Currently, at 4 years of age, the proband is on valproic acid 23 mg/kg/day and experiences no epileptic seizures. Her weight is 17 kg (50th centile), height 110 cm (97th centile), and occipito-frontal circumference 48 cm (<10th centile) She is severely developmentally delayed, although no formal psychological assessment has been performed yet. She cannot sit, there is short-duration eye-contact, response to commands is questionable. She does not speak a single word, presents with severe hypotonia with low muscle strength and a choreiform pattern of movements with stereotypies that do not wear off and do not respond to medication, although they often subside during the night. Episodes of upward eye gaze lasting up to 1 min without any epileptiform changes in EEG are observed (probably oculogyric crises). The proband wears hearing aids for bilateral congenital sensorineural deafness and has strabismus. She receives forlax daily for chronic constipation.

Head MRI performed at the age of 15 months was normal. Spinal MRI reveals occult spina bifida. Copper and ceruloplasmin levels were within normal limits. In EEG, epileptic graphoelements, burst of sharp and slow wave complexes, spikes, and polyspike-wave complexes were present. Acylcarnitines analysis in a dried blood spot by tandem mass spectrometry (MS/MS) and urine organic acids analysis by gas chromatography – mass spectroscopy (GC/MS) were normal. Amino acids in plasma and cerebrospinal fluid (CSF) showed no significant abnormalities. The CSF sample was taken according the standard protocol and the level of biogenic amine metabolites at the age of 21 months was normal.

A younger, now 10-month-old, sister of the proband was born by cesarean section at 39 weeks gestation. Birth weight was 3120 g (<50th centile), length 51 cm (>75th centile), and occipito-frontal circumference 33 cm (50th centile). Apgar scores were 10 at both 1 and 5 min. She had no major malformations or dysmorphic features at birth. At 9 months her weight was 8 kg (25th centile), height 68 cm (10th centile), and occipito-frontal circumference 44.5 cm (50th centile). At 10 months, she is hypotonic but with normal pattern of movements. The gross motor development is slightly delayed, which is most likely due to hypotonia. She started to sit with support at 9 months. She is a very social girl, she has a very good eye contact and executes plays that are nearly adequate for age (Table 1). Bilateral sensorineural deafness of moderate degree was diagnosed using auditory brainstem response testing, and hearing aids were appropriately fitted. No episodes of seizures have occurred since birth. No other typical features of SPATA5 encephalopathy have been observed. The EEG was normal and MRI of the head was not performed. Copper and ceruloplasmin levels were normal.

DNA extracted from the peripheral blood of the proband was analyzed by whole exome sequencing. The library was prepared with an Nextera DNA Library Prep Kit (Illumina; San Diego, CA) and sequencing was performed on HiSeq 1500 to the mean depth of 50x; 82% of target bases were covered at a minimum of 20x and 95% were covered at a minimum of 10x. Raw data were analyzed as previously described by Ploski et al. (2014), with Hg19 genomic build used for alignments. We found 563,432 variants passing a default quality filter. These variants were further filtered to include those affecting the coding sequence (by changing the aminoacid or splice site within 10 bp from exon/intron boundary) and having <1% of minor allele frequency in the following databases: ExAC Browser Beta (2016), 1000 Genomes (http://​www.​1000genomes.​org/), ESP6500 (http://​evs.​gs.​washington.​edu/​EVS), and an in-house database of ~500 Polish exomes. The final yield of 705 variants were screened against known mutations listed in the HGMD database (http://​www.​hgmd.​cf.​ac.​uk) as pathogenic (Table 2). In parallel, the 705 variants were searched for biallelic mutations consistent with autosomal recessive inheritance (Table 2) as well as for monoallelic variants potentially causative of an autosomal dominant or sex-linked condition. Here we considered variants not found in the available databases and predicted by SNP effect software to have a ‘high’ effect, i.e., to introduce a premature stop codon or affect a splice site. The variants finally considered are shown in Table 2.