence of recurrent LVOT obstruction. In conclusion, SSM appears to provide favorable short- and long-term relief of severe, symptomatic LVOT obstruction but may not alter progression of Fabry cardiomyopathy.

Introduction There is limited understanding of relationships between genotype, phenotype and other conditions contributing to health in neonates with medium-chain acyl-coenzyme A dehydrogenase deficiency (MCADD) identified through newborn screening. Methods Retrospective analysis of comprehensive data from a cohort of 221 newborn-screened subjects identified as affected with MCADD in the Inborn Errors of Metabolism – Information System (IBEM-IS), a long term follow-up database of the Inborn Errors of Metabolism Collaborative, was performed. Results The average age at notification of first newborn screen results to primary care or metabolic providers was 7.45 days. The average octanoylcarnitine (C8) value on first newborn screen was 11.2 μmol/L (median 8.6, range 0.36–43.91). A higher C8 level correlated with an earlier first subspecialty visit. Subjects with low birth weight had significantly lower C8 values. Significantly higher C8 values were found in symptomatic newborns, in newborns with abnormal lab testing in addition to newborn screening and/or diagnostic tests, and in subjects homozygous for the c.985A > G ACADM gene mutation or compound heterozygous for the c.985A > G mutation and deletions or other known highly deleterious mutations. Subjects with neonatal symptoms, or neonatal abnormal labs, or neonatal triggers were more likely to have at least one copy of the severe c.985A > G ACADM gene mutation. C8 and genotype category were significant predictors of the likelihood of having neonatal symptoms. Neonates with select triggers were more likely to have symptoms and laboratory abnormalities. Conclusions This collaborative study is the first in the United States to describe health associations of a large cohort of newborn-screened neonates identified as affected with MCADD. The IBEM-IS has utility as a platform to better understand the characteristics of individuals with newborn-screened conditions and their follow-up interactions with the health system.

Leigh syndrome is a mitochondrial disease caused by mutations in different genes, including ATP6A for which no known therapy is available. We report a case of adult-onset Leigh syndrome with response to immunotherapy. A twenty year-old woman with baseline learning difficulties was admitted with progressive behavioral changes, diplopia, headaches, bladder incontinence, and incoordination. Brain MRI and PET scan showed T2 hyperintensity and increased uptake in bilateral basal ganglia, respectively. Autoimmune encephalitis was suspected and she received plasmapheresis with clinical improvement. She was readmitted 4 weeks later with dysphagia and aspiration pneumonia. Plasmapheresis was repeated with resolution of her symptoms. Given the multisystem involvement and suggestive MRI changes, genetic testing was done, revealing a homoplasmic T9176C ATPase 6 gene mtDNA mutation. Monthly IVIG provided clinical improvement with worsening when infusions were delayed. Leigh syndrome secondary to mtDNA T9176C mutations could have an autoimmune mechanism that responds to immunotherapy.

Past murine studies of phenylketonuria (PKU) have documented significant effects on cerebellum at both the gross and cellular levels. The profile of neurocognitive and motor difficulties associated with early-treated PKU (ETPKU) is also consistent with potential cerebellar involvement. Previous neuroanatomical studies of cerebellum in patients with PKU, however, have yielded mixed results. The objective of the present study was to further examine potential differences in cerebellar morphometry between individuals with and without ETPKU. To this end, we analyzed high resolution T1-weighted MR images from a sample of 20 individuals with ETPKU and an age-matched comparison group of 20 healthy individuals without PKU. Measurements of whole brain volume, whole cerebellum volume, cerebellar gray matter volume, and cerebellar white matter volume were collected by means of semiautomatic volumetric analysis. Data analysis revealed no significant group differences in whole brain volume, whole cerebellar volume, or cerebellar white matter volume. A significant reduction in cerebellar gray matter volume, however, was observed for the ETPKU group compared to the non-PKU comparison group. These findings expand on previous animal work suggesting that cerebellar gray matter is impacted by PKU. It is also consistent with the hypothesis that the cognitive difficulties experienced by individuals with ETPKU may be related to disruptions in gray matter. Additional studies are needed to fully elucidate the timing and extent of the impact of ETPKU on cerebellum and the associated neurocognitive consequences., Highlights • Past animal and human work suggests cerebellum may be impacted by PKU. • Significant reduction in cerebellar gray matter volume observed for ETPKU. • Cerebellar white matter volumes similar for ETPKU and non-PKU participants.

The implementation of newborn screening for lysosomal disorders has uncovered overall poor specificity, psychosocial harm experienced by caregivers, and costly follow-up testing of false-positive cases. We report an informatics solution proven to minimize these issues. The Kentucky Department for Public Health outsourced testing for mucopolysaccharidosis type I (MPS I) and Pompe disease, conditions recently added to the recommended uniform screening panel, plus Krabbe disease, which was added by legislative mandate. A total of 55,161 specimens were collected from infants born over 1 year starting from February 2016. Testing by tandem mass spectrometry was integrated with multivariate pattern recognition software (Collaborative Laboratory Integrated Reports), which is freely available to newborn screening programs for selection of cases for which a biochemical second-tier test is needed. Of five presumptive positive cases, one was affected with infantile Krabbe disease, two with Pompe disease, and one with MPS I. The remaining case was a heterozygote for the latter condition. The false-positive rate was 0.0018% and the positive predictive value was 80%. Postanalytical interpretive tools can drastically reduce false-positive outcomes, with preliminary evidence of no greater risk of false-negative events, still to be verified by long-term surveillance.

To determine how US newborn dried bloodspot screening (NDBS) programs obtain patient-level data on clinical genetic counseling services offered to families of newborns identified through newborn NDBS and the extent to which newborns and their families receive these services. These data should serve to inform programs and lead to improved NDBS follow-up services. Collaborations were established with three state NDBS programs that reported systematically tracking genetic counseling services to newborns and their families identified through NDBS. A study protocol and data abstraction form were developed and IRB approvals obtained. Data from three state NDBS programs on a total of 151 patients indicated that genetic services are documented systematically only by metabolic clinics, most often by genetic counselors. Data from 69 endocrinology patients indicated infrequent referrals for genetic services; as expected higher for congenital adrenal hyperplasia than congenital hypothyroidism. Endocrinology patients were often counseled by physicians. While systematic tracking of genetic counseling services may be desirable for quality assurance of NDBS follow-up services, current systems do not appear conducive to this practice. Clinical records are not typically shared with NDBS programs and tracking of follow-up clinical genetic services has not been generally defined as a NDBS program responsibility. Rather, tracking of clinical services, while recognized as useful data, has been viewed by NDBS programs as a research project. The associated IRB requirements for patient-related research may pose an additional challenge. National guidance for NDBS programs that define quality genetic service indicators and monitoring responsibilities are needed. US experiences in this regard may provide information that can assist developing programs in avoiding tracking issues.

Because Cardio-facio-cutaneous (CFC) syndrome has significant phenotypic overlap with Costello syndrome, it may be difficult to establish the diagnosis on a clinical basis. The recent discoveries of germline HRAS mutations in patients with Costello syndrome and mutations in BRAF, MEK1, and MEK2 in CFC syndrome uncovered the biologic mechanism for the shared phenotypic findings based on the close interaction of the affected gene products within the MAP kinase pathway. We evaluated a series of patients who were either clinically diagnosed with Costello syndrome, or in whom the diagnoses of both Costello and CFC syndromes were considered. After excluding mutations in HRAS, we identified eight changes in BRAF and five in MEK1. Five mutations are novel, and all changes occurred de novo among those triads tested. A review of the clinical abnormalities showed important differences between patients with either a BRAF or MEK1 mutation, and those previously reported with an HRAS mutation. Statistical significance was achieved, despite the relatively small number of patients with BRAF and MEK1 mutations reported here, for polyhydramnios, growth hormone deficiency and the presence of more than one papilloma, which were less common in CFC compared to HRAS mutation positive patients. Although both CFC and Costello syndrome are characterized by cardiac abnormalities in about three-fourths of patients, the pattern of congenital heart defects (CHD), hypertrophic cardiomyopathy (HCM), and tachycardia differs somewhat. CHD, especially pulmonic stenosis associated with a secundum-type atrial septal defect, are more common in CFC than Costello syndrome (P = 0.02). Atrial tachycardia is less frequent in CFC patients with BRAF or MEK1 mutations, compared to Costello syndrome patients with HRAS mutation (P = 0.04). Chaotic atrial rhythm or multifocal atrial tachycardia was observed only in Costello syndrome. Malignant tumors have been viewed as characteristic for Costello syndrome due to HRAS mutations, however, we report here on a MEK1 mutation in a patient with a malignant tumor, a hepatoblastoma. Although this indicates that the presence of a tumor is not specific for Costello syndrome with HRAS mutation, it is noteworthy that the tumor histology differs from those commonly seen in Costello syndrome. Based on these clinical differences we suggest that patients with BRAF and MEK mutations should be diagnosed with CFC syndrome, and the diagnosis of Costello syndrome be reserved for patients with HRAS mutations.

The most widely-reported neurologic finding in individuals with early-treated phenylketonuria (PKU) is abnormality in the white matter of the brain. In contrast, much less is known regarding the impact of PKU on cortical gray matter (GM) structures. Presently, we applied advanced morphometric methods to the analysis of high-resolution structural MRI images from a sample of 19 individuals with early-treated PKU and an age- and gender-matched comparison group of 22 healthy individuals without PKU. Data analysis revealed decreased GM volume in parietal cortex for the PKU group compared with the non-PKU group. A similar trend was observed for occipital GM volume. There was no evidence of group-related differences in frontal or temporal GM volume. Within the PKU group, we also found a significant relationship between blood phenylalanine levels and GM volume for select posterior cortical sub-regions. Taken together with previous research on white matter and gray matter abnormalities in PKU, the present findings point to the posterior cortices as the primary site of neurostructural changes related to early-treated PKU.

We appreciate Dr. Finsterer et al. [1] interest in our case report [2]. They express doubts about the diagnosis of Leigh syndrome (LS). Our patient's diagnosis was based on the presence of a known ATPase 6 gene pathogenic mutation (T9176C), previously described in other LS cases [3], [4], [5], [6], [7], [8] , and also in a form of hereditary spastic paraplegia [9] leukodystrophy [10] and bilateral striatal necrosis [11].This mutation has been shown to affect ATP synthesis and the assembly or stability of complex V [9], [12]. There was no consanguinity in her family. The mother and other relatives refused genetic testing for personal (life insurance) and financial reasons. Patient and family also refused to have muscle biopsy done as she clinically improved. Magnetic resonance spectroscopy (MRS) was not performed because genetic testing found a known pathogenic mtDNA mutation. Regarding follow up imaging, brain MRI done 1 year later showed improvement in bilateral basal ganglia and periaqueductal region T2 hyperintensities. Patient did have a staring spell, with decreased oxygen saturation and was started on levetiracetam after her second admission. There was no family history of epilepsy. We agree with Dr. Finsterer et al. comprehensive list of POLG mutation-associated diseases. Regarding treatment effect, we want to reiterate that the exact mechanism by which plasmapheresis or immunoglobulin (IVIG) had beneficial effect in our patient is unknown; however as mentioned in the article, we think that reduced ATP production can increase the mitochondrial transmembrane potential (with resultant hyperpolarization) and increase reactive oxygen species (ROS) production [13], which can in turn activate the necroptosis pathway [14]. Impaired ATP synthesis can also induce necroptosis [15]. Resultant cell necrosis can lead to release of immunogenic material and activate immune and inflammation pathways (inflammasome) [14], [16]. IVIG has been shown to suppress inflammasome-mediated neuronal death in ischemic stroke models [17]. Of note, in mice, mitochondrial complex I mutations have been shown to cause inflammation and retinal ganglion cell death [18], further highlighting the possible relationship between mitochondrial mutations/diseases and inflammation. Additionally, a case of mitochondrial myopathy responsive to intravenous immunoglobulin [19] and patients with acute central nervous system inflammation possibly associated with ND4 and DARS2 mutations and responsive to plasmapheresis, steroids and rituximab have been described in a small report [20]. Further research in the role of autoimmunity and inflammation in mitochondrial disease is needed.

Previous research has documented white matter abnormalities in the brains of individuals with early-treated phenylketonuria (ETPKU). The majority of these past studies have relied on a region-based approach which focused on a limited number of spatially-defined regions within the brain. In the present study, we used diffusion tensor imaging (DTI) in conjunction with tract-based spatial statistics (TBSS) to perform an extensive examination of white matter tracts in the brains of ten individuals with ETPKU (mean age = 23.2 years) and 12 healthy non-PKU individuals (mean age = 23.5 years). Consistent with past research, we found that mean diffusivity (MD) was significantly restricted in the ETPKU group, and fractional anisotropy (FA) was comparable between the ETPKU and non-PKU groups. Moreover, we found restricted axial diffusivity (AD) and radial diffusivity (RD) in our ETPKU in numerous white matter tracts, suggesting widespread white matter compromise in ETPKU. In addition, this white matter pathology was more evident in older ETPKU participants with higher blood phenylalanine (phe) levels as compared to younger participants with lower phe levels.

Previous histological and neuroimaging studies have documented structural abnormalities in the white matter of the brain in individuals with early-treated phenylketonuria (ETPKU). It remains unclear, however, the extent to which the function of the brain’s interconnections are impacted by this condition. Presently, we utilized functional magnetic resonance imaging (fMRI) to evaluate the synchronization of neural signals (i.e., functional connectivity) among brain regions comprising the default mode network (DMN) in a sample of 11 individuals with ETPKU and 11 age- and gender-matched neurologically intact controls. The DMN is a group of interconnected brain regions that are known to be generally more active during rest than during task performance. Data analysis revealed decreased functional connectivity among DMN regions for the ETPKU group compared with the control group. Within the PKU group, we also found a significant relationship between blood phenylalanine (phe) levels and the functional connectivity between select regions of the DMN. In conjunction with findings from another recent fMRI study (Christ, Moffitt et al. 2010), the present results suggest that ETPKU-related deficiencies in functional connectivity are pervasive. The current findings also provide initial evidence that the extent of such impairment may be moderated in part by blood phe levels.

Phenylketonuria (PKU) is a genetic disorder associated with disruption of prefrontal cortex (PFC) development and executive dysfunction. To date, however, there is little evidence directly linking these two sequelae of PKU. We utilized functional magnetic resonance imaging (fMRI) to evaluate prefrontal functioning in six individuals with early-treated PKU (ETPKU) during performance of an n-back working memory task and compared results with those of six age- and gender-matched neurologically intact individuals. In addition, we evaluated the possible presence of PKU-related disruptions in functional connectivity, as might be hypothesized based on prior reports of white matter injury in individuals with ETPKU. A number of brain regions, nearly half of which were located in the PFC, were found to show atypical neural activity in individuals with ETPKU during working memory performance. We also found decreased connectivity both within the PFC as well as between the PFC and other brain regions in individuals with ETPKU compared with controls. Results from this preliminary study suggest that both prefrontal dysfunction and disruptions in functional connectivity may contribute to PKU-related executive impairment. In addition to advancing our understanding of PKU, the current findings have a broader impact in that PKU is regularly used as a model of early prefrontal dysfunction in the study of other neurodevelopmental disorders (e.g., autism).

Phenylketonuria (PKU) is a genetic disorder characterised by an inability to metabolise phenylalanine. Several studies have reported that the Corpus Callosum (CC) is one of the most severely affected structures with respect to volume loss in early treated PKU patients. In this work, we aim to detect the abnormalities of the CC in PKU from both global and local perspectives. 3D models of the CC are extracted from MRI data using a semiautomatic segmentation method. In the global analysis, raw and scaled volumes of the CC are compared between PKU patients and the controls. An oriented bounding box of the CC is constructed and its length, width and height are used as the MRI traits in our study. The raw and scaled values of these traits are compared between patients and controls. In the local analysis, shape differences at every surface point of the CC between PKU patients and the controls are computed using Hotelling T(2) two-sample metric followed by a permutation test. The height of the CC is found to be significantly shorter in the patients and significant shape abnormalities in the genu and splenium of the CC is also found in the patients.