Consensus recommendations for the assessment and management of idiopathic intracranial hypertension in children and young people.

BACKGROUND: Idiopathic intracranial hypertension (IIH) is a potentially disabling condition. There is a lack of evidence and national guidance on how to diagnose and treat paediatric IIH, leading to variation in clinical practice. We conducted a national Delphi consensus via the Children's Headache Network to propose a best-practice diagnostic and therapeutic pathway. METHODS: The Delphi process was selected as the most appropriate methodology for examining current opinion among experts in the UK. 104 questions were considered by 66 healthcare professionals, addressing important aspects of IIH care: assessment, diagnosis, treatment, follow-up and surveillance. General paediatricians, paediatric neurologists, ophthalmologists, opticians, neuroradiologists and neurosurgeons with a clinical interest or experience in IIH, were invited to take part. RESULTS: The Delphi process consisted of three rounds comprising 104 questions (round 1, 67; round 2, 24; round 3 (ophthalmological), 13) and was completed between March 2019 and August 2021. There were 54 and 65 responders in the first and second rounds, respectively. The Delphi was endorsed by the Royal College of Ophthalmologists, which engaged 59 ophthalmologists for round 3. CONCLUSIONS: This UK-based Delphi consensus process reached agreement for the management of paediatric IIH and has been endorsed by the Children's Headache Network and more broadly, the British Paediatric Neurology Association. It provides a basis for a pragmatic clinical approach. The recommendations will help to improve clinical care while minimising under and over diagnosis.

Classic ketogenic diet versus further antiseizure medicine in infants with drug-resistant epilepsy (KIWE): a UK, multicentre, open-label, randomised clinical trial.

BACKGROUND: Many infancy-onset epilepsies have poor prognosis for seizure control and neurodevelopmental outcome. Ketogenic diets can improve seizures in children older than 2 years and adults who are unresponsive to antiseizure medicines. We aimed to establish the efficacy of a classic ketogenic diet at reducing seizure frequency compared with further antiseizure medicine in infants with drug-resistant epilepsy. METHODS: In this phase 4, open-label, multicentre, randomised clinical trial, infants aged 1-24 months with drug-resistant epilepsy (defined as four or more seizures per week and two or more previous antiseizure medications) were recruited from 19 hospitals in the UK. Following a 1-week or 2-week observation period, participants were randomly assigned using a computer-generated schedule, without stratification, to either a classic ketogenic diet or a further antiseizure medication for 8 weeks. Treatment allocation was masked from research nurses involved in patient care, but not from participants. The primary outcome was the median number of seizures per day, recorded during weeks 6-8. All analyses were by modified intention to treat, which included all participants with available data. Participants were followed for up to 12 months. All serious adverse events were recorded. The trial is registered with the European Union Drug Regulating Authorities Clinical Trials Database (2013-002195-40). The trial was terminated early before all participants had reached 12 months of follow-up because of slow recruitment and end of funding. FINDINGS: Between Jan 1, 2015, and Sept 30, 2021, 155 infants were assessed for eligibility, of whom 136 met inclusion criteria and were randomly assigned; 75 (55%) were male and 61 (45%) were female. 78 infants were assigned to a ketogenic diet and 58 to antiseizure medication, of whom 61 and 47, respectively, had available data and were included in the modifified intention-to-treat analysis at week 8. The median number of seizures per day during weeks 6-8, accounting for baseline rate and randomised group, was similar between the ketogenic diet group (5 [IQR 1-16]) and antiseizure medication group (3 [IQR 2-11]; IRR 1·33, 95% CI 0·84-2·11). A similar number of infants with at least one serious adverse event was reported in both groups (40 [51%] of 78 participants in the ketogenic diet group and 26 [45%] of 58 participants in the antiseizure medication group). The most common serious adverse events were seizures in both groups. Three infants died during the trial, all of whom were randomly assigned a ketogenic diet: one child (who also had dystonic cerebral palsy) was found not breathing at home; one child died suddenly and unexpectedly at home; and one child went into cardiac arrest during routine surgery under anaesthetic. The deaths were judged unrelated to treatment by local principal investigators and confirmed by the data safety monitoring committee. INTERPRETATION: In this phase 4 trial, a ketogenic diet did not differ in efficacy and tolerability to a further antiseizure medication, and it appears to be safe to use in infants with drug-resistant epilepsy. A ketogenic diet could be a treatment option in infants whose seizures continue despite previously trying two antiseizure medications. FUNDING: National Institute for Health and Care Research.

Genome sequencing and comprehensive rare-variant analysis of 465 families with neurodevelopmental disorders.

Despite significant progress in unraveling the genetic causes of neurodevelopmental disorders (NDDs), a substantial proportion of individuals with NDDs remain without a genetic diagnosis after microarray and/or exome sequencing. Here, we aimed to assess the power of short-read genome sequencing (GS), complemented with long-read GS, to identify causal variants in participants with NDD from the National Institute for Health and Care Research (NIHR) BioResource project. Short-read GS was conducted on 692 individuals (489 affected and 203 unaffected relatives) from 465 families. Additionally, long-read GS was performed on five affected individuals who had structural variants (SVs) in technically challenging regions, had complex SVs, or required distal variant phasing. Causal variants were identified in 36% of affected individuals (177/489), and a further 23% (112/489) had a variant of uncertain significance after multiple rounds of re-analysis. Among all reported variants, 88% (333/380) were coding nuclear SNVs or insertions and deletions (indels), and the remainder were SVs, non-coding variants, and mitochondrial variants. Furthermore, long-read GS facilitated the resolution of challenging SVs and invalidated variants of difficult interpretation from short-read GS. This study demonstrates the value of short-read GS, complemented with long-read GS, in investigating the genetic causes of NDDs. GS provides a comprehensive and unbiased method of identifying all types of variants throughout the nuclear and mitochondrial genomes in individuals with NDD.

Refinements and considerations for trio whole-genome sequence analysis when investigating Mendelian diseases presenting in early childhood.

To facilitate early deployment of whole-genome sequencing (WGS) for severely ill children, a standardized pipeline for WGS analysis with timely turnaround and primary care pediatric uptake is needed. We developed a bioinformatics pipeline for comprehensive gene-agnostic trio WGS analysis of children suspected of having an undiagnosed monogenic disease that included detection and interpretation of primary genetic mechanisms of disease, including SNVs/indels, CNVs/SVs, uniparental disomy (UPD), imprinted genes, short tandem repeat expansions, mobile element insertions, SMN1/2 copy number calling, and mitochondrial genome variants. We assessed primary care practitioner experience and competence in a large cohort of 521 families (comprising 90% WGS trios). Children were identified by primary practitioners for recruitment, and we used the UK index of multiple deprivation to confirm lack of patient socio-economic status ascertainment bias. Of the 521 children sequenced, 176 (34%) received molecular diagnoses, with rates as high as 45% for neurology clinics. Twenty-three of the diagnosed cases (13%) required bespoke methods beyond routine SNV/CNV analysis. In our multidisciplinary clinician user experience assessment, both pediatricians and clinical geneticists expressed strong support for rapid WGS early in the care pathway, but requested further training in determining patient selection, consenting, and variant interpretation. Rapid trio WGS provides an efficacious single-pass screening test for children when deployed by primary practitioners in clinical settings that carry high a priori risk for rare pediatric disease presentations.

Fifteen-minute consultation: The efficient investigation of infantile and childhood epileptic encephalopathies in the era of modern genomics.

The investigation of children presenting with infantile and childhood epileptic encephalopathies (ICEE) is challenging due to diverse aetiologies, overlapping phenotypes and the relatively low diagnostic yield of MRI, electroencephalography (EEG) and biochemical investigations. Careful history and thorough examination remain essential as these may identify an acquired cause or indicate more targeted investigation for a genetic disorder. Whole exome sequencing (WES) with analysis of a panel of candidate epilepsy genes has increased the diagnostic yield. Whole genome sequencing (WGS), particularly as a trio with both parents' DNA, is likely to supersede WES. Modern genomic investigation impacts on the timing and necessity of other testing. We propose a structured approach for children presenting with ICEE where there is diagnostic uncertainty, emphasising the importance of WGS or, if unavailable, WES early in the investigative process. We note the importance of expert review of all investigations, including radiology, neurophysiology and biochemistry, to confirm the technique used was appropriate as well as the results. It is essential to counsel families on the risks associated with the procedures, the yield of the procedures, findings that are difficult to interpret and implication of 'negative' results. Where children remain without a diagnosis despite comprehensive investigation, we note the importance of ongoing multidisciplinary care.

Novel compound heterozygous STN1 variants are associated with Coats Plus syndrome.

AIM: Coats plus syndrome (CP) is a rare autosomal recessive disorder, characterised by retinal telangiectasia exudates (Coats disease), leukodystrophy, distinctive intracranial calcification and cysts, as well as extra-neurological features including abnormal vasculature of the gastrointestinal tract, portal hypertension and osteopenia with a tendency to fractures. CP most frequently occurs due to loss-of-function mutations in CTC1. The encoded protein CTC1 constitutes part of the CST (CTC1-STN1-TEN1) complex, and three patients have been described with CP due to biallelic mutations in STN1. Together with the identification of homozygosity for a specific loss-of-function mutation in POT1 in a sibling pair, these observations highlight a defect in the maintenance of telomere integrity as the cause of CP, although the precise mechanism leading to the micro-vasculopathy seen at a pathological level remains unclear. Here, we present the investigation of a fourth child who presented to us with retinal exudates, intracranial calcifications and developmental delay, in keeping with a diagnosis of CP, and later went on to develop pancytopenia and gastrointestinal bleeding. Genome sequencing revealed compound heterozygous variants in STN1 as the likely genetic cause of CP in this present case. METHODS: We assessed the phenotype to be CP and undertook targeted sequencing. RESULTS: Whilst sequencing of CTC1 and POT1 was normal, we identified novel compound heterozygous variants in STN1 (previous gene symbol OBFC1): one loss-of-function--c.894dup (p.(Asp299Argfs*58)); and one missense--c.707T>C (p.(Leu236Pro)). CONCLUSION: Given the clinical phenotype and identified variants we suggest that this is only the fourth patient reported to date with CP due to mutations in STN1.

Interaction of cannabidiol with other antiseizure medications: A narrative review.

OBJECTIVE: Cannabidiol is efficacious as an adjunctive treatment in children with epilepsy associated with Dravet and Lennox-Gastaut syndromes. As its role is currently adjunctive, we reviewed the interactions of cannabidiol with other antiseizure medications (ASMs). METHODS: A search of Cochrane, Pubmed and Embase databases from January 2015 to April 2020 was performed. All original research papers discussing interactions between cannabidiol and ASMs were included. Bibliographies of review articles were searched to identify further papers. Adverse events and side effects were excluded. RESULTS: Cannabidiol interacts with ASMs through both pharmacokinetic and pharmacodynamic mechanisms. Thirty studies were identified (eighteen observational cohort studies, two randomised-control trials, three case reports/series, three animal studies, two briefing reports, an analysis of cohort data and a clinical trial simulation). There is potential for pharmacokinetic interactions between CBD and brivaracetam, clobazam, eslicarbazepine, lacosamide, gabapentin, oxcarbazepine, phenobarbital, potassium bromide, pregabalin, rufinamide, sirolimus/everolimus, stiripentol, tiagabine, topiramate and zonisamide. Pharmacodynamic interactions were identified for clobazam, valproate and levetiracetam. An animal study identified that the brain concentration of ASMs may be altered while the serum concentration remains the same. CONCLUSION: Pharmacokinetic and pharmacodynamic interactions exist between cannabidiol and ASMs. The cytochrome p450 system in particular has been implicated in pharmacokinetic interactions, although not exclusively. The existing literature is limited for some ASMs by studies having relatively small cohorts. As increasing numbers of patients use cannabidiol, specialists need to monitor closely for interactions clinically and with blood levels when required.

The training and organization of Paediatric Neurology in Europe: Special report of the European Paediatric Neurology Society & Committee of National Advisors.

BACKGROUND: Paediatric Neurology (PN) is a discipline focused on diagnosis, comprehensive management and research into diseases of the central and peripheral nervous system from fetal life to transition into adulthood. The European Paediatric Neurology Society first designed and published the European PN training programme in the European Paediatric Neurology Syllabus in 2002. This was important in gaining recognition for the sub-specialty from the European Academy of Paediatrics and the European Academy of Neurology and in 2003 PN was recognized as a sub-specialty of paediatrics and neurology by the Board of the European Union of Medical Specialties. In 2004, the EPNS founded the Committee of National Advisors (CNA) that comprised representatives from national Paediatric Neurology societies, in order to further enhance Europe wide standards in training and practice., The EPNS Training Advisory Board (TAB) offers nation specific advice/support to PN societies on developing training and care systems. In 2019, the 2nd revision of the Paediatric Neurology Syllabus was approved by the EPNS Board and CNA. We aim to give an overview of the training of Paediatric Neurology (PN) specialists (i.e. Paediatric Neurologists), the relevant professional bodies and the current practice of Paediatric Neurology in Europe, as defined geographically by the World Health Organization. METHODS: A structured online data collection form was completed by CNA representatives from European countries. The data included training routes and structure of training, epidemiological data, nature of professional societies, organization of Paediatric Neurology care, research, academic life and recognition of the specialty. RESULTS: Data was collected from 43 European countries of which 38 have a national PN Society. In 10 (6 European Union (EU) and 4 non-EU countries) PN is recognized as a core specialty. In 26 countries PN is recognized as a sub-specialty of Paediatrics, Neurology or both (15 EU-11 non-EU). PN is not recognized as a core or sub-specialty in 7 countries (4 EU and 3 non-EU). In 35 countries paediatric neurologists begin their training from Paediatrics, but in 19 countries PN training from Neurology is also possible or the preferred route. Training in PN differs, but in over 50% of countries the three main training modules named in the 2019 2nd revision of the European PN Syllabus (PN, Paediatrics and adult Neurology) are included. Many countries have already adapted their curriculum to the suggestions in the European PN syllabus. CONCLUSIONS: There is diversity among European countries in terms of professional organization and PN training. The European PN syllabus has had impact on the development of PN training throughout Europe, independent of duration of training or route from paediatrics or neurology. The syllabus provides a basis for the future development of PN training, the recognition of PN as a (sub) specialty in individual countries and for improving the care of children with neurological disorders in Europe.

Phenotypic and genetic spectrum of epilepsy with myoclonic atonic seizures.

OBJECTIVE: We aimed to describe the extent of neurodevelopmental impairments and identify the genetic etiologies in a large cohort of patients with epilepsy with myoclonic atonic seizures (MAE). METHODS: We deeply phenotyped MAE patients for epilepsy features, intellectual disability, autism spectrum disorder, and attention-deficit/hyperactivity disorder using standardized neuropsychological instruments. We performed exome analysis (whole exome sequencing) filtered on epilepsy and neuropsychiatric gene sets to identify genetic etiologies. RESULTS: We analyzed 101 patients with MAE (70% male). The median age of seizure onset was 34 months (range = 6-72 months). The main seizure types were myoclonic atonic or atonic in 100%, generalized tonic-clonic in 72%, myoclonic in 69%, absence in 60%, and tonic seizures in 19% of patients. We observed intellectual disability in 62% of patients, with extremely low adaptive behavioral scores in 69%. In addition, 24% exhibited symptoms of autism and 37% exhibited attention-deficit/hyperactivity symptoms. We discovered pathogenic variants in 12 (14%) of 85 patients, including five previously published patients. These were pathogenic genetic variants in SYNGAP1 (n = 3), KIAA2022 (n = 2), and SLC6A1 (n = 2), as well as KCNA2, SCN2A, STX1B, KCNB1, and MECP2 (n = 1 each). We also identified three new candidate genes, ASH1L, CHD4, and SMARCA2 in one patient each. SIGNIFICANCE: MAE is associated with significant neurodevelopmental impairment. MAE is genetically heterogeneous, and we identified a pathogenic genetic etiology in 14% of this cohort by exome analysis. These findings suggest that MAE is a manifestation of several etiologies rather than a discrete syndromic entity.

Clinical and radiological characterization of novel FIG4-related combined system disease with neuropathy.

Variants in the FIG4 gene, which encodes a phosphatidylinositol-3,5-bisphosphatase lead to obstruction of endocytic trafficking, causing accumulation of enlarged vesicles in murine peripheral neurons and fibroblasts. Bi-allelic pathogenic variants in FIG4 are associated with neurological disorders including Charcot-Marie-Tooth disease type-4J (CMT4J) and Yunis-Varón syndrome (YVS). We present four probands from three unrelated families, all homozygous for a recurrent FIG4 missense variant c.506A>C p.(Tyr169Ser), with a novel phenotype involving features of both CMT4J and YVS. Three presented with infant-onset dystonia and one with hypotonia. All have depressed lower limb reflexes and distal muscle weakness, two have nerve conduction studies (NCS) consistent with severe sensorimotor demyelinating peripheral neuropathy and one had NCS showing patchy intermediate/mildly reduced motor conduction velocities. All have cognitive impairment and three have swallowing difficulties. MRI showed cerebellar atrophy and bilateral T2 hyperintense medullary swellings in all patients. These children represent a novel clinicoradiological phenotype and suggest that phenotypes associated with FIG4 missense variants do not neatly fall into previously described diagnoses but can present with variable features. Analysis of this gene should be considered in patients with central and peripheral neurological signs and medullary radiological changes, providing earlier diagnosis and informing reproductive choices.

Correlation between the total number of features of paediatric pseudotumour cerebri syndrome and cerebrospinal fluid pressure.

PURPOSE: Accurate diagnosis of pseudotumour cerebri syndrome (PTCS) in children is challenging. We aimed to see if the clinical and radiological assessment that is carried out before lumbar puncture could predict subsequently recorded CSF pressures, and thus whether it could be used to increase diagnostic certainty of paediatric PTCS. METHODS: We used internationally recognised diagnostic criteria to derive a list of clinical, brain neuroimaging and venography features that were accepted to be associated with a diagnosis of PTCS. We performed a retrospective cohort study of children referred to our centre with suspected PTCS, identifying the presence or absence of those features for each child at initial presentation. The sum total scores of the features that were present were correlated with the child's recorded CSF pressure. RESULTS: The sum total scores were significantly positively correlated with recorded CSF pressures. The positive correlation was seen when clinical and brain neuroimaging features were included alone, and the correlation was slightly stronger when venography features were included in addition. CONCLUSION: Calculating the sum total of clinical, brain neuroimaging and venography features (where venography is performed) present at initial presentation can help in the management of children under investigation for PTCS. Children with high scores are more likely to have severely raised CSF pressures and thus may warrant more urgent LP investigations. By contrast, in children with subtle abnormalities in optic disc appearance such that disc oedema cannot be ruled out, a low score may add further reassurance and less urgency to proceed to LP.

Fifteen-minute consultation: Efficient investigation of the child with early developmental impairment in the era of genomic sequencing.

The investigation of children with early developmental impairment (EDI) is challenging in terms of selecting investigations and supporting families through the diagnostic pathway. Modern genomic sequencing has the potential to greatly improve yield of investigation, but produces challenges in terms of timing and explaining its strengths/weaknesses to families. We present an evidence-based and practical guideline to help the paediatrician through all stages of investigation. We emphasise the importance of a really good history and examination, allowing targeted investigation for specific disorders and outline an approach for isolated EDI when this is not possible. This prioritises genetic investigation- after appropriate counselling to families, and balances the very low yield of biochemical/radiological investigations in isolated EDI, with the need to detect extremely rare, but potentially treatable disorders. Collaboration with both families and regional specialists to ensure appropriate testing is likely to reduce parental and clinician anxiety.

PEHO syndrome: the endpoint of different genetic epilepsies.

BACKGROUND: Progressive encephalopathy, hypsarrhythmia and optic atrophy (PEHO) has been described as a clinically distinct syndrome. It has been postulated that it is an autosomal recessive condition. However, the aetiology is poorly understood, and the genetic basis of the condition has not been fully elucidated. Our objective was to discover if PEHO syndrome is a single gene disorder. METHOD: Children with PEHO and PEHO-like syndrome were recruited. Clinical, neurological and dysmorphic features were recorded; EEG reports and MRI scans were reviewed. Where possible, exome sequencing was carried out first to seek mutations in known early infantile developmental and epileptic encephalopathy (DEE) genes and then to use an agnostic approach to seek novel candidate genes. We sought intra-interfamilial phenotypic correlations and genotype-phenotype correlations when pathological mutations were identified. RESULTS: Twenty-three children were recruited from a diverse ethnic background, 19 of which were suitable for inclusion. They were similar in many of the core and the supporting features of PEHO, but there was significant variation in MRI and ophthalmological findings, even between siblings with the same mutation. A pathogenic genetic variant was identified in 15 of the 19 children. One further girl's DNA failed analysis, but her two affected sisters shared confirmed variants. Pathogenic variants were identified in seven different genes. CONCLUSIONS: We found significant clinical and genetic heterogeneity. Given the intrafamily variation demonstrated, we question whether the diagnostic criteria for MRI and ophthalmic findings should be altered. We also question whether PEHO and PEHO-like syndrome represent differing points on a clinical spectrum of the DEE. We conclude that PEHO and PEHO-like syndrome are clinically and genetically diverse entities-and are phenotypic endpoints of many severe genetic encephalopathies.

Genome annotation for clinical genomic diagnostics: strengths and weaknesses.

The Human Genome Project and advances in DNA sequencing technologies have revolutionized the identification of genetic disorders through the use of clinical exome sequencing. However, in a considerable number of patients, the genetic basis remains unclear. As clinicians begin to consider whole-genome sequencing, an understanding of the processes and tools involved and the factors to consider in the annotation of the structure and function of genomic elements that might influence variant identification is crucial. Here, we discuss and illustrate the strengths and weaknesses of approaches for the annotation and classification of important elements of protein-coding genes, other genomic elements such as pseudogenes and the non-coding genome, comparative-genomic approaches for inferring gene function, and new technologies for aiding genome annotation, as a practical guide for clinicians when considering pathogenic sequence variation. Complete and accurate annotation of structure and function of genome features has the potential to reduce both false-negative (from missing annotation) and false-positive (from incorrect annotation) errors in causal variant identification in exome and genome sequences. Re-analysis of unsolved cases will be necessary as newer technology improves genome annotation, potentially improving the rate of diagnosis.

Ketogenic diet in the treatment of epilepsy in children under the age of 2 years: study protocol for a randomised controlled trial.

BACKGROUND: The incidence of epilepsy is greatest in the first 2 years of life, an age group where there is generally a poor prognosis for both seizure control and neurodevelopmental outcome. Early control of seizures can be associated with better developmental outcome but many of the epilepsies presenting in infancy are poorly responsive to antiepileptic medication. The ketogenic diet (KD) is a high-fat, low-carbohydrate diet designed to mimic the effects of starvation on the body. Dietary fat is converted into ketones in the body and used as an energy source by the brain. The KD has been shown to be successful in controlling seizures in many observational studies, and in two randomised controlled trials (RCTs) in older children. However, little evidence is available in the very young. METHODS/DESIGN: An open-label RCT where eligible children (age 3 months to 2 years with epilepsy who have failed two antiepileptic drugs (AEDs)) undergo baseline assessment, including medical and seizure history. Participants then start an observation period (7 or 14 days) with documentation of seizure frequency. Randomisation will occur on day 8 or day 15 to receive the KD or a further AED; the allocated treatment will commence on day 15, with instruction and training. A second assessment (4 weeks after start of treatment) will include a clinical review and tolerability questionnaire (modified Hague Scale of Side Effects - for those allocated to the KD group). Assessments will be repeated at 8 weeks after the start of treatment including biochemical investigations, after which, according to patient response, KD (diet group) or AED (standard AED group) will then be continued or changed. Those in the AED group who have failed to achieve seizure control at the 8-week assessment will then be offered KD outside the context of the trial. Those in the KD arm who fail to achieve seizure control will be changed to standard clinical management. All patients will be followed up for 12 months from randomisation for retention, seizure outcome, quality of life and neurodevelopmental status. DISCUSSION: The slow rate of recruitment is an ongoing practical issue. There is a limitation to the number of eligible patients compared to what was predicted, mainly due to the nature of this patient group. After a substantial amendment to widen inclusion criteria and reduce the baseline period to 7 days for patients with a high seizure burden, the rate of recruitment steadily increased. A number of operational concerns regarding dietetic time were also highlighted impacting on the recruitment rate. However, the combination of a low dropout rate and the opening of further centres, the trial should successfully meet the final recruitment target. All nine centres are now recruiting and we hope to open further centres within the UK. TRIAL REGISTRATION: ClinicalTrials.gov, identifier: NCT02205931 . Registered on 16 December 2013.

Mutations in SLC25A22: hyperprolinaemia, vacuolated fibroblasts and presentation with developmental delay.

Mutations in SLC25A22 are known to cause neonatal epileptic encephalopathy and migrating partial seizures in infancy. Using whole exome sequencing we identified four novel SLC25A22 mutations in six children from three families. Five patients presented clinical features similar to those in the literature including hypotonia, refractory neonatal-onset seizures and developmental delay. However, the sixth patients presented atypically with isolated developmental delay, developing late-onset (absence) seizures only at 7 years of age. Abnormal metabolite levels have not been documented in the nine patients described previously. One patient in our series was referred to the metabolic clinic because of persistent hyperprolinaemia and another three had raised plasma proline when tested. Analysis of the post-prandial plasma amino acid response in one patient showed abnormally high concentrations of several amino acids. This suggested that, in the fed state, when amino acids are the preferred fuel for the liver, trans-deamination of amino acids requires transportation of glutamate into liver mitochondria by SLC25A22 for deamination by glutamate dehydrogenase; SLC25A22 is an important mitochondrial glutamate transporter in liver as well as in brain. Electron microscopy of patient fibroblasts demonstrated widespread vacuolation containing neutral and phospho-lipids as demonstrated by Oil Red O and Sudan Black tinctorial staining; this might be explained by impaired activity of the proline/pyrroline-5-carboxylate (P5C) shuttle if SLC25A22 transports pyrroline-5-carboxylate/glutamate-γ-semialdehyde as well as glutamate.