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BACKGROUND: Breathing and aerobic exercises are the most recommended nonpharmacological treatments to improve asthma control and quality of life in patients with asthma. However, the benefits of combining both interventions in these outcomes have never been tested. OBJECTIVE: We aimed to evaluate the effects of the combination of aerobic and breathing exercises on asthma control in individuals with moderate-to-severe asthma. METHODS: Individuals with moderate-to-severe asthma were randomly assigned to the aerobic + muscle-stretching group (Control group (CG), n=25) or aerobic + breathing exercises group (Experimental group (EG), n=26). The aerobic exercise was performed using constant-load training and breathing exercises using the Buteyko technique. The training program lasted 20 sessions. All individuals were blindly assessed before and after the interventions for asthma control, health-related quality of life, anxiety and depression symptoms, sleep quality, hyperventilation, exercise capacity, lung function, levels of physical activity, and thoracoabdominal kinematics. Group x time interactions were tested using a two-way repeated measures analysis of variance. Pearson's correlation test was used to test the association between outcomes. RESULTS: The groups were similar at baseline (all p>0.05). After the intervention, there were no between-group differences for all outcomes comparing EG and CG. CONCLUSION: These results suggest that a combination of aerobic and breathing exercises did not improve asthma control, psychosocial symptoms, sleep quality, hyperventilation, exercise capacity, lung function, levels of daily physical or thoracoabdominal kinematics compared with aerobic exercise alone in individuals with moderate-to-severe asthma. CLINICAL TRIAL REGISTRATION: ClinicalTrials.gov, NCT04412720.

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Electrochemical paper-based analytical devices represent an important platform for portable, low-cost, affordable, and decentralized diagnostics. For this kind of application, chemical functionalization plays a pivotal role to ensure high clinical performance by tuning surface properties and the area of electrodes. However, controlling different surface properties of electrodes by using a single functionalization route is still challenging. In this work, we attempted to tune the wettability, chemical composition, and electroactive area of carbon-paper-based devices by thermally treating polydopamine (PDA) at different temperatures. PDA films were deposited onto pyrolyzed paper (PP) electrodes and thermally treated in the range of 300-1000 °C. After deposition of PDA, the surface is rich in nitrogen and oxygen, it is superhydrophilic, and it has a high electroactive area. As the temperature increases, the surface becomes hydrophobic, and the electroactive area decreases. The surface modifications were followed by Raman, X-ray photoelectron microscopy (XPS), laser scanning confocal microscopy (LSCM), contact angle, scanning electron microscopy (SEM-EDS), electrical measurements, transmission electron microscopy (TEM), and electrochemical experiments. In addition, the chemical composition of nitrogen species can be tuned on the surface. As a proof of concept, we employed PDA-treated surfaces to anchor [AuCl4]- ions. After electrochemical reduction, we observed that it is possible to control the size of the nanoparticles on the surface. Our route opens a new avenue to add versatility to electrochemical interfaces in the field of paper-based electrochemical biosensors.

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Staphylococcus aureus integrate the list of highly virulent and antibiotic resistant pathogens, mainly due to the mecA gene, associated with methicillin resistance. Given the ubiquity of this species, the aim of this study was to investigate whether closely related mecA+S. aureus found in the environment can be also thrive as clinical isolates and if the respective accessory genome may suggest bacterial adaptation. The genomes of environmental (water, animal facilities, food products, n = 111) isolates were compared with closely related genomes of clinical origin (human patients, n = 103). These genomes, available in the public database NCBI, were analysed for phylogeny, accessory genome, and presence of selected clinically relevant genes (n = 104). The genomes of environmental isolates belonged to 18 multi-locus sequence types (MLSTs), 11 of which also included clinical genomes, a result confirmed based on core-genome analysis. Genes significantly (p ≤ 0.05) more frequent among environmental genomes were related with resistance to ß-lactams (blaI, blaPCI), aminoglycosides (ant(6)-Ia), macrolides (mph(C), erm(B)), enterotoxins (seg, sei, sem, sen, seo, seu) and serine protease functions (splB), among others. Genes significantly more frequent among clinical genomes were associated with resistance to macrolides (erm(C)), phenicols (fexA), fosfomycin (murA), the leucocidin virulence gene (lukS-PV), and serine protease functions (splA, splE). It is suggested that mecA+S. aureus can be exchanged between clinical and environmental settings, with accessory traits (particularly antibiotic resistance, virulence and stress response) possibly being associated with the habitat. The interplay between phylogeny and accessory genome is an interesting contribution to better understanding the ecology and evolution of mecA+S. aureus.

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Diphenyl ditelluride (DPDT) is an organotellurium (OT) compound with pharmacological properties, including antioxidant, antigenotoxic and antimutagenic activities when applied at low concentrations. However, DPDT as well as other OT compounds also show cytotoxicity against mammalian cells when treatments occur at higher drug concentrations. Considering that the underlying mechanisms of toxicity of DPDT against tumor cells have been poorly explored, the objective of our study was to investigate the effects of DPDT against both human cancer and non-tumorigenic cells. As a model, we used the colonic HCT116 cancer cells and the MRC5 fibroblasts. Our results showed that DPDT preferentially targets HCT116 cancer cells when compared to MRC5 cells with IC50 values of 2.4 and 10.1 µM, respectively. This effect was accompanied by the induction of apoptosis and a pronounced G2/M cell cycle arrest in HCT116 cells. Furthermore, DPDT induces DNA strand breaks at concentrations below 5 µM in HCT116 cells and promotes the occurrence of DNA double strand breaks mostly during S-phase as measured by γ-H2AX/EdU double staining. Finally, DPDT forms covalent complexes with DNA topoisomerase I, as observed by the TARDIS assay, with a more prominent effect observed in HCT116 than in MRC5 cells. Taken together, our results show that DPDT preferentially targets HCT116 colon cancer cells likely through DNA topoisomerase I poisoning. This makes DPDT an interesting molecule for further development as an anti-proliferative compound in the context of cancer.

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While pyrolyzed paper (PP) is a green and abundant material that can provide functionalized electrodes with wide detection windows for a plethora of targets, it poses long-standing challenges against sensing assays such as poor electrical conductivity, with resistivities generally higher than 200.0 mΩ cm (e.g., gold and silver show resistivities 1000-fold lower, ∼0.2 mΩ cm). In this regard, the fundamental hypothesis that drives this work is whether a scalable, cost-effective, and eco-friendly strategy is capable of significantly reducing the resistivity of PP electrodes toward the development of sensitive electrochemical sensors, whether faradaic or capacitive. We address this hypothesis by simply annealing PP under an isopropanol atmosphere for 1 h, reaching resistivities as low as 7 mΩ cm. Specifically, the annealing of PP at 800 or 1000 °C under isopropanol vapor leads to the formation of a highly graphitic nanolayer (∼15 nm) on the PP surface, boosting conductivity as the delocalization of π electrons stemming from carbon sp2 is favored. The reduction of carbonyl groups and the deposition of dehydrated isopropanol during the annealing process are hypothesized herein as the dominant PP graphitization mechanisms. Electrochemical analyses demonstrated the capability of the annealed PP to increase the charge-transfer kinetics, with the optimum heterogeneous standard rate constant being roughly 3.6 × 10-3 cm s-1. This value is larger than the constants reported for other carbon electrodes and indium tin oxide. Furthermore, freestanding fingers of the annealed PP were prototyped using a knife plotter to fabricate impedimetric on-leaf electrodes. These wearable sensors ensured the real-time and in situ monitoring of the loss of water content from soy leaves, outperforming non-annealed electrodes in terms of reproducibility and sensitivity. Such an application is of pivotal importance for precision agriculture and development of agricultural inputs. This work addresses the foundations for the achievement of conductive PP in a scalable, low-cost, simple, and eco-friendly way, i.e. without producing any liquid chemical waste, providing new opportunities to translate PP-based sensitive electrochemical devices into practical use.

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The World Health Organization released a statement warning of increased risk for the incidence of multidrug resistant microorganisms and the absence of new drugs to control such infections soon. Since the beginning of the COVID-19 pandemic, the prescription of antimicrobial agents has increased and may have accelerated the emergence of multidrug resistant (MDR) bacteria. This study aimed to evaluate maternal and pediatric infections within a hospital from January 2019 to December 2021. An observational retrospective cohort study was performed at a quaternary referral hospital in a metropolitan area of Niteroi city, Rio de Janeiro state, Brazil. A total of 196 patients' medical records were analyzed. The data from 90 (45.9%) patients were collected before the SARS-CoV-2 pandemic, 29 (14.8%) from the 2020 pandemic period, and 77 (39.3%) from the 2021 pandemic period. A total of 256 microorganisms were identified during this period. Out of those, 101 (39.5%) were isolated in 2019, 51 (19.9%) in 2020, and 104 (40.6%) in 2021. Antimicrobial susceptibility tests were performed on 196 (76.6%) clinical isolates. The exact binomial test showed that the distribution of Gram-negative bacteria was predominant. The most common microorganism was Escherichia coli (23%; n = 45), followed by Staphylococcus aureus (17.9%, n = 35), Klebsiella pneumoniae (12.8%, n = 25), Enterococcus faecalis (7.7%, n = 15), Staphylococcus epidermidis (6.6%, n = 13) and Pseudomonas aeruginosa (5.6%, n = 11). Staphylococcus aureus was the predominant species among resistant bacteria. Among the antimicrobial agents tested, the following were resistant, presented on a descending scale: penicillin (72.7%, p = 0.001, Binomial test), oxacillin (68.3%, p = 0.006, Binomial test), ampicillin (64.3%, p = 0.003, Binomial test), and ampicillin/sulbactam (54.9%, p = 0.57, Binomial test). Infections with S. aureus were 3.1 times greater in pediatrics and maternal units than in other hospital wards. Despite the global reduction in the incidence of MRSA, we observed an increase in MDR S. aureus in this study. No changes were observed in the frequency of resistance profiles of the clinical isolates after the establishment of the global SARS-CoV-2 pandemic. More comprehensive studies are needed to understand the impact of the global SARS-CoV-2 pandemic on the resistance levels of bacteria associated with neonate and pediatric patients.

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Since its discovery in 2007, polydopamine nanofilms have been widely used in many areas for surface functionalization. The simple and low-cost preparation method of the nanofilms with tunable thickness can incorporate amine and oxygen-rich chemical groups in virtually any interface. The remarkable advantages of this route have been successfully used in the field of electrochemical sensors. The self-adhesive properties of polydopamine are used to attach nanomaterials onto the electrode's surface and add chemical groups that can be explored to immobilize recognizing species for the development of biosensors. Thus, the combination of 2D materials, nanoparticles, and other materials with polydopamine has been successfully demonstrated to improve the selectivity and sensitivity of electrochemical sensors. In this review, we highlight some interesting properties of polydopamine and some applications where polydopamine plays an important role in the field of electrochemical sensors.

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The Klebsiella pneumoniae complex is comprised of ubiquitous bacteria that can be found in soils, plants or water, and as humans' opportunistic pathogens. This study aimed at inferring common and distinctive features in clinical and environmental K. pneumoniae. Whole genome sequences of members of the K. pneumoniae complex (including K. variicola, n = 6; and K. quasipneumoniae, n = 7), of clinical (n = 78) and environmental (n = 61) origin from 21 countries were accessed from the GenBank. These genomes were compared based on phylogeny, pangenome and selected clinically relevant traits. Phylogenetic analysis based on 2704 genes of the core genome showed close relatedness between clinical and environmental strains, in agreement with the multi-locus sequence typing. Eight out of the 62 sequence types (STs) identified, included both clinical and environmental genomes (ST11, ST14, ST15, ST37, ST45, ST147, ST348, ST437). Pangenome-wide association studies did not evidence significant differences between clinical and environmental genomes. However, the genomes of clinical isolates presented significantly more exclusive genes related to antibiotic resistance/plasmids, while the environmental isolates yielded significantly higher allelic diversity of genes related with functions such as efflux or oxidative stress. The study suggests that K. pneumoniae can circulate among the natural environment and clinical settings, probably under distinct adaptation pressures.

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A rare and difficult to diagnose case of subacute infective endocarditis caused by Bacillus cereus in a patient with systemic lupus erythematosus and Libman-Sacks endocarditis has been reported. Our aim is to highlight the importance of molecular methods such as MALDI-TOF and PCR to explain clinical and epidemiological issues about infections caused by unusual pathogen.

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Staphylococcus aureus have been progressively identified in farm animals and in humans with direct contact with these animals showing that S. aureus may be a major zoonotic pathogen. Therefore, we aimed to isolate S. aureus from cows, their handlers, and their immediate surroundings, and to investigate the antimicrobial resistance and genetic lineages of the isolates. Mouth and nose swabs of 244 healthy cows (195 Maronesa, 11 Holstein-Friesians, and 28 crossbreeds), 82 farm workers, 53 water and 63 soil samples were collected. Identification of species was carried out by MALDI-TOF MS Biotyper. The presence of antimicrobial resistance genes and virulence factors was assessed based on gene search by PCR. All isolates were typed by multilocus sequence typing and spa-typing. From 442 samples, 33 (13.9%), 24 (29.3%), 1 (2%), and 1 (2%) S. aureus were recovered from cows, farm workers, water, and soil samples, respectively. Most of the isolates showed resistance only to penicillin. S. aureus isolates were ascribed to 17 sequence types (STs) and 26 spa-types. Some clonal lineages were common to both cows and farm workers such as ST30-t9413, ST72-t148, and ST45-t350. Through a One Health approach, this study revealed that there is a great diversity of clonal lineages of S. aureus in cows and their handlers. Furthermore, some S. aureus lineages are common to cows and handlers, which may suggest a possible transmission.

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Impedimetric wearable sensors are a promising strategy for determining the loss of water content (LWC) from leaves because they can afford on-site and nondestructive quantification of cellular water from a single measurement. Because the water content is a key marker of leaf health, monitoring of the LWC can lend key insights into daily practice in precision agriculture, toxicity studies, and the development of agricultural inputs. Ongoing challenges with this monitoring are the on-leaf adhesion, compatibility, scalability, and reproducibility of the electrodes, especially when subjected to long-term measurements. This paper introduces a set of sensing material, technological, and data processing solutions that overwhelm such obstacles. Mass-production-suitable electrodes consisting of stand-alone Ni films obtained by well-established microfabrication methods or ecofriendly pyrolyzed paper enabled reproducible determination of the LWC from soy leaves with optimized sensibilities of 27.0 (Ni) and 17.5 kΩ %-1 (paper). The freestanding design of the Ni electrodes was further key to delivering high on-leaf adhesion and long-term compatibility. Their impedances remained unchanged under the action of wind at velocities of up to 2.00 m s-1, whereas X-ray nanoprobe fluorescence assays allowed us to confirm the Ni sensor compatibility by the monitoring of the soy leaf health in an electrode-exposed area. Both electrodes operated through direct transfer of the conductive materials on hairy soy leaves using an ordinary adhesive tape. We used a hand-held and low-power potentiostat with wireless connection to a smartphone to determine the LWC over 24 h. Impressively, a machine-learning model was able to convert the sensing responses into a simple mathematical equation that gauged the impairments on the water content at two temperatures (30 and 20 °C) with reduced root-mean-square errors (0.1% up to 0.3%). These data suggest broad applicability of the platform by enabling direct determination of the LWC from leaves even at variable temperatures. Overall, our findings may help to pave the way for translating "sense-act" technologies into practice toward the on-site and remote investigation of plant drought stress. These platforms can provide key information for aiding efficient data-driven management and guiding decision-making steps.

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Hospital wastewaters often carry multidrug-resistant bacteria and priority pathogens, such as methicillin-resistant Staphylococcus aureus (MRSA). Pathogens and antibiotic resistance genes present in wastewaters may reach the natural environment facilitating their spread. Thus, we aimed to isolate MRSA from wastewater of 3 hospitals located in the north of Portugal and to characterize the isolates regarding the antimicrobial resistance and genetic lineages. A total of 96 wastewater samples were collected over six months. The water was filtered, and the filtration membrane was immersed in BHI broth supplemented with 6.5% of NaCl and incubated. The inoculum was streaked in ORSAB agar plates for MRSA isolation. The isolates susceptibility testing was performed against 14 antimicrobial agents. The presence of resistance and virulence genes was accessed by PCR. Molecular typing was performed in all isolates. From the 96 samples, 28 (29.2%) were MRSA-positive. Most isolates had a multidrug-resistant profile and carried the mecA, blaZ, aac(6')-Ie-aph(2″)-Ia, aph(3')-IIIa, ermA, ermB, ermC, tetL, tetM, dfrA dfrG and catpC221 genes. Most of the isolates were ascribed to the immune evasion cluster (IEC) type B. The isolates belonged to ST22-IV, ST8-IV and ST105-II and spa-types t747, t1302, t19963, t6966, t020, t008 and tOur study shows that MRSA can be found over time in hospital wastewater. The wastewater treatment processes can reduce the MRSA load. The great majority of the isolates belonged to ST22 and spa-type t747 which suggests the fitness of these genetic lineages in hospital effluents.

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BACKGROUND: Klebsiella pneumoniae are ubiquitous bacteria and recognized multidrug-resistant opportunistic pathogens that can be released into the environment, mainly through sewage, where they can survive even after wastewater treatment. A major question is if once released into wastewater, the selection of lineages missing clinically-relevant traits may occur. Wastewater (n = 25) and clinical (n = 34) 3rd generation cephalosporin-resistant K. pneumoniae isolates were compared based on phenotypic, genotypic and genomic analyses. RESULTS: Clinical and wastewater isolates were indistinguishable based on phenotypic and genotypic characterization. The analysis of whole genome sequences of 22 isolates showed that antibiotic and metal resistance or virulence genes, were associated with mobile genetic elements, mostly transposons, insertion sequences or integrative and conjugative elements. These features were variable among isolates, according to the respective genetic lineage rather than the origin. CONCLUSIONS: It is suggested that once acquired, clinically relevant features of K. pneumoniae may be preserved in wastewater, even after treatment. This evidence highlights the high capacity of K. pneumoniae for spreading through wastewater, enhancing the risks of transmission back to humans.

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OBJECTIVE: Over the past few decades, life expectancy in Brazil has increased from 48 years in 1950s to 76 years in 2017. The aim of this study was to investigate the impact of ageing on: (1) the frequency of hospitalisations due to bloodstream infection (BSI); (2) the incidence of hospital-acquired BSI (H-BSI); (3) the incidence of BSI caused by multidrug-resistant (MDR) agents and (4) the mortality rate of BSI in a public hospital. METHODS: A hospital-based case-cohort study was conducted between 1 December 2013 and 31 December 2015. The data were analysed using multivariable logistic regression. RESULTS: A total of 500 BSI episodes were detected, among 11,102 hospitalizations. The incidence of hospitalisations resulting from BSI was significantly higher in older than younger patients (3.7/100 vs. 2.0/100, p < 0.01). Similarly, the incidence of hospital-acquired BSI was significantly higher in older patients (2.7/100 vs. 0.9/100, p < 0.01). Klebsiella pneumoniae (15.9%), Staphylococcus aureus (14.3%), Escherichia coli (13.1%) and Acinetobacter spp. (12.1%) were the most common agents isolated. MDR agents caused 37.6% of the BSI episodes; enteric Gram-negative bacilli resistant to third- or fourth-generation cephalosporins (9.7%) and carbapenem-resistant Acinetobacter spp. (9.2%) were the most common MDR agents. The following complications were independently associated with ageing: Charlson comorbidity index (OR = 1.16; 95% CI = 1.09-1.24); BSI secondary to urinary tract infection (OR = 2.14; 95% CI = 1.29-3.55); BSI secondary to pneumonia (OR = 1.77; 95% CI = 1.07-2.93) and 30-day mortality following BSI (OR = 2.19; 95% CI = 1.43-3.36). CONCLUSIONS: These data suggest ageing has a significant impact on hospitalisations due to BSI, H-BSI incidence and mortality from BSI in older patients attending a Brazilian public hospital. Age was not significantly associated with MDR-related BSI. These results indicate that age plays an important role in the increase in morbidities and mortality resulting from BSI in Brazil and that with the increased life expectancy observed over recent decades in Brazil, the burden of BSI will be expected to continue to increase. This dynamic needs to be better understood with additional studies.

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Graphene oxide (GO) microfibers with controlled and homogeneous shapes and tunable diameters were fabricated using the 3 dimensional (3D) hydrodynamic focusing concept on a microfluidic device. Thermal and microwave treatments are used to obtain reduced graphene oxide (rGO) microfibers with outstanding electrical properties, thus enabling the development of ionic liquid-gate field-effect transistors (FET) based on graphene derivative microfibers.

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Infections due to multidrug resistant Gram-negative pathogens are of great concern worldwide, as they are frequently associated with high mortality and morbidity rates. The occurrence of Pseudomonas spp. producing Klebsiella pneumoniae carbapenemases (KPCs) imposes a great challenge through treatment course of bloodstream infections (BSIs). Pseudomonas putida has been recognized as an emerging pathogen of healthcare associated infections (HAIs). Therefore, we aimed to report a case of a non-fatal case of peripheral line associated BSI (PLA-BSI) in an immunocompromised host due to P. putida harboring blaKPC-2 gene in Brazil. A P. putida isolate was recovered from a blood culture of a 72-year-old man admitted at a University Hospital, identified by BD Phoenix™ 100 (Becton, Dickinson and Company), causing PLA-BSI. The species identification was confirmed by Matrix-Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry (MALDI-TOF MS) and resistance to carbapenems were confirmed by Epsilometer test (E-test®). Additionally, the presence of important carbapenemases genes (blaKPC, blaNDM, blaOXA-48-like, blaSPM, blaIMP, blaVIM) was investigated by Polymerase Chain Reaction. The bacterial isolate was confirmed as meropenem resistant P. putida harboring blaKPC-2 gene.Thereofre, these fidings suggest that P. putida can work as a reservoir for resistance genes as this bacterium has the ability to disseminate through water-fluids inside hospital and community settings. Moreover, this paper highlights that a frequent and worldwide disseminated mechanism of resistance (blaKPC-2) is currently occurring among uncommon agents of BSI.

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ABSTRACT This study addresses the effects of the art of clowning on the quality of life and wellbeing of a woman presented with aphasia and depression. She was discharged from individual Speech-Language Therapy due to improved language skills, but she continued expressing emotional complaints associated with communicative situations. The participant joined Palhafasia, a community project that welcomes aphasics for group clowning sessions. She participated for six months in once-a-week meetings that lasted three hours each. The comparison of pre- and post-intervention measures detected improvements in her quality of life, including communication and subjective well-being of the participant. This innovative case study provides a preliminary contribution that must be further investigated. The study points to the potential benefits of the practice of clowning to promote quality of life and well-being of people with aphasia who suffer from depression.

RESUMO Este relato de caso discute o efeito da arte clownesca na qualidade de vida e bem-estar de uma mulher com afasia predominantemente expressiva e depressão. Esta havia recebido alta da terapia fonoaudiológica individual devido à melhora na linguagem, mas continuava com queixas emocionais ligadas a situações comunicativas. A participante, então, ingressou no Palhafasia, um projeto de extensão que acolhe afásicos para a realização de sessões de arte clownesca em grupo. Participou uma vez por semana de encontros com duração de três horas durante seis meses. A comparação de medidas pré e pós intervenção detectou melhoras na sua qualidade de vida - incluindo aspectos de comunicação - e bem-estar subjetivo. Esse estudo de caso traz evidências preliminares sobre uma abordagem inovadora que deve ser investigada mais amplamente. O estudo aponta para potenciais benefícios que a prática da arte clownesca pode ter no que se refere à promoção de maior qualidade de vida e bem-estar de pessoas com afasia que sofrem de depressão.

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Standardized methods are needed to support monitoring of antibiotic resistance in environmental samples. Culture-based methods target species of human-health relevance, while the direct quantification of antibiotic resistance genes (ARGs) measures the antibiotic resistance potential in the microbial community. This study compared measurements of tetracycline-, sulphonamide-, and cefotaxime-resistant presumptive total and fecal coliforms and presumptive enterococci versus a suite of ARGs quantified by quantitative polymerase chain reaction (qPCR) across waste-, recycled-, tap-, and freshwater. Cross-laboratory comparison of results involved measurements on samples collected and analysed in the US and Portugal. The same DNA extracts analysed in the US and Portugal produced comparable qPCR results (variation <28%), except for blaOXA-1 gene (0%-57%). Presumptive total and fecal coliforms and cefotaxime-resistant total coliforms strongly correlated with blaCTX-M and intI1 (0.725 ≤ R2 ≤ 0.762; p < 0.0001). Further, presumptive total and fecal coliforms correlated with the Escherichia coli-specific biomarkers, gadAB, and uidA, suggesting that both methods captured fecal-sourced bacteria. The genes encoding resistance to sulphonamides (sul1 and sul2) were the most abundant, followed by genes encoding resistance to tetracyclines (tet(A) and tet(O)) and ß-lactams (blaOXA-1 and,blaCTX-M), which was in agreement with the culture-based enumerations. The findings can help inform future application of methods being considered for international antibiotic resistance surveillance in the environment.

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Klebsiella pneumoniae is an important human pathogen, able to accumulate and disseminate a variety of antimicrobial resistance genes. Resistance to colistin, one of the last therapeutic options for multi-drug-resistant bacteria, has been reported increasingly. Colistin-resistant K. pneumoniae (ColRKp) emerged in two hospitals in Rio de Janeiro state, Brazil in 2016. The aim of this study was to investigate if these ColRKp isolates were clonally related when compared between hospitals, to identify the molecular mechanisms of colistin resistance, and to describe other antimicrobial resistance genes carried by isolates. Twenty-three isolates were successively recovered, and the whole-genome sequence was analysed for 10, each of a different pulsed-field gel electrophoresis (PFGE) type. Although some PFGE clusters were found, none of them included isolates from both hospitals. Half of the isolates were assigned to CC258, three to ST152 and two to ST15. One isolate was pandrug resistant, one was extensively drug resistant, and the others were multi-drug resistant. Colistin resistance was related to mutations in mgrB, pmrB, phoQ and crrB. Eleven new mutations were found in these genes, including two nucleotide deletions in mgrB. All isolates were carbapenem resistant, and seven were associated with carbapenemase carriage (blaKPC-2 in six isolates and blaOXA-370 in one isolate). All isolates had a blaCTX-M, and two had a 16S ribosomal RNA methyltransferase encoding gene (armA and rmtB). ColRKp were composed of epidemic clones, but cross-dissemination between hospitals was not detected. Colistin resistance emerged with several novel mutations amid highly resistant strains, further restricting the number of drugs available and leading to pandrug resistance.

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Antimicrobial resistance represents one of the greatest challenges of the twenty-first century, and it is globally recognized that addressing this problem requires a concerted One Health approach involving humans, animals, and the environment. Methicillin-resistant Staphylococcus aureus (MRSA) currently represents a global burden; it is resistant to almost all beta-lactams and some MRSA strains are highly multiresistant. S. aureus infection in cattle results in major economic losses in the food industry. Moreover, cases of livestock-associated MRSA strains responsible for invasive life-threatening infections have been reported among human patients in contact with infected or colonized animals. The autochthonous Maronesa cattle breed is a threatened rustic traditional Portuguese breed of mountain cattle of high importance for the Vila Real region. It has been used for centuries as motive power in all kinds of agricultural work and also for meat production, which is its current dominant use and the main source of economic value, being the Maronesa meat commercialized with PDO - Protected Designation of Origin. This study aimed to determine the prevalence and transmission of MRSA in cattle of the Maronesa breed, through a concerted One Health approach comprising human, water, and soil samples of the animals' handlers and environments. In a total of 195, 63, 40, and 43 cattle, human, water, and soil samples screened in selective ORSAB media supplemented with 2 mg/L oxacillin; only one human sample harbored a MRSA isolate which was ascribed to spa-type t9413 and to ST30, one of the most common genetic lineages associated with community-acquired MRSA. Considering the increasing reports of MRSA isolation from cattle and handlers in Europe, the absence of this major human and animal pathogen in Maronesa cattle and their production systems represents a serendipitous result, valuing this important autochthonous breed. To our knowledge, this is the first study to determine MRSA prevalence and transmission in Maronesa cattle. Through a concerted One Health approach, this study revealed that the Maronesa cattle and their surrounding environments do not represent reservoirs for Methicillin-resistant Staphylococcus aureus.