Molecular and Cellular Basis of Microvascular Perfusion Deficits Induced by Clostridium perfringens and Clostridium septicum

Reduced tissue perfusion leading to tissue ischemia is a central component of the pathogenesis of myonecrosis caused by Clostridium perfringens. The C. perfringens α-toxin has been shown capable of inducing these changes, but its potential synergy with perfringolysin O (θ-toxin) is less well understood. Similarly, Clostridium septicum is a highly virulent causative agent of spontaneous gas gangrene, but its effect on the microcirculation has not been examined. Therefore, the aim of this study was to use intravital microscopy to examine the effects of C. perfringens and C. septicum on the functional microcirculation, coupled with the use of isogenic toxin mutants to elucidate the role of particular toxins in the resultant microvascular perfusion deficits. This study represents the first time this integrated approach has been used in the analysis of the pathological response to clostridial toxins. Culture supernatants from wild-type C. perfringens induced extensive cell death within 30 min, as assessed by in vivo uptake of propidium iodide. Furthermore, significant reductions in capillary perfusion were observed within 60 min. Depletion of either platelets or neutrophils reduced the alteration in perfusion, consistent with a role for these blood-borne cells in obstructing perfusion. In addition, mutation of either the α-toxin or perfringolysin O structural genes attenuated the reduction in perfusion, a process that was reversed by genetic complementation. C. septicum also induced a marked reduction in perfusion, with the degree of microvascular compromise correlating with the level of the C. septicum α-toxin. Together, these data indicate that as a result of its ability to produce α-toxin and perfringolysin O, C. perfringens rapidly induces irreversible cellular injury and a marked reduction in microvascular perfusion. Since C. septicum induces a similar reduction in microvascular perfusion, it is postulated that this function is central to the pathogenesis of clostridial myonecrosis, irrespective of the causative bacterium

A transcriptomic analysis of gene expression in the venom gland of the snake Bothrops alternatus (urutu)

<p>Abstract</p> <p>Background</p> <p>The genus <it>Bothrops </it>is widespread throughout Central and South America and is the principal cause of snakebite in these regions. Transcriptomic and proteomic studies have examined the venom composition of several species in this genus, but many others remain to be studied. In this work, we used a transcriptomic approach to examine the venom gland genes of <it>Bothrops alternatus</it>, a clinically important species found in southeastern and southern Brazil, Uruguay, northern Argentina and eastern Paraguay.</p> <p>Results</p> <p>A cDNA library of 5,350 expressed sequence tags (ESTs) was produced and assembled into 838 contigs and 4512 singletons. BLAST searches of relevant databases showed 30% hits and 70% no-hits, with toxin-related transcripts accounting for 23% and 78% of the total transcripts and hits, respectively. Gene ontology analysis identified non-toxin genes related to general metabolism, transcription and translation, processing and sorting, (polypeptide) degradation, structural functions and cell regulation. The major groups of toxin transcripts identified were metalloproteinases (81%), bradykinin-potentiating peptides/C-type natriuretic peptides (8.8%), phospholipases A₂(5.6%), serine proteinases (1.9%) and C-type lectins (1.5%). Metalloproteinases were almost exclusively type PIII proteins, with few type PII and no type PI proteins. Phospholipases A₂were essentially acidic; no basic PLA₂were detected. Minor toxin transcripts were related to L-amino acid oxidase, cysteine-rich secretory proteins, dipeptidylpeptidase IV, hyaluronidase, three-finger toxins and ohanin. Two non-toxic proteins, thioredoxin and double-specificity phosphatase Dusp6, showed high sequence identity to similar proteins from other snakes. In addition to the above features, single-nucleotide polymorphisms, microsatellites, transposable elements and inverted repeats that could contribute to toxin diversity were observed.</p> <p>Conclusions</p> <p><it>Bothrops alternatus </it>venom gland contains the major toxin classes described for other <it>Bothrops </it>venoms based on trancriptomic and proteomic studies. The predominance of type PIII metalloproteinases agrees with the well-known hemorrhagic activity of this venom, whereas the lower content of serine proteases and C-type lectins could contribute to less marked coagulopathy following envenoming by this species. The lack of basic PLA₂agrees with the lower myotoxicity of this venom compared to other <it>Bothrops </it>species with these toxins. Together, these results contribute to our understanding of the physiopathology of envenoming by this species.</p

Muscle Tissue Damage Induced by the Venom of Bothrops asper: Identification of Early and Late Pathological Events through Proteomic Analysis

Citation: Herrera C, Macêdo JKA, Feoli A, Escalante T, Rucavado A, Gutiérrez JM, et al. (2016) Muscle Tissue Damage Induced by the Venom of Bothrops asper: Identification of Early and Late Pathological Events through Proteomic Analysis. PLoS Negl Trop Dis 10(4): e0004599. doi:10.1371/journal. pntd.0004599The time-course of the pathological effects induced by the venom of the snake Bothrops asper in muscle tissue was investigated by a combination of histology, proteomic analysis of exudates collected in the vicinity of damaged muscle, and immunodetection of extracellular matrix proteins in exudates. Proteomic assay of exudates has become an excellent new methodological tool to detect key biomarkers of tissue alterations for a more integrative perspective of snake venom-induced pathology. The time-course analysis of the intracellular proteins showed an early presence of cytosolic and mitochondrial proteins in exudates, while cytoskeletal proteins increased later on. This underscores the rapid cytotoxic effect of venom, especially in muscle fibers, due to the action of myotoxic phospholipases A2, followed by the action of proteinases in the cytoskeleton of damaged muscle fibers. Similarly, the early presence of basement membrane (BM) and other extracellular matrix (ECM) proteins in exudates reflects the rapid microvascular damage and hemorrhage induced by snake venom metalloproteinases. The presence of fragments of type IV collagen and perlecan one hour after envenoming suggests that hydrolysis of these mechanically/structurally-relevant BM components plays a key role in the genesis of hemorrhage. On the other hand, the increment of some ECM proteins in the exudate at later time intervals is likely a consequence of the action of endogenous matrix metalloproteinases (MMPs) or of de novo synthesis of ECM proteins during tissue remodeling as part of the inflammatory reaction. Our results offer relevant insights for a more integrative and systematic understanding of the time-course dynamics of muscle tissue damage induced by B. asper venom and possibly other viperid venoms.Universidad de Costa Rica/[741-B4-660]/UCR/Costa RicaUniversidad de Costa Rica/[741-B6-125]/UCR/Costa RicaUCR::Vicerrectoría de Investigación::Unidades de Investigación::Ciencias de la Salud::Instituto Clodomiro Picado (ICP

Cross-recognition of a pit viper (Crotalinae) polyspecific antivenom explored through high-density peptide microarray epitope mapping

Snakebite antivenom is a 120 years old invention based on polyclonal mixtures of antibodies purified from the blood of hyper-immunized animals. Knowledge on antibody recognition sites (epitopes) on snake venom proteins is limited, but may be used to provide molecular level explanations for antivenom cross-reactivity. In turn, this may help guide antivenom development by elucidating immunological biases in existing antivenoms. In this study, we have identified and characterized linear elements of B-cell epitopes from 870 pit viper venom protein sequences by employing a high-throughput methodology based on custom designed high-density peptide microarrays. By combining data on antibody-peptide interactions with multiple sequence alignments of homologous toxin sequences and protein modelling, we have determined linear elements of antibody binding sites for snake venom metalloproteases (SVMPs), phospholipases A2s (PLA2s), and snake venom serine proteases (SVSPs). The studied antivenom antibodies were found to recognize linear elements in each of the three enzymatic toxin families. In contrast to a similar study of elapid (non-enzymatic) neurotoxins, these enzymatic toxins were generally not recognized at the catalytic active site responsible for toxicity, but instead at other sites, of which some are known for allosteric inhibition or for interaction with the tissue target. Antibody recognition was found to be preserved for several minor variations in the protein sequences, although the antibody-toxin interactions could often be eliminated completely by substitution of a single residue. This finding is likely to have large implications for the cross-reactivity of the antivenom and indicate that multiple different antibodies are likely to be needed for targeting an entire group of toxins in these recognized sites.Novo Nordisk Foundation/[NNF13OC0005613]/NNF/DinamarcaNovo Nordisk Foundation/[NNF16OC0019248]/NNF/DinamarcaUCR::Vicerrectoría de Investigación::Unidades de Investigación::Ciencias de la Salud::Instituto Clodomiro Picado (ICP

COVID-19 Tracheostomy: Experience in a University Hospital With Intermediate Follow-up

The benefits of percutaneous dilational tracheostomy (PDT) placement have been well documented in patients requiring prolonged mechanical ventilation. However, the data regarding the benefit of PDT in coronavirus-2019 (COVID-19) patients are scarce. The objective of this study is to evaluate the outcomes of a cohort of 37 patients who underwent tracheostomy as part of their COVID-19 care. Retrospective data from a series for 37 patients undergoing tracheostomy was collected using chart review. Primary outcomes included 30 and 60 day mortality, weaning rate, and decannulation rate. Secondary outcomes collected included admission demographics, comorbidities, and procedural information. Thirty-seven (37) patients requiring prolonged mechanical ventilation due to COVID-19. Of these 37 patients, 35 were alive 60 days post-PDT placement, 33 have been weaned from mechanical ventilation and 18 have been decannulated. The low mortality and high decannulation rates in this cohort in is a promising development in the care of critically ill COVID-19 patients. Of note, all participating physicians underwent routine polymerase chain reaction (PCR) testing for infection with the severe acute respiratory syndrome coronavirus-2 virus and no physician contracted COVID-19 as a result of their involvement. Overall, this case series describes the modified PDT technique used by our team and discusses the feasibility and potential benefit to PDT placement in COVID-19 patients requiring long-term mechanical ventilation

Exploring mHealth applications for self-management of chronic low back pain:a survey of features and benefits

Abstract The adoption of Mobile Health (mHealth) for self-management is growing. mHealth solutions are commonly used in public healthcare and health services, where they are appreciated for their ease of use, broad reach, and wide acceptance. Chronic Low Back Pain (CLBP) is one of the most common health problems and a leading cause of disability. As such, it imposes a tremendous burden on patients and society. Studies have proposed that mHealth self-management solutions, such as mobile applications, can supplement traditional care methods and benefit patients, particularly in self-managing CLBP easier. To this end, the number of available mobile applications for CLBP has increased. This paper i) provides an overview of scientific studies on mobile applications for CLBP management from three different viewpoints: researchers, health professionals, and patients, ii) uncovers the application features that were seen as beneficial in the studies, and iii) contrasts the currently available applications for CLBP in Google Play Store and Apple App Store against the discovered features. The findings show that “Personalization and customization” is the most significant feature as it is beneficial from stakeholders’ viewpoint and is represented by most applications. In contrast, “Gamification” and “Artificial intelligence” are the least significant features, indicating a lack of attention from application creators and researchers in this area

Characterization women diagnosed with cervical cancer in two medicals institutions in Ibague Tolima. 2010-2012

70 P?ginasRecurso Electr?nicoEl c?ncer de cuello uterino es la patolog?a neopl?sica maligna de mayor indecencia y mortalidad entre las mujeres alrededor del mundo, considerada principalmente como una enfermedad de trasmisi?n sexual, se presenta m?s frecuentemente en pa?ses en v?a de desarrollo y est? asociada a m?ltiples factores pero principalmente asociado a el virus del papiloma humano, el objetivo del presente estudio es caracterizar las mujeres con c?ncer de c?rvix que asistieron a un consultorio privado y el hospital de referencia de alta complejidad en la ciudad de Ibagu? ? Tolima en el periodo comprendido entre los a?os 2010 -2012 Materiales y M?todos: Se realiz? un Estudio Observacional Descriptivo Retrospectivo para el an?lisis de caracter?sticas de las mujeres con diagn?stico de c?ncer de cuello uterino que asistieron a la consulta de ginecolog?a oncol?gica en la ciudad de Ibagu? ?Tolima entre los a?os 2010 a 2012. Resultados: Las mujeres con c?ncer de c?rvix valoradas en los dos centros de atenci?n medica de Ginecolog?a Oncol?gica se presentaron en su mayor?a entre los 25 y 89 a?os, uni?n libre, de procedencia el 36,4% en el municipio de Ibagu? y el 57,5% pertenec?an a otros municipios dentro de los cuales los m?s frecuentes fueron Guamo, Planadas y Purificaci?n. El carcinoma escamo celular con un 86% aproximadamente fue el tipo histol?gico predominante en la poblaci?n de mujeres que asistieron a consulta por ginecolog?a oncol?gica; el carcinoma estadio IIIB fue el m?s frecuente con un 32%. Conclusiones: El promedio de la edad de las mujeres del estudio fue de 50 a?os, al igual que se presentaron casos de carcinoma in situ en mujeres desde los 25 a?os; en la poblaci?n estudiada se evidencia la presencia de la enfermedad en estadio IIIB en un 32% seguido por carcinoma in situ en 25,9% sin embargo tambi?n se presentan casos de estadios avanzados (IVA, IVB) que tienen un mayor riesgo de muerte.ABSTRACT Cervical cancer is the main malignant neoplastic pathology with the highest incidence and mortality among women around the world. Manly considered as a sexually transmitted disease. Occurs frequently in developing countries and is associated with multiples factors. Indeed, strongly associated with human papilloma virus infection .The purpose of this study is to describe women with cervical cancer who assisted at a private medical office and a high complexity reference hospital in Ibague - Tolima Colombia between 2010-2012. Method: An observational descriptive retrospective study was conducted in order to analyze the characteristics of women with cervical cancer diagnosis who assisted at the gynecological oncology consultation in Ibague-Tolima between 2010 and 2012. Results: Patients? ages were between 25 and 89 years old. Mostly in a common law marriage. Demographically, 36, 4% were from Ibague and 57, 5 % from other towns such as Guamo, Planadas, and Purificaci?n, which had the highest frequency. Squamous cell carcinoma was the most predominant histologic type whit 86%. While, stage IIIB cervical cancer was the most frequent with 32%. Conclusions: The average age of the women studied was 50 years old like cases of carcinoma in situ in women showed since age 25. In our study population the presence of stage IIIB disease is evident at 32% followed by in situ carcinoma in 25.9% of cases however cases of advanced stages (IVA, IVB) that have a higher risk for death also presented.INTRODUCCION 13 1. PLANTEAMIENTO DEL PROBLEMA Y JUSTIFICACI?N 14 2. MARCO TE?RICO 15 2.2. DEFINICI?N Y ETIOLOG?A DEL C?NCER DE C?RVIX 15 2.3. FACTORES DE RIESGO 15 2.3.1. VIRUS DEL PAPILOMA HUMANO 16 2.4. HISTORIA NATURAL 18 2.5. CLASIFICACI?N CL?NICA 19 2.6. TRATAMIENTO POR ESTADIOS 19 2.7. PANORAMA DEL C?NCER DE C?RVIX EN EL MUNDO, LATINOAM?RICA, COLOMBIA Y TOLIMA 19 3. OBJETIVOS 22 3.1. OBJETIVO GENERAL 22 3.2. OBJETIVOS ESPEC?FICOS 22 4. METODOLOG?A 23 4.1. TIPO DEL ESTUDIO 23 4.2. UNIDAD DE AN?LISIS 23 4.3. POBLACI?N DE ESTUDIO Y MUESTREO 23 4.4. CRITERIOS DE INCLUSI?N Y EXCLUSI?N 24 4.5. M?TODOS E INSTRUMENTOS PARA LA RECOPILACI?N DE DATOS 24 4.6. FUENTES DE INFORMACI?N 24 4.7. RECOPILACI?N DE DATOS 24 4.8. PROCESAMIENTO Y AN?LISIS DE DATOS 25 4.9. CONSIDERACIONES ?TICAS 25 5. RESULTADOS 27 5.1. CARACTER?STICAS GENERALES 27 5.2. CARACTER?STICAS GINECOL?GICAS 29 5.3. CARACTER?STICAS PATOL?GICAS 33 6. DISCUSI?N 38 7. CONCLUSIONES 42 BIBLIOGRAF?

Comparison of a nontoxic variant of Clostridium perfringens alpha-toxin with the toxic wild-type strain

The alpha-toxin produced by Clostridium perfringens is one of the best-studied examples of a toxic phospholipase C. In this study, a nontoxic mutant protein from C. perfringens strain NCTC8237 in which Thr74 is substituted by isoleucine (T74I) has been characterized and is compared with the toxic wild-type protein. Thr74 is part of an exposed loop at the proposed membrane-interfacing surface of the toxin. The mutant protein had markedly reduced cytotoxic and myotoxic activities. However, this substitution did not significantly affect the catalytic activity towards water-soluble substrate or the overall three-dimensional structure of the protein. The data support the proposed role of the 70-90 loop in the recognition of membrane phospholipids. These findings also provide key evidence in support of the hypothesis that the hydrolysis of both phosphatidylcholine and sphingomyelin are required for the cytolytic and toxic activity of phospholipases