Optical Photothermal Infrared - Fluorescence In Situ Hybridization (OPTIR-FISH).

Understanding the metabolic activities of individual cells within complex communities is critical for unraveling their role in human disease. Here, we present a comprehensive protocol for simultaneous cell identification and metabolic analysis with the OPTIR-FISH platform by combining rRNA-tagged FISH probes and isotope-labeled substrates. Fluorescence imaging provides cell identification by the specific binding of rRNA-tagged FISH probes, while OPTIR imaging provides metabolic activities within single cells by isotope-induced red shift on OPTIR spectra. Using bacteria cultured with 13C-glucose as a test bed, the protocol outlines microbial culture with isotopic labeling, fluorescence in situ hybridization (FISH), sample preparation, optimization of the OPTIR-FISH imaging setup, and data acquisition. We also demonstrate how to perform image analysis and interpret spectral data at the single-cell level with high throughput. This protocol's standardized and detailed nature will greatly facilitate its adoption by researchers from diverse backgrounds and disciplines within the broad single-cell metabolism research community.

2D vs. 3D Ultrasound Diagnosis of Pediatric Supracondylar Fractures.

Supracondylar fractures are common injuries in children. Diagnosis typically relies on radiography, which can involve long wait times in the ED, emits ionizing radiation, and can miss non-displaced fractures. Ultrasound (US) has the potential to be a safer, more convenient diagnostic tool, especially with new highly portable handheld 2D point of care US (POCUS). This study aimed to determine the reliability of 2D POCUS for the detection of supracondylar fractures and elbow joint effusions, to contrast the accuracy of 2D POCUS vs. 3DUS vs. radiographs, and to determine whether blinded image interpretation could produce similar results to non-blinded real-time imaging. Fifty-seven children were scanned with 2D POCUS and 3DUS on the affected elbow. US scans were then read by three blinded readers, and the results were compared to gold-standard radiographs. Compared to a gold standard of 30-day radiographic diagnosis, readers of 2D POCUS detected supracondylar fracture and effusion with sensitivities of 0.91 and 0.97, respectively, which were both higher than with 3DUS. Inter-rater reliability of fracture detection was moderate for 2D POCUS (k = 0.40) and 3DUS (k = 0.53). Consensus sensitivities, although high, were lower than reports from some non-blinded studies, indicating that clinical presentation serves as an important factor in detection rates. Our results from consensus US diagnosis support the validity of using 2D POCUS in children for supracondylar fracture and elbow effusion diagnosis.

The Parkinson's drug entacapone disrupts gut microbiome homeostasis via iron sequestration.

Increasing evidence shows that many human-targeted drugs alter the gut microbiome, leading to implications for host health. However, much less is known about the mechanisms by which drugs target the microbiome and how drugs affect microbial function. Here we combined quantitative microbiome profiling, long-read metagenomics, stable isotope probing and single-cell chemical imaging to investigate the impact of two widely prescribed nervous system-targeted drugs on the gut microbiome. Ex vivo supplementation of physiologically relevant concentrations of entacapone or loxapine succinate to faecal samples significantly impacted the abundance of up to one third of the microbial species present. Importantly, we demonstrate that the impact of these drugs on microbial metabolism is much more pronounced than their impact on abundances, with low concentrations of drugs reducing the activity, but not the abundance of key microbiome members like Bacteroides, Ruminococcus or Clostridium species. We further demonstrate that entacapone impacts the microbiome due to its ability to complex and deplete available iron, and that microbial growth can be rescued by replenishing levels of microbiota-accessible iron. Remarkably, entacapone-induced iron starvation selected for iron-scavenging organisms carrying antimicrobial resistance and virulence genes. Collectively, our study unveils the impact of two under-investigated drugs on whole microbiomes and identifies metal sequestration as a mechanism of drug-induced microbiome disturbance.

2D/3D ultrasound diagnosis of pediatric distal radius fractures by human readers vs artificial intelligence.

Wrist trauma is common in children and generally requires radiography for exclusion of fractures, subjecting children to radiation and long wait times in the emergency department. Ultrasound (US) has potential to be a safer, faster diagnostic tool. This study aimed to determine how reliably US could detect distal radius fractures in children, to contrast the accuracy of 2DUS to 3DUS, and to assess the utility of artificial intelligence for image interpretation. 127 children were scanned with 2DUS and 3DUS on the affected wrist. US scans were then read by 7 blinded human readers and an AI model. With radiographs used as the gold standard, expert human readers obtained a mean sensitivity of 0.97 and 0.98 for 2DUS and 3DUS respectively. The AI model sensitivity was 0.91 and 1.00 for 2DUS and 3DUS respectively. Study data suggests that 2DUS is comparable to 3DUS and AI diagnosis is comparable to human experts.

Mid-Infrared Photothermal-Fluorescence In Situ Hybridization for Functional Analysis and Genetic Identification of Single Cells.

Simultaneous identification and metabolic analysis of microbes with single-cell resolution and high throughput are necessary to answer the question of "who eats what, when, and where" in complex microbial communities. Here, we present a mid-infrared photothermal-fluorescence in situ hybridization (MIP-FISH) platform that enables direct bridging of genotype and phenotype. Through multiple improvements of MIP imaging, the sensitive detection of isotopically labeled compounds incorporated into proteins of individual bacterial cells became possible, while simultaneous detection of FISH labeling with rRNA-targeted probes enabled the identification of the analyzed cells. In proof-of-concept experiments, we showed that the clear spectral red shift in the protein amide I region due to incorporation of 13C atoms originating from 13C-labeled glucose can be exploited by MIP-FISH to discriminate and identify 13C-labeled bacterial cells within a complex human gut microbiome sample. The presented methods open new opportunities for single-cell structure-function analyses for microbiology.

Atividade in vitro de extratos e frações de Copernicia prunifera (Arecaceae) contra bactérias de importância clínica em saúde pública / In vitro activity of extracts and fractions of Copernicia prunifera (Arecaceae) against bacteria of clinical importance in public health / Actividad in vitro de extractos y fracciones de Copernicia prunifera (Arecaceae) frente a bacterias de importancia clínica en salud pública

Introdução: O aumento contínuo da resistência bacteriana aos antibióticos convencionais é um problema de importância global. Encontrar produtos como alternativas terapêuticas naturais é essencial. As plantas medicinais possuem uma composição química muito rica, que podem ser estruturalmente otimizadas e processadas em novos antimicrobianos. Objetivo: Avaliar o potencial antibacteriano frente a microrganismos humanos potencialmente patogênicos do extrato etanólico e frações de Copernicia prunifera. Metodologia: A triagem fitoquímica de plantas foi realizada usando métodos de precipitação e coloração e a atividade antibacteriana utilizando o método de difusão em disco e microdiluição em caldo contra cepas padronizadas de Klebsiella pneumoniae, Escherichia coli, Pseudomonas aeruginosa e Staphylococcus aureus. Resultados: A triagem fitoquímica revela a presença de taninos, flavonoides, esteroides, triterpernóides, saponinas e alcaloides. Os extratos etanólico e frações da casca do caule e folhas tiveram atividade inibitória contra S. aureus e K. pneumonie com zona de inibição que variou de 7,0±1,73 a 9,33±0,58 mm pelo método de difusão em disco. Pelo método de microdiluição em caldo os extratos foram satisfatórios somente contra K. pneumoniae (CIM = 125 a 1000 µg/mL) S. aureus, P. aeruginosa e E. coli se mostraram resistentes aos testes (CIM > 1000 µg/mL). Conclusão: Esses resultados fornecem uma base para futuras investigações em modelos in vivo, para que os compostos de C. prunifera possam ser aplicados no desenvolvimento de novos agentes antimicrobianos contra K. pneumoniae.

Introduction: The continuous increase in bacterial resistance to conventional antibiotics is a problem of global importance. Finding products as natural therapeutic alternatives is essential. Medicinal plants have a very rich chemical composition, which can be structurally optimized and processed into novel antimicrobials. Objective: To evaluate the antibacterial potential against potentially pathogenic human microorganisms of the ethanolic extract and fractions of Copernicia prunifera. Methodology: Phytochemical screening of plants was performed using precipitation and staining methods and antibacterial activity using the disk diffusion and broth microdilution method against standardized strains of Klebsiella pneumoniae, Escherichia coli, Pseudomonas aeruginosa and Staphylococcus aureus. Results: Phytochemical screening reveals the presence of tannins, flavonoids, steroids, triterpernoids, saponins and alkaloids. The ethanolic extracts and fractions of stem bark and leaves had inhibitory activity against S. aureus and K. pneumonie with zone of inhibition ranging from 7.0±1.73 to 9.33±0.58 mm by disc diffusion method. By broth microdilution method the extracts were satisfactory only against K. pneumoniae (MIC = 125 to 1000 µg/mL) S. aureus, P. aeruginosa and E. coli were resistant to the tests (MIC > 1000 µg/mL). Conclusion: These results provide a basis for further investigation in in vivo models, so that compounds from C. prunifera can be applied in the development of new antimicrobial agents against K. pneumoniae.

Introducción: El continuo aumento de la resistencia bacteriana a los antibióticos convencionales es un problema de importancia mundial. Es esencial encontrar productos como alternativas terapéuticas naturales. Las plantas medicinales tienen una composición química muy rica, que puede optimizarse estructuralmente y transformarse en nuevos antimicrobianos. Objetivo: Evaluar el potencial antibacteriano frente a microorganismos humanos potencialmente patógenos del extracto etanólico y fracciones de Copernicia prunifera. Metodología: Se realizó el cribado fitoquímico de las plantas mediante los métodos de precipitación y tinción y la actividad antibacteriana mediante el método de difusión en disco y microdilución en caldo frente a cepas estandarizadas de Klebsiella pneumoniae, Escherichia coli, Pseudomonas aeruginosa y Staphylococcus aureus. Resultados: El cribado fitoquímico revela la presencia de taninos, flavonoides, esteroides, triterpernoides, saponinas y alcaloides. Los extractos etanólicos y las fracciones de la corteza del tallo y las hojas presentaron actividad inhibitoria contra S. aureus y K. pneumonie con una zona de inhibición que osciló entre 7,0±1,73 y 9,33±0,58 mm por el método de difusión en disco. Por el método de microdilución en caldo, los extractos sólo fueron satisfactorios frente a K. pneumoniae (CMI = 125 a 1000 µg/mL). S. aureus, P. aeruginosa y E. coli fueron resistentes a las pruebas (CMI > 1000 µg/mL). Conclusiones: Estos resultados proporcionan una base para futuras investigaciones en modelos in vivo, de modo que los compuestos de C. prunifera puedan aplicarse en el desarrollo de nuevos agentes antimicrobianos contra K. pneumoniae.

Participación de mujeres en roles académicos. Facultad de Enfermería y Obstetricia. San Lorenzo-Paraguay. 2021

La perspectiva de género en la universidad refleja la poca inserción laboral en puestos de alta jerarquía, así como en la oportunidad de desarrollo y proyección profesional a mujeres. Su incorporación e inserción en el ámbito académico son importantes para lograr representatividad y participación. El objetivo fue determinar la participación de mujeres en roles académicos en la Facultad de Enfermería y Obstetricia de la Universidad Nacional de Asunción, San Lorenzo-Paraguay en el 2021. Estudio observacional, descriptivo de corte transversal, con muestreo no probabilístico por conveniencia, en docentes de ambos sexos. Para la recolección de datos y posterior análisis se utilizó la base de datos del personal docente del 2021, se analizó e interpretó por estadística descriptiva e inferencial. Las variables cualitativas se describieron por medio de tablas, se exploraron y analizaron posibles asociaciones utilizando la prueba de chi cuadrado con un nivel de significancia de 0,05. De 468 docentes estudiados, 78,19% eran de sexo femenino, edades entre 37 a 46 años, antigüedad de 1 a 7 años, departamento de enfermería, casa central; la mayoría contaba con formación académica de enfermería, en postgrado en especialización, 74,31% correspondió al sexo femenino. La función docente correspondió a encargado de cátedra y coordinador docente sin diferencia de sexo; en participación de mujeres el 100% ocupaba distintos roles y menos del 20% estaba en cargo académico/administrativo. No se encontraron asociaciones estadísticamente significativas entre sexo, rol académico y cargo académico/administrativo. Se determinó que la mayoría de las mujeres participaban en roles académicos.

The gender perspective in the university reflects the lack of labor insertion in high hierarchy positions, as well as in the opportunity for development and professional projection for women. Their incorporation and insertion in the academic field are important to achieve representativeness and participation. The objective was to determine the participation of women in academic roles in the Faculty of Nursing and Obstetrics of the National University of Asuncion, San Lorenzo-Paraguay in 2021. Observational, descriptive, cross-sectional study with non-probabilistic sampling by convenience in teachers of both sexes. For data collection and subsequent analysis, the 2021 teaching staff database was used, analyzed and interpreted by descriptive and inferential statistics. The qualitative variables were described by means of tables, and possible associations were explored and analyzed using the chi-square test with a significance level of 0.05. Of 468 teachers studied, 78.19% was female, ages between 37 and 46 years, seniority from 1 to 7 years, department of nursing, central house; the majority had academic training in nursing, in postgraduate specialization, 74.31% corresponded to the female sex; the teaching function corresponded to head of department and teaching coordinator without difference of sex; for the participation of women 100% occupied different roles and less than 20% was in academic/administrative position. No statistically significant associations were found between sex, academic role and academic/administrative position. It was determined that most women participate in academic roles.

Phytochemical analysis and antimicrobial potential of extracts and fractions of brosimum gaudichaudii trécul against bacteria of clinical importance / Análise fitoquímica e potencial antimicrobiano de extratos e frações de brosimum gaudichaudii trécul contra bactérias de importância clínica / Análisis fitoquímico y potencial antimicrobiano de extractos y fracciones de brosimum gaudichaudii trécul contra bacterias de importancia clínica

Medicinal plants constitute an arsenal of products with different potentials to be explored. Therefore, the objective of this paper was to assess the antimicrobial potential of the ethanolic extract from Brosimum gaudichaudii leaves and fractions against clinically important bacteria. The crude extracts and fractions from the leaves and stem bark were used against Escherichia coli, Klesbsiella pneumoniae, Pseudomonas aeruginosa and Staphylococcus aureus strains. The crude extracts and the fractions were obtained by means of maceration in ethanol and chemically characterized. In the results of the phytochemical screening, the presence of a variety of secondary metabolites was verified, such as flavonoids, steroids, saponins, alkaloids, tannins and coumarins. The extracts and their fractions showed inhibitory activity for all three bacteria tested. The inhibition halo varied from 8±0.00 to 14±0.00 mm fir K. pneumoniae, from 8±0.00 to 10±0.00 mm for P. aeruginosa and from 8±0.00 to 9±0.00 mm for S. aureus. Among the fractions tested, the ethyl acetate fraction from both the stem and the leaves presented the best inhibition potential. This indicates that the Brosimum gaudichaudii Trécul vegetable extracts present antimicrobial potential. Such being the case, it is suggested to isolate the metabolites present in this fraction to delimit the main compounds responsible for the antimicrobial action.

As plantas medicinais constituem um arsenal de produtos com diferentes potenciais a serem explorados. Portanto, o objetivo deste trabalho foi avaliar o potencial antimicrobiano do extrato etanólico das folhas e frações de Brosimum gaudichaudii contra bactérias de importância clínica. Os extratos brutos e as frações das folhas e da casca do caule foram usados contra cepas de Escherichia coli, Klesbsiella pneumoniae, Pseudomonas aeruginosa e Staphylococcus aureus. Os extratos brutos e as frações foram obtidos por meio de maceração em etanol e caracterizados quimicamente. Nos resultados da triagem fitoquímica, foi verificada a presença de uma variedade de metabólitos secundários, como flavonoides, esteroides, saponinas, alcaloides, taninos e cumarinas. Os extratos e suas frações apresentaram atividade inibitória para todas as três bactérias testadas. O halo de inibição variou de 8±0,00 a 14±0,00 mm para K. pneumoniae, de 8±0,00 a 10±0,00 mm para P. aeruginosa e de 8±0,00 a 9±0,00 mm para S. aureus. Entre as frações testadas, a fração de acetato de etila do caule e das folhas apresentou o melhor potencial de inibição. Isso indica que os extratos vegetais de Brosimum gaudichaudii Trécul apresentam potencial antimicrobiano. Sendo assim, sugere-se o isolamento dos metabólitos presentes nessa fração para delimitar os principais compostos responsáveis pela ação antimicrobiana.

Las plantas medicinales constituyen un arsenal de productos con diferentes potenciales por explorar. Por lo tanto, el objetivo de este estudio fue evaluar el potencial antimicrobiano del extracto etanólico de hojas de Brosimum gaudichaudii y fracciones contra bacterias clínicamente importantes. Los extractos crudos y las fracciones de las hojas y la corteza del tallo se utilizaron contra cepas de Escherichia coli, Klesbsiella pneumoniae, Pseudomonas aeruginosa y Staphylococcus aureus. Los extractos crudos y las fracciones se obtuvieron por maceración en etanol y se caracterizaron químicamente. Los resultados del cribado fitoquímico verificaron la presencia de diversos metabolitos secundarios, como flavonoides, esteroides, saponinas, alcaloides, taninos y cumarinas. Los extractos y sus fracciones presentaron actividad inhibitoria para las tres bacterias ensayadas. El halo de inhibición varió de 8±0,00 a 14±0,00 mm para K. pneumoniae, de 8±0,00 a 10±0,00 mm para P. aeruginosa y de 8±0,00 a 9±0,00 mm para S. aureus. Entre las fracciones probadas, la fracción de acetato de etilo del tallo y las hojas mostró el mejor potencial de inhibición. Esto indica que los extractos vegetales de Brosimum gaudichaudii Trécul tienen potencial antimicrobiano. Así, se sugiere el aislamiento de los metabolitos presentes en esta fracción para determinar los principales compuestos responsables de la acción antimicrobiana.

Nutrient niche specificity for glycosaminoglycans is reflected in polysaccharide utilization locus architecture of gut Bacteroides species.

Introduction: Glycosaminoglycans (GAGs) present in the mucosal layer can be used as nutrients by certain intestinal bacteria, particularly members of the Bacteroides. GAG abundances are altered in some diseases such as inflammatory bowel diseases, which may affect microbial composition and activity, and it is therefore important to understand GAG utilization by members of the gut microbiota. Methods: We used growth assays, transcriptomics, and comparative genomics to evaluate chondroitin sulfate (CS) and hyaluronan (HA) degradation ability by multiple gut Bacteroides species. Results and discussion: We found that not all Bacteroides species able to degrade CS could also degrade HA, despite having lyases which act on both compounds. We propose that in the model organism Bacteroides thetaiotaomicron, the lyase BT_3328 in combination with surface binding proteins BT_3329 and BT_3330 and potentially BT_4411 are involved in HA breakdown. Furthermore, degradation of both compounds provides public goods for other Bacteroides, including non-degraders, suggesting that cooperative degradation as well as cross-feeding may be widespread in the mucosal glycan utilization clade.

SRS-FISH: A high-throughput platform linking microbiome metabolism to identity at the single-cell level.

One of the biggest challenges in microbiome research in environmental and medical samples is to better understand functional properties of microbial community members at a single-cell level. Single-cell isotope probing has become a key tool for this purpose, but the current detection methods for determination of isotope incorporation into single cells do not allow high-throughput analyses. Here, we report on the development of an imaging-based approach termed stimulated Raman scattering-two-photon fluorescence in situ hybridization (SRS-FISH) for high-throughput metabolism and identity analyses of microbial communities with single-cell resolution. SRS-FISH offers an imaging speed of 10 to 100 ms per cell, which is two to three orders of magnitude faster than achievable by state-of-the-art methods. Using this technique, we delineated metabolic responses of 30,000 individual cells to various mucosal sugars in the human gut microbiome via incorporation of deuterium from heavy water as an activity marker. Application of SRS-FISH to investigate the utilization of host-derived nutrients by two major human gut microbiome taxa revealed that response to mucosal sugars tends to be dominated by Bacteroidales, with an unexpected finding that Clostridia can outperform Bacteroidales at foraging fucose. With high sensitivity and speed, SRS-FISH will enable researchers to probe the fine-scale temporal, spatial, and individual activity patterns of microbial cells in complex communities with unprecedented detail.

Differential Modulation of the European Sea Bass Gut Microbiota by Distinct Insect Meals.

The aquaculture industry is one of the fastest-growing sectors in animal food production. However, farming of carnivorous fish strongly relies on the use of wild fish-based meals, a practice that is environmentally and economically unsustainable. Insect-based diets constitute a strong candidate for fishmeal substitution, due to their high nutritional value and low environmental footprint. Nevertheless, data on the impact of insect meal (IM) on the gut microbiome of farmed fish are so far inconclusive, and very scarce in what concerns modulation of microbial-mediated functions. Here we use high-throughput 16S rRNA gene amplicon sequencing and quantitative PCR to evaluate the impact of different IMs on the composition and chitinolytic potential of the European sea bass gut digesta- and mucosa-associated communities. Our results show that insect-based diets of distinct origins differently impact the gut microbiota of the European sea bass (Dicentrarchus labrax). We detected clear modulatory effects of IM on the gut microbiota, which were more pronounced in the digesta, where communities differed considerably among the diets tested. Major community shifts were associated with the use of black soldier fly larvae (Hermetia illucens, HM) and pupal exuviae (HEM) feeds and were characterized by an increase in the relative abundance of the Firmicutes families Bacillaceae, Enterococcaceae, and Lachnospiraceae and the Actinobacteria family Actinomycetaceae, which all include taxa considered beneficial for fish health. Modulation of the digesta community by HEM was characterized by a sharp increase in Paenibacillus and a decrease of several Gammaproteobacteria and Bacteroidota members. In turn, a mealworm larvae-based diet (Tenebrio molitor, TM) had only a modest impact on microbiota composition. Further, using quantitative PCR, we demonstrate that shifts induced by HEM were accompanied by an increase in copy number of chitinase ChiA-encoding genes, predominantly originating from Paenibacillus species with effective chitinolytic activity. Our study reveals an HEM-driven increase in chitin-degrading taxa and associated chitinolytic activity, uncovering potential benefits of adopting exuviae-supplemented diets, a waste product of insect rearing, as a functional ingredient.

Expectativas de estudiantes sobre su formación universitaria. Carrera de Obstetricia. Instituto Dr. Andrés Barbero. San Lorenzo-Paraguay. 2018 / Student expectations about their university education. Obstetrics career. Instituto "Dr. Andrés Barbero". San Lorenzo-Paraguay. 2018

Desde el ingreso a la universidad cada estudiante tiene ideas y creencias en cuanto a su formación académica, con respecto a la elección de la carrera, a la institución y especialmente sobre las expectativas en cuanto a su futuro profesional, donde la calidad de su formación prevalecerá para la inserción laboral. El objetivo general fue determinar expectativas del estudiante sobre su formación universitaria en la carrera de Obstetricia del Instituto Dr. Andrés Barbero en San Lorenzo-Paraguay en 2018. El estudio tuvo un diseño observacional, descriptivo, corte transversal enfoque cuantitativo. La población original era de 253 estudiantes, pero la muestra fueron todos los estudiantes matriculados (25) del primer curso. El método usado fue la entrevista, la técnica una encuesta, y el instrumento un cuestionario impreso. Con el visto bueno de la máxima autoridad, se contactó con los sujetos de estudio, a fin de realizar la recolección de la información. Los datos fueron cargados en Excel, y analizados con EPIINFO 7.2 usando estadística descriptica. Casi la totalidad correspondió a mujeres del 17 a 19 años, poco más de la mitad era de procedencia urbana, y menos de la mitad tenía una familia nuclear. En cuanto a la motivación poco menos de la mitad ingresó al Instituto Dr. Andrés Barbero por razones personales, optó por la carrera de Obstetricia por motivos profesionales y en cuanto a las expectativas de los estudiantes sobre su formación universitaria predominaron las razones personales

Since entering the university, each student has ideas and beliefs regarding their academic training, regarding the choice of career, the institution and especially about the expectations regarding their professional future, where the quality of their training will prevail. for job placement. The general objective was to determine the student's expectations about their university training in the Obstetrics career of the Instituto "Dr. Andrés Barbero" in San Lorenzo-Paraguay in 2018. The study had an observational, descriptive, cross-sectional and quantitative approach design. The original population was 253 students, but the sample was all students enrolled (25) in the first year. The method used was the interview, the technique a survey, and the instrument a printed questionnaire. With the approval of the highest authority, the study subjects were contacted in order to collect the information. The data were loaded into Excel, and analyzed with EPIINFO 7.2 using descriptive statistics. Almost all of them corresponded to women between the ages of 17 and 19, just over half were of urban origin, and less than half had a nuclear family. Regarding motivation, just under half entered the Instituto "Dr. Andrés Barbero" for personal reasons, opted for a career in Obstetrics for professional reasons and regarding the expectations of the students about their university training, personal reasons predominated

Outcome measures and motion capture systems for assessing lower limb orthosis-based interventions after stroke: a systematic review.

PURPOSE: To review and categorize, according to the International Classification of Functioning, the outcome measures, and motion capture systems for studying the evidence-based practice of orthotic-based interventions in post-stroke gait rehabilitation. METHODS: An electronic literature search was conducted up to February 2018 in Web of Science, Scopus, MEDLINE and Physiotherapy Evidence Database. Randomized trials measuring activity, impairment or participation outcome measures for studying the evidence-based practice of orthoses in gait rehabilitation after an acute or chronic stroke were identified. The studies were assessed through the Cochrane risk-of-bias tool by three authors. Information about stroke's stage, assessment protocol (goal, timing and motion capture system), orthosis configuration and outcome measures were extracted. RESULTS: Eighteen randomized trials, including 387 post-stroke adults, mostly in the chronic stage, were selected. They assessed 39 outcomes, mainly activity outcome measures such as spatiotemporal (72.2%), kinematic (44.4%) and functional (33.3%) outcomes. Gait speed was the primary outcome in most studies. Participation (22.2%) and impairment (16.7%) outcome measures were less explored. Mostly, non-portable motion capture systems were employed opposing the freely-use of the wearable orthosis. The detection bias risk and the shortage of baseline and follow-up outcome measures affected the studies' assessment quality. CONCLUSIONS: Studies showed heterogeneity in selecting outcomes and timings for assessment. There is evidence for assessing the evidence of orthosis-based gait rehabilitation after stroke through activity outcome measures, primarily the gait speed, recorded by non-wearable motion capture systems. A unified methodology considering wearable sensors for tracking baseline and follow-up measures is needed.Implications for rehabilitationThere is evidence on use activity outcome measures to assess the meaningful evidence-based practice of orthosis-based gait rehabilitation in post-stroke.Gait speed was the primary outcome measure most reported.There is limited evidence on use impairment and participation outcome measures to measure meaningful changes due to orthotic-based rehabilitation.Outcome measures were mainly recorded by non-wearable motion capture systems.

Optofluidic Raman-activated cell sorting for targeted genome retrieval or cultivation of microbial cells with specific functions.

Stable isotope labeling of microbial taxa of interest and their sorting provide an efficient and direct way to answer the question "who does what?" in complex microbial communities when coupled with fluorescence in situ hybridization or downstream 'omics' analyses. We have developed a platform for automated Raman-based sorting in which optical tweezers and microfluidics are used to sort individual cells of interest from microbial communities on the basis of their Raman spectra. This sorting of cells and their downstream DNA analysis, such as by mini-metagenomics or single-cell genomics, or cultivation permits a direct link to be made between the metabolic roles and the genomes of microbial cells within complex microbial communities, as well as targeted isolation of novel microbes with a specific physiology of interest. We describe a protocol from sample preparation through Raman-activated live cell sorting. Subsequent cultivation of sorted cells is described, whereas downstream DNA analysis involves well-established approaches with abundant methods available in the literature. Compared with manual sorting, this technique provides a substantially higher throughput (up to 500 cells per h). Furthermore, the platform has very high sorting accuracy (98.3 ± 1.7%) and is fully automated, thus avoiding user biases that might accompany manual sorting. We anticipate that this protocol will empower in particular environmental and host-associated microbiome research with a versatile tool to elucidate the metabolic contributions of microbial taxa within their complex communities. After a 1-d preparation of cells, sorting takes on the order of 4 h, depending on the number of cells required.

Rational design of a microbial consortium of mucosal sugar utilizers reduces Clostridiodes difficile colonization.

Many intestinal pathogens, including Clostridioides difficile, use mucus-derived sugars as crucial nutrients in the gut. Commensals that compete with pathogens for such nutrients are therefore ecological gatekeepers in healthy guts, and are attractive candidates for therapeutic interventions. Nevertheless, there is a poor understanding of which commensals use mucin-derived sugars in situ as well as their potential to impede pathogen colonization. Here, we identify mouse gut commensals that utilize mucus-derived monosaccharides within complex communities using single-cell stable isotope probing, Raman-activated cell sorting and mini-metagenomics. Sequencing of cell-sorted fractions reveals members of the underexplored family Muribaculaceae as major mucin monosaccharide foragers, followed by members of Lachnospiraceae, Rikenellaceae, and Bacteroidaceae families. Using this information, we assembled a five-member consortium of sialic acid and N-acetylglucosamine utilizers that impedes C. difficile's access to these mucosal sugars and impairs pathogen colonization in antibiotic-treated mice. Our findings underscore the value of targeted approaches to identify organisms utilizing key nutrients and to rationally design effective probiotic mixtures.

Case 268: Bilateral Adrenal Hemorrhage in the Context of Sepsis.

History A 65-year-old man presented to the emergency department with a 1-week history of constipation, which was associated with increasing abdominal distention and not passing flatus. Four weeks prior to the current admission he had been diagnosed with metastatic primary adenocarcinoma of the appendix. One week ago, he had been hospitalized with small-bowel obstruction, for which he required laparotomy and loop ileostomy. His medical history included basal cell carcinoma, rheumatoid arthritis, and Barrett esophagus. Physical examination revealed a distended abdomen with tenderness at palpation within the right upper quadrant and lower abdomen and reduced bowel sounds at auscultation. Initial plain-film radiography of the abdomen at admission revealed dilated gas-filled small-bowel loops, suggestive of obstruction. His small-bowel obstruction was managed conservatively on this occasion. Nine days after admission, the patient became unwell and reported a productive cough. He became tachycardic, tachypneic, and hypotensive. Relevant blood tests at this stage revealed a C-reactive protein level of 206 mg/L (normal range, 0-10 mg/L), a white blood cell count of 24.5 × 109/L (normal range, [4.0-11.0] × 109/L), a red blood cell count of 3.39 × 1012/L (normal range, [4.5-5.5] × 1012/L), a hemoglobin level of 93 g/L (normal range, 130-170 g/L), and a hematocrit level of 0.27 (normal range, 0.4-0.5). CT of the abdomen and pelvis with intravenous contrast material (100 mL Omnipaque 350; GE Healthcare, Oslo, Norway) was performed.

Publisher Correction: An automated Raman-based platform for the sorting of live cells by functional properties.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

An automated Raman-based platform for the sorting of live cells by functional properties.

Stable-isotope probing is widely used to study the function of microbial taxa in their natural environment, but sorting of isotopically labelled microbial cells from complex samples for subsequent genomic analysis or cultivation is still in its early infancy. Here, we introduce an optofluidic platform for automated sorting of stable-isotope-probing-labelled microbial cells, combining microfluidics, optical tweezing and Raman microspectroscopy, which yields live cells suitable for subsequent single-cell genomics, mini-metagenomics or cultivation. We describe the design and optimization of this Raman-activated cell-sorting approach, illustrate its operation with four model bacteria (two intestinal, one soil and one marine) and demonstrate its high sorting accuracy (98.3 ± 1.7%), throughput (200-500 cells h-1; 3.3-8.3 cells min-1) and compatibility with cultivation. Application of this sorting approach for the metagenomic characterization of bacteria involved in mucin degradation in the mouse colon revealed a diverse consortium of bacteria, including several members of the underexplored family Muribaculaceae, highlighting both the complexity of this niche and the potential of Raman-activated cell sorting for identifying key players in targeted processes.

Case 268.

History A 65-year-old man presented to the emergency department with a 1-week history of constipation, which was associated with increasing abdominal distention and not passing flatus. Four weeks prior to the current admission he had been diagnosed with metastatic primary adenocarcinoma of the appendix. One week ago, he had been hospitalized with small-bowel obstruction, for which he required laparotomy and loop ileostomy. His medical history included basal cell carcinoma, rheumatoid arthritis, and Barrett esophagus. Physical examination revealed a distended abdomen with tenderness at palpation within the right upper quadrant and lower abdomen and reduced bowel sounds at auscultation. Initial plain-film radiography of the abdomen at admission revealed dilated gas-filled small-bowel loops, suggestive of obstruction. His small-bowel obstruction was managed conservatively on this occasion. Nine days after his admission, the patient became unwell and reported a productive cough. He became tachycardic, tachypneic, and hypotensive. Relevant blood tests at this stage revealed a C-reactive protein level of 206 mg/L (normal range, 0-10 mg/L), a white blood cell count of 24.5 × 109/L (normal range, [4.0-11.0] × 109/L), a red blood cell count of 3.39 × 1012/L (normal range, [4.5-5.5] × 1012/L), a hemoglobin level of 93 g/L (normal range, 130-170 g/L), and a hematocrit level of 0.27 (normal range, 0.4-0.5). CT of the abdomen and pelvis with intravenous contrast material (100 mL Omnipaque 350; GE Healthcare, Oslo, Norway) was performed ( Figs 1 , 2 ). Figure 1a: (a) Axial and (b) curved reformatted contrast-enhanced CT images of the upper abdomen. Figure 1b: (a) Axial and (b) curved reformatted contrast-enhanced CT images of the upper abdomen. Figure 2a: (a) Axial and (b) coronal contrast-enhanced CT images of the upper abdomen obtained 12 days before the CT images shown in Figures 1a and 1b , respectively. Figure 2b: (a) Axial and (b) coronal contrast-enhanced CT images of the upper abdomen obtained 12 days before the CT images shown in Figures 1a and 1b , respectively.