Facial clues to the photosensitive trichothiodystrophy phenotype in childhood.

Among genodermatoses, trichothiodystrophies (TTDs) are a rare genetically heterogeneous group of syndromic conditions, presenting with skin, hair, and nail abnormalities. An extra-cutaneous involvement (craniofacial district and neurodevelopment) can be also a part of the clinical picture. The presence of photosensitivity describes three forms of TTDs: MIM#601675 (TTD1), MIM#616390 (TTD2) and MIM#616395 (TTD3), that are caused by variants afflicting some components of the DNA Nucleotide Excision Repair (NER) complex and with more marked clinical consequences. In the present research, 24 frontal images of paediatric patients with photosensitive TTDs suitable for facial analysis through the next-generation phenotyping (NGP) technology were obtained from the medical literature. The pictures were compared to age and sex-matched to unaffected controls using 2 distinct deep-learning algorithms: DeepGestalt and GestaltMatcher (Face2Gene, FDNA Inc., USA). To give further support to the observed results, a careful clinical revision was undertaken for each facial feature in paediatric patients with TTD1 or TTD2 or TTD3. Interestingly, a distinctive facial phenotype emerged by the NGP analysis delineating a specific craniofacial dysmorphic spectrum. In addition, we tabulated every single detail within the observed cohort. The novelty of the present research includes the facial characterization in children with the photosensitive types of TTDs through the 2 different algorithms. This result can become additional criteria for early diagnosis, and for subsequent targeted molecular investigations as well as a possible tailored multidisciplinary personalized management.

What people really change after genetic testing (GT) performed in private labs: results from an Italian study.

Despite the widespread diffusion of direct-to-consumer genetic testing (GT), it is still unclear whether people who learn about their genetic susceptibility to a clinical condition change their behaviors, and the psychological factors involved. The aim of the present study is to investigate long-term changes in health-related choices, individual tendencies and risk attitudes in an Italian sample of GT users. In the context of the Mind the Risk study, which investigated a sample of Italian adults who underwent GT in a private laboratory, 99 clients participated in the follow up assessment. They completed a self-administered questionnaire investigating: (a) clinical history and motivation for testing, (b) lifestyle and risk behaviors, (c) individual tendencies toward health, and (d) risk-taking attitude and risk tolerance. Such variables were measured at three different time-points: T0-before GT, T1-at 6 months after genetic results, and T2-at 1 year from results. Results showed that, at baseline, participants who stated they intended to modify their behavior after GT results, effectively did so over time. This result held both for participants who received a positive or negative test result. In general, a healthier diet was the most frequently observed long-term behavioral change. As regards psychological variables, a risk-taking attitude and risk tolerance did not seem to affect the decision to change the lifestyle. Finally, we found an overall reduction in anxiety and worry over health over time, but also a reduction in the motivation for health promotion and prevention, health esteem, and positive expectations for their health in the future.

Health orientation and individual tendencies of a sample of Italian genetic testing consumers.

BACKGROUND: Over the last decade, genetic testing (GT) had markedly spread in European countries and struggled the debate concerning the psychological effects on the population. The aim of this study was to investigate the individual tendencies of GT consumers in a sample of Italian citizens. METHODS: A total of 152 Italian clients from GenomaLab, a private genetic company, were enrolled from February 2016 to September 2018 and completed an ad hoc survey. RESULTS: Results showed that GT consumers were motivated to preserve their well-being, they felt responsible for their health, they were neither pessimistic nor optimistic toward negative occurrences, and poorly inclined to take high risks in their lives. Participants who had suffered from a disease in the past appear to be less tolerant to the uncertainty for future negative events. CONCLUSION: Our results depict Italian GT consumers as health-oriented, focused on prevention, who do not have a pessimistic perception of their condition but do not like to "bet" on their health, and probably their intention (and belief) is to acquire genetic information in order to reduce uncertainty and increase their decision-making "power" related to their health. Taken together, all these results contribute to describe the population of GT users in European countries, to regulate the provision of GT results and to entail the communication of genetic risk information based on a consumers' personal profile.

Extent of chromosomal mosaicism influences the clinical outcome of in vitro fertilization treatments.

OBJECTIVE: To assess whether the extent of chromosomal mosaicism can influence the success rate of IVF treatments. DESIGN: Prospective study. SETTING: Private genetic and assisted reproduction centers. PATIENT(S): The transfer of mosaic embryos was offered to 77 women for which IVF resulted in no euploid embryos available for transfer. INTERVENTION(S): All embryos were cultured to blastocyst stage; trophectoderm biopsy was performed on day 5/6 of development. Comprehensive chromosome screening was performed using either next-generation sequencing or array-comparative genomic hybridization methodologies. MAIN OUTCOME MEASURE(S): The clinical outcome obtained after transfer of mosaic embryos with low (<50%) and high (≥50%) aneuploidy percentage was compared with that resulting from a control group of 251 euploid blastocysts. RESULT(S): A significantly higher implantation rate (48.9% vs. 24.2%), clinical pregnancy rate/ET (40.9% vs. 15.2%), and live-birth rate (42.2% vs. 15.2%) were observed comparing embryos with mosaicism <50% and ≥50%. Mosaic embryos with high aneuploidy percentage (≥50%) showed a significantly lower clinical pregnancy rate/ET (15.2% vs. 46.4%), implantation rate (24.4% vs. 54.6%), and live-birth rate (15.2% vs. 46.6%) than euploid blastocysts. In contrast, embryos with lower aneuploidy percentage (<50%) have a clinical outcome similar to euploid embryos. CONCLUSION(S): The results of this study further confirm that mosaic embryos can develop into healthy euploid newborns. We demonstrated that the extent of mosaicism influences the IVF success rate. Mosaic embryos with low aneuploidy percentage have higher chances of resulting in the birth of healthy babies compared with embryos with higher mosaicism levels.

The clinical utility of genome-wide non invasive prenatal screening.

OBJECTIVE: In this study, we expanded conventional cell-free fetal DNA (cfDNA)-based non-invasive prenatal testing (NIPT) to cover the entire genome. We aimed to compare the performance of the two tests in a large general population of pregnant women, in order to assess the clinical utility of the genome-wide screening. METHOD: Genome-wide cfDNA analysis was offered to 12 114 pregnant women undergoing NIPT for common fetal aneuploidy. Sequencing data were analyzed using an algorithm optimized to identify aneuploidies and subchromosomal aberrations. RESULTS: Genome-wide screening allowed detection of 12 (7.4%) potentially viable clinically relevant chromosomal abnormalities, which would have remained overlooked if only conventional NIPT had been performed. This resulted in a statistically significant higher sensitivity (100% vs 92.64%, p < 0.001) than did standard screening. This was achieved without sacrificing the specificity of the test, which resulted similar to that obtained with standard cfDNA testing (99.87% vs 99.77%, p = 0.064). CONCLUSION: Genome-wide cfDNA analysis represents an enhanced screening tool for prenatal detection of chromosomal abnormalities, allowing identification of clinically relevant imbalances that are not detectable by conventional cfDNA testing. The results of this study demonstrate the clinical utility of genome-wide cfDNA analysis. This level of screening provides a significant higher sensitivity compared to standard screening while maintaining a high specificity, with the potential to improve overall pregnancy management. © 2017 John Wiley & Sons, Ltd.

The importance of determining the limit of detection of non-invasive prenatal testing methods.

OBJECTIVE: Several non-invasive prenatal testing (NIPT) methods, which analyze circulating fetal cell-free DNA (cfDNA) in maternal plasma, suggest a fetal fraction (FF) ≥ 4% for a reportable result, with the assumption that fetal aneuploidies may not be detectable at lower FF. This study determined the actual limit of detection (LOD) of a massively parallel sequencing-based NIPT method and evaluated its performance in testing samples with low FF. METHOD: An experimental model, involving the creation of artificial plasma mixtures with a final aneuploid FF ranging from 1% to 4%, simulated samples at different proportions of fetal cfDNA. We then analyzed 7103 blood samples, from pregnant women undergoing NIPT, to assess the impact of low FF on the performance of cfDNA testing. RESULTS: Detection of common aneuploidies in samples with an FF as low as 2% is well within the ability of this technology. Of 105 pregnancies confirmed chromosomally abnormal, 25 (23.8%) involving a 2% < FF < 4% were consistently detected. These high-risk pregnancies would have not been identified using the suggested 4% FF cut-off. CONCLUSION: This study underscores the importance of determining the actual LOD for each specific NIPT methodology. It may reduce the incidence of test cancelations and shorten the time required for the diagnosis of aneuploidy. © 2016 John Wiley & Sons, Ltd.

Seasonal differences in climate in the Chianti region of Tuscany and the relationship to vintage wine quality.

Climatic factors and weather type frequencies affecting Tuscany are examined to discriminate between vintages ranked into the upper- and lower-quartile years as a consensus from six rating sources of Chianti wine during the period 1980 to 2011. These rankings represent a considerable improvement on any individual publisher ranking, displaying an overall good consensus for the best and worst vintage years. Climate variables are calculated and weather type frequencies are matched between the eight highest and the eight lowest ranked vintages in the main phenological phases of Sangiovese grapevine. Results show that higher heat units; mean, maximum and minimum temperature; and more days with temperature above 35 °C were the most important discriminators between good- and poor-quality vintages in the spring and summer growth phases, with heat units important during ripening. Precipitation influences on vintage quality are significant only during veraison where low precipitation amounts and precipitation days are important for better quality vintages. In agreement with these findings, weather type analysis shows good vintages are favoured by weather type 4 (more anticyclones over central Mediterranean Europe (CME)), giving warm dry growing season conditions. Poor vintages all relate to higher frequencies of either weather type 3, which, by producing perturbation crossing CME, favours cooler and wetter conditions, and/or weather type 7 which favours cold dry continental air masses from the east and north east over CME. This approach shows there are important weather type frequency differences between good- and poor-quality vintages. Trend analysis shows that changes in weather type frequencies are more important than any due to global warming.

Mediterranean climate patterns and wine quality in North and Central Italy.

Results show that the year-to-year quality variation of wines produced in North and Central Italy depends on the large-scale climate variability, and that the wine quality improvement in the last four decades is partially due to an increase of temperature and to a decrease of precipitation in West and Central Mediterranean Europe (WME; CME). In addition, wine quality is positively correlated with air temperature throughout the entire active period of the grapevine, weakly negatively correlated with precipitation in spring, and well negatively correlated in summer and fall. The month-to-month composites of the NAO anomaly show that, in years of good quality wine, this anomaly is negative in late spring, oscillates around zero in summer, and is positive in early fall; while, in years of bad quality wine, it is positive in late spring and summer, and negative in early fall, i.e. its polarity has an opposite sign in spring and fall in good versus bad years. The composite seasonal maps show that good wines are produced when the spring jet stream over the Atlantic diverts most of the weather perturbations towards North Europe, still providing a sufficient amount of rainwater to CME; when summer warming induced by southerly winds is balanced by the cooling induced by westerly winds; and when a positive geopotential anomaly over WME shelters CME from fall Atlantic storms. Bad quality wines are produced when the jet stream favors the intrusion of the Atlantic weather perturbations into the Mediterranean. Results suggest that atmospheric pattern persistencies can be used as precursors for wine quality forecast.

Sequential comprehensive chromosome analysis on polar bodies, blastomeres and trophoblast: insights into female meiotic errors and chromosomal segregation in the preimplantation window of embryo development.

STUDY QUESTION: What is the optimal stage from oocyte through preimplantation embryo development for biopsy and preimplantation genetic screening (PGS) to detect abnormal chromosome segregation patterns in eggs or embryos from advanced maternal age (AMA) patients? SUMMARY ANSWER: Testing at the polar body (PB) stage was the least accurate mainly due to the high incidence of post-zygotic events. This suggests that postponing the time of biopsy to the blastocyst stage of preimplantation embryo development may provide the most reliable results for PGS. WHAT IS KNOWN ALREADY: In the PGS field there is an ongoing debate about the optimal biopsy stage for PGS. This is a result of the lack of understanding of how aneuploidy arises in the human embryo. To date, most of the cytogenetic data obtained during PGS investigations have been derived through the analysis of cells at isolated points in the preimplantation window, thus potentially missing critical information on chromosomal segregation. Understanding the chromosome segregation patterns during preimplantation development holds the potential to significantly increase the success rates of IVF. In this study, a sequential comprehensive chromosome analysis of both the PBs and the corresponding embryos at both the cleavage and the blastocyst stages is presented. STUDY DESIGN, SIZE, DURATION: This is a prospective longitudinal cohort study performed between October 2009 and August 2011 involving 9 infertile couples and 21 sets of complete comprehensive chromosomal screening data, including PB1, PB2, corresponding blastomeres and trophectoderm (TE) samples. PARTICIPANTS/MATERIALS, SETTING, METHODS: Infertile couples undergoing IVF cycles with PGS where the female partner was older than 40 years and with a good response to controlled ovarian stimulation (>10 MII oocytes retrieved) were enrolled into the study. The exclusion criteria were (i) patients presenting with abnormal karyotype; (ii) specific ovarian pathologies including polycystic ovary syndrome, endometriosis grade III or higher and premature ovarian failure and (iii) severe male factor infertility (motile sperm count of <500 000/ml after preparation of a fresh ejaculate). The PBs, blastomere and TE samples were sequentially biopsied and analyzed by array comparative genomic hybridization (aCGH). The analysis of chromosome segregation patterns was performed to infer the origin of aneuploidy and to investigate the diagnostic accuracy of both PB and cleavage-stage PGS strategies. MAIN RESULTS AND THE ROLE OF CHANCE: Twenty-one sets of complete data (PB1/PB2/blastomere/TE) including 84 aCGH experiments showed a pattern of multiple meiotic errors typically caused by sister chromatid separation errors and predominantly arising in the second meiotic division. Twenty-two of the 24 (91.7%) errors in the first meiotic division arose as a consequence of premature sister chromatid predivision. In half of these cases, the second meiotic division resulted in a balancing chromosome segregation event producing a normal female complement for that chromosome in the resulting embryo. Overall, only 62 out of 78 (79.5%) of the abnormal meiotic segregations had errors in the either one or both PBs consistent with the aneuploidies observed in their resulting embryos. Ten of the 21 (47.6%) embryos had aneuploidies other than female meiotic-derived ones, most of which detected on Day 3 and confirmed on Day 5 or 6 of embryo development (20/25) with chromosomal loss being three times more frequent than gains. Notably, as high as 20% of female-derived aneuploidies detected on PBs and confirmed on Day 3 were rescued at the blastocyst stage, mainly as a result of diploidization of trisomic chromosomes. On a per chromosome basis, the sensitivity in predicting blastocyst chromosomal complement was significantly lower for PB approach, 61.7%, compared with blastomeres analysis, 86.4% (P < 0.01). LIMITATIONS, REASONS FOR CAUTION: The study was limited to the analysis of oocytes and embryos from AMA patients. Thus, these findings apply only to this patient group. Comparisons with other patient populations including patients with different indications for PGS should be made in future research. In addition, higher resolution and/or more accurate chromosomal screening tests could be used in future studies to corroborate the current findings. WIDER IMPLICATIONS OF THE FINDINGS: These findings provide critical insights into the mechanisms causing errors during female meiosis and the preimplantation embryo development period to improve the design and treatment outcome of PGS.

Chromosomal microarray analysis as a first-line test in pregnancies with a priori low risk for the detection of submicroscopic chromosomal abnormalities.

In this study, we aimed to explore the utility of chromosomal microarray analysis (CMA) in groups of pregnancies with a priori low risk for detection of submicroscopic chromosome abnormalities, usually not considered an indication for testing, in order to assess whether CMA improves the detection rate of prenatal chromosomal aberrations. A total of 3000 prenatal samples were processed in parallel using both whole-genome CMA and conventional karyotyping. The indications for prenatal testing included: advanced maternal age, maternal serum screening test abnormality, abnormal ultrasound findings, known abnormal fetal karyotype, parental anxiety, family history of a genetic condition and cell culture failure. The use of CMA resulted in an increased detection rate regardless of the indication for analysis. This was evident in high risk groups (abnormal ultrasound findings and abnormal fetal karyotype), in which the percentage of detection was 5.8% (7/120), and also in low risk groups, such as advanced maternal age (6/1118, 0.5%), and parental anxiety (11/1674, 0.7%). A total of 24 (0.8%) fetal conditions would have remained undiagnosed if only a standard karyotype had been performed. Importantly, 17 (0.6%) of such findings would have otherwise been overlooked if CMA was offered only to high risk pregnancies.The results of this study suggest that more widespread CMA testing of fetuses would result in a higher detection of clinically relevant chromosome abnormalities, even in low risk pregnancies. Our findings provide substantial evidence for the introduction of CMA as a first-line diagnostic test for all pregnant women undergoing invasive prenatal testing, regardless of risk factors.

Introducing array comparative genomic hybridization into routine prenatal diagnosis practice: a prospective study on over 1000 consecutive clinical cases.

OBJECTIVE: To assess the feasibility of offering array-based comparative genomic hybridization testing for prenatal diagnosis as a first-line test, a prospective study was performed, comparing the results achieved from array comparative genomic hybridization (aCGH) with those obtained from conventional karyotype. METHOD: Women undergoing amniocentesis or chorionic villus sampling were offered aCGH analysis. A total of 1037 prenatal samples were processed in parallel using both aCGH and G-banding for standard karyotyping. Specimen types included amniotic fluid (89.0%), chorionic villus sampling (9.5%) and cultured amniocytes (1.5%). RESULTS: Chromosomal abnormalities were identified in 34 (3.3%) samples; in 9 out of 34 cases (26.5%) aCGH detected pathogenic copy number variations that would not have been found if only a standard karyotype had been performed. aCGH was also able to detect chromosomal mosaicism at as low as a 10% level. There was complete concordance between the conventional karyotyping and aCGH results, except for 2 cases that were only correctly diagnosed by aCGH. CONCLUSIONS: This study demonstrates that aCGH represents an improved diagnostic tool for prenatal detection of chromosomal abnormalities. Although larger studies are needed, our results provide further evidence on the feasibility of introducing aCGH as a first-line diagnostic test in routine prenatal diagnosis practice.

Genome profiling of ovarian adenocarcinomas using pangenomic BACs microarray comparative genomic hybridization.

BACKGROUND: Routine cytogenetic investigations for ovarian cancers are limited by culture failure and poor growth of cancer cells compared to normal cells. Fluorescence in situ Hybridization (FISH) application or classical comparative genome hybridization techniques are also have their own limitations in detecting genome imbalance especially for small changes that are not known ahead of time and for which FISH probes could not be thus designed. METHODS: We applied microarray comparative genomic hybridization (A-CGH) using one mega base BAC arrays to investigate chromosomal disorders in ovarian adenocarcinoma in patients with familial history. RESULTS: Our data on 10 cases of ovarian cancer revealed losses of 6q (4 cases mainly mosaic loss), 9p (4 cases), 10q (3 cases), 21q (3 cases), 22q (4 cases) with association to a monosomy X and gains of 8q and 9q (occurring together in 8 cases) and gain of 12p. There were other abnormalities such as loss of 17p that were noted in two profiles of the studied cases. Total or mosaic segmental gain of 2p, 3q, 4q, 7q and 13q were also observed. Seven of 10 patients were investigated by FISH to control array CGH results. The FISH data showed a concordance between the 2 methods. CONCLUSION: The data suggest that A-CGH detects unique and common abnormalities with certain exceptions such as tetraploidy and balanced translocation, which may lead to understanding progression of genetic changes as well as aid in early diagnosis and have an impact on therapy and prognosis.

Diagnostic evaluation of women experiencing repeated in vitro fertilization failure.

OBJECTIVE: The aim of the study was to propose a set of tests to clarify the diagnosis of repeated implantation failure in patients undergoing in vitro fertilization (IVF). STUDY DESIGN: Fifty-nine patients with at least two unsuccessful IVF attempts were included in the study. Blood samples were evaluated for the presence of underlying thyroid abnormalities, antiphospholipid antibodies (aPL), increased levels of natural killer cells (NK), inherited thrombophilia and mouse embryo assay factor (MEA-f). The same tests were performed on 20 normal fertile control patients. RESULTS: Seventy-six percent of IVF patients showed at least one abnormal result. This incidence was higher with respect to that found among control patients (45%). The prevalence of thyroid abnormalities, aPL and increased NK level was higher in IVF patients whereas no differences were observed in terms of prevalence of inherited thrombophilias and MEA-f. CONCLUSIONS: A better understanding of reproductive failure mechanisms should allow an effective diagnostic flow chart and a focused therapeutic option for patients experiencing repeated IVF failure. With this objective in mind, our data provide two important results: thyroid abnormalities, aPL and increased NK levels are more prevalent in women experiencing IVF failure. No evidence was found for an association between inherited thrombophilia and MEA-f and failure to achieve pregnancy after IVF.

Short tandem repeats haplotyping of the HLA region in preimplantation HLA matching.

Recently, preimplantation genetic diagnosis (PGD) has been considered for several indications beyond its original purpose, not only to test embryos for genetic disease but also to select embryos for a nondisease trait, such as specific human leukocyte antigen (HLA) genotypes, related to immune compatibility with an existing affected child in need of a haematopoetic stem cell (HSC) transplant. We have optimized an indirect single-cell HLA typing protocol based on a multiplex fluorescent polymerase chain reaction (PCR) of short tandem repeat (STR) markers scattered throughout the HLA complex. The assay was clinically applied in 60 cycles from 45 couples. A conclusive HLA-matching diagnosis was achieved in 483/530 (91.1%) of the embryos tested. In total, 74 (15.3%) embryos revealed an HLA match with the affected siblings, 55 (11.4%) of which resulted unaffected and 46 (9.5%) have been transferred to the patients. Nine pregnancies were achieved, five healthy HLA-matched children have already been delivered and cord blood HSCs, were transplanted to three affected siblings, resulting in a successful haematopoietic reconstruction.

The use of non-physiological conditions to isolate fetal cells from maternal blood.

Fetal cells are always present in maternal blood starting in the first trimester of pregnancy, however a rapid, simple, and consistent procedure for their isolation for prenatal non-invasive genetic investigation is still lacking. Sensitivity and recovery of fetal cells is jeopardized by the minute amount of circulating fetal cells and their loss during the enrichment procedure. We report here a single-step approach to isolate fetal cells from maternal blood which relies on the use of non-physiological conditions to modify cell densities before their separation in a density gradient and in a newly developed cell separation device. Isolated fetal cells have been investigated using cytochemistry, Soret band absorption microscopy, monoclonal antibodies for epsilon- and gamma-chain-Hb, monoclonal antibody for i-antigen, and by fluorescence in situ hybridization (FISH). Fetal cells were always detected in all 105 maternal blood samples investigated and fetal aneuploidies were correctly diagnosed by FISH, in a pilot study of pathological pregnancies, in fetal cells isolated from maternal blood obtained either before or after invasive procedure.