SOD Diagnostica Genetica

Firenze, Italy

SOD Diagnostica Genetica

Firenze, Italy
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Fattorini P.,University of Trieste | Previdere C.,University of Pavia | Carboni I.,University of Florence | Marrubini G.,University of Pavia | And 5 more authors.
Electrophoresis | Year: 2017

Next generation sequencing (NGS) is the emerging technology in forensic genomics laboratories. It offers higher resolution to address most problems of human identification, greater efficiency and potential ability to interrogate very challenging forensic casework samples. In this study, a trial set of DNA samples was artificially degraded by progressive aqueous hydrolysis, and analyzed together with the corresponding unmodified DNA sample and control sample 2800 M, to test the performance and reliability of the ForenSeqTM DNA Signature Prep kit using the MiSeq Sequencer (Illumina). The results of replicate tests performed on the unmodified sample (1.0 ng) and on scalar dilutions (1.0, 0.5 and 0.1 ng) of the reference sample 2800 M showed the robustness and the reliability of the NGS approach even from sub-optimal amounts of high quality DNA. The degraded samples showed a very limited number of reads/sample, from 2.9-10.2 folds lower than the ones reported for the less concentrated 2800 M DNA dilution (0.1 ng). In addition, it was impossible to assign up to 78.2% of the genotypes in the degraded samples as the software identified the corresponding loci as "low coverage" (< 50x). Amplification artifacts such as allelic imbalances, allele drop outs and a single allele drop in were also scored in the degraded samples. However, the ForenSeqTM DNA Sequencing kit, on the Illumina MiSeq, was able to generate data which led to the correct typing of 5.1-44.8% and 10.9-58.7% of 58 of the STRs and 92 SNPs, respectively. In all trial samples, the SNP markers showed higher chances to be typed correctly compared to the STRs. This NGS approach showed very promising results in terms of ability to recover genetic information from heavily degraded DNA samples for which the conventional PCR/CE approach gave no results. The frequency of genetic mistyping was very low, reaching the value of 1.4% for only one of the degraded samples. However, these results suggest that further validation studies and a definition of interpretation criteria for NGS data are needed before implementation of this technique in forensic genetics. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Mannelli F.,University of Florence | Mannelli F.,Instituto Toscano Tumori | Bencini S.,University of Florence | Bencini S.,Instituto Toscano Tumori | And 21 more authors.
Cytometry Part B - Clinical Cytometry | Year: 2013

Background. Paroxysmal nocturnal hemoglobinuria (PNH) is a unique disorder caused by a PIG-A gene mutation in a stem cell clone. Its clinical picture can sometimes make challenging the distinction from other disorders, and especially from myelodysplastic syndromes (MDS), since both diseases correlate with cytopenias and morphological abnormalities of bone marrow (BM) cells. Recently, flow cytometry (FC) has been proposed to integrate the morphologic assessment of BM dysplasia, and thus to improve the diagnostics of MDS. Methods. In the present study, we have analyzed systematically FC data resulting from the study of BM cells from patients with PNH and MDS. Results. Our data demonstrated abnormalities in PNH beyond the deficiency of glycosylphosphatidylinositol- linked proteins and the application of a systematic approach allowed us to separate effectively MDS and PNH in a cluster analysis and to highlight disease-specific abnormalities. Indeed, the parallel evaluation of some key parameters, i.e. patterns of expression of CD45 and CD10, provided information with practical diagnostic usefulness in the distinction between PNH and MDS. Moreover, the hypo-expression of CD36 that we observed on monocytes might be related to the thrombotic tendency in PNH. Conclusions. We investigated systematically the phenotypic profile of BM cells from patients with PNH; our data provide useful antigenic patterns to solve between PNH and MDS, sometimes morphologically overlapping. Moreover, some PNH-related phenotypic changes might be involved in the physiopathology of the disease and further studies addressing this issue are warranted. © 2013 International Clinical Cytometry Society.

Tomberli B.,Centro Of Riferimento Per Le Cardiomiopatie | Girolami F.,SOD Diagnostica Genetica | Coppini R.,University of Florence | Ferrantini C.,University of Florence | And 2 more authors.
Giornale Italiano di Cardiologia | Year: 2012

The management of patients with hypertrophic cardiomyopathy (HCM) and refractory symptoms due to massive hypertrophy and severe diastolic dysfunction represents a real challenge for the clinical cardiologist. Such patients often require novel therapeutic approaches, both invasive and pharmacological, involving multidisciplinary teamwork; however, the implementation of potentially viable treatment options is hindered by lack of disease-specific evidence. We report the case of a young woman with severe HCM and restrictive physiology, who underwent extensive myectomy via the transaortic and transapical approach, followed by biventricular pacing for cardiac resynchronization, with significant but incomplete symptomatic improvement. The subsequent introduction of ranolazine, based on promising preclinical data, has led to an excellent final result. An ongoing randomized clinical trial is currently testing the efficacy of ranolazine in symptomatic HCM. © 2012 Il Pensiero Scientifico Editore.

Biagini E.,University of Bologna | Olivotto I.,Centro Of Riferimento Per Le Cardiomiopatie | Iascone M.,USSD Laboratorio Genetica Medica | Parodi M.I.,Science Laboratorio Of Genetica Umana | And 17 more authors.
American Journal of Cardiology | Year: 2014

End-stage hypertrophic cardiomyopathy (ES-HC) has an ominous prognosis. Whether genotype can influence ES-HC occurrence is unresolved. We assessed the spectrum and clinical correlates of HC-associated mutations in a large multicenter cohort with end-stage ES-HC. Sequencing analysis of 8 sarcomere genes (MYH7, MYBPC3, TNNI3, TNNT2, TPM1, MYL2, MYL3, and ACTC1) and 2 metabolic genes (PRKAG2 and LAMP2) was performed in 156 ES-HC patients with left ventricular (LV) ejection fraction (EF) <50%. A comparison among mutated and negative ES-HC patients and a reference cohort of 181 HC patients with preserved LVEF was performed. Overall, 131 mutations (36 novel) were identified in 104 ES-HC patients (67%) predominantly affecting MYH7 and MYBPC3 (80%). Complex genotypes with double or triple mutations were present in 13% compared with 5% of the reference cohort (p = 0.013). The distribution of mutations was otherwise indistinguishable in the 2 groups. Among ES-HC patients, those presenting at first evaluation before the age of 20 had a 30% prevalence of complex genotypes compared with 19% and 21% in the subgroups aged 20 to 59 and ≥60 years (p = 0.003). MYBPC3 mutation carriers with ES-HC were older than patients with MYH7, other single mutations, or multiple mutations (median 41 vs 16, 26, and 28 years, p ≤0.001). Outcome of ES-HC patients was severe irrespective of genotype. In conclusion, the ES phase of HC is associated with a variable genetic substrate, not distinguishable from that of patients with HC and preserved EF, except for a higher frequency of complex genotypes with double or triple mutations of sarcomere genes. © 2014 Elsevier Inc. All rights reserved.

Previdere C.,University of Pavia | Grignani P.,University of Pavia | Alessandrini F.,Marche Polytechnic University | Alu M.,University of Modena and Reggio Emilia | And 17 more authors.
Forensic Science International: Genetics | Year: 2013

The 2011 collaborative exercise of the ISFG Italian Working Group GeFI was aimed at validating the five ENFSI/EDNAP miniSTR loci D1S1656, D2S441, D10S1248, D12S391 and D22S1045. The protocol required to type at least 50 multilocus profiles from locally resident individuals and two blind bloodstains in duplicate (i.e., using at least two different commercial kits), and to send the electropherograms to the Organizing Committee. Nineteen laboratories distributed across Italy participated, collecting a total of 960 samples. Full concordance was found for the five new miniSTRs as observed from the comparison of 13,150 alleles. The inspection of the electropherograms allowed the identification of a very limited number of mistypings in the miniSTR genotypes thus contributing to the establishment of an high quality Italian database of frequencies. © 2012 Elsevier Ireland Ltd.

Inturri S.,University of Turin | Robino C.,University of Turin | Carboni I.,SOD Diagnostica Genetica | Ricci U.,SOD Diagnostica Genetica | Gino S.,University of Turin
Forensic Science International: Genetics Supplement Series | Year: 2011

We report the case of four women and a man, all born in an Italian village during and immediately after WWII, that recently contacted our laboratory in order to perform kinship analysis. According to their claim, the propositi were the illegitimate offspring of a country gentleman and a peasant woman, given in adoption immediately after birth. A story that curiously reminded us of Jean Jacques Rousseau, Thérèse Levasseur and their five children. Problems connected with DNA analysis in cases where all stated relationship are questioned, and a wide range of different pedigrees could be used as hypotheses in LR calculations are discussed. © 2011 Elsevier Ireland Ltd.

Marseglia G.,SOD Diagnostica genetica | Scordo M.R.,University of Florence | Pescucci C.,SOD Diagnostica genetica | Nannetti G.,SOD Diagnostica genetica | And 6 more authors.
European Journal of Medical Genetics | Year: 2012

Several cases of interstitial deletion encompassing band 18q12.3 are described in patients with mild dysmorphic features, mental retardation and impairment of expressive language. The critical deleted region contains SETBP1 gene (SET binding protein 1). Missense heterozygous mutations in this gene cause Schinzel-Giedion syndrome (SGS, MIM#269150), characterized by profound mental retardation and multiple congenital malformations. Recently, a 18q12.3 microdeletion causing SETBP1 haploinsufficiency has been described in two patients that show expressive speech impairment, moderate developmental delay and peculiar facial features. The phenotype of individual with partial chromosome 18q deletions does not resemble SGS. The deletion defines a critical region in which SETBP1 is the major candidate gene for expressive speech defect. We describe an additional patient with the smallest 18q12.3 microdeletion never reported that causes the disruption of SETBP1. The patient shows mild mental retardation and expressive speech impairment with striking discrepancy between expressive and receptive language skills. He is able to communicate using gestures and mimic expression of face and body with surprising efficacy. The significant phenotypic overlap between this patient and the cases previously reported enforce the hypothesis that SETBP1 haploinsufficiency may have a role in expressive language development. © 2012 Elsevier Masson SAS.

Gerundino F.,SOD Diagnostica Genetica | Marseglia G.,SOD Diagnostica Genetica | Pescucci C.,SOD Diagnostica Genetica | Pelo E.,SOD Diagnostica Genetica | And 5 more authors.
European Journal of Medical Genetics | Year: 2014

We describe a patient with speech impairment, global developmental delay, behavioural problems and a 186kb de novo microdeletion on 16p11.2. There are four OMIM Phenotypes entries partially overlapping with the deleted region and related to recurrent microdeletions/microduplications in 16p11.2. A detailed review of published data shows that microdeletions/microduplications boundaries' do not include genes that are deleted in the case here reported. The deletion encompasses 9 RefSeq genes and includes SRCAP (Snf2-related CREBBP activator protein, OMIM*611421), a disease causing gene. Recently, truncating mutations in the SRCAP gene have been shown to cause Floating-Harbor syndrome (FHS, OMIM#136140), a rare disorder characterized by peculiar facial features, short stature with delayed osseous maturation and speech impairment. The patient reported here shows few subtle phenotypic features resembling that of FHS, but she does not have sufficient signs and symptoms for the clinical diagnosis and a clinical classification based on facial gestalt is not possible. This is the first report of a 16p11.2 deletion completely removing one copy of SRCAP, suggesting that haploinsufficiency of this gene could be associated to speech impairment, global developmental delay, behavioural problems and few subtle phenotypic features resembling FHS. However, further evidence for the putative causative role of SRCAP isolated deletion is needed. © 2014 Elsevier Masson SAS.

PubMed | University of Pavia, University of Trieste, SOD Diagnostica Genetica and International School for Advanced Studies
Type: | Journal: Electrophoresis | Year: 2017

Next generation sequencing (NGS) is the emerging technology in forensic genomics laboratories. It offers higher resolution to address most problems of human identification, greater efficiency and potential ability to interrogate very challenging forensic casework samples. In the present study, a trial set of DNA samples was artificially degraded, by progressive aqueous hydrolysis, and analyzed together with the corresponding unmodified DNA sample and control sample 2800M, to test the performance and reliability of the ForenSeq

PubMed | SOD Laboratorio Centrale, University of Florence and SOD Diagnostica Genetica
Type: Journal Article | Journal: Experimental hematology | Year: 2015

The presence of multilineage dysplasia (MLD) by morphology at diagnosis in acute myeloid leukemia (AML) defines a separate subset in the World Health Organization classification with still-debated prognostic value. A major controversy concerns MLDs role in NPM1-mutated (NPM1) AML, which correlates with good prognosis. We used flow cytometry (FC), an emerging technique for assessing dysplasia, to investigate MLD in NPM1 AML by an immunophenotypic score (IPS), a technique previously adopted in myelodysplastic syndrome. Eighty-five intensively treated NPM1 AML cases were studied. Patients were grouped according to the combination of data in maturing cell compartments. FC-assessed dysplasia showed a significant correlation with morphology-assessed dysplasia, showing the efficacy of this method in highlighting dysplasia in AML. Except for MLD, IPS did not influence any patient- or disease-related characteristics at diagnosis. Furthermore, IPS did not influence complete remission rate, disease-free survival, or overall survival. By investigating NPM1 status on separated cell compartments, we established a correlation between FC-assessed MLD and belonging to AML clone. This study shows that dysplasia evaluated by immunophenotype has no impact on clinical-biological characteristics or on outcome of NPM1 AML. Dysplasia is part of the spectrum of NPM1 AML, and the prognostic stratification of this category of patients should not be based upon it.

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