NGS-Based Identification of Two Novel PCDH19 Mutations in Female Patients with Early-Onset Epilepsy.

Developmental and epileptic encephalopathy-9 (DEE9) is characterized by seizure onset in infancy, mild to severe intellectual impairment, and psychiatric features and is caused by a mutation in the PCDH19 gene on chromosome Xq22. The rare, unusual X-linked type of disorder affects heterozygous females and mosaic males; transmitting males are unaffected. In our study, 165 patients with epilepsy were tested by Next Generation Sequencing (NGS)-based panel and exome sequencing using Illumina technology. PCDH19 screening identified three point mutations, one indel, and one 29 bp-long deletion in five unrelated female probands. Two novel mutations, c.1152_1180del (p.Gln385Serfs*6) and c.830_831delinsAA (p.Phe277*), were identified and found to be de novo pathogenic. Moreover, among the three inherited mutations, two originated from asymptomatic mothers and one from an affected father. The PCDH19 c.1682C>T and c.1711G>T mutations were present in the DNA samples of asymptomatic mothers. After targeted parental testing, X chromosome inactivation tests and Sanger sequencing were carried out for mosaicism examination on maternal saliva samples in the two asymptomatic PCDH19 mutation carrier subjects. Tissue mosaicism and X-inactivation tests were negative. Our results support the opportunity for reduced penetrance in DEE9 and contribute to expanding the genotype-phenotype spectrum of PCDH19-related epilepsy.

Case report: Initial atypical skeletal symptoms and dental anomalies as first signs of Gardner syndrome: the importance of genetic analysis in the early diagnosis.

Background: Gardner syndrome is a rare genetic cancer predisposition disorder characterized by intestinal polyposis, multiple osteomas, and soft and hard tissue tumors. Dental anomalies are present in approximately 30%-70% of patients with Gardner syndrome and can be discovered during routine dental examinations. However, sometimes the diagnosis is challenging due to the high clinical variability and incomplete clinical picture. Herein, we report a family with various dental and bone anomalies, in which the definitive diagnosis was established with the help of a comprehensive genetic analysis based on state-of-the-art next-generation sequencing technology. Case presentation: A 17-year-old female index patient presented with dental (caries, impacted, retained and anteriorly located teeth) and atypical bone anomalies not resembling Gardner syndrome. She was first referred to our Genetic Counselling Unit at the age of 11 due to an atypical bone abnormality identified by a panoramic X-ray. Tooth 3.6 was surgically removed and the histopathology report revealed a Paget's disease-like bone metabolic disorder with mixed osteoblastic and osteoclastic activity of the mandible. A small lumbar subcutaneous tumor was discovered by physical examination. Ultrasound examination of the tumor raised the possibility of a soft tissue propagation of chondromatosis. Her sister, 2 years younger at the age of 14, had some benign tumors (multiple exostoses, odontomas, epidermoid cysts) and impacted teeth. Their mother had also skeletal symptoms. Her lower teeth did not develop, the 9th-10th ribs were fused, and she complained of intermittent jaw pain. A cranial CT scan showed fibrous dysplasia on the cranial bones. Whole exome sequencing identified a heterozygous pathogenic nonsense mutation (c.4700C>G; p.Ser1567*) in the APC gene in the index patient's DNA. Targeted sequencing revealed the same variant in the DNA of the other affected family members (the sister and the mother). Conclusion: Early diagnosis of this rare, genetically determined syndrome is very important, because of the potentially high malignant transformation of intestinal polyps. Dentists should be familiar with the typical maxillofacial features of this disorder, to be able to refer patients to genetic counseling. Dental anomalies often precede the intestinal polyposis and facilitate the early diagnosis, thereby increasing the patients' chances of survival. Genetic analysis may be necessary in patients with atypical phenotypic signs.

Identification of an NF1 Microdeletion with Optical Genome Mapping.

Neurofibromatosis type 1 (NF1) is a clinically heterogeneous neurocutaneous disorder inherited in autosomal dominant manner. Approximately 5-10% of the cases are caused by NF1 microdeletions involving the NF1 gene and its flanking regions. Microdeletions, which lead to more severe clinical manifestations, can be subclassified into four different types (type 1, 2, 3 and atypical) according to their size, the genomic location of the breakpoints and the number of genes included within the deletion. Besides the prominent hallmarks of NF1, patients with NF1 microdeletions frequently exhibit specific additional clinical manifestations like dysmorphic facial features, macrocephaly, overgrowth, global developmental delay, cognitive disability and an increased risk of malignancies. It is important to identify the genes co-deleted with NF1, because they are likely to have an effect on the clinical manifestation. Multiplex ligation-dependent probe amplification (MLPA) and microarray analysis are the primary techniques for the investigation of NF1 microdeletions. However, based on previous research, optical genome mapping (OGM) could also serve as an alternative method to identify copy number variations (CNVs). Here, we present a case with NF1 microdeletion identified by means of OGM and demonstrate that this novel technology is a suitable tool for the identification and classification of the NF1 microdeletions.

Copy Number Variations in Neuropsychiatric Disorders.

Neuropsychiatric disorders are complex conditions that represent a significant global health burden with complex and multifactorial etiologies. Technological advances in recent years have improved our understanding of the genetic architecture of the major neuropsychiatric disorders and the genetic loci involved. Previous studies mainly investigated genome-wide significant SNPs to elucidate the cross-disorder and disorder-specific genetic basis of neuropsychiatric disorders. Although copy number variations represent a major source of genetic variations, they are known risk factors in developing a variety of human disorders, including certain neuropsychiatric diseases. In this review, we demonstrate the current understanding of CNVs contributing to liability for schizophrenia, bipolar disorder, and major depressive disorder.

Copper-transporting ATPases throughout the animal evolution - From clinics to basal neuron-less animals.

Copper-transporting ATPases are a group of heavy metal-transporting proteins and which can be found in all living organisms. In animals, they are generally referred to as ATP7 proteins and are involved in many different physiological processes including the maintaining of copper homeostasis and the supply of copper to cuproenzymes. A single ATP7 gene is present in non-chordate animals while it is divided into ATP7A and ATP7B in chordates. In humans, dysfunction of ATP7 proteins can lead to severe genetic disorders, such as, Menkes disease and Wilson's disease, which are characterized by abnormal copper transport and accumulation, causing significant health complications. Therefore, there is a substantial amount of research on ATP7 genes and ATP7 proteins in humans and mice to understand pathophysiological conditions and find potential therapeutic interventions. Copper-transporting ATPases have also been investigated in some non-mammalian vertebrates, protostomes, single-cellular eukaryotes, prokaryotes, and archaea to gain useful evolutionary insights. However, ATP7 function in many animals has been somewhat neglected, particularly in non-bilaterians. Previous reviews on this topic only broadly summarized the available information on the function and evolution of ATP7 genes and ATP7 proteins and included only the classic vertebrate and invertebrate models. Given this, and the fact that a considerable amount of new information on this topic has been published in recent years, the present study was undertaken to provide an up-to-date, comprehensive summary of ATP7s/ATP7s and give new insights into their evolutionary relationships. Additionally, this work provides a framework for studying these genes and proteins in non-bilaterians. As early branching animals, they are important to understand the evolution of function of these proteins and their important role in copper homeostasis and neurotransmission.

Down-Syndrome-Related Maternal Dysbiosis Might Be Triggered by Certain Classes of Antibiotics: A New Insight into the Possible Pathomechanisms.

Down syndrome (DS) is a leading human genomic abnormality resulting from the trisomy of chromosome 21. The genomic base of the aneuploidy behind this disease is complex, and this complexity poses formidable challenges to understanding the underlying molecular basis. In the spectrum of the classic DS risk factor associations, the role of nutrients, vitamins, and, in general, the foodborne-associated background, as part of the events ultimately leading to chromosome nondisjunction, has long been recognized as a well-established clinical association. The integrity of the microbiome is a basic condition in these events, and the dysbiosis may be associated with secondary health outcomes. The possible association of DS development with maternal gut microbiota should therefore require more attention. We have hypothesized that different classes of antibiotics might promote or inhibit the proliferation of different microbial taxa; and hence, we might find associations between the use of the different classes of antibiotics and the prevalence of DS through the modification of the microbiome. As antibiotics are considered major disruptors of the microbiome, it could be hypothesized that the consumption/exposure of certain classes of antibiotics might be associated with the prevalence of DS in European countries (N = 30). By utilizing three different statistical methods, comparisons have been made between the average yearly antibiotic consumption (1997-2020) and the estimated prevalence of people living with DS for the year 2019 as a percentage of the population in European countries. We have found strong statistical correlations between the consumption of tetracycline (J01A) and the narrow-spectrum, beta-lactamase-resistant penicillin (J01CF) and the prevalence of DS.

Microhomology-Mediated Break-Induced Replication: A Possible Molecular Mechanism of the Formation of a Large CNV in FBN1 Gene in a Patient with Marfan Syndrome.

BACKGROUND: Marfan syndrome (MFS) is an autosomal dominant multisystem disorder caused by mutations in the fibrillin-1 gene (FBN1). A small portion of them is copy number variations (CNVs), which can occur through recombination-based, replication-based mechanisms or retrotransposition. Not many have been characterized precisely in MFS. METHODS: A female patient with suspected Marfan syndrome was referred for genetic testing at our institute. After systematic sequencing of FBN1, TGFBR1, and TGFBR2 genes, multiplex ligation-dependent probe amplification was applied. Long-range PCR, subsequent Sanger sequencing with designed primers, and preliminary in silico analysis were applied for the precise characterization of the breakpoints. RESULTS: Primary analysis displayed a de novo large deletion affecting exons 46 and 47 in the FBN1 gene, which resulted in the loss of the 31st and 32nd calcium-binding EGFlike domains. Further examination of the breakpoints showed a 4916 nucleotide long deletion localized in intronic regions. Surprisingly a 'TG' dinucleotide insertion was detected at the junction. We hypothesize that the CNV formation was generated by a rare event based on the known microhomology-mediated break-induced replication (MMBIR). CONCLUSION: An increasing number of CNVs are associated with Mendelian diseases and other traits. Approximately 2-7% of the cases in MFS are caused by CNVs. Up to date, hardly any model was proposed to demonstrate the formation of these genomic rearrangements in the FBN1 gene. Hereby, with the help of previous models and breakpoint analysis, we presented a potential mechanism (based on MMBIR) in the formation of this large deletion.

Correlation between large FBN1 deletions and severe cardiovascular phenotype in Marfan syndrome: Analysis of two novel cases and analytical review of the literature.

BACKGROUND: Marfan syndrome (MFS) is a clinically heterogeneous hereditary connective tissue disorder. Severe cardiovascular manifestations (i.e., aortic aneurysm and dissection) are the most life-threatening complications. Most of the cases are caused by mutations, a minor group of which are copy number variations (CNV), in the FBN1 gene. METHODS: Multiplex ligation-dependent probe amplification test was performed to detect CNVs in 41 MFS patients not carrying disease-causing mutations in FBN1 gene. Moreover, the association was analyzed between the localization of CNVs, the affected regulatory elements and the cardiovascular phenotypes among all cases known from the literature. RESULTS: A large two-exon deletion (exon 46 and 47) was identified in two related patients, which was associated with a mild form of cardiovascular phenotype. Severe cardiovascular symptoms were found significantly more frequent in patients with FBN1 large deletion compared to our patients with intragenic small scale FBN1 mutation. Bioinformatic data analyses of regulatory elements located within the FBN1 gene revealed an association between the deletion of STAT3 transcription factor-binding site and cardiovascular symptoms in five out of 25 patients. CONCLUSION: Our study demonstrated that large CNVs are often associated with severe cardiovascular manifestations in MFS and the localization of these CNVs affect the phenotype severity.

Characterization of Danube Swabian population samples on a high-resolution genome-wide basis.

BACKGROUND: German-derived ethnicities are one of the largest ethnic groups in Hungary, dating back to the formation of the Kingdom of Hungary, which took place at the beginning of the 11th century. Germans arrived in Hungary in many waves. The most significant immigration wave took place following the collapse of the Ottoman Empire in East-Central Europe which closed the 150 year long Ottoman occupation. To date, there are no comprehensive genome-wide studies investigating the genetic makeup of the Danube Swabians. Here we analyzed 47 Danube Swabian samples collected from elderly Swabian individuals living in the Dunaszekcso-Bár area, in Danube side villages of Southwest Hungary. These Swabians, according to self-declaration, did not admix with other ethnic groups for 3-6 succeeding generations. Using Illumina Infinium 720 K Beadchip genotype data, we applied allele frequency-based and haplotype-based genome-wide marker data analyses to investigate the ancestry and genetic composition of the collected Danube Swabian samples. RESULTS: Haplotype-based analyses like identity by descent segment analysis show that the investigated Danube Swabians possess significant German and other West European ancestry, but their Hungarian ancestry is also prominent. Our results suggest that their main source of ancestry can be traced back to Western Europe, presumably to the region of Germany. CONCLUSION: This is the first analysis of Danube Swabian population samples based on genome-wide autosomal data. Our results establish the basis for conducting further comprehensive research on Danube Swabians and on other German ethnicities of the Carpathian basin, which can help reconstruct their origin, and identify their major archaic genomic patterns.

[Neurofibromatosis-1 microdeletion syndrome.] / Neurofibromatosis-1 microdeletiós szindróma.

Neurofibromatosis type 1 is a clinically extremely heterogeneous neurocutaneous disorder, inherited in autosomal dominant manner. It is primarily caused by intragenic loss-of-function mutations in the NF1 gene, however, as a result of improvements in molecular diagnostics, copy number variants affecting the NF1 gene and its flanking regions are increasingly being detected. Based on genotype-phenotype analyses, two groups can be distinguished: neurofibromatosis type 1 caused by point mutations and the so-called 17q11.2 microdeletion syndrome caused by microdeletions. Microdeletions are observed in 5-10% of cases and can be divided into four different types (type 1, 2, 3 and atypical) according to the size of the deletion, the genomic location of the breakpoints and the affected gene content. Patients with microdeletions often have a more severe course of the disease, with an increased risk of malignancies. With this review, which summarizes the main characteristics and molecular genetic background of neurofibromatosis-1 microdeletion syndrome, we would like to emphasize the importance of early diagnosis of patients with microdeletion syndrome and draw attention to the importance of close follow-up. Orv Hetil. 2022; 163(51): 2041-2051.

Genome-Wide Marker Data-Based Comparative Population Analysis of Szeklers From Korond, Transylvania, and From Transylvania Living Non-Szekler Hungarians.

Genome-wide genotype data from 48 carefully selected population samples of Transylvania-living Szeklers and non-Szekler Hungarians were analyzed by comparative analysis. Our analyses involved contemporary Hungarians living in Hungary, other Europeans, and Eurasian samples counting 530 individuals altogether. The source of the Szekler samples was the commune of Korond, Transylvania. The analyzed non-Szekler Hungarian samples were collected from villages with a history dating back to the era of the Árpád Dynasty. Population structure by principal component analysis and ancestry analysis also revealed a great within-group similarity of the analyzed Szeklers and non-Szekler Transylvanian Hungarians. These groups also showed similar genetic patterns with each other. Haplotype analyses using identity-by-descent segment discovering tools showed that average pairwise identity-by-descent sharing is similar in the investigated populations, but the Korond Szekler samples had higher average sharing with the Hungarians from Hungary than non-Szekler Transylvanian Hungarians. Average sharing results showed that both groups are isolated compared to other Europeans, and pointed out that the non-Szekler Transylvanian Hungarian inhabitants of the investigated Árpád Age villages are more isolated than investigated Szeklers from Korond. This was confirmed by our autozygosity analysis as well. Identity-by-descent segment analyses and 4-population tests also confirmed that these Hungarian-speaking Transylvanian ethnic groups are strongly related to Hungarians living in Hungary.

Genotype-Phenotype Associations in Patients With Type-1, Type-2, and Atypical NF1 Microdeletions.

Neurofibromatosis type 1 is a tumor predisposition syndrome inherited in autosomal dominant manner. Besides the intragenic loss-of-function mutations in NF1 gene, large deletions encompassing the NF1 gene and its flanking regions are responsible for the development of the variable clinical phenotype. These large deletions titled as NF1 microdeletions lead to a more severe clinical phenotype than those observed in patients with intragenic NF1 mutations. Around 5-10% of the cases harbor large deletion and four major types of NF1 microdeletions (type 1, 2, 3 and atypical) have been identified so far. They are distinguishable in term of their size and the location of the breakpoints, by the frequency of somatic mosaicism with normal cells not harboring the deletion and by the number of the affected genes within the deleted region. In our study genotype-phenotype analyses have been performed in 17 mostly pediatric patients with NF1 microdeletion syndrome identified by multiplex ligation-dependent probe amplification after systematic sequencing of the NF1 gene. Confirmation and classification of the NF1 large deletions were performed using array comparative genomic hybridization, where it was feasible. In our patient cohort 70% of the patients possess type-1 deletion, one patient harbors type-2 deletion and 23% of our cases have atypical NF1 deletion. All the atypical deletions identified in this study proved to be novel. One patient with atypical deletion displayed mosaicism. In our study NF1 microdeletion patients presented dysmorphic facial features, macrocephaly, large hands and feet, delayed cognitive development and/or learning difficulties, speech difficulties, overgrowth more often than patients with intragenic NF1 mutations. Moreover, neurobehavior problems, macrocephaly and overgrowth were less frequent in atypical cases compared to type-1 deletion. Proper diagnosis is challenging in certain patients since several clinical manifestations show age-dependency. Large tumor load exhibited more frequently in this type of disorder, therefore better understanding of genotype-phenotype correlations and progress of the disease is essential for individuals suffering from neurofibromatosis to improve the quality of their life. Our study presented additional clinical data related to NF1 microdeletion patients especially for pediatric cases and it contributes to the better understanding of this type of disorder.

Identification, presence, and possible multifunctional regulatory role of invertebrate gonadotropin-releasing hormone/corazonin molecule in the great pond snail (Lymnaea stagnalis).

In the last years, our interpretation of the origin and function of the gonadotropin-releasing hormone (GnRH) neuropeptide superfamily has changed substantially. A main driver for these conceptual changes came from increased investigations into functions and evolutionary lineage of previously identified molluscan GnRH molecules. Emerging evidence suggests not only reproductive, but also diverse biological effects of these molecules and proposes they should most likely be called corazonin (CRZ). Clearly, a more global understanding requires further exploration of species-specific functions and structure of invGnRH/CRZ peptides. Towards this goal, we have identified the full-length cDNA of invGnRH/CRZ peptide in an invertebrate model species, the great pond snail Lymnaea stagnalis, termed ly-GnRH/CRZ, and characterized the transcript and peptide distribution in the central nervous system (CNS) and peripheral organs. Our results are consistent with previous data that molluscan GnRHs are more related to CRZs and serve diverse functions. Hence, our findings support the notion that peptides originally termed molluscan GnRH are multifunctional modulators and that nomenclature change should be taken into consideration.

[A rare form of ion channel gene mutation identified as underlying cause of generalized epilepsy]. / Generalizált epilepszia hátterében azonosított ioncsatorna-génmutáció ritka formája.

The advances in molecular genetic methods has lead to the discovery of the genetic alterations that underlie the etiology of most diseases previously held to be idiopathic. Targeted genetic examination of a pediatric male patient showing a normal intellect, an extended area of skin hypopigmentation, and suffering from generalized epilepsy displaying a switch in epilepsy syndrome during the course of the disease towards a neurocutaneous syndrome was unsuccessful. Whole-exome sequencing identified a heterozygous missense mutation in a potassium chloride cotransporter gene, which together with the phenotype underscores the diagnosis of an epilepsy syndrome known in the literature as idiopathic generalized epilepsy type 14. Orv Hetil. 2019; 160(21): 835-838.

Revealing the impact of the Caucasus region on the genetic legacy of Romani people from genome-wide data.

Romani people are a significant minority in Europe counting about 10 million individuals scattered throughout the continent. They are a migratory group originating from Northwestern India. Their exodus from India occurred approximately 1000-1500 years ago. The migration route of the Romani people was reconstructed with the help of cultural anthropology, linguistics and historical records. Their migration made them through Central Asia, Middle East and the Caucasus region, prior to the arriving into Europe. Yet the significance of these regions, especially of the Caucasus, in Roma ancestry was a rather neglected topic. Contribution of the Caucasus and further affected regions to the ancestry of Roma was investigated based on genome-wide autosomal marker data. 158 European Roma samples and 41 populations from the Caucasus region, from Middle East, Central Asia and from South Asia were considered in our tests. Population structure and ancestry analysis algorithms were applied to investigate the relationship of Roma with these populations. Identical by descent DNA segment analyses and admixture linkage disequilibrium based tests were also applied. Our results suggest that the Caucasus region plays also a significant role in the genetic legacy of Romani people besides the main sources, Europe and South Asia, previously investigated by other population genetic studies. The Middle East and Central Asia seems slightly less important but far from negligible in connection with the sources of Roma ancestry. Our results point out that the Caucasus region and altogether the area of the Caspian and Black Seas had a significant role in the migration of Romani people towards Europe and contributed significantly to the genetic legacy of Roma rival to the European and Indian main sources.