Chagas Disease Diagnosis with -Exclusive Epitopes in GFP.

Serological tests are critical tools in the fight against infectious disease. They detect antibodies produced during an adaptive immune response against a pathogen with an immunological reagent, whose antibody binding characteristics define the specificity and sensitivity of the assay. While pathogen proteins have conveniently served as reagents, their performance is limited by the natural grouping of specific and non-specific antibody binding sites, epitopes.

Protocol for separation of fungal extracellular vesicles using ultracentrifugation from solid medium cultures.

Extracellular vesicles (EVs) have been identified in diverse fungi, including human pathogens. In this protocol, we present two techniques for isolating and analyzing fungal EVs. The first is for high-throughput screening, and the second is for yielding concentrated samples suitable for centrifugation-based density gradients.

Analysis of Cryptococcus Extracellular Vesicles.

Extracellular vesicles (EVs) are produced by all domains of life. In fungal pathogens, they participate in virulence mechanisms and/or induce protective immunity, depending on the pathogenic species. EVs produced by pathogenic members of the Cryptococcus genus mediate virulence, antifungal resistance, as well as humoral and cell-mediated immunity.

Comprehensive characterization of extracellular vesicles produced by environmental (Neff) and clinical (T4) strains of .

We conducted a comprehensive comparative analysis of extracellular vesicles (EVs) from two strains, Neff (environmental) and T4 (clinical). Morphological analysis via transmission electron microscopy revealed slightly larger Neff EVs (average = 194.5 nm) compared to more polydisperse T4 EVs (average = 168.

Proteomics reveals that the antifungal activity of fenbendazole against Cryptococcus neoformans requires protein kinases.

Cryptococcus neoformans is responsible for over 100 000 deaths annually, and the treatment of this fungal disease is expensive and not consistently effective. Unveiling new therapeutic avenues is crucial. Previous studies have suggested that the anthelmintic drug fenbendazole is an affordable and nontoxic candidate to combat cryptococcosis.

The ecotin-like peptidase inhibitor of Trypanosoma cruzi prevents TMPRSS2-PAR2-TLR4 crosstalk downmodulating infection and inflammation.

Trypanosoma cruzi is the causative agent of Chagas disease, a chronic pathology that affects the heart and/or digestive system. This parasite invades and multiplies in virtually all nucleated cells, using a variety of host cell receptors for infection. T.

Corrected and republished from: "Extracellular Vesicle Formation in Impacts Fungal Virulence".

Small molecules are components of fungal extracellular vesicles (EVs), but their biological roles are only superficially known. is a eukaryotic gene that is required for the activity of benzimidazoles against . In this study, during the phenotypic characterization of mutants expected to lack expression, we observed a reduced EV production.

Organoselenium Has a Potent Fungicidal Effect on Cryptococcus neoformans and Inhibits the Virulence Factors.

Cryptococcosis therapy is often limited by toxicity problems, antifungal tolerance, and high costs. Studies approaching chalcogen compounds, especially those containing selenium, have shown promising antifungal activity against pathogenic species. This work aimed to evaluate the and antifungal potential of organoselenium compounds against Cryptococcus neoformans.

releases proteins during intracellular residence that affect the outcome of the fungal-macrophage interaction.

is a facultative intracellular pathogen that can replicate and disseminate in mammalian macrophages. In this study, we analyzed fungal proteins identified in murine macrophage-like cells after infection with . To accomplish this, we developed a protocol to identify proteins released from cryptococcal cells inside macrophage-like cells; we identified 127 proteins of fungal origin in infected macrophage-like cells.

Extracellular Vesicle Formation in Cryptococcus deuterogattii Impacts Fungal Virulence and Requires the Gene.

Small molecules are components of fungal extracellular vesicles (EVs), but their biological roles are only superficially known. is a eukaryotic gene that is required for the activity of benzimidazoles against Cryptococcus deuterogattii. In this study, during the phenotypic characterization of mutants lacking expression, we observed that this gene was required for EV production.

Screening of the Pandemic Response Box Reveals an Association between Antifungal Effects of MMV1593537 and the Cell Wall of , , and .

There is an urgent unmet need for novel antifungals. In this study, we searched for novel antifungal activities in the Pandemic Response Box, a collection of 400 structurally diverse compounds in various phases of drug discovery. We identified five molecules which could control the growth of Cryptococcus neoformans, Cryptococcus deuterogattii, and the emerging global threat Candida auris.

Bone marrow granulocytes downregulate IL-1β and TNF production and the microbicidal activity of inflammatory macrophages.

Macrophages play critical roles in inflammation and defense against pathogens, as well as in the return to tissue homeostasis. Macrophage subpopulations displaying antagonistic phenotypes are generally classified as proinflammatory M1, implicated in antipathogen and antitumoral activities, or as anti-inflammatory M2, associated with tissue repair. Granulocytic and monocytic myeloid-derived suppressor cells recruited from the bone marrow to tissues and phagocytosis of apoptotic neutrophils can attenuate macrophage microbicidal activity.

Extracellular Vesicles Regulate Biofilm Formation and Yeast-to-Hypha Differentiation in Candida albicans.

In this study, we investigated the influence of fungal extracellular vesicles (EVs) during biofilm formation and morphogenesis in Candida albicans. Using crystal violet staining and scanning electron microscopy (SEM), we demonstrated that C. albicans EVs inhibited biofilm formation .

Biogenesis of Fungal Extracellular Vesicles: What Do We Know?

So far, extracellular vesicles (EVs) have been described in 15 genera of fungi. They carry molecules that contribute to the interaction of fungal cells with the host. Although the number of studies on fungal EVs has increased, the mechanisms involved in their biogenesis are still poorly understood.

extracellular vesicles properties and their use as vaccine platforms.

Whereas extracellular vesicle (EV) research has become commonplace in different biomedical fields, this field of research is still in its infancy in mycology. Here we provide a robust set of data regarding the structural and compositional aspects of EVs isolated from the fungal pathogenic species and . Using cutting-edge methodological approaches including cryogenic electron microscopy and cryogenic electron tomography, proteomics, and flow cytometry, we revisited cryptococcal EV features and suggest a new EV structural model, in which the vesicular lipid bilayer is covered by mannoprotein-based fibrillar decoration, bearing the capsule polysaccharide as its outer layer.

Analysis of Cryptococcal Extracellular Vesicles: Experimental Approaches for Studying Their Diversity Among Multiple Isolates, Kinetics of Production, Methods of Separation, and Detection in Cultures of Titan Cells.

Extracellular vesicles (EVs) produced by members of the Cryptococcus genus are associated with fundamental processes of fungal physiology and virulence. However, several questions about the properties of cryptococcal EVs remain unanswered, mostly because of technical limitations. We recently described a fast and efficient protocol of high-yield EV isolation from solid medium.

Population genomic analysis of Cryptococcus Brazilian isolates reveals an African type subclade distribution.

The genomes of a large number of Cryptococcus neoformans isolates have been sequenced and analyzed in recent years. These genomes have been used to understand the global population structure of this opportunistic pathogen. However, only a small number of South American isolates have been considered in these studies, and the population structure of C.

Small Molecule Analysis of Extracellular Vesicles Produced by : Identification of a Tripeptide Controlling Cryptococcal Infection in an Invertebrate Host Model.

The small molecule (molecular mass <900 Daltons) composition of extracellular vesicles (EVs) produced by the pathogenic fungus is unknown, which limits the understanding of the functions of cryptococcal EVs. In this study, we analyzed the composition of small molecules in samples obtained from solid cultures of by a combination of chromatographic and spectrometric approaches, and untargeted metabolomics. This analysis revealed previously unknown components of EVs, including small peptides with known biological functions in other models.

Phytotoxic Tryptoquialanines Produced by Are Exported in Extracellular Vesicles.

is the most aggressive pathogen of citrus fruits. Tryptoquialanines are major indole alkaloids produced by It is unknown if tryptoquialanines are involved in the damage of citrus fruits caused by To investigate the pathogenic roles of tryptoquialanines, we initially asked if tryptoquialanines could affect the germination of seeds. Exposure of the citrus seeds to tryptoquialanine A resulted in a complete inhibition of germination and an altered metabolic response.

Pathogenic Delivery: The Biological Roles of Cryptococcal Extracellular Vesicles.

Extracellular vesicles (EVs) are produced by all domains of life. In fungi, these structures were first described in and, since then, they were characterized in several pathogenic and non-pathogenic fungal species. Cryptococcal EVs participate in the export of virulence factors that directly impact the -host interaction.

Protective effect of fungal extracellular vesicles against murine candidiasis.

Extracellular vesicles (EVs) are lipid bilayered compartments released by virtually all living cells, including fungi. Among the diverse molecules carried by fungal EVs, a number of immunogens, virulence factors and regulators have been characterised. Within EVs, these components could potentially impact disease outcomes by interacting with the host.

Fenbendazole Controls Growth, Virulence Potential, and Animal Infection in the Model.

The human diseases caused by the fungal pathogens and are associated with high indices of mortality and toxic and/or cost-prohibitive therapeutic protocols. The need for affordable antifungals to combat cryptococcal disease is unquestionable. Previous studies suggested benzimidazoles as promising anticryptococcal agents combining low cost and high antifungal efficacy, but their therapeutic potential has not been demonstrated so far.

Polysaccharide diversity in VNI isolates of Cryptococcus neoformans from Roraima, Northern Brazil.

Species of the Cryptococcus genus comprise environmental, encapsulated fungal pathogens that cause lethal meningitis in immunosuppressed individuals. In humans, fungal uptake of hypocapsular Cryptococcus by macrophages was associated with high fungal burden in the cerebrospinal fluid and long-term patient survival. On the basis of the key role of the cryptococcal capsule in disease, we analyzed the diversity of capsular structures in 23 isolates from pigeon excreta collected in the cities of Boa Vista, Bonfim and Pacaraima, in the state of Roraima (Northern Brazil).

A Novel Protocol for the Isolation of Fungal Extracellular Vesicles Reveals the Participation of a Putative Scramblase in Polysaccharide Export and Capsule Construction in .

Regular protocols for the isolation of fungal extracellular vesicles (EVs) are time-consuming, hard to reproduce, and produce low yields. In an attempt to improve the protocols used for EV isolation, we explored a model of vesicle production after growth of and on solid media. Nanoparticle tracking analysis in combination with transmission electron microscopy revealed that and produced EVs in solid media.