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Összefoglaló. A 3p25-deletiós szindróma nagyon ritka genetikai rendellenesség, a nemzetközi szakirodalom jelenleg kevesebb mint 60 esetet ír le. A kórképre általánosan jellemző a növekedési és pszichomotoros visszamaradottság, a microcephalia, a hypotonia, a veleszületett szívfejlődési rendellenesség, a ptosis és micrognathia, de nagyon ritkán előfordul klinikai tünetek nélküli megjelenése is. Általában újonnan kialakult rendellenesség, bár egyes esetekben előfordulhat familiáris formája. A kromoszomális töréspont változó helyen fordul elő. Közleményünkben egy 3p25-deletiós szindrómával született gyermek esetét mutatjuk be: a deletiót kariotipizálással és fluoreszcens in situ hibridizációval igazoltuk, majd microarray-komparatív genomhibridizálással meghatároztuk a pontos töréspontot és a hiányzó géneket. Az érintett régióban 43 OMIM-gént találtunk, melyek szerepet játszanak a megkésett pszichomotoros és növekedési elmaradásban, valamint az intellektuális zavarban. A genetikai háttér pontos karakterizálása hozzásegít a várható tünetek és a prognózis meghatározásához, egyben támpontot biztosíthat a jövőbeli terápia tervezéséhez és a személyre szabott fejlesztés kivitelezéséhez. Orv Hetil. 2022; 163(12): 478–483. Summary. The 3p25 deletion syndrome is a very rare genetic abnormality, characterized by growth and psychomotor retardation, microcephaly, hypotonia, congenital heart defects, ptosis and micrognathia. Less than 60 cases have been published in the literature so far. However, a few patients with normal or mild phenotype have also been described. The majority of the cases are de novo mutations, with variable chromosomal breakpoints. We present the case of a newborn infant with 3p25 deletion syndrome, whose genetic analysis was done by karyotyping, fluorescent in situ hybridization and array comparative genomic hybridization. The latter method enabled us to define the precise breakpoint and the genes involved in the deletion, thus we could provide information for further clinical management. We identified 43 OMIM genes in the deleted region, which may have a causative effect on the pscychomotor and developmental delay and also on the intellectual disability. Exact cytogenomic characterisation of a rare genetic syndrome may allow to employ personalised treatment. Orv Hetil. 2022; 163(12): 478–483.

Összefoglaló. Bevezetés és célkitűzés: A gyakori autoszomális trisomiák és a nemi kromoszómaeltérések a mikroszkóppal észlelhető kromoszóma-rendellenességek kb. 80–85%-át képviselik. A ritka kromoszóma-rendellenességek klinikai következménye jelentős, kimutatásukat a jelenlegi szűrővizsgálatok ugyan nem célozzák, de a teljes kromoszómaszerelvényt vizsgáló, nem invazív praenatalis tesztelés új lehetőséget nyitott a korai felismerésükre. Módszer: Retrospektív analízis (2014–2019) a mikroszkóppal kimutatható kromoszóma-rendellenességek eloszlására, a fetoplacentaris mozaikosság előfordulására, klinikai összefüggéseire a praenatalis vizsgálatok tükrében egy hazai tercier centrumban. Eredmények: 2504 invazív beavatkozást végeztünk és 200 kromoszómaeltérést mutattunk ki (8%), melyek közül újonnan kialakult, ritka rendellenesség 27 volt (13,5%). Ritka autoszomális trisomia 14, poliploidia 6, de novo szerkezeti kromoszómaeltérés 5, marker kromoszóma 2 esetben igazolódott. A fetoplacentaris mozaikosság aránya a gyakori/ritka kromoszómaeltérésekben 12,4%/77,8% volt (p = 0,001), 17/40 esetben lepényre korlátozódott. A gyakori trisomiákkal kóros tarkóredő-vastagság 58%-ban, major ultrahangeltérés 35%-ban társult, melyek jelentősen különböztek a ritka kromoszómaeltérésekben (11%, p = 0,006; 67%, p = 0,047). A ritka kromoszómaeltérések jellemző praenatalis major ultrahangeltérése a facialis dysmorphismus volt. A teljes kromoszómaszerelvényt vizsgáló praenatalis tesztelés a ritka kromoszómaeltérések 12 lepényi mozaikos esetében (44%) feltételezhetően álpozitív, 1 esetben (3,7%) álnegatív eredményt generált volna, miközben a ritka autoszomális trisomiák 2 esetében ultrahangeltérés nélkül is korán detektálta volna a ritka magzati kromoszómaeltérést (7,4%). Következtetés: A normális tarkóredő-vastagság esetén észlelt major ultrahangeltérések felhívhatják a figyelmet a döntően mozaikos ritka kromoszóma-rendellenességekre. A teljes kromoszómaszerelvényt vizsgáló, nem invazív szűrőteszt a korai diagnosztika alternatívája lehet, a mozaikosságból adódó álpozitív eredményekre azonban számítani kell. A fetoplacentaris mozaikosság ismerete fontos klinikai információt biztosít, mely befolyásolhatja a terhesség kimenetelét, a terhesség követésének módját. A pontos citogenetikai karakterizálás elengedhetetlen. Orv Hetil. 2021; 162(29): 1156–1165. Summary. Introduction and objective: To determine the prevalence of microscopically visible de novo atypical chromosomal aberrations and fetoplacental mosaicism in a prenatal tertial referral center, and to investigate the maternal and fetal characteristics in connection with genomewide non-invasive prenatal screening. Method: Retrospective cohort study from 2014 to 2019 of pregnancies with invasive genetic analysis. Results: In the cohort of 2504 cases, the proportion of CVS was 53.3%. We diagnosed 200 chromosomal aberrations (8%), including 13.5% of de novo rare chromosomal aberrations (14 rare autosomal trisomies, 6 polyploidies, 5 structural aberrations and 2 small supernumerary marker chromosomes). The rate of fetoplacental mosaicism was 12.4%/77.8% in common/atypical chromosomal aberrations (p = 0.001) and confined to placenta in 17/40 cases. Associated ultrasound abnormalities were abnormal nuchal translucency and major malformations in 58% and 35% with common trisomies and 11% (p = 0.006) and 67% (p = 0.047) with true mosaic cases of rare abnormalities, respectively. Major ultrasound malformation was facial dysmorphism with rare aberrations. Potential application of genomewide non-invasive prenatal test in atypical chromosomal aberrations presumably would have been false-positive in 12 cases (44%), false-negative in 1 case (3.7%), and would have early detected 2 cases of rare autosomal trisomies (7.4%) without ultrasound anomalies. Conclusion: Structural ultrasound malformations with normal nuchal translucency thickness may be indicative of rare chromosomal aberrations. Application of genomewide non-invasive prenatal test is an alternative of early diagnostic methods with a potential of discordant results due to mosaicism. Knowledge about the presence of fetoplacental mosaicism influences risk estimation and genetic counseling, detailed cytogenetic characterization is of utmost importance. Orv Hetil. 2021; 162(29): 1156–1165.

We report a patient with loss of chromosome region 2q14.3 encompassing exon 1 of the gene

We report a patient with loss of chromosome region 2q14.3 encompassing exon 1 of the gene CNTNAP5. The deletion occurred in association with a de novo complex chromosomal rearrangement, characterized by routine G-banding, fluorescence in situ hybridization and microarray analysis. The presented patient's phenotype is dominated by severe early childhood weight gain, severe speech delay and behavioural problems. To our knowledge, a few similar patients have been reported previously. CNTNAP5 is a member of the neurexin gene family and is associated with autism spectrum disorder and potentially other behavioural and neurodevelopmental disorders. Recent data point to its possible role in obesity and/or metabolism. The phenotype of the herein presented pediatric patient corroborates CNTNAP5's pathogenic role in human disease.

Összefoglaló. A 3p25-deletiós szindróma nagyon ritka genetikai rendellenesség, a nemzetközi szakirodalom jelenleg kevesebb mint 60 esetet ír le. A kórképre általánosan jellemző a növekedési és pszichomotoros visszamaradottság, a microcephalia, a hypotonia, a veleszületett szívfejlődési rendellenesség, a ptosis és micrognathia, de nagyon ritkán előfordul klinikai tünetek nélküli megjelenése is. Általában újonnan kialakult rendellenesség, bár egyes esetekben előfordulhat familiáris formája. A kromoszomális töréspont változó helyen fordul elő. Közleményünkben egy 3p25-deletiós szindrómával született gyermek esetét mutatjuk be: a deletiót kariotipizálással és fluoreszcens in situ hibridizációval igazoltuk, majd microarray-komparatív genomhibridizálással meghatároztuk a pontos töréspontot és a hiányzó géneket. Az érintett régióban 43 OMIM-gént találtunk, melyek szerepet játszanak a megkésett pszichomotoros és növekedési elmaradásban, valamint az intellektuális zavarban. A genetikai háttér pontos karakterizálása hozzásegít a várható tünetek és a prognózis meghatározásához, egyben támpontot biztosíthat a jövőbeli terápia tervezéséhez és a személyre szabott fejlesztés kivitelezéséhez. Orv Hetil. 2022; 163(12): 478-483. Summary. The 3p25 deletion syndrome is a very rare genetic abnormality, characterized by growth and psychomotor retardation, microcephaly, hypotonia, congenital heart defects, ptosis and micrognathia. Less than 60 cases have been published in the literature so far. However, a few patients with normal or mild phenotype have also been described. The majority of the cases are de novo mutations, with variable chromosomal breakpoints. We present the case of a newborn infant with 3p25 deletion syndrome, whose genetic analysis was done by karyotyping, fluorescent in situ hybridization and array comparative genomic hybridization. The latter method enabled us to define the precise breakpoint and the genes involved in the deletion, thus we could provide information for further clinical management. We identified 43 OMIM genes in the deleted region, which may have a causative effect on the pscychomotor and developmental delay and also on the intellectual disability. Exact cytogenomic characterisation of a rare genetic syndrome may allow to employ personalised treatment. Orv Hetil. 2022; 163(12): 478-483.

Abstract: Invasive prenatal testing and conventional G-banding chromosome analysis have been considered to be the gold standard of fetal cytogenetic diagnosis. Standard karyotyping is, however, constrained by the limits of the resolution of using a microscope. The advantage of molecular karyotyping, array based methods is the evaluation of sub-microscopic copy number changes across the whole genome in a single analysis. The application of array comparative genome hybridization has greatly increased the detection of pathogenic chromosomal abnormalities in prenatal settings. Based on available data in the international literature of the last decade, the clinical utility of arrayCGH is the recognition of some 1–2% and 5–7% additional genetical information compared to metaphase karyotype alone in fetuses without ultrasound anomaly and in fetuses with ultrasonographically detected malformations, respectively. Thus arrayCGH improves the prenatal diagnosis of genetic abnormalities mainly in fetuses with structural sonographic findings. In the present paper we review the literature of chromosomal microarray and make a proposal for the application of the method in Hungarian prenatal genetical practice. Orv Hetil. 2019; 160(13): 484–493.

Összefoglaló. Bevezetés és célkitűzés: A gyakori autoszomális trisomiák és a nemi kromoszómaeltérések a mikroszkóppal észlelhető kromoszóma-rendellenességek kb. 80-85%-át képviselik. A ritka kromoszóma-rendellenességek klinikai következménye jelentős, kimutatásukat a jelenlegi szűrővizsgálatok ugyan nem célozzák, de a teljes kromoszómaszerelvényt vizsgáló, nem invazív praenatalis tesztelés új lehetőséget nyitott a korai felismerésükre. Módszer: Retrospektív analízis (2014-2019) a mikroszkóppal kimutatható kromoszóma-rendellenességek eloszlására, a fetoplacentaris mozaikosság előfordulására, klinikai összefüggéseire a praenatalis vizsgálatok tükrében egy hazai tercier centrumban. Eredmények: 2504 invazív beavatkozást végeztünk és 200 kromoszómaeltérést mutattunk ki (8%), melyek közül újonnan kialakult, ritka rendellenesség 27 volt (13,5%). Ritka autoszomális trisomia 14, poliploidia 6, de novo szerkezeti kromoszómaeltérés 5, marker kromoszóma 2 esetben igazolódott. A fetoplacentaris mozaikosság aránya a gyakori/ritka kromoszómaeltérésekben 12,4%/77,8% volt (p = 0,001), 17/40 esetben lepényre korlátozódott. A gyakori trisomiákkal kóros tarkóredő-vastagság 58%-ban, major ultrahangeltérés 35%-ban társult, melyek jelentősen különböztek a ritka kromoszómaeltérésekben (11%, p = 0,006; 67%, p = 0,047). A ritka kromoszómaeltérések jellemző praenatalis major ultrahangeltérése a facialis dysmorphismus volt. A teljes kromoszómaszerelvényt vizsgáló praenatalis tesztelés a ritka kromoszómaeltérések 12 lepényi mozaikos esetében (44%) feltételezhetően álpozitív, 1 esetben (3,7%) álnegatív eredményt generált volna, miközben a ritka autoszomális trisomiák 2 esetében ultrahangeltérés nélkül is korán detektálta volna a ritka magzati kromoszómaeltérést (7,4%). Következtetés: A normális tarkóredő-vastagság esetén észlelt major ultrahangeltérések felhívhatják a figyelmet a döntően mozaikos ritka kromoszóma-rendellenességekre. A teljes kromoszómaszerelvényt vizsgáló, nem invazív szűrőteszt a korai diagnosztika alternatívája lehet, a mozaikosságból adódó álpozitív eredményekre azonban számítani kell. A fetoplacentaris mozaikosság ismerete fontos klinikai információt biztosít, mely befolyásolhatja a terhesség kimenetelét, a terhesség követésének módját. A pontos citogenetikai karakterizálás elengedhetetlen. Orv Hetil. 2021; 162(29): 1156-1165.To determine the prevalence of microscopically visible de novo atypical chromosomal aberrations and fetoplacental mosaicism in a prenatal tertial referral center, and to investigate the maternal and fetal characteristics in connection with genomewide non-invasive prenatal screening.Retrospective cohort study from 2014 to 2019 of pregnancies with invasive genetic analysis.In the cohort of 2504 cases, the proportion of CVS was 53.3%. We diagnosed 200 chromosomal aberrations (8%), including 13.5% of de novo rare chromosomal aberrations (14 rare autosomal trisomies, 6 polyploidies, 5 structural aberrations and 2 small supernumerary marker chromosomes). The rate of fetoplacental mosaicism was 12.4%/77.8% in common/atypical chromosomal aberrations (p = 0.001) and confined to placenta in 17/40 cases. Associated ultrasound abnormalities were abnormal nuchal translucency and major malformations in 58% and 35% with common trisomies and 11% (p = 0.006) and 67% (p = 0.047) with true mosaic cases of rare abnormalities, respectively. Major ultrasound malformation was facial dysmorphism with rare aberrations. Potential application of genomewide non-invasive prenatal test in atypical chromosomal aberrations presumably would have been false-positive in 12 cases (44%), false-negative in 1 case (3.7%), and would have early detected 2 cases of rare autosomal trisomies (7.4%) without ultrasound anomalies.Structural ultrasound malformations with normal nuchal translucency thickness may be indicative of rare chromosomal aberrations. Application of genomewide non-invasive prenatal test is an alternative of early diagnostic methods with a potential of discordant results due to mosaicism. Knowledge about the presence of fetoplacental mosaicism influences risk estimation and genetic counseling, detailed cytogenetic characterization is of utmost importance. Orv Hetil. 2021; 162(29): 1156-1165.

Acute lymphoblastic leukemia is the most common pediatric cancer characterized by a heterogeneous genomic landscape with copy number aberrations occurring at various stages of pathogenesis, disease progression, and treatment resistance. In this study, disease-relevant copy number aberrations were profiled in bone marrow samples of 91 children with B- or T-cell precursor acute lymphoblastic leukemia using digital multiplex ligation-dependent probe amplification (digitalMLPA(TM)). Whole chromosome gains and losses, subchromosomal copy number aberrations, as well as unbalanced alterations conferring intrachromosomal gene fusions were simultaneously identified with results available within 36 hours. Aberrations were observed in 96% of diagnostic patient samples, and increased numbers of copy number aberrations were detected at the time of relapse as compared with diagnosis. Comparative scrutiny of 24 matching diagnostic and relapse samples from 11 patients revealed three different patterns of clonal relationships with (i) one patient displaying identical copy number aberration profiles at diagnosis and relapse, (ii) six patients showing clonal evolution with all lesions detected at diagnosis being present at relapse, and (iii) four patients displaying conserved as well as lost or gained copy number aberrations at the time of relapse, suggestive of the presence of a common ancestral cell compartment giving rise to clinically manifest leukemia at different time points during the disease course. A newly introduced risk classifier combining cytogenetic data with digitalMLPA(TM)-based copy number aberration profiles allowed for the determination of four genetic subgroups of B-cell precursor acute lymphoblastic leukemia with distinct event-free survival rates. DigitalMLPA(TM) provides fast, robust, and highly optimized copy number aberration profiling for the genomic characterization of acute lymphoblastic leukemia samples, facilitates the decipherment of the clonal origin of relapse and provides highly relevant information for clinical prognosis assessment.

Introduction: 46,XX ovotesticular disorder of sex development (DSD), as defined by the Chicago consensus in 2006, is characterized by histologically confirmed testicular and ovarian tissue in an individual with a 46,XX karyotype and a wide phenotypic spectrum from female to male appearance. Case Presentation: We report the case of two 46,XX sex determining region Y (SRY) gene-negative siblings and their 46,XY father with an approximately 150 kilobase pair (kbp) duplication upstream of SOX9 (SRY-box 9) gene’s transcriptional start site on chromosome 17 (chr17), which involved SOX9’s minimal critical 46,XX sex reversal region. This duplication is sufficient to trigger male development in the absence of Y-chromosomal material and can lead to various degrees of masculinization in 46,XX individuals by overexpression of SOX9. Based on anamnestic information and pedigree analysis, another possible carrier of this copy number variation (CNV) could have been the father’s sister. Discussion: By comparing the duplications of our two sibling patients and previously reported similar cases, we suggest that the small differences between their breakpoints could alternatively modify the inner structure and functioning of SOX9’stopologically associated domain (TAD) due to the differing fine TAD arrangements. Our data support the phenotypic modularity impact – incomplete penetrance and variable expressivity – of very similar but non-identical CNVs, which are possibly inherited across three generations.

Terminal deletion of chromosome 4 (4q deletion syndrome) is a rare genetic condition that is characterized by a broad clinical spectrum and phenotypic variability. Diagnosis of the distinct condition can be identified by conventional chromosome analysis and small deletions by novel molecular cytogenetic methods such as microarray comparative genome hybridization (aCGH). Prenatal diagnosis is challenging; to date 10 cases have been described. We report a prenatally diagnosed case of de novo 4q deletion syndrome confirmed by conventional karyotyping and FISH due to an elevated combined risk for Down syndrome and prenatal ultrasound findings. aCGH validated the diagnosis and offered exact characterization of the disorder. Cytogenetic and microarray results described a 4q32.1qter terminal deletion of the fetus. Prenatal ultrasound detected multiple nonstructural findings (micrognathia, choroid plexus cysts, echogenic fetal bowel, short femur, and cardiac axis deviation). Pregnancy was terminated at 20 weeks. In addition to the index patient, we reviewed the 10 prenatally published cases of 4q deletion syndrome in the literature and compared these with our results. We summarize the patients' characteristics and prenatal clinical findings. Alterations of maternal serum biochemical factors, an elevated combined risk for trisomies, and distinct ultrasonographic findings can often be observed in cases of prenatal 4q deletion syndrome and may facilitate the otherwise difficult prenatal diagnosis.

The short arm of chromosome 16 (16p) is enriched for segmental duplications, making it susceptible to recurrent, reciprocal rearrangements implicated in the etiology of several phenotypes, including intellectual disability, speech disorders, developmental coordination disorder, autism spectrum disorders, attention deficit hyperactivity disorders, obesity and congenital skeletal disorders. In our clinical study 73 patients were analyzed by chromosomal microarray, and results were confirmed by fluorescence in situ hybridization or polymerase chain reaction. All patients underwent detailed clinical evaluation, with special emphasis on behavioral symptoms. 16p rearrangements were identified in 10 individuals. We found six pathogenic deletions and duplications of the recurrent regions within 16p11.2: one patient had a deletion of the distal 16p11.2 region associated with obesity, while four individuals had duplications, and one patient a deletion of the proximal 16p11.2 region. The other four patients carried 16p variations as second-site genomic alterations, acting as possible modifying genetic factors. We present the phenotypic and genotypic results of our patients and discuss our findings in relation to the available literature. info:eu-repo/semantics/publishedVersion

Invasive prenatal testing and conventional G-banding chromosome analysis have been considered to be the gold standard of fetal cytogenetic diagnosis. Standard karyotyping is, however, constrained by the limits of the resolution of using a microscope. The advantage of molecular karyotyping, array based methods is the evaluation of sub-microscopic copy number changes across the whole genome in a single analysis. The application of array comparative genome hybridization has greatly increased the detection of pathogenic chromosomal abnormalities in prenatal settings. Based on available data in the international literature of the last decade, the clinical utility of arrayCGH is the recognition of some 1-2% and 5-7% additional genetical information compared to metaphase karyotype alone in fetuses without ultrasound anomaly and in fetuses with ultrasonographically detected malformations, respectively. Thus arrayCGH improves the prenatal diagnosis of genetic abnormalities mainly in fetuses with structural sonographic findings. In the present paper we review the literature of chromosomal microarray and make a proposal for the application of the method in Hungarian prenatal genetical practice. Orv Hetil. 2019; 160(13): 484-493.Absztrakt: Az invazív mintavétel kapcsán elvégzett hagyományos magzati kromoszómavizsgálat a mai napig a praenatalis diagnosztika alapvető vizsgálómódszere. Felhasználásának a fénymikroszkópos vizsgálat felbontási képessége szab határt. A kariotipizálással nem felismerhető, szubmikroszkópos kromoszóma-rendellenességek, microdeletiók és microduplicatiók, kópiaszám-variációk (CNV-k) vizsgálatára a nagy felbontású molekuláris vizsgálóeljárások biztosítanak lehetőséget. A kromoszomális összehasonlító microarray-vizsgálat (array-komparatív genomhibridizálás – arrayCGH) alkalmas az anyai életkortól függetlenül előforduló kópiaszám-variációk prae- és postnatalis kimutatására. A módszer a fejlett országok orvosi gyakorlatában rutinszerűen alkalmazott eljárás a magzati diagnosztikában. Az elmúlt egy évtized külföldi eredményei alapján alkalmazása ultrahangeltérést nem mutató magzatok esetén körülbelül 1–2%, strukturális ultrahangeltérést mutató magzatoknál körülbelül 5–7% többlet genetikai információval szolgál a hagyományos kromoszómavizsgálattal szemben. Közleményünkben áttekintjük az arrayCGH módszerét, praenatalis alkalmazásának nemzetközi gyakorlatát, s javaslatokat és indikációs kört fogalmazunk meg a módszer praenatalis használatának magyarországi bevezetésére. Orv Hetil. 2019; 160(13): 484–493.

Contains fulltext : 204641.pdf (Publisher’s version ) (Open Access) Genetic abnormalities provide vital diagnostic and prognostic information in pediatric acute lymphoblastic leukemia (ALL) and are increasingly used to assign patients to risk groups. We recently proposed a novel classifier based on the copy-number alteration (CNA) profile of the 8 most commonly deleted genes in B-cell precursor ALL. This classifier defined 3 CNA subgroups in consecutive UK trials and was able to discriminate patients with intermediate-risk cytogenetics. In this study, we sought to validate the United Kingdom ALL (UKALL)-CNA classifier and reevaluate the interaction with cytogenetic risk groups using individual patient data from 3239 cases collected from 12 groups within the International BFM Study Group. The classifier was validated and defined 3 risk groups with distinct event-free survival (EFS) rates: good (88%), intermediate (76%), and poor (68%) (P < .001). There was no evidence of heterogeneity, even within trials that used minimal residual disease to guide therapy. By integrating CNA and cytogenetic data, we replicated our original key observation that patients with intermediate-risk cytogenetics can be stratified into 2 prognostic subgroups. Group A had an EFS rate of 86% (similar to patients with good-risk cytogenetics), while group B patients had a significantly inferior rate (73%, P < .001). Finally, we revised the overall genetic classification by defining 4 risk groups with distinct EFS rates: very good (91%), good (81%), intermediate (73%), and poor (54%), P < .001. In conclusion, the UKALL-CNA classifier is a robust prognostic tool that can be deployed in different trial settings and used to refine established cytogenetic risk groups.

Background Premature ovarian failure / primary ovarian insufficiency (POF/POI) associated with the mutations of the FMR1 (Fragile-X Mental Retardation 1) gene belongs to the group of the so-called trinucleotide expansion diseases. Our aim was to analyse the relationship between the paternally inherited premutation (PIP) and the maternally inherited premutation (MIP) by the examination of the family members of women with POF, carrying the premutation allele confirmed by molecular genetic testing. Methods Molecular genetic testing was performed in the patients of the 1st Department of Obstetrics and Gynecology with suspected premature ovarian failure. First we performed the southern blot analyses and for the certified premutation cases we used the Repeat Primed PCR. Results Due to POF/POI, a total of 125 patients underwent genetic testing. The FMR1 gene trinucleotide repeat number was examined in the DNA samples of the patients, and in 15 cases (12%) deviations (CGG repeat number corresponding to premutation or gray zone) were detected. In 6 cases out of the 15 cases the CGG repeat number fell within the range of the so-called gray zone (41–54 CGG repeat) (4.8%, 6/125), and the FMR1 premutation (55–200 CGG repeat) ratio was 7.2% (9/125). In 4 out of the 15 cases we found differences in both alleles, one was a premutation allele, and the other allele showed a repeat number belonging to the gray zone. Out of 15 cases, only maternal inheritance (MIP) was detected in 2 cases, in one case the premutation allele (91 CGG repeat number), while in the other case an allele belonging to the gray zone (41 CGG repeat number) were inherited from their mothers. In 10 out of 15 cases, the patient inherited the premutation allele only from the father (PIP). In 5 out of the 10 cases (50%) the premutation allele was inherited from the father, and the repeat number ranged from 55 to 133. Out of 125 cases, 9 patients had detectable cytogenetic abnormalities (7.2%). Conclusions The RP-PCR method can be used to define the smaller premutations and the exact CGG number. Due to the quantitative nature of the RP-PCR, it is possible to detect the mosaicism as well.

Introduction: Acute lymphoblastic leukemia (ALL) is the most common pediatric malignancy characterized by a heterogeneous genomic landscape. Copy number aberrations (CNA) emerge during the development, progression and treatment resistance of ALL, and can serve as genomic markers for prognostic classification of patients or for scrutinizing clonal evolution associated with relapse. While identification of distinct CNAs with well-characterized prognostic significance has its own value, uncovering the co-segregation of driver aberrations in individual patient samples could allow for a more personalized risk assessment and treatment response prediction. Methods: Disease-relevant CNAs were profiled in children with B- or T-cell precursor ALL using a next-generation sequencing based digital multiplex ligation-dependent probe amplification (digitalMLPA) assay containing 598 probes specific for 54 genes with key relevance in ALL. Besides the diagnostic samples of 91 patients treated according to the BFM protocols, 14 matching samples drawn at the time of first or second relapse were comparatively analyzed. Clonal relationship between B-cell precursor cell populations prevailing at different time points during the disease course was also investigated by screening immunoglobulin heavy-chain gene rearrangements in matching diagnostic and relapse samples using Illumina deep-sequencing with >20,000x coverage. Results: Whole chromosome gains and losses, subchromosomal CNAs as well as alterations conferring intrachromosomal gene fusions were simultaneously identified by digitalMLPA with results available within 36 hours. Aberrations were observed in 96% of diagnostic patient samples and increased numbers of CNAs were detected in individual samples at the time of relapse as compared to diagnosis. DigitalMLPA results were successfully validated by conventional MLPA, FISH and PCR data. Comparative scrutiny of 24 matching diagnostic and relapse samples from 11 patients harboring CNAs revealed three different patterns of clonal relationships with (i) one patient displaying identical CNA profiles at diagnosis and relapse, (ii) six patients showing clonal evolution with all lesions detected at diagnosis being present at relapse and (iii) four patients displaying conserved as well as lost or gained CNAs at the time of relapse, suggestive of the presence of a common ancestral cell compartment giving rise to clinically manifest leukemia at different time points during the disease course. Time between diagnosis and first relapse of T-ALL patients displaying altered CNA profiles suggested a prolonged time requirement of clonal evolution, and of the development of manifest leukemia from an ancestral clone compared to the quick return of an identical clone at the time of relapse. Comparison of the IGH gene rearrangements identified at diagnoses and relapse revealed identical compositions of the most abundant clonotypes in all but one B-ALL patients analyzed; hence, IGH repertoire did not reveal an additional depth of clonal history in our cohort, e.g. by demonstrating the presence of an ancestral clone as the major source of clonal expansion at disease progression in a patient with altered CNA profiles suggesting direct clonal evolution between diagnosis and relapse. Copy number profiles acquired by digitalMLPA were used for determining CNA-based risk groups (Table 1) which were combined with karyotyping and molecular cytogenetic data in order to establish an extended prognostic classifier for patients with B-cell precursor ALL. This novel classifier distinguished four combined genetic risk groups showing significantly different 5-year survival rates (GR: 97%, IR: 84%, IHR: 63% and PR: 13%). Conclusions: DigitalMLPA allows for a rapid, scalable and highly optimized copy number profiling of genomic regions recurrently altered by driver aberrations in pediatric ALL. Based on the comparison of CNA profiles at diagnosis and relapse, clonal evolution and emergence of relapse from an ancestral clone are the predominant driving mechanisms of disease progression. Comprehensive copy number profiling by digitalMLPA identifies distinct prognostic groups for risk assessment in B-cell precursor ALL. Supporting grants : LP95021, K_16 #119950, NVKP_16-1-2016-0004, KH17-126718, BO/00320/18/5, FK_19 #131476, ÚNKP-19-4-SE-77 Disclosures Benard-Slagter: MRC Holland: Employment. de Groot:MRC Holland: Employment. Savola:MRC Holland: Employment.

Background: Nephronophthisis, an autosomal recessive nephropathy, is responsible for 10% of childhood chronic renal failure. The deletion of its major gene, NPHP1, with a minor allele frequency of 0.24% in the general population, is the most common mutation leading to a monogenic form of childhood chronic renal failure. It is challenging to detect it in the heterozygous state. We aimed to evaluate the sensitivity and the specificity of the quantitative multiplex PCR of short fluorescent fragments (QMPSF) in its detection. Methods: After setting up the protocol of QMPSF, we validated it on 39 individuals diagnosed by multiplex ligation-dependent probe amplification (MLPA) with normal NPHP1 copy number (n=17), with heterozygous deletion (n=13, seven parents and six patients), or with homozygous deletion (n=9). To assess the rate of the deletions that arise from independent events, deleted alleles were haplotyped. Results: The results of QMPSF and MLPA correlated perfectly in the identification of 76 heterozygously deleted and 56 homozygously deleted exons. The inter-experimental variability of the dosage quotient obtained by QMPSF was low: control, 1.05 (median; range, 0.86−1.33, n = 102 exons); heterozygous deletion, 0.51 (0.42−0.67, n = 76 exons); homozygous deletion, 0 (0−0, n = 56 exons). All patients harboring a heterozygous deletion were found to carry a hemizygous mutation. At least 15 out of 18 deletions appeared on different haplotypes and one deletion appeared de novo. Conclusions: The cost- and time-effective QMPSF has a 100% sensitivity and specificity in the detection of NPHP1 deletion. The potential de novo appearance of NPHP1 deletions makes its segregation analysis highly recommended in clinical practice.

Muscular dystrophies are a genetically heterogeneous group of degenerative muscle disorders. This article focuses on two severe forms of muscular dystrophies and provides genetic data for a large cohort of Hungarian patients diagnosed within the last few years by the authors. The Duchenne/Becker muscular dystrophy (DMD/BMD) is caused by mutations in the dystrophin gene, which is located on chromosome Xp21. The genetic analysis of dystrophin is usually performed by multiplex polymerase chain reaction (PCR), which detects approximately 95% of all deletions but does not distinguish between one and two copies of the exons investigated. The present work, therefore, concentrates on the improvement of the diagnostic panel for the analysis of DMD/BMD in Hungary. Radioactively labelled cDNA probes, encompassing the whole dystrophin gene detect all the deletions and the analysis is quantitative. In addition, the new multiple ligation-dependent probe amplification (MLPA) technique was recently introduced that enabled more reliable and faster quantitative detection of the entire dystrophin gene. The genomic basis of facioscapulohumeral muscular dystrophy (FSHD) is associated with contraction of the D4Z4 repeat region in the subtelomere of chromosome 4q. In case of FSHD, molecular genetic criteria still have to be improved because of the complexity of the disorder.

A Duchenne-/Becker-izomdystrophia súlyos, X-kromoszómához kötött, recesszív neuromuscularis betegség, amelynek előfordulási gyakorisága 1/3500 (Duchenne-típus) és 1/30000 (Becker-típus). A betegség kialakulásáért felelős gént az Xp21-es régióban lokalizálták, amelynek terméke a 427 kD nagyságú dystrophinfehérje. Deletiókat, pontmutációkat és ritkán duplikációkat a teljes génben detektáltak, ami megnehezíti a diagnosztikát. Multiplex polimeráz láncreakció az érintett férfiak 95%-ában kimutatja a deletiót, de nem alkalmas a hordozóság detektálására. Southern-blot-analízissel, 6 cDNS-próbával a teljes 14 kb nagyságú dystrophin-gént kódoló szekvencia analizálható, és a különböző exonokhoz tartozó radioaktív jelek intenzitásának meghatározásával a hordozósági állapot is vizsgálható. A Southern-blot-analízis idő- és anyagigényes, összehasonlítva a multiplex ligációfüggő próbaamplifikációs módszerével, amely lehetővé teszi több DNS-szekvencia relatív mennyiségének meghatározását egyetlen reakcióban. Kilencvenhárom női hozzátartozó hordozósági státuszát vizsgálták Southern-blot- és multiplex ligációfüggő próbaamplifikáció analízissel összehasonlítva. Negyvenkét esetben (45%) igazolták mind a két módszerrel a hordozósági státuszt. Érdekességként, a hordozó populációban két esetben detektáltak duplikációt, két másik esetben pedig a női rokonok hordozónak bizonyultak a teljes dystrophingén hiánya tekintetében. Három további leánybeteg bizonyult hordozónak, klinikai tünetekkel (manifest carrier). A szerzők tapasztalatai alapján a multiplex ligációfüggő próbaamplifikáció analízise a Duchenne-/Becker-izomdystrophia-betegség hordozóságának szűrésében gyors és megbízható módszernek bizonyul, valamint alkalmas az érintett férfiak deletioméretének pontos meghatározására is. A pontos deletiós vagy duplikációs méret ismeretében a fenotípusra is előrejelzés adható, amely a jövőbeli terápiás eljárás kiválasztásához is segítséget nyújthat.

Background Turner syndrome, a congenital condition that affects 1/2,500 births, results from absence or structural alteration of the second sex chromosome. Turner syndrome is usually associated with short stature, gonadal dysgenesis and variable dysmorphic features. The classical 45,X karyotype accounts approximately for half of all patients, the remainder exhibit mosaicism or structural abnormalities of the X chromosome. However, complex intra-X chromosomal rearrangements involving more than three breakpoints are extremely rare. Results We present a unique case of a novel complex X chromosome rearrangement in a young female patient presenting successively a wide range of autoimmune diseases including insulin dependent diabetes mellitus, Hashimoto’s thyroiditis, celiac disease, anaemia perniciosa, possible inner ear disease and severe hair loss. For the genetic evaluation, conventional cytogenetic analysis and FISH with different X specific probes were initially performed. The complexity of these results and the variety of autoimmune problems of the patient prompted us to identify the exact composition and breakpoints of the rearranged X as well as methylation status of the X chromosomes. The high resolution array-CGH (assembly GRCh37/hg19) detected single copy for the whole chromosome X short arm. Two different sized segments of Xq arm were present in three copies: one large size of 80,3 Mb from Xq11.1 to Xq27.3 region and another smaller (11,1 Mb) from Xq27.3 to Xq28 region. An 1,6 Mb Xq27.3 region of the long arm was present in two copies. Southern blot analysis identified a skewed X inactivation with ≈ 70:30 % ratios of methylated/unmethylated fragments. The G-band and FISH patterns of the rearranged X suggested the aspect of a restructured i(Xq) chromosome which was shattered and fortuitously repaired. The X-STR genotype analysis of the family detected that the patient inherited intact maternal X chromosome and a rearranged paternal X chromosome. The multiple Xq breakages and fusions as well as inverted duplication would have been expected to cause a severe Turner phenotype. However, the patient lacks many of the classic somatic features of Turner syndrome, instead she presented multiorgan autoimmune diseases. Conclusions The clinical data of the presented patient suggest that fragmentation of the i(Xq) chromosome elevates the risk of autoimmune diseases.

Autosomal dominant facioscapulohumeral muscular dystrophy (FSHD) is caused by contraction of the D4Z4 repeat region on 4q35. In addition, epigenetic modifying factors play a role in the complex pathomechanism of the disease. Aims: Introduction of a new diagnostic panel in Hungary for the extended molecular analysis of the disease which also provides new insights into the pathomechanism. Methods: In total, DNA samples of 185 clinically diagnosed FSHD patients and 71 asymptomatic relatives were analyzed by EcoRI and BlnI restriction digestion and Southern blot technique with probe p13-E11. Further investigations of the 4q35 alleles associated with the FSHD phenotype utilized qA and qB probes and a restriction analysis of the proximal D4Z4 unit by detecting a G/C SNP and the methylation status. Results: From the patients analyzed 115 had the D4Z4 repeat contraction, whereas from 71 asymptomatic family members five harbored the pathogenic fragment size. In eight families, prenatal testing had to be offered with an outcome of four affected fetuses. Methylation test was performed in 31 genetically confirmed FSHD patients and hypomethylation status was detected in all cases. All the 115 confirmed patients had 4qA alleles with the G polymorphism. Translocation events between 4q35 and the homologous 10q26 regions were also detected. Conclusion: Molecular diagnosis of FSHD became a routine approach in Hungary thus supporting the work of the clinicians, improving quality of life and genetic counseling of the affected families. The provided results from this research suggest that FSHD is associated with complex epigenetic disease mechanisms. Orv. Hetil., 2011, 152, 1576–1585.

Recent medical genetic research has identified a number of novel, or previously known, but rare conditions, caused by private founder mutations. The Finnish and Ashkenazi Jew populations provide the best examples for identifying genes in unique genetic disorders. In these populations, research efforts and high-level medical services resulted in intense improvements of medical care and in organization of population-based screening programs. Hereditary disorders of the Roma populations are known for a long time. The genetic background of these diseases has been established by extensive molecular genetic studies. The Romas represent 6% of the Hungarian population and live under extremely bad health conditions. Therefore, our aim was to map the incidence of the hereditary neuromuscular disorders among the Hungarian Roma population. Moreover, we intended to provide proper information, genetic counseling and possible prevention strategies for the families at risk, which should represent a primer task in public health. Because of our experience in neuromuscular disorders, we choose six, frequent, autosomal recessive disorders for these clinical and genetic studies: hereditary motor and sensory neuropathy type Lom (HMSNL), hereditary motor and sensory neuropathy type Russe (HMSNR), congenital cataracts facial dysmorphism syndrome (CCFDN), limb-girdle muscular dystrophy 2C (LGMD2C), congenital myasthenic syndrome (CMS) and spinal muscular atrophy (SMA). Following identification of the founder mutations, the possibility of prenatal diagnosis and carrier screening for family members will contribute to the decrease of the recurrence risk for these severe, mostly untreatable disorders.

A comprehensive study of the Hungarian Duchenne/Becker muscular dystrophy (DMD/BMD) families is presented. Deletions in the hot spots regions were identified by multiplex PCR, whereas rare mutations were detected by Southern blot and multiplex ligation-dependent probe amplification (MLPA) techniques. DMD/BMD disease was confirmed and exact deletion borders were determined in 19 out of 135 affected males using multiplex PCR. Additional exons involved as well as rare exon deletions were identified by MLPA in 71 male patients, whereas duplications were observed in seven patients. In two DMD patients, the entire dystrophin gene and adjacent genes were deleted. Out of the 95 female relatives, 41 proved to be carriers, including three manifesting carrier females. Using MLPA method, a large portion of the Hungarian DMD/BMD patients and their female relatives were exactly genotyped. For the first time, the incidence and prevalence of asymptomatic and symptomatic female carriers in Hungary was estimated.

Multiple-drug resistance in enteric bacteria is frequently associated with integrons. To determine whether integrons may play a role in avian pathogenic Escherichia coli, isolates of extra-intestinal (n = 27) and intestinal (n = 40) E. coli from dead chicks and turkey poults were analysed for the presence of class 1 integrons and of the virulence-associated genes iss, tsh and colV. Eleven extra-intestinal strains possessed a 1.0 kb class 1 integron with a variable region of aadA1 and were resistant to tetracycline. These traits were indicative of the presence of the Tn21 transposon, which was confirmed by polymerase chain reaction. All extra-intestinal strains had the colV, iss and tsh genes, but none of these genes were cotransferred with the 1.0 kb integron when conjugal transferability was tested. The integron content of the intestinal strains showed considerable variability: one or two of four different class 1 integrons, which varied from 1.0 to 2.4 kb in size, were detected in the 11 strains. The aadA7 gene of the 1.0 kb integron, the dfrA1-aadA1 genes of the 1.6 kb integron and the folA-catB3-aadA5 genes of the 2.4 kb integron were identical to those described by other workers. However, the orfIN682-dhfrV-orfD gene cassette arrangement of the 1.5 kb integron of an intestinal strain of serogroup O5 had no similarity to any previously reported integrons. Conjugal transfer of the 1.6 and 2.4 kb integrons was successful, and in a serogroup O33 intestinal E. coli strain the iss gene was apparently cotransferred with a 1.6 kb integron. The 1.0 and 1.5 kb integrons were not transferable. Our data suggest that intestinal E. coli strains of poultry may be a reservoir for emerging multiresistant strains of avian pathogenic E. coli.

Mutations in CAV3 gene encoding the protein caveolin-3 are associated with autosomal dominant limb girdle muscular dystrophy 1C, rippling muscle disease, hyperCKemia, distal myopathy, hypertrophic cardiomyopathy and rare autosomal recessive limb girdle muscular dystrophy phenotypes. In a 57-year-old patient with asymmetric limb girdle weakness, we detected a novel homozygous intronic mutation (IVS1+2T>C) of the CAV3 gene. This is the first splicing mutation reported for CAV3 . These findings add to the clinical and genetic variability of CAV3 mutations.

Limb gird muscular dystrophies (LGMD2) are a clinically and genetically heterogeneous group of hereditary diseases with autosomal recessive trait, characterized by progressive atrophy and weakness predominantly in the proximal limb muscles. The authors present clinical, histological, immunohistochemical and immunoblot results of two sisters suffering from so far unclassified autosomal recessive limb girdle muscular dystrophy. Haplotype analysis for genes possibly involved in autosomal recessive limb girdle muscular dystrophies was performed in the genetically informative family. All of the results pointed to a molecular genetic defect of the calpain-3 (CAPN3) gene. Direct sequencing of the CAPN3 gene revealed compound heterozygous state for two mutations previously described in association with limb girdle muscular dystrophy, proving pathogenicity. The authors would like to emphasize the importance of the above described combined strategy in diagnosing limb girdle muscular dystrophies.

Background Microduplications 22q11 have been characterized as a genomic duplication syndrome mediated by nonallelic homologous recombination between region-specific low-copy repeats. Here we report on a 19 years old boy with intellectual disability having an unexpected structurally complex ring small supernumerary marker chromosome (sSMC) originated from a larger trisomy and a smaller tetrasomy of proximal 22q11 harboring additional copies of cat eye syndrome critical regions genes. Results Principal clinical features were: anorectal and urogenital malformations, total anomalous pulmonary venous return with secundum ASD, hearing defect, preauricular pits, seizure and eczema. The proband also presented some rare or so far not reported clinical findings such as hyperinsulinaemia, severe immunodeficiency and grave cognitive deficits. Chromosome analysis revealed a mosaic karyotype with the presence of a small ring-like marker in 60% of cells. Array CGH detected approximately an 1,2 Mb single and a 0,2 Mb double copy gain of the proximal long arm of chromosome 22. The 1,3 Mb intervening region of chromosome 22 from centromere to the breakpoints showed no copy alteration. The karyotype of the patient was defined as 47,XY,+mar[60]/46,XY[40].ish idic r(22)(q11.1.q11.21) × 4.arr 22q11(17,435, 645-18,656,678) × 3,(17,598,642-17,799,783) × 4 dn. Conclusions The present report is the first one with a detailed description of clinical presentation in a patient carrying an atypical size ring sSMC (22) analyzed by array CGH. The specialty of the finding is emphasized by the fact that although the patient had a mosaic sSMC and the amplified region was smaller than in typical cat eye syndrome cases, the clinical presentation was severe.

Background One of the frequent reasons for unsuccessful conception is premature ovarian failure/primary ovarian insufficiency (POF/POI) that is defined as the loss of functional follicles below the age of 40 years. Among the genetic causes the most common one involves the X chromosome, as in Turner syndrome, partial X deletion and X-autosome translocations. Here we report a case of a 27-year-old female patient referred to genetic counselling because of premature ovarian failure. The aim of this case study to perform molecular genetic and cytogenetic analyses in order to identify the exact genetic background of the pathogenic phenotype. Results For premature ovarian failure disease diagnostics we performed the Fragile mental retardation 1 gene analysis using Southern blot technique and Repeat Primed PCR in order to identify the relationship between the Fragile mental retardation 1 gene premutation status and the premature ovarion failure disease. At this early onset, the premature ovarian failure affected patient we detected one normal allele of Fragile mental retardation 1 gene and we couldn’t verify the methylated allele, therefore we performed the cytogenetic analyses using G-banding and fluorescent in situ hybridization methods and a high resolution molecular cytogenetic method, the array comparative genomic hybridization technique. For this patient applying the G-banding, we identified a large deletion on the X chromosome at the critical region (ChrX q21.31-q28) which is associated with the premature ovarian failure phenotype. In order to detect the exact breakpoints, we used a special cytogenetic array ISCA plus CGH array and we verified a 67.355 Mb size loss at the critical region which include total 795 genes. Conclusions We conclude for this case study that the karyotyping is definitely helpful in the evaluation of premature ovarian failure patients, to identify the non submicroscopic chromosomal rearrangement, and using the array CGH technique we can contribute to the most efficient detection and mapping of exact deletion breakpoints of the deleted Xq region.