Detection of a virus related to betacoronaviruses in Italian greater horseshoe bats.

SUMMARYThe association between coronaviruses and bats is a worldwide phenomenon and bats belonging to genus Rhinolophus are the reservoir host for several coronaviruses, including a large number of viruses closely related genetically to severe acute respiratory syndrome-coronavirus (SARS-CoV). We carried out a survey in colonies of Italian bats (Rhinolophus ferrumequinum) for the presence of coronaviruses. Two of 52 R. ferrumequinum captured from different Italian areas tested positive by reverse transcription–PCR for a fragment of RNA-dependent RNA polymerase (RdRp) gene of viruses related to Coronavirus. Phylogenetic analysis revealed close correlations between one of the positive samples and SARS-related CoV belonging to the genus Betacoronavirus

A Real-Time PCR Assay for Bat SARS-Like Coronavirus Detection and Its Application to Italian Greater Horseshoe Bat Faecal Sample Surveys

Bats are source of coronaviruses closely related to the severe acute respiratory syndrome (SARS) virus. Numerous studies have been carried out to identify new bat viruses related to SARS-coronavirus (bat-SARS-like CoVs) using a reverse-transcribed-polymerase chain reaction assay. However, a qualitative PCR could underestimate the prevalence of infection, affecting the epidemiological evaluation of bats in viral ecology. In this work an SYBR Green-real time PCR assay was developed for diagnosing infection with SARS-related coronaviruses from bat guano and was applied as screening tool in a survey carried out on 45 greater horseshoe bats (Rhinolophus ferrumequinum) sampled in Italy in 2009. The assay showed high sensitivity and reproducibility. Its application on bats screening resulted in a prevalence of 42%. This method could be suitable as screening tool in epidemiological surveys about the presence of bat-SARS-like CoVs, consequently to obtain a more realistic scenario of the viral prevalence in the population

Genetic complexity and multiple infections with more Parvovirus species in naturally infected cats

Parvoviruses of carnivores include three closely related autonomous parvoviruses: canine parvovirus (CPV), feline panleukopenia virus (FPV) and mink enteritis virus (MEV). These viruses cause a variety of serious diseases, especially in young patients, since they have a remarkable predilection for replication in rapidly dividing cells. FPV is not the only parvovirus species which infects cats; in addition to MEV, the new variants of canine parvovirus, CPV-2a, 2b and 2c have also penetrated the feline host-range, and they are able to infect and replicate in cats, causing diseases indistinguishable from feline panleukopenia. Furthermore, as cats are susceptible to both CPV-2 and FPV viruses, superinfection and co-infection with multiple parvovirus strains may occur, potentially facilitating recombination and high genetic heterogeneity. In the light of the importance of cats as a potential source of genetic diversity for parvoviruses and, since feline panleukopenia virus has re-emerged as a major cause of mortality in felines, the present study has explored the molecular characteristics of parvovirus strains circulating in cat populations. The most significant findings reported in this study were (a) the detection of mixed infection FPV/CPV with the presence of one parvovirus variant which is a true intermediate between FPV/CPV and (b) the quasispecies cloud size of one CPV sample variant 2c. In conclusion, this study provides new important results about the evolutionary dynamics of CPV infections in cats, showing that CPV has presumably started a new process of readaptation in feline hosts

Chloride intracellular channel 1 activity is not required for glioblastoma development but its inhibition dictates glioma stem cell responsivity to novel biguanide derivatives

Background: Chloride intracellular channel-1 (CLIC1) activity controls glioblastoma proliferation. Metformin exerts antitumor effects in glioblastoma stem cells (GSCs) inhibiting CLIC1 activity, but its low potency hampers its translation in clinical settings. Methods: We synthesized a small library of novel biguanide-based compounds that were tested as antiproliferative agents for GSCs derived from human glioblastomas, in vitro using 2D and 3D cultures and in vivo in the zebrafish model. Compounds were compared to metformin for both potency and efficacy in the inhibition of GSC proliferation in vitro (MTT, Trypan blue exclusion assays, and EdU labeling) and in vivo (zebrafish model), migration (Boyden chamber assay), invasiveness (Matrigel invasion assay), self-renewal (spherogenesis assay), and CLIC1 activity (electrophysiology recordings), as well as for the absence of off-target toxicity (effects on normal stem cells and toxicity for zebrafish and chick embryos). Results: We identified Q48 and Q54 as two novel CLIC1 blockers, characterized by higher antiproliferative potency than metformin in vitro, in both GSC 2D cultures and 3D spheroids. Q48 and Q54 also impaired GSC self-renewal, migration and invasion, and displayed low systemic in vivo toxicity. Q54 reduced in vivo proliferation of GSCs xenotransplanted in zebrafish hindbrain. Target specificity was confirmed by recombinant CLIC1 binding experiments using microscale thermophoresis approach. Finally, we characterized GSCs from GBMs spontaneously expressing low CLIC1 protein, demonstrating their ability to grow in vivo and to retain stem-like phenotype and functional features in vitro. In these GSCs, Q48 and Q54 displayed reduced potency and efficacy as antiproliferative agents as compared to high CLIC1-expressing tumors. However, in 3D cultures, metformin and Q48 (but not Q54) inhibited proliferation, which was dependent on the inhibition dihydrofolate reductase activity. Conclusions: These data highlight that, while CLIC1 is dispensable for the development of a subset of glioblastomas, it acts as a booster of proliferation in the majority of these tumors and its functional expression is required for biguanide antitumor class-effects. In particular, the biguanide-based derivatives Q48 and Q54, represent the leads to develop novel compounds endowed with better pharmacological profiles than metformin, to act as CLIC1-blockers for the treatment of CLIC1-expressing glioblastomas, in a precision medicine approach

Custom CGH array profiling of copy number variations (CNVs) on chromosome 6p21.32 (HLA locus) in patients with venous malformations associated with multiple sclerosis

<p>Abstract</p> <p>Background</p> <p>Multiple sclerosis (MS) is a complex disorder thought to result from an interaction between environmental and genetic predisposing factors which have not yet been characterised, although it is known to be associated with the HLA region on 6p21.32. Recently, a picture of chronic cerebrospinal venous insufficiency (CCSVI), consequent to stenosing venous malformation of the main extra-cranial outflow routes (VM), has been described in patients affected with MS, introducing an additional phenotype with possible pathogenic significance.</p> <p>Methods</p> <p>In order to explore the presence of copy number variations (CNVs) within the HLA locus, a custom CGH array was designed to cover 7 Mb of the HLA locus region (6,899,999 bp; chr6:29,900,001-36,800,000). Genomic DNA of the 15 patients with CCSVI/VM and MS was hybridised in duplicate.</p> <p>Results</p> <p>In total, 322 CNVs, of which 225 were extragenic and 97 intragenic, were identified in 15 patients. 234 known polymorphic CNVs were detected, the majority of these being situated in non-coding or extragenic regions. The overall number of CNVs (both extra- and intragenic) showed a robust and significant correlation with the number of stenosing VMs (Spearman: r = 0.6590, p = 0.0104; linear regression analysis r = 0.6577, p = 0.0106).</p> <p>The region we analysed contains 211 known genes. By using pathway analysis focused on angiogenesis and venous development, MS, and immunity, we tentatively highlight several genes as possible susceptibility factor candidates involved in this peculiar phenotype.</p> <p>Conclusions</p> <p>The CNVs contained in the HLA locus region in patients with the novel phenotype of CCSVI/VM and MS were mapped in detail, demonstrating a significant correlation between the number of known CNVs found in the HLA region and the number of CCSVI-VMs identified in patients. Pathway analysis revealed common routes of interaction of several of the genes involved in angiogenesis and immunity contained within this region. Despite the small sample size in this pilot study, it does suggest that the number of multiple polymorphic CNVs in the HLA locus deserves further study, owing to their possible involvement in susceptibility to this novel MS/VM plus phenotype, and perhaps even other types of the disease.</p

31st Annual Meeting and Associated Programs of the Society for Immunotherapy of Cancer (SITC 2016) : part two

Background The immunological escape of tumors represents one of the main ob- stacles to the treatment of malignancies. The blockade of PD-1 or CTLA-4 receptors represented a milestone in the history of immunotherapy. However, immune checkpoint inhibitors seem to be effective in specific cohorts of patients. It has been proposed that their efficacy relies on the presence of an immunological response. Thus, we hypothesized that disruption of the PD-L1/PD-1 axis would synergize with our oncolytic vaccine platform PeptiCRAd. Methods We used murine B16OVA in vivo tumor models and flow cytometry analysis to investigate the immunological background. Results First, we found that high-burden B16OVA tumors were refractory to combination immunotherapy. However, with a more aggressive schedule, tumors with a lower burden were more susceptible to the combination of PeptiCRAd and PD-L1 blockade. The therapy signifi- cantly increased the median survival of mice (Fig. 7). Interestingly, the reduced growth of contralaterally injected B16F10 cells sug- gested the presence of a long lasting immunological memory also against non-targeted antigens. Concerning the functional state of tumor infiltrating lymphocytes (TILs), we found that all the immune therapies would enhance the percentage of activated (PD-1pos TIM- 3neg) T lymphocytes and reduce the amount of exhausted (PD-1pos TIM-3pos) cells compared to placebo. As expected, we found that PeptiCRAd monotherapy could increase the number of antigen spe- cific CD8+ T cells compared to other treatments. However, only the combination with PD-L1 blockade could significantly increase the ra- tio between activated and exhausted pentamer positive cells (p= 0.0058), suggesting that by disrupting the PD-1/PD-L1 axis we could decrease the amount of dysfunctional antigen specific T cells. We ob- served that the anatomical location deeply influenced the state of CD4+ and CD8+ T lymphocytes. In fact, TIM-3 expression was in- creased by 2 fold on TILs compared to splenic and lymphoid T cells. In the CD8+ compartment, the expression of PD-1 on the surface seemed to be restricted to the tumor micro-environment, while CD4 + T cells had a high expression of PD-1 also in lymphoid organs. Interestingly, we found that the levels of PD-1 were significantly higher on CD8+ T cells than on CD4+ T cells into the tumor micro- environment (p < 0.0001). Conclusions In conclusion, we demonstrated that the efficacy of immune check- point inhibitors might be strongly enhanced by their combination with cancer vaccines. PeptiCRAd was able to increase the number of antigen-specific T cells and PD-L1 blockade prevented their exhaus- tion, resulting in long-lasting immunological memory and increased median survival

The Vineland-II in Preschool Children with Autism Spectrum Disorders: An Item Content Category Analysis

We investigated which item subsets of the Vineland-II can discriminate low-functioning preschoolers with ASD from matched peers with other neurodevelopmental disorders, using a regression analysis derived from a normative sample to account for cognitive and linguistic competencies. At variance with the typical profile, a pattern with Communication more impaired than Socialization was observed. The source of the frequently reported Socialization delay in ASD appears to be in Playing and Imitating skills only, not in other social adaptive behavior skills. The combination of item subsets Playing, Following instructions, Beginning to talk, and Speech skills provided the best discrimination between the two clinical groups. Evaluation of the Vineland-II score on item content categories is a useful procedure for a more efficient clinical description

Vineland-II. Vineland Adaptive Behavior Scales Second Edition- Survey Form - Standardizzazione italiana

Il volume presenta la standardizzazione italiana, su un campione normativo di 2666 individui distribuiti in 27 gruppi di età costituiti da circa 100 persone ciascuno, della versione Survey Form delle Scale Vineland-II di Sparrow. Cicchetti e Balla (2005), che valuta il comportamento adattivo in individui tra 0 e 90 anni di età, nelle aree Comunicazione, Abilità del vivere quotidiano, Socializzazione e Abilità otorie. Tale strumento e le relative norme sono utilizzabili a vari fini, sia in età evolutiva che in adulti ed anziani: 1. per effettuare diagnosi di disabilità intellettiva; 2: per delineare profili di comportamento adattivo in individui con sviluppo tipico e in diversi tipi di popolazioni cliniche (ad es. con sindrome dello spettro autistico, con ADHD, con disturbo della condotta, con demenza, con altri tipi di patologia), 3. per predisporre piani di trattamento; 4. a scopo di ricerca

Vineland Adaptive Behavior Scales, Second Edition (Vineland-II) \u2013 Survey Form. Adattamento italiano

Le Vineland Adaptive Behavior Scales \u2013 Second Edition (Vineland-II), versione Survey Interview Form, valutano il comportamento adattivo di individui con et\ue0 da 0 a 90 anni per mezzo di un\u2019intervista semistrutturata a una persona che li conosce in modo adeguato. Lo strumento \ue8 formato da undici subscale raggruppate nelle quattro scale: Comunicazione, Abilit\ue0 del vivere quotidiano, Socializzazione e Abilit\ue0 motorie. Pur avendo la medesima struttura delle Vineland ABS, ne costituisce una sostanziale revisione. La maggior parte dei nuovi item \ue8 stata aggiunta per migliorare la valutazione di bambini molto piccoli o di adulti. Altri item sono stati introdotti o modificati per migliorare l\u2019accuratezza e la completezza dell\u2019assessment di tutte le fasce d\u2019et\ue0. Le Vineland-II sono utilizzabili per la valutazione delle abilit\ue0 di vita quotidiana in contesti clinici, educativi e di ricerca. Possono essere impiegate per la misurazione del comportamento adattivo nella diagnosi di disabilit\ue0 intellettiva, e come strumento complementare nella diagnosi di vari disturbi e disabilit\ue0, quali i disturbi dello spettro autistico, sindromi genetiche, ritardi dello sviluppo, disturbi nell\u2019area emotiva e comportamentale. Sono anche uno strumento essenziale per la valutazione dello sviluppo di bambini piccoli e per la determinazione della loro ammissibilit\ue0 a interventi precoci

Vineland Adaptive Behavior Scales, Second Edition (Vineland-II) \u2013 Survey Interview Form. Standardizzazione italiana.

Le Vineland Adaptive Behavior Scales \u2013 Second Edition (Vineland-II), versione Survey Interview Form, valutano il comportamento adattivo di individui con et\ue0 da 0 a 90 anni per mezzo di un\u2019intervista semistrutturata a una persona che li conosce in modo adeguato. In questo manuale viene descritto l\u2019adattamento delle Vineland-II Survey Interview Form al contesto italiano e il processo di standardizzazione italiana. La novit\ue0 rispetto all\u2019adattamento italiano delle Vineland ABS \u2013 Intervista \u2013 Forma completa \ue8 costituita dal campione italiano composto da 2666 individui da 0 a 90 anni di et\ue0, con sviluppo tipico e disabilit\ue0 secondo le percentuali note della popolazione italiana e rappresentativo della popolazione generale per et\ue0, genere e area d\u2019Italia in cui risiede (Nord, Centro, Sud e Isole). Questa nuova versione inoltre prevede i punteggi ponderati QI di deviazione per Scala composta e scale, che consentono di rilevare la distanza in unit\ue0 di deviazione standard dalla popolazione generale. In tal modo \ue8 possibile verificare l\u2019eleggibilit\ue0 per la diagnosi di DI e, in accordo con il al DSM-5, la classificazione del disturbo in lieve, moderato, grave/estremo