Biofilm formation of multidrug-resistant MRSA strains isolated from different types of human infections

Methicillin-resistant Staphylococcus aureus (MRSA) is one of the main pathogens causing chronic infections, mainly due to its capacity to form biofilms. However, the mechanisms underlying the biofilm formation of MRSA strains from different types of human infections are not fully understood. MRSA strains isolated from distinct human infections were characterized aiming to determine their biofilm-forming capacity, the biofilm resistance to conventional antibiotics and the prevalence of biofilm-related genes, including, icaA, icaB, icaC, icaD, fnbA, fnbB, clfA, clfB, cna, eno, ebpS, fib and bbp. Eighty-three clinical MRSA strains recovered from bacteremia episodes, osteomyelitis and diabetic foot ulcers were used. The biofilm-forming capacity was evaluated by the microtiter biofilm assay and the biofilm structure was analyzed via confocal scanning laser microscopy. The antimicrobial susceptibility of 24-h-old biofilms was assessed against three antibiotics and the biomass reduction was measured. The metabolic activity of biofilms was evaluated by the XTT assay. The presence of biofilm-related genes was investigated by whole-genome sequencing and by PCR. Despite different intensities, all strains showed the capacity to form biofilms. Most strains had also a large number of biofilm-related genes. However, strains isolated from osteomyelitis showed a lower capacity to form biofilms and also a lower prevalence of biofilm-associated genes. There was a significant reduction in the biofilm biomass of some strains tested against antibiotics. Our results provide important information on the biofilm-forming capacity of clinical MRSA strains, which may be essential to understand the influence of different types of infections on biofilm production and chronic infections.This work was funded by the R&D Project CAREBIO2: Comparative assessment of antimicrobial resistance in environmental biofilms through proteomics—towards innovative thera-nostic biomarkers, with reference NORTE-01-0145-FEDER-030101 and PTDC/SAU-INF/30101/2017, financed by the European Regional Development Fund (ERDF) through the Northern Regional Operational Program (NORTE 2020) and the Foundation for Science and Technology (FCT). This work was supported by the Associate Laboratory for Green Chemistry—LAQV which is financed by national funds from FCT/MCTES (UIDB/50006/2020 and UIDP/50006/2020). Vanessa Silva is grateful to FCT (Fundação para a Ciência e a Tecnologia) for financial support through the PhD grant SFRH/BD/137947/2018.info:eu-repo/semantics/publishedVersio

Recognition Of Serous Ovarian Tumors In Human Samples By Multimodal Nonlinear Optical Microscopy.

We used a multimodal nonlinear optics microscopy, specifically two-photon excited fluorescence (TPEF), second and third harmonic generation (SHG∕THG) microscopies, to observe pathological conditions of ovarian tissues obtained from human samples. We show that strong TPEF + SHG + THG signals can be obtained in fixed samples stained with hematoxylin and eosin (H&E) stored for a very long time, and that H&E staining enhanced the THG signal. We then used the multimodal TPEF-SHG-THG microscopies in a stored file of H&E stained samples of human ovarian cancer to obtain complementary information about the epithelium∕stromal interface, such as the transformation of epithelium surface (THG) and the overall fibrillary tissue architecture (SHG). This multicontrast nonlinear optics microscopy is able to not only differentiate between cancerous and healthy tissue, but can also distinguish between normal, benign, borderline, and malignant specimens according to their collagen disposition and compression levels within the extracellular matrix. The dimensions of the layers of epithelia can also be measured precisely and automatically. Our data demonstrate that optical techniques can detect pathological changes associated with ovarian cancer.1609601

Tissue distribution of DNA-Hsp65/TDM-loaded PLGA microspheres and uptake by phagocytic cells

This study aimed to demonstrate that microspheres, used as delivery vehicle of DNA-Hsp65/TDM [plasmid DNA encoding heat shock protein 65 (Hsp65) coencapsulated with trehalose dimycolate (TDM) into PLGA microspheres], are widely spread among several organs after intramuscular administration in BALB/c mice. In general, we showed that these particles were phagocytosed by antigen presenting cells, such as macrophages and dendritic cells. Besides, it was demonstrated herein that draining lymph node cells presented a significant increase in the number of cells expressing costimulatory molecules (CD80 and CD86) and MHC class II, and also that the administration of the DNA-Hsp65/TDM and vector/TDM formulations resulted in the up-regulation of CD80, CD86 and MHC class II expression when compared to control formulations (vector/TDM and empty). Regarding the intracellular trafficking we observed that following phagocytosis, the microspheres were not found in the late endosomes and/or lysosomes, until 15 days after internalization, and we suggest that these constructions were hydrolysed in early compartments. Overall, these data expand our knowledge on PLGA [poly (lactic-co- glycolic acid)] microspheres as gene carriers in vaccination strategies, as well as open perspectives for their potential use in clinical practice

Estimating cocaine consumption in the Brazilian Federal District (FD) by sewage analysis

Este é o primeiro trabalho que relata a ocorrência de resíduos de cocaína (COC) e benzoilecgonina (BE) em amostras coletadas em seis estações de esgoto diferentes (ETE) instaladas no Distrito Federal (DF) do Brasil. As concentrações de BE nos afluentes de esgoto foram utilizadas para calcular o consumo de cocaína (kg ano-1 por 1000 habitantes) em cada uma das regiões atendidas pelas ETE, em duas campanhas de amostragem (março e junho de 2010). Dentre as ETE estudadas, amostras provenientes de Samambaia apresentaram as maiores concentrações (de 3866 a 2477 ng L-1 de BE e 805 a 579 ng L-1 de COC) e doses por habitante (mais de 13 doses habitante-1 por ano). A extrapolação para toda a população do DF indica um consumo anual alcançando 1,0 tonelada de cocaína base livre, ou 1,1 tonelada de cloridrato de cocaína. Este trabalho também aborda a influência da forma de apresentação da cocaína (base livre ou sal cloridrato) e a integração com resultados de perfil químico na busca de estimativas mais realistas, principalmente no que se refere aos pontos de vista da criminalística e da segurança pública.This is the first report on the occurrence of cocaine (COC) and benzoylecgonine (BE) residues in six samples collected from different wastewater treatment plants (WTP) located in the Brazilian Federal District (FD). Concentrations of BE in the influent sewage were used to calculate cocaine consumption (kg year-1 per 1000 inhabitants) for each region attended by the WTP from two sampling campaigns (March and June, 2010). Among the WTP studied, samples from Samambaia showed higher concentrations (from 3866 to 2477 ng L-1 of BE and 805 to 579 ng L-1 of COC) and doses per inhabitants (more than 13 doses inhabitant-1 per year). The extrapolation to the whole FD population points out to an annual consumption reaching 1.0 ton of free base cocaine, or 1.1 tons of cocaine hydrochloride. The work also addresses the influence of the cocaine presentation form (free base or hydrochloride) and the integration with chemical profiling results in a more realistic estimate, mainly concerning the viewpoints of forensics and law enforcement

Local Power as the Basis of the Understanding of the Federative Pact

The research aimed to describe the existing problems in the relationship between City, State and Federal Government, through the Brazilian Federative Pact, mainly for municipalities with population of less than 50,000 inhabitants. The research is structured from a qualitative perspective. The theoretical framework was built from the local power of the discussion based on the understanding of the federal pact and local interest and the municipality in Brazil. The paper argues that the federal pact is little debated, discussed, much less questioned by society in general, it only strengthens the lack of a legal and institutional framework for coordination and cooperation among federal entities in the country, which results in public policy fragmented the territory and without direction, causing waste of public resources

B cells Can Modulate the CD8 Memory T Cell after DNA Vaccination Against Experimental Tuberculosis

Abstract\ud \ud \ud \ud Background\ud \ud Although B cells are important as antigen presenting cells (APC) during the immune response, their role in DNA vaccination models is unknown.\ud \ud \ud \ud Methods\ud \ud In this study in vitro and in vivo experiments were performed to evaluate the ability of B cells to protect mice against Mycobacterium tuberculosis challenge.\ud \ud \ud \ud Results\ud \ud \ud In vitro and in vivo studies showed that B cells efficiently present antigens after naked plasmid pcDNA3 encoding M. leprae 65-kDa heat shock protein (pcDNA3-Hsp65) internalization and protect B knock-out (BKO) mice against Mycobacterium tuberculosis infection. pcDNA3-Hsp65-transfected B cells adoptively transferred into BKO mice rescued the memory phenotypes and reduced the number of CFU compared to wild-type mice.\ud \ud \ud \ud Conclusions\ud \ud These data not only suggest that B cells play an important role in the induction of CD8 T cells but also that they improve bacterial clearance in DNA vaccine model.We are thankful to Ana Paula Masson and Izaíra T Brandão for providing the DNA vaccine and recombinant protein. This study was supported by a FAPESP fellowship (05/030873) to LPA.We are thankful to Ana Paula Masson and Izaíra T Brandão for providing the DNA vaccine and recombinant protein. This study was supported by a FAPESP fellowship (05/03087-3) to LPA

MLL5 improves ATRA driven differentiation and promotes xenotransplant engraftment in acute promyelocytic leukemia model

Although the mixed lineage leukemia 5 (MLL5) gene has prognostic implications in acute promyelocyte leukemia (APL), the underlying mechanism remains to be elucidated. Here, we demonstrate the critical role exerted by MLL5 in APL regarding cell proliferation and resistance to drug-induced apoptosis, through mtROS regulation. Additionally, MLL5 overexpression increased the responsiveness of APL leukemic cells to all-trans retinoic acid (ATRA)-induced differentiation, via regulation of the epigenetic modifiers SETD7 and LSD1. In silico analysis indicated that APL blasts with MLL5(high) transcript levels were associated with retinoic acid binding and downstream signaling, while MLL5(low) blasts displayed decreased expression of epigenetic modifiers (such as KMT2C, PHF8 and ARID4A). Finally, APL xenograft transplants demonstrated improved engraftment of MLL5-expressing cells and increased myeloid differentiation over time. Concordantly, evaluation of engrafted blasts revealed increased responsiveness of MLL5-expressing cells to ATRA-induced granulocytic differentiation. Together, we describe the epigenetic changes triggered by the interaction of MLL5 and ATRA resulting in enhanced granulocytic differentiation

Classification and regression tree (CART) model to predict pulmonary tuberculosis in hospitalized patients

Background: Tuberculosis (TB) remains a public health issue worldwide. The lack of specific clinical symptoms to diagnose TB makes the correct decision to admit patients to respiratory isolation a difficult task for the clinician. Isolation of patients without the disease is common and increases health costs. Decision models for the diagnosis of TB in patients attending hospitals can increase the quality of care and decrease costs, without the risk of hospital transmission. We present a predictive model for predicting pulmonary TB in hospitalized patients in a high prevalence area in order to contribute to a more rational use of isolation rooms without increasing the risk of transmission. Methods: Cross sectional study of patients admitted to CFFH from March 2003 to December 2004. A classification and regression tree (CART) model was generated and validated. The area under the ROC curve (AUC), sensitivity, specificity, positive and negative predictive values were used to evaluate the performance of model. Validation of the model was performed with a different sample of patients admitted to the same hospital from January to December 2005. Results: We studied 290 patients admitted with clinical suspicion of TB. Diagnosis was confirmed in 26.5% of them. Pulmonary TB was present in 83.7% of the patients with TB (62.3% with positive sputum smear) and HIV/AIDS was present in 56.9% of patients. The validated CART model showed sensitivity, specificity, positive predictive value and negative predictive value of 60.00%, 76.16%, 33.33%, and 90.55%, respectively. The AUC was 79.70%. Conclusions: The CART model developed for these hospitalized patients with clinical suspicion of TB had fair to good predictive performance for pulmonary TB. The most important variable for prediction of TB diagnosis was chest radiograph results. Prospective validation is still necessary, but our model offer an alternative for decision making in whether to isolate patients with clinical suspicion of TB in tertiary health facilities in countries with limited resources.Fogarty/NIHFogarty/NIH [3 D43 TW000018-16S3, 5 U2R TW006883-02]CNPq [504162/2008-0, 308889/2007-0]CNP