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The X-chromosome (X-Chr) markers are more efficient than autosomes in special cases of kinship analysis. Here, we show an unusual case, where two half-sisters (AR, AP), daughters of the same woman (AL), wanted to know whether they were also half-sisters of two other women (MM, MS) and a man (CR). These latter subjects were known to be children of different fathers. DNA was also available from AR’s and AP’s maternal aunt AA. Using the undisputed relationships (AA being aunt of the half-sisters AR and AP), the X-haplotypes of AR’s and AP’s maternal grandparents could be inferred. Grandmother’s X-haplotypes were also shared by MM, MS and CR, thus supporting hypothesis 1. A Mendelian inconsistency was observed at locus DXS10134, though it could not be distinguished between mutation and intra cluster recombination. Extra cluster meiotic recombination events were also observed, as some other interesting outcomes useful for forensic genetics discussion. According to ISFG guidelines [ 1 , 2 ] the biostatistical evaluation in kinship analysis was based on a likelihood ratio approach, using FamiLinkX and Familias programs respectively on the X-chromosome and autosomes analysis. [ [3] , [4] , [5] ]

Every year thousands of migrants die during the endeavour to reach the Italian coasts, making the Mediterranean the theatre of one of the greatest tragedies of mankind. Over 60% of these victims is buried unidentified: one of the reasons behind this is related to the specific difficulties and lack of strategies concerning AM and PM data collection. The present article describes how Italy is trying to face the problem of migrant identification, thanks to the collaboration between government, the Italian national police and universities. In particular, this is the first pilot study carried out to identify the victims of the second greatest tragedy of its kind off the Italian coast, near Lampedusa, on October 3rd 2013, which caused 366 victims. The present article shows the strategies conceived to collect postmortem and especially antemortem data and to compare them to identify matches, using medicolegal, anthropological, odontological and genetic approaches. Thirty-one victims out of 53 missing sought by relatives were identified (58.5%). The type and the quality of antemortem data available, generally photos and videos, pinpoints the importance of the face and the body for identification when the bodies are well preserved and how DNA analyses may at times present difficulties. In fact, critical points emerged concerning especially the lack of genetic information of the populations to which the victims belonged, the number of genetic markers needed to reach a statistical support for the identification and the need to adopt lineage markers such as mitochondrial DNA and Y-chromosome polymorphisms to identify parental relationships. This pilot study however has proven that families continue to seek their relatives and that it is possible, as well as mandatory, to identify migrant victims in spite of the difficulties in the collection of antemortem and postmortem data. In addition, considering the peculiar scenario, novel strategies for positive identification have to be defined in each field (anthropological, odontological and genetic) as well as in combination.

On October 3rd, 2013 a boat carrying more than 500 migrants coming mostly from the Horn of Africa (Eritrea, Somalia, Ethiopia) sank near Lampedusa, a small Italian Island in the middle of the Mediterranean Sea. The recovered bodies were examined by a forensic team, and post mortem data (anthropological and odontological records, and DNA) were collected for identification. Genetic profiles based on 16 autosomal STRs were acquired from both victims and putative relatives recruited following an international call. The final genetic database included 363 victims and 43 reference persons from 36 independent families recruited until mid-2017, who were missing 35 first-degree and 6 second-degree relatives. A pairwise blind search approach was used to identify familial relationships within the victims and between victims and putative relatives. Two statistics were calculated, the Identity by State (IBS) and the Identity by Descent (IBD), the latter by using the DVI module of the FAMILIAS3 software to compute LRs and posterior probabilities. The putative identifications were confirmed in pedigree analysis using the information provided by the relatives. In selected cases, additional autosomal and lineage (Y-chromosome and mtDNA) markers were typed. Some critical points were highlighted: the lack for accurate allele and haplotype frequencies in African populations, especially for the lineage markers, and the need for a shared approach to the biostatistical interpretation of the results in DVI. In the end, 29 first-degree (parent-child and full sibs) out of 35 missing (83%), and 3 out of 6 of second-degree relatives (50%) showed a high statistical confidence for a positive identification. This study represents the first attempt to systematically deal with the genetic identification of African migrants who died in the Mediterranean Sea. The methodological and statistical approach used in this study was proved to be reliable and appropriate for future genetic identifications in other similar mass disasters.

We report here the identification of a gene associated with the hyperparathyroidism-jaw tumor (HPT-JT) syndrome. A single locus associated with HPT-JT (HRPT2) was previously mapped to chromosomal region 1q25-q32. We refined this region to a critical interval of 12 cM by genotyping in 26 affected kindreds. Using a positional candidate approach, we identified thirteen different heterozygous, germline, inactivating mutations in a single gene in fourteen families with HPT-JT. The proposed role of HRPT2 as a tumor suppressor was supported by mutation screening in 48 parathyroid adenomas with cystic features, which identified three somatic inactivating mutations, all located in exon 1. None of these mutations were detected in normal controls, and all were predicted to cause deficient or impaired protein function. HRPT2 is a ubiquitously expressed, evolutionarily conserved gene encoding a predicted protein of 531 amino acids, for which we propose the name parafibromin. Our findings suggest that HRPT2 is a tumor-suppressor gene, the inactivation of which is directly involved in predisposition to HPT-JT and in development of some sporadic parathyroid tumors.

Background PancPRO is a computer program that estimates the risk of pancreatic cancer for asymptomatic individuals based on a genetic model of susceptibility and the familial incidence of cancer. Aim To evaluate the distribution of the familial risk in a series of incident cases of pancreatic adenocarcinoma. Materials and methods The lifetime risk of pancreatic cancer was calculated by PancPro for a hypothetical 40-year-old son of 570 consecutive probands with pancreatic cancer. Results The 570 risk values were included between 1% and 13%. The distribution was bimodal, with the antimode located at risk = 7.5%. Considering a 10-fold risk over the general population as a threshold for including a subject in a surveillance program, 19 families (3.3%) would be selected, totalling 92 first-degree relatives with age >40 years. Conclusions PancPro is a valid instrument to rank families based on risk of pancreatic cancer.

Estimating the age of founder mutations may contribute to improve our knowledge of population genetics and evolutionary history of diseases. Previous haplotype analysis suggested that the BRCA1*1499insA mutation was a founder allele, probably originated in Tuscany (Italy). Here, we collected additional pedigrees carrying this mutation, and applied a phylogenetic method for estimating mutation age. A chromosome segment of about 25 cM, including 37 short tandem repeats (STRs) on both sides of the BRCA1 gene (DeCode map), was typed in 50 subjects (28 mutation carriers) from 14 unrelated families. The time to the most recent common ancestor (MRCA) of the mutation carriers was estimated by the length of the shared haplotype between all possible pairs of individuals. A function relating the length of the shared haplotype to the time to the MRCA was obtained by a computer simulation. This approach gives results comparable with those of other existing mutation-dating methods, but does not depend explicitly on population-specific parameters such as allele frequencies, provides narrower confidence intervals (CI), and allows one to build an extended genealogical tree of all mutation carriers. The 1499insA mutation shared by the investigated subjects was estimated to be present in an individual living about 30 generations ago (95% CL 22-56), or 750 years (95% CL 550-1,400).

Mutation-predicting models can be useful when deciding on the genetic testing of individuals at risk and in determining the cost effectiveness of screening strategies at the population level. The aim of this study was to evaluate the performance of a newly developed genetic model that incorporates tumor microsatellite instability (MSI) information, called the AIFEG model, and in predicting the presence of mutations in MSH2 and MLH1 in probands with suspected hereditary non-polyposis colorectal cancer. The AIFEG model is based on published estimates of mutation frequencies and cancer penetrances in carriers and non-carriers and employs the program MLINK of the FASTLINK package to calculate the proband's carrier probability. Model performance is evaluated in a series of 219 families screened for mutations in both MSH2 and MLH1, in which 68 disease-causing mutations were identified. Predictions are first obtained using family history only and then converted into posterior probabilities using information on MSI. This improves predictions substantially. Using a probability threshold of 10% for mutation analysis, the AIFEG model applied to our series has 100% sensitivity and 71% specificity.

Many X-chromosome short tandem repeats (X-STRs) have been validated for forensic use even if further studies are needed on allele frequencies and mutation rates to evaluate the extent of polymorphism in different populations and to establish reference databases useful for forensic applications and for anthropological studies. A single multiplex reaction of seven X-STRs, which includes the DXS6789, HUMARA, DXS10011, DXS7423, HPRTB, DXS6807, DXS101 loci, is presented and their allele frequency distribution in a large population sample including 556 subjects (268 females and 288 males) analysed by five forensic laboratories of Central and Northern Italy is shown. Our results demonstrate the feasibility of a single amplification/detection reaction involving seven markers of the X chromosome, which can be fruitfully used in complex kinship analysis.

Mutations in BRCA1 and BRCA2 show different expressivity with respect to cancer risk, and allelic heterogeneity may be present in both genes. We collected 179 pedigrees with identified germline mutation (104 BRCA1 and 75 BRCA2), ascertained in six collaborating centers of the Italian Consortium for Hereditary Breast and Ovarian Cancer. Significant heterogeneity was detected for several variables, and a logistic regression model including age of diagnosis in the proband, presence of ovarian cancer in the family, presence of prostate or pancreatic cancer in the family, and presence of male breast cancer in the family proved to be effective in predicting the presence of a mutation in a gene rather than the other. Excess of familial aggregation of both breast and ovarian cancer was observed in both genes. Proportion of ovarian cancer was increased in the 5' portion of BRCA1, and presence of prostate or pancreatic cancer in a family was correlated with presence of ovarian cancer in BRCA2.

BARD1 (BRCA1-associated RING domain) was identified by yeast two-hybrid screening as a protein interacting with BRCA1. Somatic and germline mutations of BARD1 have been detected in sporadic breast, ovarian, and endometrial cancers. The present study represents the first description of BARD1 germline mutations in hereditary breast and breast/ovarian cancer patients. We analyzed the BARD1 gene in 40 families with hereditary breast and breast/ovarian cancer, tested negative for BRCA1 and BRCA2 mutations. A mutational analysis by PCR-SSCP on the coding region and the exon-intron splice boundaries of the BARD1 gene yielded four different germline mutations. A group of 20 patients diagnosed with sporadic breast cancer below the age of 40 was also examined and only one germline mutation was found. A study of loss of heterozygosity at the BARD1 locus in neoplastic tissues from patients with BARD1 germline mutations was carried out. In all cases, we were unable to find any evidence for allelic deletions. The involvement of BARD1 mutations in the susceptibility to hereditary breast and breast/ovarian cancer is discussed.

Summary A novel HLA-A allele, HLA-A*68:105, was detected by sequence-based typing (SBT) in an Italian bone marrow donor. It differs from HLA-A*68:01:02 at five nucleotides, three intronic, nt 699 T->G (intron 2), nt 705 T->C (intron 2) and nt 2770 G->A (intron 7), and two located in exon 3, at positions 726 A-G (codon 94 Ile->Val) and 733 T-G (codon 97 Arg->Met), respectively.

TT virus (TTV) is a recently identified widespread DNA virus of humans that produces persistent viremia in the absence of overt clinical manifestations. In an attempt to shed light on the dynamics of chronic infection, we measured the levels of TTV in the plasma of 25 persistently infected patients during the first 3 months of alpha interferon (IFN-α) treatment for concomitant hepatitis C virus (HCV) infection. The first significant decline of TTV loads was observed at day 3 versus day 1 for HCV. Subsequently, the loads of TTV became progressively lower in most patients, but some initial responders relapsed before the end of the follow-up, suggesting that at least in some subjects the effects of IFN on TTV can be very short-lived. No correlation between the responses of TTV and HCV to therapy was found. Fitting the viremia data obtained during the first week of treatment into previously developed mathematical models showed that TTV sustains very active chronic infections, with over 90% of the virions in plasma cleared and replenished daily and a minimum of approximately 3.8 × 10 10 virions generated per day. Low levels of TTV were occasionally detected in the peripheral blood mononuclear cells of patients who had cleared plasma viremia, thus corroborating previous results showing that these cells may support TTV replication and/or persistence.

OBJECTIVE: Evaluation of school attainments in children with congenital hypothyroidism (CH) detected by neonatal screening and treated early in life. PATIENTS AND METHODS: Text comprehension, mathematics, reading, writing and verbal and spatial memory, as indices of school learning, were evaluated in nineteen 5- to 10-year-old children with CH attending nursery or elementary school. l-Thyroxine substitution (starting dose 8-10 microg/kg body weight per day) was initiated at a mean age of 30+/-10 days of life. The control group included 298 unaffected children matched with the CH children for age and school grade. Thirty per cent of controls were classmates of CH children. Intelligence quotients (IQ), language performances and motor development were evaluated in CH children at age 5 years, and were related to their school attainments. School performances of CH children were also compared with their neonatal serum thyroxine (T4) concentration, and with the social-cultural level of the family. RESULTS: Four out of 19 (21%) children with CH, 3 in the nursery and 1 in the elementary school, displayed a generalized learning disorder. Symbol copy, geometric copy, phrase repetition, dictation writing and spontaneous writing were particularly defective in nursery school CH children, while orthographic error recognition was defective in elementary school CH children. School learning disorders in CH children were significantly correlated with a borderline-low IQ, poor language performances and a low social-cultural level of the family, but not with motor skills or neonatal T4 concentration. CONCLUSION: School attainments of early treated CH children were within the normal range in most affected cases. However, about 20% of CH children, most of them attending nursery school, showed a generalized learning disorder. Low IQ scores and poor language performances at age 5 years were associated with defective learning, mainly in CH children living in a poor social-cultural environment. In this subset of CH children, prompt initiation of speech and psychomotor rehabilitation therapy is recommended in order to prevent subsequent school learning disorders.

Higher brain dopamine content depending on lower activity of Catechol-O-Methyltransferase (COMT) in subjects with high hypnotisability scores (highs) has been considered responsible for their attentional characteristics. However, the results of the previous genetic studies on association between hypnotisability and the Catechol-O-Methyltransferase (COMT) single nucleotide polymorphism (SNP) rs4680 (Val158Met) were inconsistent. Here, we used a selective genotyping approach to re-evaluate the association between hypnotisability and COMT in the context of a two-SNP haplotype analysis, considering not only the Val158Met polymorphism, but also the closely located rs4818 SNP. An Italian sample of 53 highs, 49 low hypnotizable subjects (lows) and 57 controls, were genotyped for a segment of 805 bp of the COMT gene, including Val158Met and the closely located rs4818 SNP. Our selective genotyping approach had 97.1% power to detect the previously reported strongest association at the significance level of 5%. We found no evidence of association at the SNP, haplotype and diplotype levels. Thus, our results challenge the dopamine-based theory of hypnosis and indirectly support recent neuropsychological and neurophysiological findings reporting the lack of any association between hypnotisability and focused attention abilities.

The present-day Sherpa are thought to descend from a small number of ancestors that set- tled in Nepal several centuries ago, coming from the Eastern Tibetan region of Kham. A generally ac- cepted ethnographic theory involves the out-of-Kham migration of four proto-clans between the 15th and 16th centuries. Traditional Sherpa society is still divided into clans, called ru, which are patrilin- early transmitted. Ru therefore roughly correspond to the surnames of Western societies: males of the same ru are expected to share identical Y-chromosome haplotypes. However, multiple origins of ru and/or frequent gene flow of male lineages from neighbouring populations can complicate the genea- logical structure. In the present work, 25 male Sherpas of the Solukhumbu district were typed for the 17 Y-chromosomal short tandem repeats included in the AmpFlSTR ® Yfiler™ kit. Seventeen different haplotypes were found; 12 were unique. A phylogenetic tree was then drawn from the pairwise muta- tional distance matrix with a neighbour-joining algorithm. Branching reliability was also assessed through bootstrap analysis. Two macro-clusters of haplotypes were found, ascribable on the whole to two out of four of the presumed Tibetan proto-ru, the Thimmi and the Minyagpa. However, the Min- yagpa macro-cluster was found to be bipartite in terms of haplogroups, being composed by two distinct haplotype clusters. Clustering of the contributors by birthplace was also performed, suggesting a dif- ferential ru spatial distribution between upper Khumbu and lower Solu. Khumbu seems predominantly populated by newer clans and putative descendants of the Thimmi proto-ru, whereas Solu is mostly inhabited by members of the Minyagpa proto-ru.

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.

Adenomatous polyposis coli (APC) is an autosomal dominant disease characterized by the development of hundreds of colorectal adenomatous polyps during the first decades of life. The expression of the disease varies, as the age of onset of colonic cancer and the severity of extracolonic manifestations often differ between affected families. An attenuated form of APC has also been described in which a small number of polyps and a later age of onset of colonic cancer is observed. Cloning of the APC gene has allowed disease-causing mutations in APC families to be identified. Here, we report a novel splice site mutation (a G to T transversion at position +5 of the splice donor site in intron 9) in the APC gene of affected individuals in an Italian family. Characterization of the transcription products from this mutant APC allele revealed that normal splicing was disrupted: a shorter mRNA was expressed in which exon 8 was connected directly to exon 10. This created a shift in the reading frame and the introduction of a stop codon at position 1358. In addition, some normal APC transcript was produced from the mutant allele in lymphoblastoid cells. A comparison of the clinical features of affected members of this family with four unrelated Italian APC kindreds, in which the same AAAAG deletion at position 3926 has been found, showed a significant difference in the onset of disease symptoms and in the age of death attributable to colorectal cancer. Inefficient exon skipping may be, at least in part, responsible for the delay in the development of the disease in the reported family.

The Bracco Italiano is one of the oldest breeds of Italian pointing dogs, used for hunting ever since Renaissance times. After the Second World War it was included among the breeds officially recorded by the ENCI (the Italian Cynological Club), and since 1970 more than 23,000 animals have been registered; there are currently approximately 750 births per year. In this paper, we present the breed characterization of the population at the molecular level using 21 STR markers from the panels recommended for the 2006, 2008 and 2010 ISAG canine comparison test. Number of alleles, allele frequencies, deviations from Hardy-Weinberg proportions, linkage disequilibrium among loci, genetic similarity, genetic distances and molecular co-ancestry-based parameters were calculated. The number of alleles ranged from 3 to 9 (mean 6.43) whereas the expected heterozygosity ranged from 0.44 to 0.81 (mean 0.64). There was a high genetic similarity within the whole population (0.455) showing the great homogeneity of the sampled animals, as confirmed also by the small kinship distance (0.336), by the high values of the self molecular coancestry (0.703) and of the inbreeding coefficient (0.406). These results suggest the need for a careful genetic management of the population in order to avoid the risk of an excessive increase in the inbreeding level which would result in significant inbreeding depression and in significant loss of genetic variation.

We collected published and unpublished data from 17 contributing groups participating in the GeFI (Italian Forensic Geneticists). The total number of typed subjects was 1114 males and 777 females, coming from 11 regions of North, Centre, and South Italy, and Sardinia. Individual's multilocus genotypes included 4–12 loci. The total number of typed markers was 29, scattered along the X-chromosome genetic map in several clusters; the most used marker was DXS7423 (2429 gene copies); the mean number of subjects typed per marker was 336 for males and 208 for females. Data are available online.

The dramatic size contraction of local cattle breeds due to replacement with cosmopolite improved breeds highlights the need for native genetic resources conservation. In 1985, the Anagraphic Register of local cattle breeds and small-size ethnic groups was established by the Italian Ministry of Agriculture and Forestry. Calvana, Pontremolese and Garfagnina are among the included breeds. They are all native from Tuscany. Present breeding area covers the provinces of Firenze, Prato, Pistoia and Siena for the Calvana breed (around 280 heads), while it is restricted to the province of Lucca for both Garfagnina (around 180 heads) and Pontremolese. This latter breed consists, nowadays, of less than 40 heads, while being around 15000 in 1940s.The characterization of the genetic structure and variability via molecular markers could provide useful information for breed management and conservation. In the present study, a total of 149 animals, evenly distributed among the three breeds, were genetically char...

Within an Italian collaborative exercise on the extended haplotype of the Y-chromosome, 1288 subjects were typed by the AmpFlSTR YFiler Amplification Kit (AB Applied Biosystems) and other 526 were typed by the PowerPlex Y ® System (Promega). The sampling scheme included either a "regional" or a "local" recruitment, the first referring to individuals born in the region of the participating lab, the second referring to individuals coming from small villages. Total sample sizes were N =954 and 860, respectively. A significant decrease of haplotype diversity was found in the local samples. The results may be of interest in forensic applications of the Y-chromosome.

The Anagraphic Register of the Amiata donkey breed has been established in 1993, and is maintained by the association of breeders of the Grosseto province (APA-GR). Being an endangered breed, subjects with unknown pedigree have been allowed to be recorded, provided that females scored at least 60 and males at least 75 points on morphological evaluation. Recording is still open for females, whereas it has been closed in 1997 for stallions. In the present work, we have investigated the morphological structure of the Amiata donkey through the data reported in the Anagraphic Register.This last was started recently and was maintained on paper only, meaning that it inevitably includes a certain degree of incompleteness. The morphological records of 74 unrelated Amiata donkeys (62 females and 12 males) were collected at the APA-GR in 2005. For each animal the following biometrical data was available: 10 individual scores (from 1 to 10) for coat colour, head shape, neck, withers, shoulders, back, back-lum...

The current study presents the results of a preliminary haplotype association analysis at the bovine PRKAG3 locus with meat quality traits in the Chianina breed. No significant association was shown between haploid haplotypes (or diplotypes) and phenotypical traits after applying a Bonferroni correction for multiple comparison. Nonetheless, data from Longissimus dorsi muscle suggest the presence of a statistically non-significant trend toward an influence of the PRKAG3 haploid haplotypes on meat colour (a*) and water holding capacity (M/T) traits, as confirmed also by diplotype-based association analysis. A less clear set of results was observed for the Triceps brachii and Semitendinosus muscles.

Results from a collaborative exercise with proficiency testing conducted by 20 Italian laboratories on the 15 loci included in the Identifiler ® kit were analyzed by allele sharing methods and by standard population genetics tests. The validated database, including about 1500 subjects, was merged with that of a previous exercise conducted on nine loci, and the resulting allele frequencies, subdivided by Italian region, were published on-line.

In 239 torquetenovirus-positive people, multiple-genogroup infections were common and associated with higher viral loads than would be expected from simple additive effects. The latter observation was restricted to the infections which included both genogroups 1 and 3, pointing to the possible existence of some kind of infection facilitation between these genogroups.

Background The haplotypes of the X chromosome are accessible to direct count in males, whereas the diplotypes of the females may be inferred knowing the haplotype of their sons or fathers. Here, we investigated: 1) the possible large-scale haplotypic structure of the X chromosome in a Caucasian population sample, given the single-nucleotide polymorphism (SNP) maps and genotypes provided by Illumina and Affimetrix for Genetic Analysis Workshop 14, and, 2) the performances of widely used programs in reconstructing haplotypes from population genotypic data, given their known distribution in a sample of unrelated individuals. Results All possible unrelated mother-son pairs of Caucasian ancestry (N = 104) were selected from the 143 families of the Collaborative Study on the Genetics of Alcoholism pedigree files, and the diplotypes of the mothers were inferred from the X chromosomes of their sons. The marker set included 313 SNPs at an average density of 0.47 Mb. Linkage disequilibrium between pairs of markers was computed by the parameter D', whereas for measuring multilocus disequilibrium, we developed here an index called D*, and applied it to all possible sliding windows of 5 markers each. Results showed a complex pattern of haplotypic structure, with regions of low linkage disequilibrium separated by regions of high values of D*. The following programs were evaluated for their accuracy in inferring population haplotype frequencies: 1) ARLEQUIN 2.001; 2) PHASE 2.1.1; 3) SNPHAP 1.1; 4) HAPLOBLOCK 1.2; 5) HAPLOTYPER 1.0. Performances were evaluated by Pearson correlation (r) coefficient between the true and the inferred distribution of haplotype frequencies. Conclusion The SNP haplotypic structure of the X chromosome is complex, with regions of high haplotype conservation interspersed among regions of higher haplotype diversity. All the tested programs were accurate (r = 1) in reconstructing the distribution of haplotype frequencies in case of high D* values. However, only the program PHASE realized a high correlation coefficient (r > 0.7) in conditions of low linkage disequilibrium.

Germ-line mutations in the BRCA1 gene cause hereditary predisposition to breast and ovarian cancer. BRCA1 and BRCA2 mutations account for about 40% of high-risk families. Mutation-screening methods generally focus on genomic DNA and are usually PCR based; they enable the detection of sequence alterations such as point mutations and small deletions and insertions. However, they do not allow the detection of partial or entire exon(s) loss, because the presence of the homologous allele results in a positive PCR signal, giving rise to a false-negative result. Identification of unusual haplotypes in patient samples by an expectation maximisation algorithm has recently been suggested as a method for identifying hemizygous regions caused by large intragenic deletions. Using a similar approach, we identified a novel BRCA1 genomic rearrangement in a breast/ovarian cancer family negative at the first mutation screening; we detected a deletion encompassing exons 14–19, probably due to replication slippage between Alu sequences.

Children with bronchopneumonia have considerably higher Torque tenovirus (TTV) loads than do children with milder acute respiratory diseases (ARDs). Moreover, in children with ARDs, high TTV loads correlate with low percentages of circulating CD3 + and CD4 + T cells and with elevated percentages of B cells, suggesting that TTV might be immunomodulatory. Here, we show that, in children with ARDs, the presence of TTV and TTV load correlate with concentrations of serum eosinophil cationic protein. The possible mechanisms whereby TTV infection might lead to augmented activity of eosinophils and the implications for pathogenesis are discussed.

Accurate estimates of breast and ovarian cancer penetrance in BRCA1/2 mutation carriers are crucial in genetic counseling. Estimation is difficult because of the low frequency of mutated alleles and the often-uncertain mechanisms of family ascertainment. We estimated the penetrances of breast and ovarian cancers in carriers of BRCA1/2 mutations by maximizing the retrospective likelihood of the genetic model, given the observed test results, in 568 Italian families screened for germline mutations. The software BRCAPRO was used as a probability calculation tool in a Markov Chain Monte Carlo approach. Breast cancer penetrances were 27% (95% CI 20-34%) at age 50 years and 39% (27-52%) at age 70 in BRCA1 carriers, and 26% (0.18-0.34%) at age 50 and 44% (29-58%) at age 70 in BRCA2 carriers, and ovarian cancer penetrances were 14% (7-22%) at age 50 and 43% (21-66%) at age 70 in BRCA1 carriers and 3% (0-7%) at age 50 and 15% (4-26%) at age 70 in BRCA2 carriers. The new model gave a better fit than the current default in BRCAPRO, the likelihood being 70 log units greater; in addition, the observed numbers of mutations in families stratified by gene and by cancer profile were not significantly different from those expected. Our new penetrance functions are appropriate for predicting breast cancer risk, and for determining the probability of carrying BRCA1/2 mutations, in people who are presently referred to genetic counseling in Italy. Our approach could lead to country-customized versions of the BRCAPRO software by providing appropriate population-specific estimates.

Four X-chromosome markers (DXS101, HPRTB, DXS8377, and STRX-1) were typed in a population sample from Tuscany, Italy, using a single amplification reaction. Resolution of a deficiency paternity case involving two women (either they were half-sisters or non-relatives) was made possible by typing these marker in addition to 16 conventional autosomal markers.

The natural history and pathogenic potential of the recently identified TT virus (TTV) are currently a matter of intensive investigation. In an attempt to shed some light on these issues, nasal and blood specimens of 1- to 24-month-old children hospitalized with a clinical diagnosis of acute respiratory disease (ARD) were examined for the presence, load, and genetic characteristics of TTV. The results have indicated that at least in young children, the respiratory tract not only represents a route by which abundant TTV can be shed into the environment but also may be a site of primary infection and continual replication. Although we found no compelling evidence that TTV was the direct cause of ARD in some of the children studied, the average loads of TTV were considerably higher in patients with bronchopneumonia (BP) than in those with milder ARD, raising interesting questions about the pathophysiological significance of TTV at this site. Furthermore, group 4 TTV was detected almost exclusively in children with BP.

TT virus (TTV) produces chronic plasma viremia in around 90% of healthy individuals of all ages and has, therefore, been proposed as a commensal human virus. We recently demonstrated that in children hospitalized for acute respiratory diseases high TTV loads were associated with severe forms of disease. Here, we report that in such children TTV loads showed an inverse correlation with the percentage of circulating total T and helper T cells and a direct correlation with the percentage of B cells. Thus, florid TTV replication might contribute to lymphocyte imbalances and, possibly, immunosuppressive effects, thus resembling related animal viruses. TT virus (TTV) has genome properties similar to those of animal circoviruses, such as chicken anemia virus and porcine circovirus. Because it produces chronic plasma viremia in around 90% of healthy individuals of all ages worldwide and no associated disease has yet been identified, it has been suggested that TTV might be totally apathogenic (for reviews, see references 1 and 11). Recently, we tested for the presence of TTV and assessed viral load by universal untranslated-region real-time PCR in 157 children under 2 years old with acute respiratory diseases (ARD) on the day of hospital admission and obtained convincing data that TTV might replicate in the respiratory tract (5). Also, although we found no evidence that TTV might be the direct cause of ARD, TTV loads in both nasal swabs and plasma samples were substantially higher in subjects with bronchopneumonia (BP) than in the subjects with milder ARD (laryngitis, bronchitis, and bronchiolitis), suggesting among other possibilities that TTV could be locally or systemically immunosuppressive and aggravate disease induced by other agents (5). However, there is no information on this matter except for recent reports showing an inverse relationship between TTV burdens and CD4 cell counts in patients with human immunodeficiency virus type 1 (2, 10, 13). In this study, we examined, with informed parental consent, peripheral leukocyte and lymphocyte subset counts in 40 randomly chosen children with ARD from the above study (5). Of the 40 children, 18 had an X-ray-confirmed diagnosis of BP. The same blood samples used for TTV assays were analyzed. Features similar to those described for the entire sample (5) were confirmed in this subset. In particular, mean TTV loads in plasma were significantly higher in BP patients than in children with milder ARD (7.9 1.2 versus 5.6 2.3 log10 per ml [P 0.001]). Interestingly, although all absolute cell counts were within the values reported for healthy children of similar age (3) with unknown TTV status (but presumably mostly TTV positive, based on what is currently known of TTV epidemiol

Hereditary non-polyposis colorectal cancer (HNPCC) is a dominantly inherited syndrome linked to DNA-mismatch-repair (MMR) gene defects, which also account for microsatellite instability (MSI) in tumour tissues. Diagnosis is based mainly on family history, according to widely accepted criteria (Amsterdam Criteria: AC). Aim of this work was to assess MSI in colorectal-cancer patients with suspected genetic predisposition, and to verify whether MSI represents a tool to manage MMR gene (hMSH2 and hMLH1) mutation analysis. We investigated 13 microsatellites (including the 5 NCI/ICG-HNPCC markers) in 45 patients with suspected hereditary predisposition (including 16 subjects from HNPCC families fulfilling the AC). We found MSI-H (high frequency of instability, i.e., in ≥30% of the markers) in 85% of the HNPCC patients and in 16% of the non-HNPCC subjects. The 5 NCI/ICG-HNPCC microsatellites proved to be the most effective in detecting MSI, being mononucleotide repeats the most unstable markers. We investigated the association between hMSH2- and hMLH1 gene mutations and MSI. Our results indicate that AC are highly predictive both of tumour instability and of MMR-gene mutations. Therefore, as the most likely mutation carriers, HNPCC subjects might be directly analyzed for gene mutations, while to test for MSI in selected non-HNPCC patients and to further investigate MMR genes in MSI-H cases, appears to be a cost-effective way to identify subjects, other than those from kindred fulfilling AC, who might benefit from genetic testing. Int. J. Cancer (Pred. Oncol.) 89:87–91, 2000. © 2000 Wiley-Liss, Inc.

Conflicting data exist on the prognosis of hereditary colorectal cancer. HNPCC patients, in particular, are often reported to have a better survival. We examined 2,340 colorectal-cancer patients treated in our Institution: 144 HNPCC patients (Amsterdam Criteria), 161 FAP patients and 2,035 patients with sporadic cancer. Data on hereditary-cancer patients treated between 1980 and 1995 was collected in a registry. The 2,035 sporadic colorectal-cancer patients (controls) included all new cases treated in the Department of Gastrointestinal-Tract Surgery during the same period. Observed survival was estimated using the Kaplan-Meier method. Cumulative survival probability was estimated at 5 years within each group and stratified by various clinical and pathological variables. The age distribution at diagnosis of sporadic patients was significantly higher than that of FAP and HNPCC patients (median 60 years vs. 43 and 49 years; p < 0.0001). In the HNPCC group, 40% had a right cancer location, vs. 14% in the FAP group and 13% in the sporadic-cancer group. In the sporadic group, 51% were early-stage cancers (Dukes A or B) vs. 48.4% and 52.1% in the FAP and HNPCC groups respectively. In the HNPCC, FAP and sporadic-cancer groups, the 5-year cumulative survival rate was 56.9%, 54.4% and 50.6% respectively. Survival analysis by the Cox proportional-hazards method revealed no substantial survival advantage for HNPCC and FAP patients compared with the sporadic group, after adjustment for age, gender, stage and tumor location. The hazard ratio for HNPCC was 1.01 (95% CI 0.72-1.39) and 1.27 (95% CI 0.95-1.7) for FAP patients compared with the sporadic-colorectal-cancer group.

Background Genetic improvement of fibre-producing animal species has often induced transition from double coated to single coated fleece, accompanied by dramatic changes in skin follicles and hair composition, likely implying variation at multiple loci. Huacaya, the more common fleece phenotype in alpaca (Vicugna pacos), is characterized by a thick dense coat growing perpendicularly from the body, whereas the alternative rare and more prized single-coated Suri phenotype is distinguished by long silky fibre that grows parallel to the body and hangs in separate, distinctive pencil locks. A single-locus genetic model has been proposed for the Suri-Huacaya phenotype, where Huacaya is recessive. Results Two reciprocal experimental test-crosses (Suri × Huacaya) were carried out, involving a total of 17 unrelated males and 149 unrelated females. An additional dataset of 587 offspring of Suri × Suri crosses was analyzed. Segregation ratios, population genotype frequencies, and/or recombination fraction under different genetic models were estimated by maximum likelihood. The single locus model for the Suri/Huacaya phenotype was rejected. In addition, we present two unexpected observations: 1) a large proportion (about 3/4) of the Suri animals are segregating (with at least one Huacaya offspring), even in breeding conditions where the Huacaya trait would have been almost eliminated; 2) a model with two different values of the segregation ratio fit the data significantly better than a model with a single parameter. Conclusions The data support a genetic model in which two linked loci must simultaneously be homozygous for recessive alleles in order to produce the Huacaya phenotype. The estimated recombination rate between these loci was 0.099 (95% C.L. = 0.029-0.204). Our genetic analysis may be useful for other species whose breeding system produces mainly half-sib families.

Background The traditional exact method for inferring relationships between individuals from genetic data is not easily applicable in all situations that may be encountered in several fields of applied genetics. This study describes an approach that gives affordable results and is easily applicable; it is based on the probabilities that two individuals share 0, 1 or both alleles at a locus identical by state.

The possible genetic effects produced by styrene have been investigated by means of different methodologies in several biological organisms: (a) the induction of point mutation has been investigated in Salmonella typhimurium (reverse mutation), in the yeast Schizosaccharomyces pombe (forward mutation), both in vitro and in vivo, in the host-mediated assay of mice, and in the Chinese hamster cell line grown in vitro (V-79) (forward mutation); (b) the induction of chromosome mutation has been investigated in vivo, in mice, through the analysis of the presence of chromosome aberrations in bone marrow cells of treated animals; (c) the production of DNA (deoxyribonucleic acid) damage and the stimulation of DNA repair synthesis have been evaluated from measurements of unscheduled DNA synthesis in a heteroploid human cell line (EUE) and gene-conversion produced in the yeast Saccharomyces cerevisiae treated in vitro and in vivo (host-mediated assay). All the in vitro studies have been developed by the testing of the styrene in the presence of a metabolic activating system obtained with a mouse liver microsomal preparation. Styrene oxide, one of the in vivo metabolites of styrene with electrophilic properties towards DNA molecules, have also been tested in similar systems. Styrene was not mutagenic in all the systems tested; styrene oxide, on the contrary, was shown to be an active mutagen, independently of the genetic system under evaluation.

Deleterious mutations of the BRCA1 and BRCA2 genes are a major risk factor for the development of breast and ovarian cancers.1–4 Mutation tests for these two genes commonly are now offered in specialised clinics.5,6 As a result, a large number of women with personal or family histories of breast or ovarian cancer seek genetic counselling. Accurate evaluation of the probability that a woman carries a germline pathogenic mutation at BRCA1 or BRCA2 therefore is essential to help counsellors and those being counselled to decide whether testing is appropriate. In this context, the questions of practical interest are: Given the pedigree, what is the chance of a mutation being present? and What is the chance of the DNA laboratory finding a mutation? After testing became available, several models were developed to assess the pre-test probability of identifying carriers of mutations. Broadly speaking, two different approaches have been used to develop predictive models: the “empirical approach” and the “Mendelian approach”.7 In empirical models, families are stratified according to variables that describe their family history; regression or other approaches are used to predict the results of Mendelian testing. In some cases, this approach simply consists of observing the proportion of mutations found in different strata. Mendelian models, in contrast, address the probability that a proband is a mutation carrier on the basis of explicit assumptions about the genetic parameters (allele frequencies and cancer penetrances in carriers and non-carriers) and the Mendelian rules of gene transmission. A consequence of the two different strategies is that the Mendelian models evaluate the probability that a proband is a gene carrier, whereas the empirical models evaluate the probability of identifying a mutation. The main purpose of this study was to compare the performances of published models in predicting mutation test results in …

A case-control study was carried out to determine whether residential exposure to environmental pollutants increased risk for canine lymphoma in pet dogs. One hundred one cases with cytologically or histologically confirmed lymphoma diagnosed at a veterinary teaching hospital between the middle of 1996 and the middle of 1998 were examined. Controls were obtained by choosing twice the number of dogs without neoplastic disease, with overlapping distributions of province of residence, age, sex, and breed. Information regarding animal management, residence type, professional or hobby use of chemicals by owners, and treatment with herbicides or other pesticides in the area frequently visited by the dogs was obtained with a multiple-choice questionnaire by telephone interview. Two variables were positively and independently associated with the disease, namely residency in industrial areas (odds ratio [OR]; = 8.5; 95% confidence interval [CI], 2.3-30.9) and use of chemicals by owners, specifically paints or solvents (OR = 4.6; 95% CI, 1.7-12.6). A significantly lower value of the mean age of disease onset was found in the group of dogs at risk in comparison with the group of all other dogs (6.1 +/- 0.4 years, n = 36 versus 7.5 +/- 0.4 years, n = 65, respectively; P = .008). Variables describing animal care and pesticide use were either not associated with the disease or were uninformative. We suggest that canine lymphoma may be considered a sentinel of potentially hazardous situations for humans, because of the relatively short latency between exposure and disease onset.