{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,26]],"date-time":"2026-10-26T16:03:54Z","timestamp":1793030634304,"version":"4.5.0"},"reference-count":35,"publisher":"MDPI AG","issue":"8","license":[{"start":{"date-parts":[[2025,7,29]],"date-time":"2025-07-29T00:00:00Z","timestamp":1753747200000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"EEA Grant 2014\u20132021","award":["6\/2019"],"award-info":[{"award-number":["6\/2019"]}]},{"name":"EEA Grant 2014\u20132021","award":["2022\u20132027"],"award-info":[{"award-number":["2022\u20132027"]}]},{"name":"EEA Grant 2014\u20132021","award":["10N\/01.01.2023"],"award-info":[{"award-number":["10N\/01.01.2023"]}]},{"name":"EEA Grant 2014\u20132021","award":["PN 23.16.02.03"],"award-info":[{"award-number":["PN 23.16.02.03"]}]},{"name":"Core Program within the National Research, Development, and Innovation Plan","award":["6\/2019"],"award-info":[{"award-number":["6\/2019"]}]},{"name":"Core Program within the National Research, Development, and Innovation Plan","award":["2022\u20132027"],"award-info":[{"award-number":["2022\u20132027"]}]},{"name":"Core Program within the National Research, Development, and Innovation Plan","award":["10N\/01.01.2023"],"award-info":[{"award-number":["10N\/01.01.2023"]}]},{"name":"Core Program within the National Research, Development, and Innovation Plan","award":["PN 23.16.02.03"],"award-info":[{"award-number":["PN 23.16.02.03"]}]},{"name":"MCID","award":["6\/2019"],"award-info":[{"award-number":["6\/2019"]}]},{"name":"MCID","award":["2022\u20132027"],"award-info":[{"award-number":["2022\u20132027"]}]},{"name":"MCID","award":["10N\/01.01.2023"],"award-info":[{"award-number":["10N\/01.01.2023"]}]},{"name":"MCID","award":["PN 23.16.02.03"],"award-info":[{"award-number":["PN 23.16.02.03"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Genes"],"abstract":"<jats:p>Background\/Objectives: Autism spectrum disorders (ASDs) are neurodevelopmental conditions with early onset of clinical manifestations. ASD etiology is highly heterogeneous, with genetic factors being strong determinants of the behavioral problems and neurodevelopmental deficits. Fragile X syndrome (FXS) (OMIM #300624), caused by the transcriptional silencing of the FMR1 gene, represents the most common monogenic cause of autism. Our study included 226 boys with a diagnosis of ASD, for a systematic screening of genetic and epigenetic defects in the FMR1 gene promoter in a Romanian pediatric cohort. Methods: The methods, methylation-specific multiplex ligation-dependent probe amplification (MS-MLPA) and triplet-primed PCR (TP-PCR)\/melt curve analysis (MCA), were chosen for their ability to detect the methylation anomalies (the former) as well as repeat expansions in the FMR1 promoter (the latter). Results: Both methods used in our screening generated concordant results, detecting FMR1 full mutation in 4 out of 226 patients (~1.8%). This yield is similar to data obtained in larger studies. Three out of four boys presented the typical clinical features, in correlation with genetic findings. Conclusions: The combined use of MS-MLPA and TP-PCR\/MCA-based assay was, in our experience, useful to fully describe the genetic defects responsible for FXS. A significant variability of clinical presentations was observed in our small group of children with FXS, from mild to severe intellectual disability and from atypical to characteristic dysmorphic features, as well as various behavioral problems.<\/jats:p>","DOI":"10.3390\/genes16080903","type":"journal-article","created":{"date-parts":[[2025,7,29]],"date-time":"2025-07-29T08:00:41Z","timestamp":1753776041000},"page":"903","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":2,"title":["FMR1 Methylation Pattern and Repeat Expansion Screening in a Cohort of Boys with Autism Spectrum Disorders: Correlation of Genetic Findings with Clinical Presentations"],"prefix":"10.3390","volume":"16","author":[{"given":"Maria","family":"Dobre","sequence":"first","affiliation":[{"name":"Victor Babes National Institute of Pathology, 050096 Bucharest, Romania"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0009-0007-2582-3248","authenticated-orcid":false,"given":"Gisela","family":"Gaina","sequence":"additional","affiliation":[{"name":"Victor Babes National Institute of Pathology, 050096 Bucharest, Romania"},{"name":"Faculty of Medicine, Carol Davila University of Medicine and Pharmacy, 050474 Bucharest, Romania"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Alina","family":"Erbescu","sequence":"additional","affiliation":[{"name":"Victor Babes National Institute of Pathology, 050096 Bucharest, Romania"},{"name":"Doctoral School, Faculty of Biology, University of Bucharest, 030018 Bucharest, Romania"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Adelina","family":"Glangher","sequence":"additional","affiliation":[{"name":"Prof. Dr. Alexandru Obregia Clinical Hospital of Pathology, 041914 Bucharest, Romania"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-1697-9132","authenticated-orcid":false,"given":"Florentina Ionela","family":"Linca","sequence":"additional","affiliation":[{"name":"Prof. Dr. Alexandru Obregia Clinical Hospital of Pathology, 041914 Bucharest, Romania"},{"name":"Faculty of Psychology and Education-Sciences, University of Bucharest, 050663 Bucharest, Romania"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Doina","family":"Ioana","sequence":"additional","affiliation":[{"name":"Prof. Dr. Alexandru Obregia Clinical Hospital of Pathology, 041914 Bucharest, Romania"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-3901-615X","authenticated-orcid":false,"given":"Emilia Maria","family":"Severin","sequence":"additional","affiliation":[{"name":"Faculty of Medicine, Carol Davila University of Medicine and Pharmacy, 050474 Bucharest, Romania"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Florina","family":"Rad","sequence":"additional","affiliation":[{"name":"Faculty of Medicine, Carol Davila University of Medicine and Pharmacy, 050474 Bucharest, Romania"},{"name":"Prof. Dr. Alexandru Obregia Clinical Hospital of Pathology, 041914 Bucharest, Romania"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Mihaela Catrinel","family":"Iliescu","sequence":"additional","affiliation":[{"name":"Faculty of Medicine, Carol Davila University of Medicine and Pharmacy, 050474 Bucharest, Romania"},{"name":"Prof. Dr. Alexandru Obregia Clinical Hospital of Pathology, 041914 Bucharest, Romania"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-1352-3535","authenticated-orcid":false,"given":"Sorina Mihaela","family":"Papuc","sequence":"additional","affiliation":[{"name":"Victor Babes National Institute of Pathology, 050096 Bucharest, Romania"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-7740-9336","authenticated-orcid":false,"given":"Mihail Eugen","family":"Hinescu","sequence":"additional","affiliation":[{"name":"Victor Babes National Institute of Pathology, 050096 Bucharest, Romania"},{"name":"Faculty of Medicine, Carol Davila University of Medicine and Pharmacy, 050474 Bucharest, Romania"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-9315-5604","authenticated-orcid":false,"given":"Aurora","family":"Arghir","sequence":"additional","affiliation":[{"name":"Victor Babes National Institute of Pathology, 050096 Bucharest, Romania"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Magdalena","family":"Budi\u0219teanu","sequence":"additional","affiliation":[{"name":"Victor Babes National Institute of Pathology, 050096 Bucharest, Romania"},{"name":"Prof. Dr. Alexandru Obregia Clinical Hospital of Pathology, 041914 Bucharest, Romania"},{"name":"Faculty of Medicine, Titu Maiorescu University, 031593 Bucharest, Romania"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2025,7,29]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"1287","DOI":"10.1016\/j.cell.2019.07.037","article-title":"Getting to the Cores of Autism","volume":"178","author":"Yakoucheva","year":"2019","journal-title":"Cell"},{"key":"ref_2","doi-asserted-by":"crossref","unstructured":"Rylaarsdam, L., and Guemez-Gamboa, A. (2019). Genetic Causes and Modifiers of Autism Spectrum Disorder. Front. Cell. Neurosci., 13.","DOI":"10.3389\/fncel.2019.00385"},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"173","DOI":"10.1038\/s41431-020-00711-x","article-title":"Genetic modifiers in rare disorders: The case of fragile X syndrome","volume":"29","author":"Crawford","year":"2021","journal-title":"Eur. J. Hum. Genet."},{"key":"ref_4","doi-asserted-by":"crossref","unstructured":"American Psychiatric Association (2013). Diagnostic and Statistical Manual of Mental Disorders, American Psychiatric Association. [5th ed.].","DOI":"10.1176\/appi.books.9780890425596"},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"5","DOI":"10.1038\/s41572-019-0138-4","article-title":"Autism spectrum disorder","volume":"6","author":"Lord","year":"2020","journal-title":"Nat. Rev. Dis. Primers"},{"key":"ref_6","unstructured":"(2025, March 11). Online Mendelian Inherintance in Man. Available online: https:\/\/omim.org\/."},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"905","DOI":"10.1016\/0092-8674(91)90397-H","article-title":"Identification of a gene (FMR-1) containing a CGG repeat coincident with a breakpoint cluster region exhibiting length variation in fragile X syndrome","volume":"65","author":"Verkerk","year":"1991","journal-title":"Cell"},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"401","DOI":"10.1007\/s00415-011-6161-3","article-title":"Fragile X-associated disorders: A clinical overview","volume":"259","author":"Gallagher","year":"2012","journal-title":"J. Neurol."},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"584","DOI":"10.1097\/01.GIM.0000182468.22666.dd","article-title":"Fragile X syndrome: Diagnostic and carrier testing","volume":"7","author":"Sherman","year":"2005","journal-title":"Genet. Med."},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"17065","DOI":"10.1038\/nrdp.2017.65","article-title":"Fragile X syndrome","volume":"3","author":"Hagerman","year":"2017","journal-title":"Nat. Rev. Dis. Primers"},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"2063","DOI":"10.1002\/ajmg.a.31416","article-title":"Diagnostic yield of various genetic approaches in patients with unexplained developmental delay or mental retardation","volume":"140","author":"Rauch","year":"2006","journal-title":"Am. J. Med. Genet. A"},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"229","DOI":"10.1016\/j.gde.2012.03.002","article-title":"Genetic architecture in autism spectrum disorder","volume":"22","author":"Devlin","year":"2012","journal-title":"Curr. Opin. Genet. Dev."},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"454","DOI":"10.1136\/jmg.30.6.454","article-title":"Population studies of the fragile X: A molecular approach","volume":"30","author":"Jacobs","year":"1993","journal-title":"J. Med. Genet."},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"226","DOI":"10.1002\/ajmg.10427","article-title":"Prevalence of the fragile X syndrome in African-Americans","volume":"110","author":"Crawford","year":"2002","journal-title":"Am. J. Med. Genet."},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"503","DOI":"10.1016\/j.ajhg.2009.09.007","article-title":"Incidence of fragile X syndrome by newborn screening for methylated FMR1 DNA","volume":"85","author":"Coffee","year":"2009","journal-title":"Am. J. Hum. Genet."},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"5840","DOI":"10.1016\/j.cell.2023.11.019","article-title":"Spatially coordinated heterochromatinization of long synaptic genes in fragile X syndrome","volume":"186","author":"Malachowski","year":"2023","journal-title":"Cell"},{"key":"ref_17","doi-asserted-by":"crossref","unstructured":"Savatt, J.M., and Myers, S.M. (2021). Genetic Testing in Neurodevelopmental Disorders. Front. Pediatr., 9.","DOI":"10.3389\/fped.2021.526779"},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"121","DOI":"10.1111\/dmcn.13985","article-title":"Epigenetics of Fragile X Syndrome and Fragile X-related Disorders","volume":"61","author":"Kraan","year":"2019","journal-title":"Dev. Med. Child. Neurol."},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"1358","DOI":"10.1002\/ajmg.a.32261","article-title":"Mosaic FMR1 deletion causes fragile X syndrome and can lead to molecular misdiagnosis: A case report and review of the literature","volume":"146","author":"Coffee","year":"2008","journal-title":"Am. J. Med. Genet. A"},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"1185","DOI":"10.1038\/ejhg.2013.311","article-title":"Fragile X syndrome due to a missense mutation","volume":"22","author":"Myrick","year":"2014","journal-title":"Eur. J. Hum. Genet."},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"799","DOI":"10.1038\/s41436-021-01115-y","article-title":"Laboratory testing for fragile X, 2021 revision: A technical standard of the American College of Medical Genetics and Genomics (ACMG)","volume":"23","author":"Spector","year":"2021","journal-title":"Genet. Med."},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"365","DOI":"10.1146\/annurev-pharmtox-052120-090147","article-title":"Syndrome: Lessons Learned and What New Treatment Avenues Are on the Horizon","volume":"62","author":"Hagerman","year":"2022","journal-title":"Annu. Rev. Pharmacol. Toxicol."},{"key":"ref_23","unstructured":"World Health Organization (2019, November 20). ICD-10: International Statistical Classification of Diseases and Related Health Problems 10th Revision. Available online: https:\/\/icd.who.int\/."},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"e128","DOI":"10.1093\/nar\/gni127","article-title":"Methylation-specific MLPA (MS-MLPA): Simultaneous detection of CpG methylation and copy number changes of up to 40 sequences","volume":"33","author":"Nygren","year":"2005","journal-title":"Nucleic Acids Res."},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"496","DOI":"10.2353\/jmoldx.2008.080053","article-title":"Methylation-specific multiplex ligation-dependent probe amplification enables a rapid and reliable distinction between male FMR1 premutation and full-mutation alleles","volume":"10","author":"Nygren","year":"2008","journal-title":"J. Mol. Diagn."},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"719","DOI":"10.1016\/j.jmoldx.2016.05.002","article-title":"Defining the performance parameters of a rapid screening tool for FMR1 CGG-repeat expansions based on direct triplet-primed PCR and melt curve analysis","volume":"18","author":"Lian","year":"2016","journal-title":"J. Mol. Diagn."},{"key":"ref_27","doi-asserted-by":"crossref","unstructured":"Tan, V.J., Lian, M., Faradz, S.M.H., Winarni, T.I., and Chong, S.S. (2018). A Single Common Assay for Robust and Rapid Fragile X Mental Retardation Syndrome Screening From Dried Blood Spots. Front. Genet., 9.","DOI":"10.3389\/fgene.2018.00582"},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"1047","DOI":"10.1016\/0092-8674(91)90283-5","article-title":"Variation of the CGG repeat at the fragile X site results in genetic instability: Resolution of the Sherman paradox","volume":"67","author":"Fu","year":"1991","journal-title":"Cell"},{"key":"ref_29","first-page":"e35505","article-title":"Neurological, Psychiatric, and Multisystemic Involvement of Fragile X Syndrome Along With Its Pathophysiology, Methods of Screening, and Current Treatment Modalities","volume":"15","author":"Ranjan","year":"2023","journal-title":"Cureus"},{"key":"ref_30","doi-asserted-by":"crossref","unstructured":"Nobile, V., Pucci, C., Chiurazzi, P., Neri, G., and Tabolacci, E. (2021). DNA Methylation, Mechanisms of FMR1 Inactivation and Therapeutic Perspectives for Fragile X Syndrome. Biomolecules, 11.","DOI":"10.3390\/biom11020296"},{"key":"ref_31","doi-asserted-by":"crossref","unstructured":"Tabolacci, E., Nobile, V., Pucci, C., and Chiurazzi, P. (2022). Mechanisms of the FMR1 Repeat Instability: How Does the CGG Sequence Expand?. Int. J. Mol. Sci., 23.","DOI":"10.3390\/ijms23105425"},{"key":"ref_32","doi-asserted-by":"crossref","unstructured":"Usdin, K., and Kumari, D. (2015). Repeat-mediated epigenetic dysregulation of the FMR1 gene in the fragile X-related disorders. Front. Genet., 6.","DOI":"10.3389\/fgene.2015.00192"},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"407","DOI":"10.1016\/j.molcel.2014.03.023","article-title":"Fragile X mental retardation protein regulates translation by binding directly to the ribosome","volume":"54","author":"Chen","year":"2014","journal-title":"Mol. Cell"},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"417","DOI":"10.1016\/S0896-6273(03)00034-5","article-title":"RNA cargoes associating with FMRP reveal deficits in cellular functioning in Fmr1 null mice","volume":"37","author":"Miyashiro","year":"2003","journal-title":"Neuron"},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"1070","DOI":"10.1016\/j.neuron.2019.02.041","article-title":"A Synaptic Perspective of Fragile X Syndrome and Autism Spectrum Disorders","volume":"101","author":"Bagni","year":"2019","journal-title":"Neuron"}],"container-title":["Genes"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/2073-4425\/16\/8\/903\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,9]],"date-time":"2025-10-09T18:17:40Z","timestamp":1760033860000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/2073-4425\/16\/8\/903"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2025,7,29]]},"references-count":35,"journal-issue":{"issue":"8","published-online":{"date-parts":[[2025,8]]}},"alternative-id":["genes16080903"],"URL":"https:\/\/doi.org\/10.3390\/genes16080903","relation":{},"ISSN":["2073-4425"],"issn-type":[{"value":"2073-4425","type":"electronic"}],"subject":[],"published":{"date-parts":[[2025,7,29]]}}}