Functional dna nanostructures for in vitro biosensing in live cells

DNA is a fascinating biomolecule that is well known for its genetic role in living systems. The emerging area of DNA nanotechnology provides an alternative view that exploits unparallel self-assembly ability of DNA molecules for material use of DNA. Although many reports exist on the results of DNA self-assembling systems, still few of them focus on the in vitro study about the function of such DNA nanostructures in live cells. Due to this, there are still a limited research about the in vitro functionality of such designs. To address an aspect of this issue, we have designed, synthesized and characterized two multifunctional fluorescencent nanobiosensors by DNA self-assembling. Each structure was designed and implemented to be introduced in live cells in order to give information on their functioning in real-time. Computational tools were used in order to design a graphic model of two new DNA motifs and also to obtain the specific sequences to all the ssDNA molecules. By thermal self-assembly techniques we have successfully synthesized the structure and corroborate their formation by the PAGE technique. In addition, we have established the conditions to characterize their structural conformation change when they perform their sensor response. The sensing behavior was also accomplished by fluorescence spectroscopy techniques; FRET evaluation and fluorescence microscopy imaging. Providing the evidence about their adequate sensing performance outside and inside the cells detected in real-time. In a preliminary evaluation we have tried to show the in vitro functionality of our structures in different cancer cell lines with the ability to perform local sensing responses. Our findings suggest that DNA sensor nanostructures could serve as a platform to exploit further therapeutic achievements in live cells.

L'interpretazione a distanza: presentazione della piattaforma veasyt live!, case study italiano e proposta di seminario.

Il presente elaborato offre una panoramica generale del Remote Interpreting (RI) con l'obiettivo di strutturare un ipotetico seminario da tenere alla SLLTI (Scuola di Lingue e Letterature, Traduzione e Interpretazione, ex SSLMIT) di Forlì, in collaborazione con l'azienda VEASYT, spin-off dell’università Ca' Foscari di Venezia, per introdurre l'argomento agli studenti del corso di laurea in Interpretazione di Conferenza. A tal scopo la tesi è stata suddivisa in dieci capitoli, ognuno dei quali analizza una sfaccettatura dell'interpretazione da remoto; la prima parte verte sull'evoluzione storica delle nuove tecnologie nel mondo dell'interpretariato, con particolare attenzione all'introduzione degli impianti per l'interpretazione simultanea e delle Tecnologie dell'Informazione e della Comunicazione (TIC); si presentano inoltre i principali tipi di interpretazione a distanza, seguendo la loro comparsa in ordine cronologico, e i dispositivi necessari per effettuare una sessione di video-interpretazione. Nella seconda parte si descrivono alcuni ambiti di applicazione del RI (RI per non udenti, in ambito giuridico, in ambito medico-sanitario) e si illustra la situazione a livello mondiale passando in rassegna, a titolo di esempio, alcune aziende presenti in Australia, Stati Uniti, Germania, Austria e Gran Bretagna. Nella terza parte si presentano alcuni regolamenti e disposizioni raccomandati da associazioni di categoria e dalle istituzioni europee e si analizzano i principali vantaggi e svantaggi del RI. Nella quarta parte si mettono a confronto le caratteristiche e il tipo di formazione di un interprete che lavora in situ con quelle di un interprete che lavora a distanza, illustrando alcune strategie sviluppate dagli interpreti durante le sessioni di RI. L'ultima parte riguarda l'azienda VEASYT, la quale viene presentata come case study italiano, descrivendo la sua piattaforma per il servizio di interpretariato da remoto e la modalità di reclutamento degli interpreti. In conclusione si trova la proposta di seminario da tenere ipoteticamente alla SLLTI di Forlì, che viene articolata seguendo la successione dei capitoli precedenti.

Vaccine hypersensitivity--update and overview

Concerns about possible reactions to vaccines or vaccinations are frequently raised. However, the rate of reported vaccine-induced adverse events is low and ranges between 4.8-83.0 per 100,000 doses of the most frequently used vaccines. The number of true allergic reactions to routine vaccines is not known; estimations range from 1 per 500,000 to 1 per 1,000,000 doses for most vaccines. When allergens such as gelatine or egg proteins are components of the formulation, the rate for serious allergic reactions may be higher. Nevertheless, anaphylactic, potentially life-threatening reactions to vaccines are still a rare event (approximately 1 per 1,500,000 doses). The variety of reported vaccine-related adverse events is broad. Most frequently, reactions to vaccines are limited to the injection site and result from a non specific activation of the inflammatory system by, for example, aluminium salts or the active microbial components. If allergy is suspected, an accurate examination followed by algorithms is the key for correct diagnosis, treatment and the decision regarding revaccination in patients with immediate-type reactions to vaccines.

Multiple colonization with S. pneumoniae before and after introduction of the seven-valent conjugated pneumococcal polysaccharide vaccine

Simultaneous carriage of more than one strain of Streptococcus pneumoniae promotes horizontal gene transfer events and may lead to capsule switch and acquisition of antibiotic resistance. We studied the epidemiology of cocolonization with S. pneumoniae before and after introduction of the seven-valent conjugated pneumococcal vaccine (PCV7).

Confocal imaging protocols for live/dead staining in three-dimensional carriers

In tissue engineering, a variety of methods are commonly used to evaluate survival of cells inside tissues or three-dimensional (3D) carriers. Among these methods confocal laser scanning microscopy opened accessibility of 3D tissue using live cell imaging into the tissue or 3D scaffolds. However, although this technique is ideally applied to 3D tissue or scaffolds with thickness up to several millimetres, this application is surprisingly rare and scans are often done on slices with thickness <20 μm. Here, we present novel protocols for the staining of 3D tissue (e.g. intervertebral disc tissue) and scaffolds, such as fibrin gels or alginate beads.

Comparing pneumococcal conjugate vaccine schedules based on 3 and 2 primary doses: systematic review and meta-analysis

Background Pneumococcal conjugate vaccines (PCV) were first licensed for use with 3 primary doses in infancy and a booster dose. The evidence for the effects of different schedules was examined in this systematic review and meta-analysis. Methods We searched 12 databases and trial registers up to March 2010. We selected randomised controlled trials (RCTs), cohort and case–control studies making direct comparisons between PCV schedules with (2p) or (3p) primary doses, with (+1) or without (+0) a booster dose. We extracted data on clinical, nasopharyngeal carriage and immunological outcomes and used meta-analysis to combine results where appropriate. Results Seropositivity levels (antibody concentration ≥0.35 μg/ml) following 3p and 2p PCV schedules were high for most serotypes (5 RCTs). Differences between schedules were generally small and tended to favour 3p schedules, particularly for serotypes 6B and 23F; between-study heterogeneity was high. Seropositivity levels following 3p+1 and 2p+1 schedules were similar but small differences favouring 3p+1 schedules were seen for serotypes 6B and 23F. We did not identify any RCTs reporting clinical outcomes for these comparisons. In 2 RCTs there was weak evidence of a reduction in carriage of S. pneumoniae serotypes included in the vaccine when 3p+0 schedules were compared to 2p+0 at 6 months of age. Conclusions Most data about the relative effects of different PCV schedules relate to immunological outcomes. Both 3p and 2p schedules result in high levels of seropositivity. The clinical relevance of differences in immunological outcomes between schedules is not known. There is an absence of clinical outcome data from RCTs with direct comparisons of any 2p with any 3p PCV schedule.

Measles vaccination in HIV-infected children: systematic review and meta-analysis of safety and immunogenicity

Background. Measles control may be more challenging in regions with a high prevalence of HIV infection. HIV-infected children are likely to derive particular benefit from measles vaccines because of an increased risk of severe illness. However, HIV infection can impair vaccine effectiveness and may increase the risk of serious adverse events after receipt of live vaccines. We conducted a systematic review to assess the safety and immunogenicity of measles vaccine in HIV-infected children. Methods. The authors searched 8 databases through 12 February 2009 and reference lists. Study selection and data extraction were conducted in duplicate. Meta-analysis was conducted when appropriate. Results. Thirty-nine studies published from 1987 through 2008 were included. In 19 studies with information about measles vaccine safety, more than half reported no serious adverse events. Among HIV-infected children, 59% (95% confidence intervals [CI], 46–71%) were seropositive after receiving standard-titer measles vaccine at 6 months (1 study), comparable to the proportion of seropositive HIV-infected children vaccinated at 9 (8 studies) and 12 months (10 studies). Among HIV-exposed but uninfected and HIV-unexposed children, the proportion of seropositive children increased with increasing age at vaccination. Fewer HIV-infected children were protected after vaccination at 12 months than HIV-exposed but uninfected children (relative risk, 0.61; 95% CI, .50–.73). Conclusions. Measles vaccines appear to be safe in HIV-infected children, but the evidence is limited. When the burden of measles is high, measles vaccination at 6 months of age is likely to benefit children of HIV-infected women, regardless of the child's HIV infection status.

An attenuated Salmonella enterica serovar Typhimurium strain lacking the ZnuABC transporter induces protection in a mouse intestinal model of Salmonella infection

Salmonella enterica serovar Typhimurium has long been recognised as a zoonotic pathogen of economic significance in animals and humans. Attempts to protect humans and livestock may be based on immunization with vaccines aimed to induce a protective response. We recently demonstrated that the oral administration of a Salmonella enterica serovar Typhimurium strain unable to synthesize the zinc transporter ZnuABC is able to protect mice against systemic salmonellosis induced by a virulent homologous challenge. This finding suggested that this mutant strain could represent an interesting candidate vaccine for mucosal delivery. In this study, the protective effect of this Salmonella strain was tested in a streptomycin-pretreated mouse model of salmonellosis that is distinguished by the capability of evoking typhlitis and colitis. The here reported results demonstrate that mice immunized with Salmonella enterica serovar Typhimurium (S. Typhimurium) SA186 survive to the intestinal challenge and, compared to control mice, show a reduced number of virulent bacteria in the gut, with milder signs of inflammation. This study demonstrates that the oral administration a of S. Typhimurium strain lacking ZnuABC is able to elicit an effective immune response which protects mice against intestinal S. Typhimurium infection. These results, collectively, suggest that the streptomycin-pretreated mouse model of S. typhimurium infection can represent a valuable tool to screen S. typhimurium attenuated mutant strains and potentially help to assess their protective efficacy as potential live vaccines.