Next Generation Sequencing for the Analysis of Parvovirus B19 Genomic Diversity

Parvovirus B19 (B19V) is a ssDNA human virus, responsible for an ample range of clinical manifestations. Sequencing of B19V DNA from clinical samples is frequently reported in the literature to assign genotype (genotypes 1-3) and for finer molecular epidemiological tracing. The increasing availability of Next Generation Sequencing (NGS) with its depth of coverage potentially yields information on intrinsic sequence heterogeneity; however, integration of this information in analysis of sequence variation is not routinely obtained. The present work investigated genomic sequence heterogeneity within and between B19V isolates by application of NGS techniques, and by the development of a novel dedicated bioinformatic tool and analysis pipeline, yielding information on two newly defined parameters. The first, alpha-diversity, is a measure of the amount and distribution of position-specific, normalised Shannon Entropy, as a measure of intra-sample sequence heterogeneity. The second, sigma-diversity, is a measure of the amount of inter-sample sequence heterogeneity, also incorporating information on alpha-diversity. Based on these indexes, further cluster analysis can be performed. A set of 24 high-titre viraemic samples was investigated. Of these, 23 samples were genotype 1 and one sample was genotype 2. Genotype 1 isolates showed low alpha-diversity values, with only a few samples showing distinct position-specific polymorphisms; a few genetically related clusters emerged when analysing inter-sample distances, correlated to the year of isolation; the single genotype 2 isolate showed the highest alpha-diversity, even if not presenting polymorphisms, and was an evident outlier when analysing inter-sample distance. In conclusion, NGS analysis and the bioinformatic tool and pipeline developed and used in the present work can be considered effective tools for investigating sequence diversity, an observable parameter that can be incorporated into the quasispecies theory framework to yield a better insight into viral evolution dynamics

Genetic Characterization of Cancer of Unknown Primary Using Liquid Biopsy Approaches

Cancers of unknown primary (CUPs) comprise a heterogeneous group of rare metastatic tumors whose primary site cannot be identified after extensive clinical\u2013pathological investigations. CUP patients are generally treated with empirical chemotherapy and have dismal prognosis. As recently reported, CUP genome presents potentially druggable alterations for which targeted therapies could be proposed. The paucity of tumor tissue, as well as the difficult DNA testing and the lack of dedicated panels for target gene sequencing are further relevant limitations. Here, we propose that circulating tumor cells (CTCs) and circulating tumor DNA (ctDNA) could be used to identify actionable mutations in CUP patients. Blood was longitudinally collected from two CUP patients. CTCs were isolated with CELLSEARCH\uae and DEPArrayTM NxT and Parsortix systems, immunophenotypically characterized and used for single-cell genomic characterization with Ampli1TM kits. Circulating cell-free DNA (ccfDNA), purified from plasma at different time points, was tested for tumor mutations with a CUP-dedicated, 92-gene custom panel using SureSelect Target Enrichment technology. In parallel, FFPE tumor tissue was analyzed with three different assays: FoundationOne CDx assay, DEPArray LibPrep and OncoSeek Panel, and the SureSelect custom panel. These approaches identified the same mutations, when the gene was covered by the panel, with the exception of an insertion in APC gene. which was detected by OncoSeek and SureSelect panels but not FoundationOne. FGFR2 and CCNE1 gene amplifications were detected in single CTCs, tumor tissue, and ccfDNAs in one patient. A somatic variant in ARID1A gene (p.R1276 17) was detected in the tumor tissue and ccfDNAs. The alterations were validated by Droplet Digital PCR in all ccfDNA samples collected during tumor evolution. CTCs from a second patient presented a pattern of recurrent amplifications in ASPM and SEPT9 genes and loss of FANCC. The 92-gene custom panel identified 16 non-synonymous somatic alterations in ccfDNA, including a deletion (I1485Rfs 1719) and a somatic mutation (p. A1487V) in ARID1A gene and a point mutation in FGFR2 gene (p.G384R). Our results support the feasibility of non-invasive liquid biopsy testing in CUP cases, either using ctDNA or CTCs, to identify CUP genetic alterations with broad NGS panels covering the most frequently mutated genes

Parvovirus B19 DNA CpG Dinucleotide Methylation and Epigenetic Regulation of Viral Expression

CpG DNA methylation is one of the main epigenetic modifications playing a role in the control of gene expression. For DNA viruses whose genome has the ability to integrate in the host genome or to maintain as a latent episome, a correlation has been found between the extent of DNA methylation and viral quiescence. No information is available for Parvovirus B19, a human pathogenic virus, which is capable of both lytic and persistent infections. Within Parvovirus B19 genome, the inverted terminal regions display all the characteristic signatures of a genomic CpG island; therefore we hypothesised a role of CpG dinucleotide methylation in the regulation of viral genome expression

Socioeconomic differences and persistent segregation of Italian territories during COVID-19 pandemic

Lockdowns implemented to address the COVID-19 pandemic have disrupted human mobility flows around the globe to an unprecedented extent and with economic consequences which are unevenly distributed across territories, firms and individuals. Here we study socioeconomic determinants of mobility disruption during both the lockdown and the recovery phases in Italy. For this purpose, we analyze a massive data set on Italian mobility from February to October 2020 and we combine it with detailed data on pre-existing local socioeconomic features of Italian administrative units. Using a set of unsupervised and supervised learning techniques, we reliably show that the least and the most affected areas persistently belong to two different clusters. Notably, the former cluster features significantly higher income per capita and lower income inequality than the latter. This distinction persists once the lockdown is lifted. The least affected areas display a swift (V-shaped) recovery in mobility patterns, while poorer, most affected areas experience a much slower (U-shaped) recovery: as of October 2020, their mobility was still significantly lower than pre-lockdown levels. These results are then detailed and confirmed with a quantile regression analysis. Our findings show that economic segregation has, thus, strengthened during the pandemic

Sistemi motivazionali interpersonali in adolescenza

In this work we describe two evaluation methods (PIGA and AIMIT) we used in analysis of interactions between adolescents and their own parents. In our opinion it’s very important in Adolescence to take in account and analyze how interpersonal motivational systems can define the parent-adolescent interactions to understand what kind of situation can improve and subtain the adoescent’s development

Drivers and Barriers to Online Shopping: The Interaction of Product, Consumer, and Retailer Factors

Through a review of the literature, this chapter focuses on three key influences on purchase behavior on the Internet: product, consumer, and retailer factors. Product characteristics and branding not only influence many consumer-related factors (e.g. the need to handle the product and risk perceptions), but also affect retailers\u2019 strategic and tactical online decisions (e.g. the balance between offline and online retail provision and the breadth and depth of products and brands selection). This chapter also examines how consumer-related factors (e.g. consumers\u2019 expertise, attitudes toward the Internet, and shopping orientation) affect online purchasing and the implications for e-retailers. Finally, the chapter discusses how e-retailers marketing efforts (retailer factors) can be used to overcome the barriers to web purchasing resulting from specific product and consumer related characteristics. Clear and easy to implement recommendations to managers are offered

Le ricerche sui comportamenti d’acquisto online: l’orientamento edonico e utilitarista

Da un lato la letteratura ha sottolineato i numerosi vantaggi che Internet potrebbe offrire come canale distributivo, d’altro lato però la diffusione effettiva del canale elettronico è ancora assai lontana dall’eguagliare quella dei canali offline. Dunque, oltre a sottolineare i punti di forza e le potenzialità di Internet, è importante anche cercare di comprendere quali fattori influenzino la propensione dei consumatori a fare acquisti utilizzando il canale elettronico. In questa analisi si considera l’impatto dell’orientamento edonico e utilitarista, il ruolo del prodotto, del rischio percepito e del bisogno di un contatto fisico con il bene prima del suo acquisto. I risultati indicano che 1-il diverso orientamento all’acquisto comporta differenze significative nell’intenzione di ricorrere al canale elettronico, 2-la categoria merceologica gioca un ruolo rilevante, 3-i consumatori preferiscono acquistare online dopo avere interagito fisicamente col prodotto nel canale offline, piuttosto che acquistare direttamente da Internet, anche quando la marca è a loro nota. Si discutono quindi le implicazioni teoriche e manageriali