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Introduction: Phenylalanine hydroxylase deficiency (E.C.1.14.16.1) causes in patients biochemical and clinical phenotype termed as phenylketonuria (PKU). Based on data presented by Slovak neonatal screening centre, the incidence of all forms of hyperphenylalaninemia is 1:6500. High efficiency of neonatal screening programme enables early therapy and discrimination of the production of toxic metabolites which are in a causal relation to brain impairment and mental retardation. Thanks to screening programme it is practically impossible to come across a patient with expressed pathological metabolomic profile typical for phenylketonuria. However we had an opportunity to experience this rare situation and to see clear metabolic changes in two brothers with non-treated disease. Material and Methods: Determination of amino acids in serum and urine was performed by thin layer chromatography (TLC) and quantitatively by high performance liquid chromatography (HPLC). The profile of organic acids in urine and determination of pathological metabolites was achieved by gas chromatography coupled with mass spectrometry (GC/MS). Molecular genetic methods included isolation of DNA from blood, PCR amplification, and sequence analysis of PAH gene. Results: Positive Fölling test was demonstrated in patients' urine samples. In TLC test of serum and urine samples intensive zones of phenylalanine were present. Quantitative determination of serum amino acids revealed high values of phenylalanine in both patients - 2503 µmol/l and 1594 µmol/l respectively. GC/MS analysis of urine organic acid shows typical picture of phenylketonuria - extremely high values of phenyllactate, phenylpyruvate and presence of other typical metabolites. We have found secondary metabolomic changes as a result of impaired phenylalanine hydroxylation. In PAH gene, two mutations were detected R408W and IVS12nt1. Conclusion: In two patients - brothers with non-treated phenylketonuria - we describe metabolomic changes. We present the findings as an educative model of hereditary metabolic disorder, which has not so important consequences for health today, thanks to neonatal screening and effective therapy. [ABSTRACT FROM AUTHOR]

Introduction: Rickets is a failure of enchondral mineralisation of new formed bone in a growing persons. There are three groups of causes of rickets - the failure of bone mineralisation, disorders of phosphate homeostasis, and disorders related to vitamin D. The Vitamin D aquires its biological activity after the activation in two steps. The microsomal 25-hydroxylase provides course of first step in liver. The deficiency of this enzyme causes vitamin D dependent rickets type 1B with low serum level of 25-hydroxyvitamin D and with normal serum level of 1,25(OH)2D. The rate-limiting step of biological activation of vitamin D is the reaction catalysed by the mitochondrial 1a-hydroxylase in kidney. Mutations of CYP27B1 (gene for 1a-hydroxylase) causing deficiency of 1a-hydroxylase enzymatic activity lead to decreased serum level of biological active form of vitamin D (with normal serum level of recirculated form of the vitamin D - 25OHvitD) and cause the vitamin D dependent rickets type 1A. The defect of nuclear receptor for vitamin D (VDR) or the defect of another member of nuclear hormonal receptors family (retinoid X receptor) forming heterodimer with VDR cause the vitamin D dependent rickets type 2. This type is associated with increased level of serum 1,25(OH)2D. Case report: We present the case of a 2 yrs old girl with the diagnosis of vitamin D dependent rickets type 1A. She had typical clinical features (dental problems, caput quadratum, rachitic rosary, widening of wrist, bowed legs, bone pain, muscle weakness, retardation of growth), and the following laboratory findings: hypocalcemia, hypophosphatemia, elevation of serum alkaline phosphatese and parathyroid hormone. Radiografic findings showed skeletal deformities, cupping and fraying of metaphyseal region. Material and metods: The DNA was isolated from periferal blood leukocytes and all exons of CYP27B1 gene were analysed by DNA sequencing. Results: The mutation c.1166G>A (p.Arg389His) was detected in homozygous state in our patient. Conclusion: The molecular analysis of rickets helps to determine the precise type of the vitamin D dependent rickets and to start the treatment in childhood early. [ABSTRACT FROM AUTHOR]

Introduction: Pompe disease is autosomal recessive disorder caused by deficiency of the lysosomal enzyme acid α-glucosidase. Accumulation of glycogen in various tissues of the body and impairment of organ function is a consequence of this metabolic disorder. Pompe disease is classified into infantile, juvenile and adult types based on the onset and clinical manifestation of the disorder. Classic infantile form is the most severe form manifested with prominent cardiomyopathy, hypotonia and death before 12 month of life due to cardiorespiratory failure. The late-onset forms, referred to as juvenile and adult onset, have symptoms that are generally limited to skeletal muscles, with a slowly progressive proximal myopathy and a marked clinical involvement of respiratory muscles. The gene encoding acid α-glucosidase (GAA) is located on chromosome 17q25.2-q25.3 and contains 20 exons. Material and methods: α-glucosidase activity in dried blood spots was measured using fluorogenic substrate 4- methylumbelliferyl-α-D-glucopyranoside and acarbose, inhibitor that eliminate isoenzyme interference of maltaseglucoamylase. Results: By determination of α-glucosidase activity in dried blood spots, we have detected two patients with Pompe disease. Diagnosis was confirmed by determination of α-glucosidase in leukocytes. Molecular analysis revealed the presence of common mutation IVS1 (-13T→G). Conclusion: The use of dried blood spots (DBS) for enzyme assays offers particular advantage over using conventional samples. In addition, it allows for large scale neonatal screening and high-risk population screening. Enzyme replacement therapy has been used as a treatment option, improving the quality of life and prognosis of patients. Early diagnosis and treatment of Pompe disease are considered to be critical for maximum efficacy of the enzyme replacement therapy. [ABSTRACT FROM AUTHOR]

Introduction: Achondroplasia is a type of skeletal dysplasia inherited in an autosomal dominant pattern with complete penetrance and prevalence rate between 1:30000 and 1:10000. 80-90% of cases are sporadic, resulting from de novo mutation. This disorder is usually caused by gain of function mutations in exon 10 of FGFR3 (4p16; 19 exons), gene encoding fibroblast growth factor receptor 3, which plays an important role in the metabolism of connective tissues. These mutations cause receptor activation and inhibition of chondrocyte proliferation, which leads to abnormalities of epiphyseal plate and bone tissues. The clinical manifestations with characteristic radiographic findings include disproportionally small stature (rhizomelic shortening of limbs), disproportionally small and long chest, large head with prominent forehead, midfacial hypoplasia and trident hand, and exaggerated lumbar lordosis later in infancy. Neurological complications and upper airway obstruction may also occur. More than 99% of achondroplasia is caused by a glycine-to-arginine substitution at codon 380. About 98% of these substitution cases is based on a G-to-A transistion and about 1% of cases on a G-to-C transversion at nucleotide 1138. The homozygous state is often manifested with a more severe phenotype. Material and Methods: The molecular analysis of exon 10 of FGFR3 in our laboratory includes two molecular-genetics methods - PCR-RFLP and sequencing. In indicated cases (abnormal ultrasound findings during the third trimester of pregnancy), it is possible to perform molecular-genetics diagnostics also in amniotic fluid samples. Results: Using this methods, we screened genomic DNA isolated from the leucocytes from patients with clinical features similar to achondroplasia. To date, we have confirmed the suspected diagnosis and found the 1138 G-to- A mutation in 4 patients. Conclusion: We have developed a reliable diagnostics for the detection of most frequent mutations in suspected diagnoses of achondroplasia. [ABSTRACT FROM AUTHOR]

Single-membrane surrounded organelles, the peroxisomes, are present in all eukaryotic cells. Main function of peroxisomes is the breakdown of very long chains of fatty acids through β-oxidation; synthesis of plasmalogens, which is the most abundant phospholipid in myelin; metabolisms of bile acids; and also detoxification of hydrogen peroxide. Peroxisomes arise by division of pre-existing peroxisomes and also by de novo biogenesis from endoplasmatic reticulum. A set of approximately thirty proteins, known as peroxins or PEX proteins, have been identified as factors critical for correct peroxisomal biogenesis and maintenance of functional peroxisome. Because peroxisomes do not contain endogenous DNA, all peroxins involved in peroxisomal biogenesis must be encoded by nuclear PEX genes. Subsequently, all these proteins required for assembly and function of peroxisomes are synthesized on free polyribosomes in the cytosol before post-translational import into the peroxisome. Transportation of peroxisomal proteins requires the presence of specific import sequences known as peroxisomal targeting sequences (PTS1 and PTS2). These PTSs sequences are recognized by different receptors, which direct peroxisomal proteins to the membrane. At present, about twenty genetically determined peroxisome diseases were described. Depending on the extent of damaged metabolic functions, they are divided into diseases of generalized loss of peroxisomal functions (for example Zellweger syndrome) and deficiencies of individual peroxisome enzymes (most commonly XAdrenoleukodystrophy). Most of disorders are autosomal recessive; however, the commonest peroxisomal disorder X-adrenoleukodystrophy has an X-linked mode of inheritance. In the first step, human patients are correctly classified in the group of disease by detection of their metabolites and subsequently by detection of biochemical abnormalities using molecular genetic techniques to identify the exact cause of the disease. [ABSTRACT FROM AUTHOR]

Introduction: Hypophosphatasia is a rare hereditary metabolic disease caused by the deficit of tissue nonspecific serum alkaline phosphatase (TNSALP). The incidence of all forms of hypophosphatasia is estimated at around 1:100 000 liveborns. Recent studies estimate the prevalence of mild autosomaldominant form at 1: 6370. During prenatal developement, this disease may cause severe damage of a fetus and even intrauterine death. During childhood, this disease manisfests with defects of mineralization with rickets signs. The following hypercalcaemia and hypercalciuria may even lead to death. In adults, the main manifestations are osteomalatia, skeletal deformities, fractures and early arthritis. Material and methods: We present the clinical course of hypophosphatasia in two sisters and the clinical features in an unrelated adult woman. Extremely low values of TNSALP were observed (0.06 μkat/l; 0.08 µkat/l; 0.03 µkat/l). To find causal mutations, sequencing of ALPL gene was performed. Results: Diagnosis of hypophosphatasia was confirmed in all three patients. We found a 436G>A and a 1183insT mutation in the adult woman. Very mild phenotype was observed and both mutations were previously not described. In silico 3D modeling of TNSALP was used to demonstrate the mutation causality. Segregation analysis was useful for genetic counselling in affected family. We found a 299C>T and a 571G>A mutation in two sisters, who were the first documented cases with childhood form of the disease in Slovakia. Conclusion: Defects of mineralization of bones and teeth (in the presence of low or moderately decrased values of ALP) is a reason for considering hypophosphatasia. The molecular genetic diagnostics opens the possibility of determinig the molecular cause of hypophosphatasia in Slovak patients. The proper diagnosis is essential for an adequate therapy management, like avoiding the treatment with biphosphonates and excess of vitamin D, where higher risk of pseudofractures in adult patients with osteomalatia was described. After determining the causal mutation, genetic counselling and eventual prenatal genetic testing are recommended. [ABSTRACT FROM AUTHOR]

Introduction: Congenital hypofibrinogenemia (CH) and congenital dysfibrinogenemia (CD) are rare coagulation disorders caused by quantitative or qualitative defects in the fibrinogen gene. The aim of this study was to characterize the genetic background and the clinical manifestations of congenital fibrinogen disorders in the patients from Slovakia registered at the National Haemophilia Centre., Materials and Methods: Results of genetic analysis of the fibrinogen genes FGA, FGB and FGG using polymerase chain reaction followed by direct sequencing were evaluated in 36 patients., Results: Molecular-genetic analysis revealed six novel variants - FGA c.923_968dup p.(Gly324Lysfs*44) and FGG c.1105C>T p.(His369Tyr) were identified in CD patients. In CH patients, in the FGG gene c.8G>A p.(Trp3*), c.823G>T p.(Glu275*) and c.323C>A p.(Ala108Asp) variants were detected. In the FGB gene c.1427C>T p.(Ser476Leu) was identified., Conclusion: This study is a positive contribution towards expanding knowledge about genetic variants in patients with congenital fibrinogen disorders., Competing Interests: The authors report no conflicts of interest in this work.

Here, we present newly derived in vitro model for modeling Duchenne muscular dystrophy. Our new cell line was derived by reprogramming of peripheral blood mononuclear cells (isolated from blood from pediatric patient) with Sendai virus encoding Yamanaka factors. Derived iPS cells are capable to differentiate in vitro into three germ layers as verified by immunocytochemistry. When differentiated in special medium, our iPSc formed spontaneously beating cardiomyocytes. As cardiomyopathy is the main clinical complication in patients with Duchenne muscular dystrophy, the cell line bearing the dystrophin gene mutation might be of interest to the research community., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 The Authors. Published by Elsevier B.V. All rights reserved.)

Background: Charcot-Marie-Tooth 1C (CMT1C) is a rare form of dominantly inherited CMT1 neuropathy caused by a mutated gene encoding lipopolysaccharide-induced tumour necrosis alpha factor (LITAF)., Case Presentation: We report a 56-year-old patient with an atypical clinical phenotype of CMT1C, which started as progressive weakness of a single upper limb resembling acquired inflammatory neuropathy. Nerve conduction studies (NCS) and temporarily limited and partial effects of immunotherapy supported the diagnosis of inflammatory neuropathy. Significant progression of polyneuropathy, despite intensive long-lasting immunotherapy, together with repeatedly negative auxiliary investigations (CSF, MRI and antibodies) and genetic testing results finally led to the diagnosis of CMT1C neuropathy., Conclusions: CMT1C should be added to the list of inherited neuropathies that need to be considered in suspected cases of inflammatory demyelinating neuropathy., (© 2021. The Author(s).)

Background: Transthyretin-related familial amyloid polyneuropathy (ATTR-FAP) is a rare, progressive, hereditary, highly disabling multisystem disorder. ATTR-FAP phenotypes differ according to the type of TTR mutation, geographic region and other as yet unidentified factors. The aim of this study was to establish the clinical and genetic characteristics of Polish patients., Methods and Patients: Clinical data and necessary examinations were collected from patients diagnosed with ATTR-FAP at the Department of Neurology of Medical University of Warsaw between 1970 and 2019., Results: 16 patients from eight unrelated families with five different TTR mutations were identified. The family with Val71Ala TTR mutation presented with early onset severe progressive polyneuropathy, with marked visual symptoms in a few patients. The next family with Ile73Val TTR mutation developed symptoms in middle age, and presented with mixed neuropathic and cardiologic phenotype. Four unrelated families were found to have the Phe33Leu TTR mutation with mixed neuropathic and cardiologic phenotype and late onset of symptoms. Other TTR mutations identified were: Val30Met and Asp38Val, both with late onset sensory, motor and autonomic neuropathy., Conclusion: Polish ATTR-FAP cases presented with heterogeneity typical for non-endemic areas. Phe33Leu TTR mutation was the most common, found in four unrelated families.

Impairment of saposin B causes rare atypical metachromatic leukodystrophy (MLD). It is encoded (together with saposin A, C, and D) by the PSAP gene. Only ten pathogenic variants were described in the PSAP gene in MLD patients to date. We report on two novel variants in the PSAP gene - c.679_681delAAG in the saposin B encoding exon 6 and c.1268delT in the saposin D encoding exon 11 in a patient with MLD. We discuss the fact, that variants resulting in PSAP null allele can be shared in patients with the deficit of other saposins (A-D) or whole prosaposin. The patient's phenotype depends then on the nature of the second allele - atypical Gaucher disease in case of saposin A, MLD in case of saposin B, and Krabbe disease in case of saposin C impairing mutations. The clinically most severe prosaposin deficit is caused by the presence of two PSAP null alleles. Thus, the assessment of a variant impact is needed to prevent delayed diagnosis or misdiagnosis in patients with PSAP mutations.

Phelan-McDermid syndrome (PMD) is a rare genetic condition with only a few cases describing patients diagnosed as adults. We describe a long diagnostic odyssey of a 30-year-old woman who was diagnosed with Phelan-McDermid syndrom. Array comparative genomic hybridization analysis confirmed a 22q13.33 deletion, encompassing exon 9-23 of the SHANK3 gene and exon 1 of the ACR gene. We provide an uncommon feature of the disease, where psychotic alteration is repeatedly triggered by the same physical factor in our patient - mild fever episodes.

Background: Short-chain acyl-CoA dehydrogenase deficiency (SCADD) represents a rare autosomal recessive inborn metabolic disorder of mitochondrial β-oxidation of monocarboxylic acids. Clinical symptoms can vary from a severe life-threatening condition to an asymptomatic state, reported in the majority of cases. Since the expansion of newborn screenings, more than three hundred probands were admitted for molecular-genetic analysis, most selected because of elevated values of C4-acylcarnitine detected in newborn screenings in Slovakia. Searching for the principal genomic changes led us to the selection of sixty-two patients in whom the presence of sequence variants in the ACADS gene was analysed and correlated with the available biochemical and clinical data., Methods: Biochemical and molecular genetic tests were performed. Acylcarnitine profiles focused on an elevated level of C4-acylcarnitine, which was analysed via tandem mass spectrometry. Urinary organic acids, specifically a quantity of ethylmalonic acid, were determined by gas chromatography/mass spectrometry. The entire coding region of the ACADS gene was sequenced. A low-cost restriction fragment length polymorphism of PCR amplified fragments analysis (PCR-RFLP) of pathogenic variants was introduced and implemented for the molecular-genetic algorithm appropriate for the Slovak population., Results: Our molecular genetic study was performed on sixty-two patients with a pathological biochemical pattern related to short-chain acyl-CoA dehydrogenase deficiency. In this cohort, we discovered a high occurrence of two rare pathogenic variants-the deletion c.310_312delGAG and the substitution c.1138C>T, with allelic frequencies of 64% and 31%, respectively. Up to 86% of investigated individuals belong to the Roma ethnic group., Conclusions: Analogous to other countries, SCADD is not included in the newborn screening programme. Based on the exceeded levels of the specific biomarker C4-acylcarnitine as well as ethylmalonic acid, we revealed a high prevalence of short-chain acyl-CoA dehydrogenase deficiency cases, confirmed by the findings of two rare pathogenic variants. A deletion c.310_312delGAG and c.1138C > T substitution in the ACADS gene appear with a high frequency in the Roma ethnic group of Slovakia. Due to the uncertainty of the pathogenicity and clinical consequences, it is important to follow up the morbidity and mortality in these patients over time and evaluate SCADD in relation to clinical outcomes and preventive healthcare recommendations.

Background: Primary hyperoxaluria type 2 is a rare monogenic disorder inherited in an autosomal recessive pattern. It results from the absence of the enzyme glyoxylate reductase/hydroxypyruvate reductase (GRHPR). As a consequence of deficient enzyme activity, excessive amounts of oxalate and L-glycerate are excreted in the urine, and are a source for the formation of calcium oxalate stones that result in recurrent nephrolithiasis and less frequently nephrocalcinosis., Case Presentation: We report a case of a 10-month-old patient diagnosed with urolithiasis. Screening of inborn errors of metabolism, including the performance of GC/MS urine organic acid profiling and HPLC amino acid profiling, showed abnormalities, which suggested deficiency of GRHPR enzyme. Additional metabolic disturbances observed in the patient led us to seek other genetic determinants and the elucidation of these findings. Besides the elevated excretion of 3-OH-butyrate, adipic acid, which are typical marks of ketosis, other metabolites such as 3-aminoisobutyric acid, 3-hydroxyisobutyric acid, 3-hydroxypropionic acid and 2-ethyl-3-hydroxypropionic acids were observed in increased amounts in the urine. Direct sequencing of the GRHPR gene revealed novel mutation, described for the first time in this article c.454dup (p.Thr152Asnfs*39) in homozygous form. The frequent nucleotide variants were found in AGXT2 gene., Conclusions: The study presents metabolomic and molecular-genetic findings in a patient with PH2. Mutation analysis broadens the allelic spectrum of the GRHPR gene to include a novel c.454dup mutation that causes the truncation of the GRHPR protein and loss of its two functional domains. We also evaluated whether nucleotide variants in the AGXT2 gene could influence the biochemical profile in PH2 and the overproduction of metabolites, especially in ketosis. We suppose that some metabolomic changes might be explained by the inhibition of the MMSADH enzyme by metabolites that increase as a consequence of GRHPR and AGXT2 enzyme deficiency. Several facts support an assumption that catabolic conditions in our patient could worsen the degree of hyperoxaluria and glyceric aciduria as a consequence of the elevated production of free amino acids and their intermediary products.

Unlabelled: We present 25-year experience with inhibitors in previously untreated patients (PUPs) with severe hemophilia A in Slovakia, where safe factor VIII (FVIII) concentrates have been used since 1990. A prospective study focused on inhibitor incidence in PUPs was established in 1997. Out of a total 61 PUPs born between January 1997 and October 2015, 59 were eligible for evaluation; 50 and 9 were treated with > 20 exposure days (ED) of plasma-derived FVIII (pdFVIII) and recombinant FVIII (rFVIII) products, respectively. In the entire group 13/59 (22%) PUPs developed inhibitors; i.e. 7/50 (14%) and 6/9 (67%) treated with pdFVIII and rFVIII, respectively. Univariate analysis of inhibitor risk factors in patient groups with and without inhibitors showed the rFVIII and serious/recurrent infections within the first 50 EDs to be associated with inhibitor development (OR of 12.3 [95% CI 2.48-60.83; p = 0.002] and 5.0; [95% CI 1.16-21.9; p = 0.03), respectively]). Also, in multivariate Cox regression analysis, peak treatment ≥ 5 EDs reached statistical significance. The hazard ratio (HR) was 7.15 (95% CI 1.65-31.36) p = 0.0086 for rFVIII and 4.38 (95% CI 1.02-18.67) p = 0.046 for intensive treatment. Between 1993 and 2015, 21 immune tolerance inductions (ITIs) in 19 inhibitor patients were performed in the two largest hemophilia centers in Slovakia. In all but one ITI courses pdFVIII containing von Willebrand factor (FVIII/VWF) was used with preferred use of high-dose ITI (HD ITI) in high responders (HRs). Complete or partial success was achieved in 17/19 (89.5%) patients. Evaluating only the patients who already completed ITI, the success rate was even higher (15/16; 94%), including 7/7 low responders and 8/9 HR., Conclusion: Our national prospective study comprising entire group of PUPs with severe hemophilia A showed higher incidence of inhibitors in patients treated with rFVIII and those with intensive therapy within first 50 EDs. However, our experience is limited to small numbers of patients; thus, our results must be interpreted cautiously. High success rate of the ITI in our inhibitor patients has been achieved with FVIII/VWF concentrates and preferred use of HD ITI in HR patients., (Thieme Medical Publishers 333 Seventh Avenue, New York, NY 10001, USA.)

Objectives: Molecular-genetic analysis is a determining step in setting the diagnosis of spinal and bulbar muscular atrophy (SBMA). We present the first nation-wide study and experience with this disease and its diagnosis in Slovakia. The study is enriched by comparison of genetic findings from Slovak patients to patients from other countries., Methods: Molecular-genetic analysis was performed for patients suspected of SBMA. Data of patients with confirmed diagnosis were statistically evaluated. In addition, the detection rate and the prevalence of the disease for Slovakia were estimated., Results: In 40 patients with confirmed diagnosis of SBMA, average values were observed at 44.7 CAG repeats and 52.5 years at the time of molecular-genetic diagnosis. The detection rate represents approximately 23% and an estimated prevalence is of 1 : 41,700., Conclusion: Concerning the population of Slovakia with 5,420,000 inhabitants, we document a relatively large cohort of SBMA patients. This is obvious when comparing similar studies from other countries, while this is the only study representing the Central Europe. Our findings prove that molecular-genetic analyses for the detection of this neuromuscular disorder show high efficiency. This fact underlines the necessity of such testing and may serve as a guide for clinicians from other countries in setting the right diagnosis for these patients (Tab. 1, Fig. 2, Ref. 29).

Objectives: The clinical, biochemical and genetic findings in two Slovak patients with glutaric aciduria type I (GAI) are presented., Background: GAI is a rare autosomal recessive neuro-metabolic disorder caused by deficiency of glutaryl-CoA dehydrogenase, which is involved in the catabolic pathways of lysine, hydroxylysine and tryptophan. This enzymatic defect gives rise to elevated levels of glutaric acid (GA), 3-hydroxyglutaric acid (3-OH-GA) and glutarylcarnitine (C5DC) in body fluids., Methods: Biochemical and molecular-genetic tests were performed. Urinary organic acids were analysed by Gas Chromatography/Mass Spectrometry (GC/MS) and the entire coding region of the GCDH gene, including flanking parts, was sequenced., Results: We found the presence of typical metabolic profile and novel causal pathogenic variants in both GAI patients., Conclusion: We present the first report of two Slovak patients with GAI, which differed in the clinical and biochemical phenotype significantly. They were diagnosed by two distinct approaches - selective and newborn screening. Their diagnosis was complexly confirmed by biochemical and later on molecular-genetic examinations. Though we agreed with a thesis that early diagnostics might positively influenced patient's health outcome, contradictory facts should be considered. Supposed extremely low prevalence of GAI patients in the general population and/or the existence of asymptomatic individuals with a questionable benefit of the applied therapeutic intervention for them lead to doubts whether the inclusion of disease into the newborn screening programme is justified well enough (Tab. 1, Fig. 3, Ref. 41).

The peroxisomal biogenesis disorders are autosomal recessive diseases morphologically characterised by lacking peroxisomes, biochemically by generalised deficiency of peroxisomal constituent and clinically manifested by serious health problems. Genes involved in the peroxisomal biogenesis are defined as the PEX genes encoding proteins called the peroxins. These peroxins are required for function in assembly of the peroxisomal membrane or in import of the enzymes into the peroxisomes. In this study we present a full overview of the clinical presentation, biochemical and molecular data of patient with Zellweger syndrome from Slovakia. We investigated biochemical metabolites using gas chromatography/mass spectrometry. The presence of causal ins/del mutations we identified by a Sanger sequencing and RFLP. We reported that the patient was a compound heterozygote for mutations in the gene PEX12: a 2-bp insertion (c.767_768dupAT) and a 2-bp deletion (c.887_888delTC). The first one mentioned is a novel mutation, which has not been reported before. Both mutations create a frameshift of the open reading frame which result a premature STOP codon and generate a complete loss of the C-terminal RING finger domain that is crucial for the correct import of proteins into peroxisomes. We found causal mutations responsible for a severe phenotype, and moreover we noted a novel mutation c.767_768dupAT that has not been reported before. The presence of mutations was studied in all family members, and the resulting data were successfully utilized for prenatal diagnosis.