Positive predictive values and outcomes for uninformative cell-free DNA tests: An Italian multicentric Cytogenetic and cytogenomic Audit of diagnOstic testing (ICARO study).

OBJECTIVES: To establish the positive predictive values (PPV) of cfDNA testing based on data from a nationwide survey of independent clinical cytogenetics laboratories. METHODS: Prenatal diagnostic test results obtained by Italian laboratories between 2013 and March 2020 were compiled for women with positive non-invasive prenatal tests (NIPT), without an NIPT result, and cases where there was sex discordancy between the NIPT and ultrasound. PPV and other summary data were reviewed. RESULTS: Diagnostic test results were collected for 1327 women with a positive NIPT. The highest PPVs were for Trisomy (T) 21 (624/671, 93%) and XYY (26/27, 96.3%), while rare autosomal trisomies (9/47, 19.1%) and recurrent microdeletions (8/55, 14.5%) had the lowest PPVs. PPVs for T21, T18, and T13 were significantly higher when diagnostic confirmation was carried out on chorionic villi (97.5%) compared to amniotic fluid (89.5%) (p < 0.001). In 19/139 (13.9%), of no result cases, a cytogenetic abnormality was detected. Follow-up genetic testing provided explanations for 3/6 cases with a fetal sex discordancy between NIPT and ultrasound. CONCLUSIONS: NIPT PPVs differ across the conditions screened and the tissues studied in diagnostic testing. This variability, issues associated with fetal sex discordancy, and no results, illustrate the importance of pre- and post-test counselling.

Evaluation of Two-Assay Serological Testing Strategies for Anti-HCV Screening in Italian Populations: A Dual Screening Approach.

(1) Background: Hepatitis C virus (HCV) screening mostly uses a one-assay anti-HCV testing approach, which has a higher probability of false-positive results in populations with low HCV prevalence. (2) Methods: In this investigation, 17,926 participants were screened for HCV, and the reactives were tested using a two-assay anti-HCV approach: Elecsys ElectroChemiLuminescence (ECL) and a ChemiLuminescence ImmunoAssay (CLIA), respectively. A recombinant immunoblot assay (RIBA) was performed to confirm anti-HCV positivity. Statistical analysis was performed. (3) Results: A total of 350 specimens were reactive in the ECL screening, of which CLIA retesting showed that 292 (83.4%) were anti-HCV reactive (283 positives, 9 indeterminate, none negative by RIBA), but 58 (16.6%) were not anti-HCV reactive (15 positive, 12 indeterminate, 31 negatives by RIBA). The two-assay strategy significantly improved the positive predictive value (PPV: 95.00%) with χ2: 7.59 (p < 0.01) compared to the PPV assessed by one assay (PPV: 90.6%) with χ2: 34.51 (p < 0.001). The ROC curve defined a sensibility and specificity for the dual approach of 99.66% and 100.00%. (4) Conclusions: Compared with a one-assay testing strategy, the two-assay testing strategy may significantly reduce false positives in anti-HCV testing and identify inactive HCV infection in low seroprevalence populations.

Efficacy of Fluorecare SARS-CoV-2 Spike Protein Test Kit for SARS-CoV-2 detection in nasopharyngeal samples of 121 individuals working in a manufacturing company.

OBJECTIVES: The aim of this study was to evaluate the clinical performance of the Fluorecare SARS-CoV-2 Spike Protein Test Kit, a rapid immunochromatographic assay for SARS-CoV-2 detection. Moreover, we sought to point out the strategy adopted by a local company to lift the lockdown without leading to an increase in the number of COVID-19 cases, by performing a precise and timely health surveillance. METHODS: The rapid Fluorecare SARS-CoV-2 Spike Protein Test was performed immediately after sampling following the manufacturer's instructions. RT-PCRs were performed within 24 hours of specimen collection. A total amount of 253 nasopharyngeal samples from 121 individuals were collected between March 16 and April 2, 2021 and tested. RESULTS: Of 253 nasopharyngeal samples, 11 (9.1%) were positive and 242 (90.9%) were negative for SARS-CoV-2 RNA by RT-PCR assays. The rapid SARS-CoV-2 antigen detection test's mean sensitivity and specificity were 84,6% (95% CI, 54.6-98.1%) and 100% (95% CI, 98.6-100%), respectively. Two false negative test results were obtained from samples with high RT-PCR cycle threshold (Ct). CONCLUSION: Our study suggested that Fluorecare SARS-CoV-2 Spike Protein Test can be introduced into daily diagnostic practice, as its mean sensitivity and specificity follow the standards recommended by WHO and IFCC Task Force. In addition, we underlined how the strategy adopted by a local company to risk assessment and health surveillance was appropriate for infection containment. This real-life scenario gave us the possibility to experience potential approaches aimed to preserve public health and work activities.

Cytogenetic and Array-CGH Characterization of a Simple Case of Reciprocal t(3;10) Translocation Reveals a Hidden Deletion at 5q12.

Chromosome deletions, including band 5q12, have rarely been reported and have been associated with a wide range of clinical manifestations, such as postnatal growth retardation, intellectual disability, hyperactivity, nonspecific ocular defects, facial dysmorphism, and epilepsy. In this study, we describe for the first time a child with growth retardation in which we identified a balanced t(3;10) translocation by conventional cytogenetic analysis in addition to an 8.6 Mb 5q12 deletion through array-CGH. Our results show that the phenotypic abnormalities of a case that had been interpreted as "balanced" by conventional cytogenetics are mainly due to a cryptic deletion, highlighting the need for molecular investigation in subjects with an abnormal phenotype before assuming the cause is an apparently simple cytogenetic rearrangement. Finally, we identify PDE4D and PIK3R1 genes as the two major candidates responsible for the clinical features expressed in our patient.

Risks of COVID-19 transmission in blood and serum during surgery A prospective cross-sectional study from a single dedicated COVID-19 center.

The present pandemic caused by the SARS COV-2 coronavirus is still ongoing, although it is registered a slowdown in the spread for new cases. The main environmental route of transmission of SARS-CoV-2 is through droplets and fomites or surfaces, but there is a potential risk of virus spread also in smaller aerosols during various medical procedures causing airborne transmission. To date, no information is available on the risk of contagion from the peritoneal fluid with which surgeons can come into contact during the abdominal surgery on COVID-19 patients. We have investigated the presence of SARS-CoV-2 RNA in the peritoneal cavity of patients affected by COVID-19, intraoperatively and postoperatively. KEY WORDS: Covid-19, Laparotomy, Surgery.

Insertional translocation involving an additional nonchromothriptic chromosome in constitutional chromothripsis: Rule or exception?

BACKGROUND: Chromothripsis, which is the local massive shattering of one or more chromosomes and their reassembly in a disordered array with frequent loss of some fragments, has been mainly reported in association with abnormal phenotypes. We report three unrelated healthy persons, two of which parenting a child with some degree of intellectual disability, carrying a chromothripsis involving respectively one, two, and three chromosomes, which was detected only after whole-genome sequencing. Unexpectedly, in all three cases a fragment from one of the chromothripsed chromosomes resulted to be inserted within a nonchromothripsed one. METHODS: Conventional cytogenetic techniques, paired-end whole-genome sequencing, polymerase chain reaction, and Sanger sequencing were used to characterize complex rearrangements, copy-number variations, and breakpoint sequences in all three families. RESULTS: In two families, one parent was carrier of a balanced chromothripsis causing in the index case a deletion and a noncontiguous duplication at 3q in case 1, and a t(6;14) translocation associated with interstitial 14q deletion in case 2. In the third family, an unbalanced chromothripsis involving chromosomes 6, 7, and 15 was inherited to the proband by the mosaic parent. In all three parents, the chromothripsis was concurrent with an insertional translocation of a portion of one of the chromothriptic chromosomes within a further chromosome that was not involved in the chromothripsis event. CONCLUSION: Our findings show that (a) both simple and complex unbalanced rearrangements may result by the recombination of a cryptic parental balanced chromothripsis and that (b) insertional translocations are the spy of more complex rearrangements and not simply a three-breakpoint event.

Molecular cytogenetics characterization of seven small supernumerary marker chromosomes derived from chromosome 19: Genotype-phenotype correlation and review of the literature.

Only a few subjects carrying supernumerary marker chromosomes derived from 19 chromosome (sSMC(19)) have been described to date and for a small portion of them the genic content has been defined at the molecular level. We present seven new different sSMCs(19) identified in eight individuals, seven of whom unrelated. The presence of the sSMC is associated with a clinical phenotype in five subjects, while the other three carriers, two of whom related, are normal. All sSMCs(19) have been characterized by means of conventional and molecular cytogenetics. We compare the sSMCs(19) carriers with a clinical phenotype to already described patients with gains (sSMCs or microduplications) of overlapping genomic regions with the aim to deepen the pathogenicity of the encountered imbalances and to assess the role of the involved genes on the phenotype. The present work supports the correlation between the gain of some chromosome 19 critical regions and specific phenotypes.

Testis development in the absence of SRY: chromosomal rearrangements at SOX9 and SOX3.

Duplications in the ~2 Mb desert region upstream of SOX9 at 17q24.3 may result in familial 46,XX disorders of sex development (DSD) without any effects on the XY background. A balanced translocation with its breakpoint falling within the same region has also been described in one XX DSD subject. We analyzed, by conventional and molecular cytogenetics, 19 novel SRY-negative unrelated 46,XX subjects both familial and sporadic, with isolated DSD. One of them had a de novo reciprocal t(11;17) translocation. Two cases carried partially overlapping 17q24.3 duplications ~500 kb upstream of SOX9, both inherited from their normal fathers. Breakpoints cloning showed that both duplications were in tandem, whereas the 17q in the reciprocal translocation was broken at ~800 kb upstream of SOX9, which is not only close to a previously described 46,XX DSD translocation, but also to translocations without any effects on the gonadal development. A further XX male, ascertained because of intellectual disability, carried a de novo cryptic duplication at Xq27.1, involving SOX3. CNVs involving SOX3 or its flanking regions have been reported in four XX DSD subjects. Collectively in our cohort of 19 novel cases of SRY-negative 46,XX DSD, the duplications upstream of SOX9 account for ~10.5% of the cases, and are responsible for the disease phenotype, even when inherited from a normal father. Translocations interrupting this region may also affect the gonadal development, possibly depending on the chromatin context of the recipient chromosome. SOX3 duplications may substitute SRY in some XX subjects.

Silver-Russell syndrome and Beckwith-Wiedemann syndrome phenotypes associated with 11p duplication in a single family.

Genomic imprinting is an epigenetic phenomenon resulting in differential expression of maternal and paternal alleles of a subset of genes. In the mouse, mutation of imprinted genes often results in contrasting phenotypes, depending on parental origin. The overgrowth-associated Beckwith-Wiedemann syndrome (BWS) and the growth restriction-associated Silver-Russell syndrome (SRS) have been linked with a variety of epigenetic and genetic defects affecting a cluster of imprinted genes at chromosome 11p15.5. Paternally derived and maternally derived 11p15.5 duplications represent infrequent findings in BWS and SRS, respectively. Here, we report a case in which a 6.5 Mb duplication of 11p15.4-pter resulted in SRS and BWS phenotypes in a child and her mother, respectively. Molecular analyses demonstrated that the duplication involved the maternal chromosome 11p15 in the child and the paternal chromosome 11p15 in the mother. This observation provides a direct demonstration that SRS and BWS represent specular images, both at the clinical and molecular levels.

De novo balanced chromosome rearrangements in prenatal diagnosis.

OBJECTIVE: We surveyed the datasheets of 29 laboratories concerning prenatal diagnosis of de novo apparently balanced chromosome rearrangements to assess the involvement of specific chromosomes, the breakpoints distribution and the impact on the pregnancy outcome. METHOD: By means of a questionnaire, data on 269.371 analyses performed from 1983 to 2006 on amniotic fluid, chorionic villus and fetal blood samples were collected. RESULTS: A total of 246 balanced anomalies were detected at frequencies of 72% for reciprocal translocations, 18% for Robertsonian translocations, 7% for inversions and 3% for complex chromosome rearrangements. The total frequencies of balanced rearrangements were 0.09%, 0.08% and 0.05% on amniotic fluid, chorionic villus and fetal blood samples. CONCLUSION: A preferential involvement of chromosomes 22, 7, 21, 3, 9 and 11 and a less involvement of chromosomes X, 19, 12, 6 and 1 was observed. A nonrandom distribution of the breakpoints across chromosomes was noticed. Association in the location of recurrent breakpoints and fragile sites was observed for chromosomes 11, 7, 10 and 22, while it was not recorded for chromosome 3. The rate of pregnancy termination was about 20%, with frequencies decreasing from complex chromosomal rearrangements (33%), reciprocal translocations (24%) to inversions (11%) and Robertsonian translocations (3%).