Maternal mosaicism underlies the inheritance of a rare germline AKT3 variant which is responsible for megalencephaly-polymicrogyria-polydactyly-hydrocephalus syndrome in two Roma half-siblings.

Megalencephaly-polymicrogyria-polydactyly-hydrocephalus (MPPH) syndrome is a developmental brain disorder characterized by an enlarged brain size with bilateral perisylvian polymicrogyria and a variable degree of ventriculomegaly. MPPH syndrome is associated with oromotor dysfunction, epilepsy, intellectual disability and postaxial hexadactyly. The molecular diagnosis of this disorder is established by the identification of a pathogenic variant in either AKT3, CCND2 or PIK3R2. Previously reported AKT3 variants are associated with various brain abnormalities and may lead to megalencephaly. MPPH syndrome is usually due to germline pathogenic AKT3 variants. Somatic mosaic pathogenic variants associated with hemimegalencephaly, which is similar to MPPH, have also been observed. A Hungarian Roma family with two half-siblings, which present with intellectual disability, dysmorphic features, epilepsy, brain malformations, and megalencephaly was studied. Whole exome sequencing (WES) analysis was performed. WES analysis revealed a heterozygous c.1393C > T p.(Arg465Trp) pathogenic missense AKT3 variant in both affected half-siblings. The variant was verified via Sanger sequencing and was not present in the DNA sample from the healthy mother, which was derived from peripheral blood, suggesting maternal germline mosaicism. In conclusion, this is the first report in which maternal germline mosaicism of a rare pathogenic AKT3 variant leads to autosomal dominantly inherited MPPH syndrome.

Intrafamilial variability of limb-girdle muscular dystrophy, LGMD1D type.

LGMD1D is an autosomal dominant limb girdle muscular dystrophy caused by variants in the DNAJB6 gene. This is typically an adult-onset disorder characterized by moderately progressive proximal muscle weakness without respiratory or bulbar involvement; however phenotypic variability is often observed with some individuals having earlier onset and more severe symptoms. Here, we present a family with a novel NM_005494.2:c.271T > G p.(Phe91Val) variant in DNAJB6 with a late-onset, mild and slowly progressive form of the disease, including one individual, who in her 7th decade of life has subclinical LGMD1D with only mild features on muscle biopsy and MRI. Unlike previously reported cases where missense variants affecting the Phe91 amino acid residue are associated with a more severe form of the disease, this family represents the mild end of the LGMD1D clinical spectrum. Therefore, this family adds further complexity to the genotype-phenotype correlation in DNAJB6-associated muscular dystrophies.

Revealing the Genetic Impact of the Ottoman Occupation on Ethnic Groups of East-Central Europe and on the Roma Population of the Area.

History of East-Central Europe has been intertwined with the history of Turks in the past. A significant part of this region of Europe has been fallen under Ottoman control during the 150 years of Ottoman occupation in the 16-17th centuries. The presence of the Ottoman Empire affected this area not only culturally but also demographically. The Romani people, the largest ethnic minority of the East-Central European area, share an even more eventful past with Turkish people from the time of their migration throughout Eurasia and they were a notable ethnic group in East-Central Europe in the Ottoman era already. The relationship of Turks with East-Central European ethnic groups and with regional Roma ethnicity was investigated based on genome-wide autosomal single nucleotide polymorphism data. Population structure analysis, ancestry estimation, various formal tests of admixture and DNA segment analyses were carried out in order to shed light to the conclusion of these events on a genome-wide basis. Analyses show that the Ottoman occupation of Europe left detectable impact in the affected East-Central European area and shaped the ancestry of the Romani people as well. We estimate that the investigated European populations have an average identity-by-descent share of 0.61 with Turks, which is notable, compared to other European populations living in West and North Europe far from the affected area, and compared to the share of Sardinians, living isolated from these events. Admixture of Roma and Turks during the Ottoman rule show also high extent.

Hearing impairment locus heterogeneity and identification of PLS1 as a new autosomal dominant gene in Hungarian Roma.

Roma are a socially and culturally distinct isolated population with genetically divergent subisolates, residing mainly across Central, Southern, and Eastern Europe. We evaluated the genetic etiology of hearing impairment (HI) in 15 Hungarian Roma families through exome sequencing. A family with autosomal dominant non-syndromic HI segregating a rare variant in the Calponin-homology 2 domain of PLS1, or Plastin 1 [p.(Leu363Phe)] was identified. Young adult Pls1 knockout mice have progressive HI and show morphological defects to their inner hair cells. There is evidence that PLS1 is important in the preservation of adult stereocilia and normal hearing. Four families segregated the European ancestral variant c.35delG [p.(Gly12fs)] in GJB2, and one family was homozygous for p.(Trp24*), an Indian subcontinent ancestral variant which is common amongst Roma from Slovakia, Czech Republic, and Spain. We also observed variants in known HI genes USH1G, USH2A, MYH9, MYO7A, and a splice site variant in MANBA (c.2158-2A>G) in a family with HI, intellectual disability, behavioral problems, and respiratory inflammation, which was previously reported in a Czech Roma family with similar features. Lastly, using multidimensional scaling and ADMIXTURE analyses, we delineate the degree of Asian/European admixture in the HI families understudy, and show that Roma individuals carrying the GJB2 p.(Trp24*) and MANBA c.2158-2A>G variants have a more pronounced South Asian background, whereas the other hearing-impaired Roma display an ancestral background similar to Europeans. We demonstrate a diverse genetic HI etiology in the Hungarian Roma and identify a new gene PLS1, for autosomal dominant human non-syndromic HI.

Refining the South Asian Origin of the Romani people.

BACKGROUND: Recent genetic studies based on genome-wide Single Nucleotide Polymorphism (SNP) data further investigated the history of Roma and suggested that the source of South Asian ancestry in Roma originates most likely from the Northwest region of India. METHODS: In this study, based also on genome-wide SNP data, we attempted to refine these findings using significantly larger number of European Roma samples, an extended dataset of Indian groups and involving Pakistani groups into the analyses. Our Roma data contained 179 Roma samples. Our extended Indian data consisted of 51 distinct Indian ethnic groups, which provided us a higher resolution of the population living on the Indian subcontinent. We used in this study principal component analysis and other ancestry estimating methods for the study of population relationships, several formal tests of admixture and an improved algorithm for investigating shared IBD segments in order to investigate the main sources of Roma ancestry. RESULTS: According to our analyses, Roma showed significant IBD sharing of 0.132 Mb with Northwest Indian ethnic groups. The most significant IBD sharings included ethnic groups of Punjab, Rajasthan and Gujarat states. However, we found also significant IBD sharing of 0.087 Mb with ethnic groups living in Pakistan, such as Balochi, Brahui, Burusho, Kalash, Makrani, Pashtun and Sindhi. CONCLUSION: Our results show that Northwest India could play an important role in the South Asian ancestry of Roma, however, the origin of Romani people might include the area of Pakistan as well.

Marked Differences of Haplotype Tagging SNP Distribution, Linkage, and Haplotype Profile of APOA5 Gene in Roma Population Samples.

Roma people are underprivileged, neglected population worldwide, with severe healthcare problems. They have significantly increased prevalence of cardiovascular morbidity, presumably related to their poor social status, alcohol consumption and smoking habits. Assuming that genetic background also plays a role in their susceptibility for cardiovascular diseases, we hypothesized that APOA5 gene polymorphisms, an important role-player in lipid metabolism and in the development of metabolic syndrome and cardio/cerebrovascular events, may also be involved. We examined four APOA5 polymorphisms in 363 Roma and 404 Hungarian DNA samples. For rs662799, rs2266788, rs207560 and rs3135506 we found elevated plasma triglyceride levels in the risk allele carriers compared to non-carriers in both populations. At least a two-fold significant increase was detected in minor allele frequencies in Roma when compared to Hungarians, except the rs2266788 variant. Haplotype analysis revealed significant increase of APOA5*2, APOA5*4 in Roma, as opposed to the higher levels of APOA5*5 found in Hungarians. Different linkage disequilibrium was found between rs207560 and rs3135506 variants in Roma compared to Hungarians. The profound differences observed in almost all APOA5 polymorphisms in Roma require special attention, since these variants are known to associate with cardio/cerebrovascular susceptibility.

Interethnic variability of CYP4F2 (V433M) in admixed population of Roma and Hungarians.

AIMS: Pharmacogenetic based dosing recommendations are provided in FDA-approved warfarin label for Caucasians. Evidence of notable difference in dosing algorithms of under-represented populations forced us to explore the genetic variability of CYP4F2 gene in Roma and Hungarian populations. PATIENTS AND METHODS: 484 Roma, 493 Hungarian untreated subjects were genotyped for the CYP4F2*3 (rs2108622) variant by PCR-RFLP assay. RESULTS AND DISCUSSION: We firstly report, that frequencies of the CYP4F2 rs2108622 GG, GA, AA genotypes and A allele in the Roma population were 46.5%, 42.6%, 10.9% and 32.2%; in Hungarians 50.1%, 42.2%, 7.7% and 22.8%, respectively. Bearing of two minor alleles of CYP4F2 missense variant (AA genotype) modestly explains inter-ethnic differences of studied populations (p<0.08). CYP4F2*3 (V433M) risk allele frequency of Roma (0.32) was in higher range, and of Hungarians (0.23) in lower range, as compared with other world populations. CONCLUSIONS: Roma have an elevated chance for higher mean warfarin dose, besides a decreased risk of major bleeding events in long-term warfarin use.

Significant interethnic differencies in functional variants of PON1 and P2RY12 genes in Roma and Hungarian population samples.

Antiplatelet therapy with clopidogrel is one of the most common therapies given to patients worldwide. However, the clinical efficacy and toxicity of clopidogrel is not constant in every patient due to interindividual variations. There are several factors that contribute to these interindividual differencies such as SNPs in genes of specific receptors and enzymes. PON1 (paraoxonase 1) plays an important role in the bioactivation of clopidogrel. Single nucleotide polymorphisms of this gene decrease the activity of paraoxonase enzyme and lead to an unefficient clopidogrel effect. P2RY12 (purinergic receptor P2Y, G-protein coupled, 12) gene is coding a receptor, which is situated on the surface of the platelets and plays a role in ADP-induced platelet aggregation. In this study we investigated 2 functional SNPs of PON1 gene (rs662 and rs854560) and 3 variants of the P2RY12 gene (rs2046934, rs6798347, rs6801273) in samples pooled from average Hungarian Roma and Hungarian population samples with PCR-RFLP method. For the PON1 variants we detected that the R allele frequency was significantly lower in the Roma group compared to the Hungarian population. (0.249 vs 0.318 p < 0.001). By contrast, the frequency of the M allele was significantly higher in Roma than in Hungarians (0.332 vs 0.290 p < 0.05). For the 3 P2RY12 variants we could find significant differencies only in rs2046934: the frequency of the CC genotype is 7 times higher in Hungarians than in Romas (1.4 vs 0.2 %, p < 0.05). The data presented here represent a unique genetic profile in Roma people that has not been reported for other populations.

Hodgkin disease therapy induced second malignancy susceptibility 6q21 functional variants in roma and hungarian population samples.

Patients treated successfully for pediatric Hodgkin's lymphoma are known to develop secondary malignancies; care is already taken in treatment to prevent this adverse effect. Recent GWAS study identified rs4946728 and rs1040411 noncoding SNPs located between PRDM1 and ATG1 genes on chromosome 6q21 as risk factors for secondary malignancies in patients formerly treated with radiotherapy for pediatric Hodgkin disease. We investigated the allele frequencies of these two SNPs in biobanked, randomly selected DNA of average, apparently healthy Hungarians (n = 277) and in samples of Roma (n = 279) population living Hungary. The risk allele frequency for rs4946728 was 79.4 % in Hungarian and 83.5 % in Roma samples, while for rs1040411 it was 56.4 % in Hungarian and 55.8 % in Roma samples. These values are quite similar in the two populations, and are rather high. The values are higher than those frequencies observed in the controls (rs4946728: 59.1 % and rs1040411: 39.6 %, p < 0.05), and are in the range of the cases (86 % and 68.2 %, respectively) of the above original GWAS study. Our findings suggest, that beside the already taken precautions, genetic characterization of Hungarian pediatric Hodgkin patients seems to be advantageous prior to the treatment of their disease.

Marked differences of haplotype tagging SNP distribution, linkage, and haplotype profile of IL23 receptor gene in Roma and Hungarian population samples.

Polymorphisms of the interleukin-23 receptor (IL23R) gene have been found to play an important role in the development of several autoimmune diseases. We examined five susceptible (rs10889677, rs1004819, rs2201841, rs11805303, rs11209032), one protective (rs7517847) and two neutral variants (rs7530511, rs1884444) of the IL23R gene in pooled DNA of healthy Roma (Gipsy) and Hungarian population samples. Our aim was to determine the genetic variability of the major haplotype tagging polymorphisms, and the haplotype profile of IL23R between the two groups. We analyzed 273 healthy Roma and 253 Hungarian DNA samples using PCR/RFLP assay. Comparing the five susceptible conferring alleles, there were significant increase (p<0.05), while in the protective alleles, there were decrease in the allele frequencies in Roma population (p<0.05). One of the neutral alleles showed increase, the another one did not differ between the two groups. The haplotype analysis of the SNPs revealed fundamentally different association types of SNPs in the two groups; moreover, the frequencies of the various haplotypes also exhibited strong differences, as of ht4 and ht5 haplotypes were significantly higher, whereas the frequencies of ht2 and ht3 haplotypes were significantly lower in the Roma population than in Hungarians (p<0.05). The data presented here show profound differences in the IL23R genetic profiles in the Roma population, that likely has also clinical implications in respect their possible role in the development of certain immunological diseases.

Differences in circulating carnitine status of preterm infants fed fortified human milk or preterm infant formula.

OBJECTIVE: The aim of the study was to compare plasma carnitine profiles in fortified human milk (HM)-fed preterm infants or formula-fed preterm infants. METHODS: Plasma acylcarnitine concentrations were determined in 20 formula-fed and 18 HM-fed preterm infants (birth weights between 1000 and 2200 g) by isotope dilution ESI MS/MS technique on study days 0, 14, and 28. RESULTS: Concentrations of free carnitine (FC) and different acylcarnitines did not change during the 4 weeks of the study in infants fed HM. In contrast, in infants fed formula FC increased markedly (day 0: 29.989 [16.646] µmol/L, median [interquartile range], day 14: 43.972 [8.455], P < 0.05) along with increases of short-chain esters (C2 day 0: 5.300 [3.272], day 14: 6.773 [2.127], P < 0.05; C3 day 0: 0.070 [0.059], day 14: 0.110 [0.069], P < 0.05). In contrast, some medium-chain (C8:1, C12) and long-chain esters (C14, C16) decreased significantly in infant formula by day 14, whereas FC and C2 and C3 esters increased further by day 28 (FC: 47.672 [14.753], C2: 7.430 [4.688], C3: 0.107 [0.047]). CONCLUSIONS: The altered carnitine ester profile likely reflects active involvement of the carnitine molecule in the buffering, metabolism, and elimination of nonphysiological acyl moieties.

High prevalence of CYP2C19*2 allele in Roma samples: study on Roma and Hungarian population samples with review of the literature.

The purpose of our study was to characterise the CYP2C19*2 and CYP2C19*3 alleles in healthy Roma and Hungarian populations. DNA of 500 Roma and 370 Hungarian subjects were genotyped for CYP2C19*2 (G681A, rs4244285) and CYP2C19*3 (G636A, rs4986893) by PCR-RFLP assay and direct sequencing. Significant differences were found comparing the Roma and Hungarian populations in CYP2C19 681 GG (63.6 vs. 75.9%), GA (31.8 vs. 23.0%), AA (4.6 vs. 1.1%), GA+AA (36.4 vs. 24.1%) and A allele frequencies (0.205 vs. 0.125) (p<0.004). Striking differences were found between Roma and Hungarian samples in CYP2C19*1 (79.5 vs. 87.4%) and CYP2C19*2 (20.5 vs. 12.6%) alleles, respectively (p<0.001). None of the subjects was found to carry the CYP2C19*3 allele. Frequencies of the intermedier metabolizer phenotype defined by the *1/*2 genotype (0.318 vs. 0.230, p<0.005) and poor metabolizer predicted by the *2/*2 genotype (0.046 vs. 0.011, p<0.005) was significantly higher in Roma than in Hungarians, respectively. Genotype distribution of the Roma population was similar to those of the population of North India, however, a major difference was found in the frequency of the CYP2C19*2 allele, which is likely a result of admixture with European lineages. In conclusion, the frequencies of the CYP2C19 alleles, genotypes and corresponding extensive, intermediate and poor metabolizer phenotypes studied here in the Hungarian population are similar to those of other European Caucasian populations, but display clear differences when compared to the Roma population.

Reconstructing Roma history from genome-wide data.

The Roma people, living throughout Europe and West Asia, are a diverse population linked by the Romani language and culture. Previous linguistic and genetic studies have suggested that the Roma migrated into Europe from South Asia about 1,000-1,500 years ago. Genetic inferences about Roma history have mostly focused on the Y chromosome and mitochondrial DNA. To explore what additional information can be learned from genome-wide data, we analyzed data from six Roma groups that we genotyped at hundreds of thousands of single nucleotide polymorphisms (SNPs). We estimate that the Roma harbor about 80% West Eurasian ancestry-derived from a combination of European and South Asian sources-and that the date of admixture of South Asian and European ancestry was about 850 years before present. We provide evidence for Eastern Europe being a major source of European ancestry, and North-west India being a major source of the South Asian ancestry in the Roma. By computing allele sharing as a measure of linkage disequilibrium, we estimate that the migration of Roma out of the Indian subcontinent was accompanied by a severe founder event, which appears to have been followed by a major demographic expansion after the arrival in Europe.

Non-syndromic Hearing Impairment in a Hungarian Family with the m.7510T>C Mutation of Mitochondrial tRNA(Ser(UCN)) and Review of Published Cases.

The m.7510T>C mitochondrial DNA (mtDNA) mutation is a tRNA(Ser(UCN)) alteration leading to matrilineal isolated hearing impairment. The current paper reviews the available reports on the m.7510T>C mtDNA mutation, with special attention to phenotypic variations and haplogroup background. A Hungarian family, the fourth family reported in the literature, is presented, in which analysis of three generations with bilateral isolated hearing loss revealed the m.7510T>C tRNA(Ser(UCN)) mutation in homoplasmic form in the affected members. Haplogroup analysis verified an unnamed subgroup of mitochondrial haplogroup H. Previously reported Spanish and North American Caucasian families belong to different subgroups of haplogroup H. Analyzing our biobank of Hungarian patients with sensorineural hearing loss, we did not detect this mutation in any other patient, nor was it found in Caucasian haplogroup H control samples. Comparing the cases reported so far, there is interfamilial variablity in the age of onset, accompanying symptoms, and haplogroup background. Our case adds further genetic evidence for the pathogenicity of the m.7510T>C mutation and underlines the need to include full mtDNA sequencing in the screening for unexplained hearing loss.

Marked diversity of IL23R gene haplotype variants in rheumatoid arthritis comparing with Crohn's disease and ankylosing spondylitis.

Haplotype tagging SNPs of interleukin-23 receptor gene rs1004819, rs7517847, rs7530511, rs2201841, rs1343151 and rs10889677 were determined in 396 patients with rheumatoid arthritis, 190 patients with Crohn's disease, 206 patients with ankylosing spondylitis and 182 controls. Using regression analysis models the rs1004819, rs2201841, and rs10889677 SNPs were found to confer risk for Crohn's disease and ankylosing spondylitis, while rs1343151 had a protective effect in both of these diseases, and the rs2201841 and rs10889677 SNPs showed susceptibility nature for rheumatoid arthritis. Using these SNPs we could study the susceptibility haplotype profiles in these diseases with special attention to the rheumatoid arthritis, first in the literature. Seven different haplotypes could be differentiated. We found that the SNPs exert their susceptibility character in specific haplotype blocks: thus, for rheumatoid arthritis the rs1343151 SNP was risk factor only in a specific haplotype surrounding; this can explain the controversial results published so far about this variant. More importantly, we observed, that while a specific haplotype can confer risk for rheumatoid arthritis, the same haplotype tended to protect against the development of the other two diseases. The data presented here serve evidence for the need of haplotype analysis instead of just single standing SNP analysis when susceptibility to or protection against a certain disease are interpreted.

Difference of interleukin-23 receptor gene haplotype variants in ulcerative colitis compared to Crohn's disease and psoriasis.

OBJECTIVE: Polymorphisms of the interleukin-23 receptor (IL23R) gene have been found to play a role in the development of several autoimmune diseases. Our aim was to examine the possible effect of not only simple individual variants, but of haplotypes composed of them. SUBJECTS: We analysed 263 patients with psoriasis, 199 patients with Crohn's disease (CD), 282 patients with ulcerative colitis (UC), and 253 controls for rs1884444, rs11805303, rs7517847, rs2201841, rs10889677 and rs11209032 variants. METHODS: The genotypes were determined by using PCR/RFLP assay. Logistic regression analysis was used to compare the genotype distribution of the polymorphisms and haplotypes between the examined autoimmune diseases and healthy controls. RESULTS: Rs1884444 was found to confer risk for UC and psoriasis, rs10889677 for CD and psoriasis, while rs2201841 and rs7517847 had effect only in CD. Using these SNPs we could study the susceptibility haplotype profiles in these diseases with special attention to UC. Eight different haplotypes could be differentiated. We found that the SNPs exert their susceptibility character in specific haplotype blocks, and the frequency of one haplotype differed significantly in UC compared with both other diseases and also with healthy controls. This haplotype conferred risk for UC, even while it had a somewhat lower frequency in the other diseases than in controls. CONCLUSIONS: The data presented here serve as evidence for the need of haplotype analysis instead of just single standing SNP analysis when susceptibility is interpreted.