Biology subjects

Araujo, G.

Publications and source records attributed to Araujo, G..

4 recordsLinked to original sources

bioRxiv2022

Phospholipase B is critical for Cryptococcus neoformans survival in the central nervous system

Cryptococcus neoformans (Cn) is an opportunistic, encapsulated, yeast-like fungus that causes severe meningoencephalitis, especially in countries with high HIV prevalence. In addition to its well-known polysaccharide capsule, Cn has other virulence factors such as phospholipases, a heterogeneous group of enzymes that hydrolyze ester linkages in glycerophospholipids. Phospholipase B (PLB1) has been demonstrated to play a key role in Cn pathogenicity. In this study, we used a PLB1 mutant (plb1) and its reconstituted strain (Rec1) to assess the importance of this enzyme on Cn brain infection in vivo and in vitro. Mice infected with plb1 strain survive significantly longer, have lower central nervous system (CNS) fungal load, and fewer and smaller cryptococcomas or biofilm-like brain lesions compared to H99- and Rec1-infected animals. plb1 cryptococci are significantly more phagocytosed and killed by NR-9460 microglia-like cells. plb1 cells have altered capsular polysaccharide biophysical properties that impair their ability to stimulate glia cell responses or morphological changes. We provide significant evidence demonstrating that Cn phospholipase is an important virulence factor for fungal colonization of and survival in the CNS as well as in the progression of cryptococcal meningitis. These findings may potentially help fill in a gap of knowledge in our understanding of cerebral cryptococcosis and may provide novel research avenues in Cn pathogenesis. IMPORTANCECryptococcal meningoencephalitis is a serious disease caused by infection of the neurotropic fungal pathogen Cryptococcus neoformans (Cn). Due to the increasing number of cases in HIV-infected individuals, as well as the limited therapies available, investigation into potential targets for new therapeutics has become critical. Phospholipase B (PLB1) is an enzyme synthesized by Cn that confers virulence to the fungus through capsular enlargement, immunomodulation, and intracellular replication. In this study, we examined the properties of PLB1 by comparing infection of Cn PLB1 mutant strain with both the wild-type and a PLB1 reconstituted strain. We show that PLB1 augments the survival and proliferation of the fungus in the CNS and strengthens virulence through modulation of the immune response and enhancement of specific biophysical properties of the fungus. The implications of PLB1 inhibition reveal its involvement in Cn infection and suggest that it may be a possible molecular target in the development of antifungal therapies. The results of this study support additional investigation into the mechanism of PLB1 to further understand the intricacies of Cn infection.

microbiology↗

bioRxiv2021

Straintables: An application that extracts sequences fromgenome assemblies and generates dissimilarity matrices

Background and ObjectivesThe dissimilarity matrix (DM) is an important component of phylogenetic analysis, and many software packages exist to build and show DMs. However, as the common input for this type of software are sequences in FASTA file format, the process of extracting and aligning each set of sequences to produce a big number of matrices can be laborious. Additionally, existing software do not facilitate the comparison of clusters of similarity across several DMs built for the same group of individuals, using different genomic regions. To address our requirements of such a tool, we designed Straintables to extract specific genomic region sequences from a group of intraspecies genomic assemblies, using extracted sequences to build dissimilarity matrices. MethodsA Python module with executable scripts was developed for a study on genetic diversity across strains of Toxoplasma gondii, being a general purpose system for DM calculation and visualization for preliminary phylogenetic studies. For automatic region sequence extraction from genomic assemblies we assembled a system that designs virtual primers using reference sequences located at genomic annotations, then matches those primers on genome files by using regex patterns. Extracted sequences are then aligned using Clustal Omega and compared to generate matrices. ResultsUsing this software saves the user from manual preparation and alignment of the sequences, a process that can be laborious when a large number of assemblies or regions are involved. The automatic sequence extraction process can be checked against BLAST results using extracted sequence as queries, where correct results were observed for same-species pools for various organisms. The package also contains a matrix visualization tool focused on cluster visualization, capable of drawing matrices into image files with custom settings, and features methods of reordering matrices to facilitate the comparison of clustering patterns across two or more matrices. ConclusionStraintables may replace and extend the functionality of existing matrix-oriented phylogenetic software, featuring automatic region extraction from genomic assemblies and enhanced matrix visualization capabilities emphasizing cluster identification. This module is open source, available at GitHub (https://github.com/Gab0/straintables) under a MIT license and also as a PIPY package. HighlightsSimple in-silico protocol for generation, visualization and comparison of dissimilarity matrices. Accurate automatic sequence extraction from multiple genomic assemblies by using virtual primers built from reference sequences in an annotation file. Draws matrices as images, with enhanced cluster visualization and customized options. Supports reordering of matrix indices to better visualize clustering pattern conservation across multiple regions.

bioinformatics↗

bioRxiv2021

Extracellular vesicles regulate yeast growth, biofilm formation, and yeast-to-hypha differentiation in Candida albicans

The ability to undergo morphological changes during adaptation to distinct environments is exploited by Candida albicans and has a direct impact on virulence. In this study, we investigated the influence of fungal extracellular vesicles (EVs) during yeast growth, biofilm formation, and morphogenesis in C. albicans. Addition of C. albicans EVs (Ca EVs) to the culture medium positively affected yeast growth. Using crystal violet staining and scanning electron microscopy (SEM), we demonstrated that Ca EVs inhibited biofilm formation by C. albicans in vitro. By time-lapse microscopy and SEM, we showed that Ca EV-treatment stops filamentation promoting pseudohyphae formation with multiple sites for yeast budding. The ability of Ca EVs to regulate dimorphism was further compared to EVs isolated from different C. albicans strains, Saccharomyces cerevisiae, and Histoplasma capsulatum. Ca EVs from distinct strains robustly inhibited yeast-to-hyphae differentiation with morphological changes occurring in less than 4 hours. A minor inhibitory effect was promoted by EVs from S. cerevisiae and H. capsulatum only after 24 hours of incubation. The inhibitory effect of Ca EVs was promoted by a combination of lipid compounds identified by gas chromatography-tandem mass spectrometry analysis as sesquiterpenes, diterpenes, and fatty acids. Remarkably, Ca EVs were also able to reverse filamentation, transforming hyphal growth to yeast forms. Transcriptomic analysis demonstrated that treatment with Ca EVs modified the expression of more than 300 genes. The most effectively upregulated pathways were related to DNA metabolism. The downregulated genes were mostly associated with extracellular and adhesion proteins. Finally, yeast cells treated with Ca EVs for 24 hours lost their agar invasive ability and were avirulent when inoculated in Galleria mellonella larvae. In summary, our results indicate that fungal EVs can profoundly modify C. albicans growth and regulate yeast-to-hypha differentiation inhibiting biofilm formation and virulence.

microbiology↗

bioRxiv2020

Assessment of a weak mode of bacterial adhesion by applying an electric field

Microbial attachment to surfaces is ubiquitous in nature. Most species of bacteria attach and adhere to surfaces via special appendages such as pili and fimbriae, the roles of which have been extensively studied. Here we report an experiment on pilus-less mutants of Caulobacter crescentus weakly attached to a plastic surface and subjected to an electric field parallel to the surface. We find that some individual cells transiently but repeatedly adhere to the surface in a stick-slip fashion in the presence of an electric field. Even while transiently detached, these bacteria move significantly slower than the unattached ones in the same field of view undergoing electrophoretic motion. We refer this behavior of repeated and transient attachment as "quasi-attachment". The speed of the quasi-attached bacteria exhibits large variations, frequently dropping close to zero for short intervals of time. This study suggests applying electric field as a useful method to investigate bacteria-surface interaction, which is significant in broader contexts such as infection and environmental control. SignificanceInteraction between bacteria and surfaces occur widely in nature, including those in industrial, environmental, and medical settings. It is therefore important to understand various mechanisms and factors that affect numerous forms of bacterium-surface interaction, particularly those resulting in adhesion or attachment, be they strong or weak, permanent or transient. This work takes a unique approach to identify a transient and reversible mode of bacterial attachment to a solid surface, by applying an electric field to exert a force for detachment. The force thus exerted proves to reach the amplitude required to detach bacteria of a pilus-less strain that weakly attach to a plastic surface. The method may be applied broadly to investigate bacteria-surface interaction.

microbiology↗