Silvia Guarducci

Fragile X Premutation Is a Significant Risk Factor for Premature Ovarian Failure: The International Collaborative POF in Fragile X Study—Preliminary Data

The preliminary results of an international collaborative study examining premature menopause in fragile X carriers are presented. A total of 760 women from fragile X families was surveyed about their fragile X carrier status and their menstrual and reproductive histories. Among the subjects, 395 carried a premutation, 128 carried a full mutation, and 237 were noncarriers. Sixty-three (16%) of the premutation carriers had experienced menopause prior to the age of 40 compared with none of the full mutation carriers and one (0.4%) of the controls. Based on these preliminary data, there is a significant association between fragile X premutation carrier status and premature menopause.

Premature ovarian failure (POF) and fragile X premutation females : From POF to fragile X carrier identification, from fragile X carrier diagnosis to POF association data

Early menopause in the fragile X carriers has been well documented in several reports. All surveys demonstrated that 13-25% of fragile X carriers experienced premature ovarian failure (POF), defined as menopause before the age of 40 years. In 1995 we started screening two groups of subjects as a part of a Fragile X Research Program: 1) women previously diagnosed as fragile X carriers from the register of our center and 2) women with POF and without a family history of fragile X or other forms of mental retardation. In this study we report the preliminary data collected from 75 fragile X families; in 30 of them, POF was present in one or several subjects, all of whom had a fragile X premutation. None of the women with a full mutation experienced POF in our series of patients. We also identified 89 families without a family history of fragile X or mental retardation, and there were 108 subjects who experienced POF, of which 6.5% had a fragile X premutation. This is 70-fold higher than the background prevalence of fragile X premutation in the Italian population and suggests an association with POF. These data confirm the results of other surveys.

Infrared fluorescent automated detection of thirteen short tandem repeat polymorphisms and one gender-determining system of the CODIS core system.

We used an infrared (IR) automated fluorescence monolaser sequencer for the analysis of 13 autosomal short tandem repeat (STR) systems (TPOX, D3S1358, FGA, CSF1PO, D5S818, D7S820, D8S1179, TH01, vWA, D13S317, D16S359, D18S51, D21S11) and the X‐Y homologous gene amelogenin system. These two systems represent the core of the combined DNA index systems (CODIS). Four independent multiplex reactions, based on the polymerase chain reaction (PCR) technique and on the direct labeling of the forward primer of every primer pair, with a new molecule (IRDyeTM800), were set up, permitting the exact characterization of the alleles by comparison with ladders of specific sequenced alleles. This is the first report of the whole analysis of the STRs of the CODIS core using an IR automated DNA sequencer. The protocol was used to solve paternity/maternity tests and for population studies. The electrophoretic system also proved useful for the correct typing of those loci differing in size by only 2 bp. A sensibility study demonstrated that the test can detect an average of 10 pg of undegraded human DNA. We also performed a preliminary study analyzing some forensic samples and mixed stains, which suggested the usefulness of using this analytical system for human identification as well as for forensic purposes.

Predictive factors associated with the acceptance of pandemic and seasonal influenza vaccination in health care workers and students in Tuscany, Central Italy

Assessing the beliefs and attitudes of Health Care Workers (HCW) to influenza and influenza vaccination can be useful in overcoming low compliance rates. The purpose of our study is to evaluate the opinion of HCW and students regarding influenza, influenza vaccine and the factors associated with vaccination compliance. A survey was conducted between October 2010 and April 2011 in the Florence metropolitan area. A questionnaire was administered to HCW in three local healthcare units and at Careggi University Teaching Hospital. Students matriculating in health degree programs at Florence University were also surveyed. The coverage with vaccination against seasonal and pandemic influenza is generally low, and it is lower in students than in HCW (12.5% vs 15% for the seasonal vaccination, 8.5% vs 18% for the pandemic vaccination). Individuals comply with vaccination offer mainly to protect themselves and their contacts. Individuals not receiving vaccination did not consider themselves at risk, had never been vaccinated before or believed that pandemic influenza was not a public health concern. Physicians had the highest compliance to vaccination and women were less frequently vaccinated than men. HCW do not appear to perceive their possible influenza infections as a risk for patients: HCW receive vaccination mainly as a form of personal protection. Low compliance to vaccination is determined by various factors and therefore requires a multi-faceted strategy of response. This should include short-term actions to overcome organizational barriers, in addition to long-term interventions to raise HCW’s level of knowledge about influenza and influenza vaccination.

Spectrum and distribution of MECP2 mutations in 64 Italian Rett syndrome girls: tentative genotype/phenotype correlation

We report a direct DNA sequencing analysis of the MECP2 gene undertaken on a further 64 Italian patients with Rett syndrome by using a LICOR 4200 Automated Sequencer. All of the girls entering the study had a consistent clinical diagnosis for this disorder. All coding regions and the flanking intronic splice site sequences were amplified as three non-overlapping fragments by using both forward and reverse primers. The results were then compared to the MECP2 reference sequences published in GenBank. Mutations of the MECP2 gene were identified in 64 of 75 (85.33%) unrelated sporadic Rett syndrome girls. Genotype/phenotype correlation studies, in particular in groups of patients with the same mutation, did not offer definitive and interesting data.

Analysis of 13 tetrameric short tandem repeat loci in a population of Tuscany (Central Italy) performed by means of an automated infrared sequencer

Allele frequencies for the 13 STRs of the Combined DNA Index System (CODIS) core were obtained from a sample of 188 unrelated individuals living in the area of Florence, Prato and Pistoia (Tuscany, Central Italy).

Chromosome Mapping of Miller-Diecker, Smith-Magenis and RARA Loci in Non-Human Primates: Implications in the Evolution of Human Chromosome 17

Molecular cytogenetics allows to verify chromosomal homologies previously hypothesised on the base of banding pattern comparison in different species. So far only the chromosome painting technique has been extensively used in studies of chromosomal evolution. This technique allows to detect only interchromosomal rearrangements. Human and Great Apes chromosomes basically differ by intrachromosomal rearrangements, in particular inversions; with chromosome painting it has just been possible to confirm the origin by fusion of human chromosome 2 and a reciprocal translocation in Gorilla, involving the homologous of chromosome 5 and 17. In order to verify intrachromosomal rearrangements in human chromosomal evolution, chromosome mapping of human loci in non-human primates is a useful approach. We mapped Miller-Diecker, Smith-Magenis and RARA loci localised on human chromosome 17, in Gorilla gorilla, Pongo pygmaeus, Macaca fascicularis and Cercopithecus aethiops. On the base of the obtained results it was possible to verify chromosomal rearrangements previously identified by banding, to achieve new informations about the controversial evolution of human chromosome 17, and to detect the occurrence of a paracentric inversion in the homologous in Cercopithecus aethiops.

Twin zygosity studies with the formula from DNA-View's Kinship Module after molecular analyses by polymorphic markers

Abstract Human Monozygotic (MZ) Twins estimated to occur once in 250 live births, result from an abnormal behavior by embryonic cell(s) to develop as separate embryos. The MZ twins are considered genetically identical and the reduced phenotypic, genetic or chromosomal concordance represents a very important challenge for geneticists. Determination of zygosity by molecular studies usefully provides a superior replacement for the conventional studies of placenta and chorionic membranes. We utilized a protocol based on the CODIS polymorphic markers battery, by using an infrared LI-COR 4200 instrument (LICOR, Nebraska, USA), and we associated the statistical analysis method with formulae derived by the DNA-Views Kinship Module to better evaluate the genetic identity in a group of MZ twins.

A new duplex PCR system for YCAII and DXYS156Y microsatellites analysis

Abstract The Y-chromosome polymorphisms represent an important tool in forensic genetics for different purposes: paternity testing, special problems of medical genetics, such as incest causing autosomal recessive diseases, analysis of mixed stains and other biological samples in criminal events. The number of Y-chromosome polymorphisms is huge, but only a well-defined number of them are used in official protocols of the international forensic genetics community. Recently, we presented a definitive haplotype of Y-STR in 107 unrelated individuals living in a Tuscany population (Central Italy) [Forensic Sci. Int. 120 (2001) 210]. To study this “minimal haplotype” we used the markers DYS19, DYS389I, DYS389II, DYS390, DYS391, DYS392, DYS385I/II [Forensic Sci. Int. 114 (2000) 31]. Subsequently, we introduced in our laboratory the use of an automated infrared DNA sequencer (LI-COR 4200). We applied this technology to study the hypervariable locus YCAII [Hum. Mol. Genet. 3 (1994) 115] for the “extended haplotype” and the polymorphism DXYS156 [Int. J. Legal Med. 110 (1997) 125], suggested by Kayser et al. [Hum. Biol. 70 (1998) 979]. We set up a new duplex PCR system for the simultaneous analysis of these loci with the automatic technology. This system was employed to obtain frequency data in the Tuscany population and to solve paternity dispute in which STR mutations were detected. A preliminary sensitivity study was also performed to evaluate the efficiency of this new duplex for stains analysis.

Analysis of mitochondrial DNA with an infrared automated DNA sequencer in a Tuscan population (Central Italy)

Abstract In our laboratory, we performed a study of the D-loop region of mtDNA in unrelated individuals, natives of Tuscany (Central Italy), using an Infrared Automated DNA Sequencer (LI-COR 4200). Taking advantage of the long read characteristics of the IR Automated Sequencer, we adopted a new strategy based on a single fragment amplification and sequencing, including the entire HVI and HVII regions. Preliminary results of the variability in the population we studied were also presented.