Prion diseases are a group of rapidly progressing neurodegenerative disorders caused by the misfolding of the endogenous prion protein (PrP
) into a pathogenic form (PrP
). This process, despite ...being the central event underlying these disorders, remains largely unknown at a molecular level, precluding the prediction of new potential outbreaks or interspecies transmission incidents. In this work, we present a method to generate bona fide recombinant prions de novo, allowing a comprehensive analysis of protein misfolding across a wide range of prion proteins from mammalian species. We study more than 380 different prion proteins from mammals and classify them according to their spontaneous misfolding propensity and their conformational variability. This study aims to address fundamental questions in the prion research field such as defining infectivity determinants, interspecies transmission barriers or the structural influence of specific amino acids and provide invaluable information for future diagnosis and therapy applications.
Coxiella burnetii is an intracellular pathogenic bacterium and etiological agent of Q fever in humans. Recently, the bacterium has been set free from the strictly intracellular condition by ...successful cultivation in acidified citrate cysteine medium. Here, we report a bacterial cell counting method that allows rapid quantification of the absolute or relative number of live cells of C. burnetii in a high throughput manner. The method utilizes TaqMan-based quantitative polymerase chain reaction (qPCR) targeting a single dotA gene for determination of genome equivalent (GE) presented either as DNA or complementary DNA (cDNA) synthesized via reverse transcription. The assay was shown to be specific, sensitive and efficiently reproducible. The quantification was linear over a range of 30 to 3x108 copies. Since there is only one copy of the dotA gene per Coxiella chromosome, the calculated dotA copy numbers can be compared to the number of bacterial cells. Finally, we demonstrated the potential of the method to assess effects of antibiotic on cell viability and to determine the antibiotic-tolerant fraction within a cell population. Keywords: Coxiella burnetii; Q fever; real-time polymerase chain reaction; copy number; antibiotic; axenic media; dotA gene.
•The prevalence of OCBs in a cohort of Cuban MS patients was 87%.•OCB frequency in Cuban MS patients was slightly lower, but close to Caucasians.•A relationship between OCB prevalence and latitude ...might be present in Latin America.
Background. The diagnostic sensitivity of CSF specific oligoclonal bands (OCBs) in multiple sclerosis (MS), using state of the art methods, has been clearly established to be over 95% in patients with a predominantly Caucasian background. This is not the case for other geographical regions, where reports of OCB prevalence can be much lower, and a relationship between OCB frequency and latitude has been suggested. Objective. The aim of the present study was to assess the frequency of OCBs in a cohort of MS patients evaluated at the Institute of Neurology and Neurosurgery (Havana, Cuba), and to review the scientific literature in order to investigate the possible relationship between OCB status and latitude in the region of Latin America. Methods. Fifty-three patients (47 with definite MS and 6 with clinically isolated syndrome – CIS) were included. Isoelectric focusing (IEF) with IgG immunoblotting for OCB analyses, was performed placing paired CSF and serum samples in the same analytical run. PubMed, Scielo and Google Scholar were searched for papers containing information concerning CSF OCB status (employing isoelectric focusing with IgG immunoblotting) in patients with definite MS in Latin America and the Caribbean. Results. In Cuban patients with definite MS, an OCB prevalence of 87% was observed, while the frequency in CIS patients was lower (67%). The prevailing pattern was that of OCBs restricted to the CSF (type 2), which was observed in 71% of definite MS patients and in all CIS patients with intrathecal IgG synthesis. OCB prevalence was slightly lower, but very close to that reported in Caucasian populations. Comparison with other Latin American countries revealed a significant correlation between OCB prevalence and latitude. Conclusions. A prevalence of CSF restricted OCBs of 87% was observed in definite MS patients, a frequency which was slightly lower, but similar to that reported in Caucasian populations. The analysis of OCB frequency in Latin American countries revealed a possible relationship between OCB prevalence and latitude, but this must be further investigated in more countries and larger samples of patients.
Hydrogels are polymeric biomaterials characterised by their promising biological and biomechanical properties, which make them potential alternatives for use in tendon repair. The aim of the present ...study was to generate in vitro, and determine the therapeutic efficacy in vivo, of novel nanostructured fibrin-based hydrogels to be used as an augmentation strategy for the surgical repair of rat Achilles tendon injuries. Fibrin, fibrin-agarose and fibrin-collagen nanostructured hydrogels (NFH, NFAH and NFCH, respectively) were generated and their biomechanical properties and cell-biomaterial interactions characterised ex vivo. Achilles tendon ruptures were created in 24 adult Wistar rats, which were next treated with direct repair (control group) or direct repair augmented with the generated biomaterials (6 rats/group). After 4 and 8 weeks, the animals were euthanised for macroscopical and histological analyses. Biomechanical characterisation showed optimal properties of the biomaterials for use in tendon repair. Moreover, biological analyses confirmed that tendon-derived fibroblasts were able to adhere to the surface of the generated biomaterials, with high levels of viability and functionality. In vivo studies demonstrated successful tendon repair in all groups. Lastly, histological analyses disclosed better tissue and extracellular matrix organisation and alignment with biomaterial-based augmentation strategies than direct repair, especially when NFAH and NFCH were used. The present study demonstrated that nanostructured fibrin-collagen hydrogels can be used to enhance the healing process in the surgical repair of tendon ruptures.
In general, cultivation and purification of intracellular pathogenic rickettsiae represents a risk for laboratory personnel due to exposure to highly infectious aerosol or accidental inoculation ...during these procedures. In this study, we describe an alternative, effective and time saving technique for rickettsial purification using digitonin to release intracellular bacteria from host cell without physical disruption. No significant differences were noted in yield and infectivity between digitonin treated rickettsiae and rickettsiae purified by sonication. This is the first report of using digitonin in purification of pathogenic rickettsiae and this approach might be effective for other intracellular pathogenic bacteria.
Cluster immunotherapy is becoming an alternative to conventional IT due to its shorter schedule, but the safety of such schedule is still controversial. At present, only few studies assess the risk ...of immunotherapy in a prospective manner, in well-controlled patients, using the same extract and intending to evaluate a single schedule. The aim of the present study is to evaluate the safety of a cluster immunotherapy administration regimen. A total of 91 outpatients (41 male and 50 female), with a mean age of 25 years old (range: 16-50) were included. Sixty-one patients were diagnosed mild to moderate asthma and 30 rhinoconjunctivitis. Forty-six of the enrolled patients were sensitised to pollen (Lolium perenne and/or Olea europea), 38 to Dermatophagoides pteronyssinus and 7 to Alternaria alternata. Patients received specific immunotherapy following a five-week cluster schedule. It was considered as a preseasonal treatment, that is, it was accomplished before olive and grass initial pollinating months in this area (March--April). A total 1029 injections were administered during the induction phase. Adverse reactions were assessed and classified according to the EAACI criteria. The average number of visits to maximum dose was 6 (range 2-10), and 70 patients (77%) reached the maximum between 5-7 visits. In each of the visits an average of 2 (range 1-3) injections were administered. Eighty-one of the 91 initially enrolled patients (89%) completed the cluster schedule. The total number of reactions were 47 (24 local and 23 systemic). No fatal reactions were observed. Since the total number of administered injections was 1029, the relative frequency of adverse reactions was 4.6% (2.3% local and 2.2% systemic). The percentage of patients affected by systemic reaction was 18% and by local reaction 14%. No relationship can be shown between adverse reactions and gender or disease. However, a clear relationship with the composition of immunotherapy has been shown, with a lower risk of adverse reactions associated with the extract of D. pteronyssinus. The shorter period required to achieve the maintenance dose, with a similar frequency of adverse events, leads to the conclusion that the proposed administration regimen can be an alternative to conventional schedule to increase patient compliance.
RESUMEN El vidrio tiene, generalmente, como óxido formador el dióxido de silicio, el cual se acompaña de otros óxidos metálicos que permiten darles brillo, transparencia, estabilidad química, ...insolubilidad en agua y color. Es el material inventado por el hombre más versátil que existe, y se destaca entre otros materiales, porque puede reciclarse infinitamente, permitiendo el ahorro de recursos naturales, energía y costes. Históricamente ha desempeñado importantes funciones en la arquitectura, la automoción, los artículos para el hogar y los envases. Hoy es un elemento esencial en sectores clave como la energía, la biomedicina, la agricultura, la electrónica, las telecomunicaciones, la óptica y el sector aeroespacial. El presente trabajo se suma a los esfuerzos de las Naciones Unidas y la comunidad científica internacional, para que se declarara el 2022 Año Internacional del Vidrio, conmemoración que celebra el pasado, el presente y el futuro del material más transformador de la historia de la humanidad.
El vidrio tiene, generalmente, como óxido formador el dióxido de silicio, el cual se acompaña de otros óxidos metálicos que permiten darles brillo, transparencia, estabilidad química, insolubilidad ...en agua y color. Es el material inventado por el hombre más versátil que existe, y se destaca entre otros materiales, porque puede reciclarse infinitamente, permitiendo el ahorro de recursos naturales, energía y costes. Históricamente ha desempeñado importantes funciones en la arquitectura, la automoción, los artículos para el hogar y los envases. Hoy es un elemento esencial en sectores clave como la energía, la biomedicina, la agricultura, la electrónica, las telecomunicaciones, la óptica y el sector aeroespacial. El presente trabajo se suma a los esfuerzos de las Naciones Unidas y la comunidad científica internacional, para que se declarara el 2022 Año Internacional del Vidrio, conmemoración que celebra el pasado, el presente y el futuro del material más transformador de la historia de la humanidad.
To date, only three rickettsial species have been found in ticks in Slovakia by serological and/or molecular-biological techniques, namely Rickettsia slovaca, Candidatus rickettsia IRS, and ...Rickettsia raoultii. Recently, we succeeded in isolation of the forth species, Rickettsia helvetica from Ixodes ricinus, the most frequent tick in Slovakia. The isolation, positive for 10% of tested ticks, was performed on XTC cells by the shell-vial technique, Gimenez staining and light microscopy. The infected cell cultures contained rod-shaped particles morphologically identical to rickettsiae. The isolation was confirmed by direct detection of a fragment of the R. helvetica gene for citrate synthase in the positive ticks by PCR and its subsequent cloning, sequencing and comparison with the database.
Rickettsia helvetica; isolation; Ixodes ricinus; Slovakia.
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