Molecular biology of the dimorphic fungi Paracoccidioides spp

Paracoccidioidomycosis (PCM) is a mycotic disease caused by the Paracoccidioides species, a group of thermally dimorphic fungi that grow in mycelial form at 25 °C and as budding yeasts when cultured at 37 °C or when parasitizing the host tissues. PCM occurs in a large area of Latin America, and the most critical regions of endemicity are in Brazil, Colombia, and Venezuela. The clinical diagnosis of PCM needs to be confirmed through laboratory tests. Although classical laboratory techniques provide valuable information due to the presence of pathognomonic forms of Paracoccidioides spp., nucleic acid-based diagnostics gradually are replacing or complementing culture-based, biochemical, and immunological assays in routine microbiology laboratory practice. Recently, taxonomic changes driven by whole-genomic sequencing of Paracoccidioides have highlighted the need to recognize species boundaries, which could better ascertain Paracoccidioides taxonomy. In this scenario, classical laboratory techniques do not have significant discriminatory power over cryptic agents. On the other hand, several PCR-based methods can detect polymorphisms in Paracoccidioides DNA and thus support species identification. This review is focused on the recent achievements in molecular diagnostics of paracoccidioidomycosis, including the main advantages and pitfalls related to each technique. We discuss these breakthroughs in light of taxonomic changes in the Paracoccidioides genus.

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Paracoccin overexpression in Paracoccidioides brasiliensis enhances fungal virulence by remodeling chitin properties of the cell wall

The Journal of Infectious Diseases ◽

10.1093/infdis/jiaa707 ◽

2020 ◽

Author(s):

Relber Aguiar Gonçales ◽

Rafael Ricci-Azevedo ◽

Vanessa C S Vieira ◽

Fabrício F Fernandes ◽

Sandra M de O Thomaz ◽

...

Keyword(s):

Cell Wall ◽

Paracoccidioides Brasiliensis ◽

Chitinase Activity ◽

Growth Media ◽

Lung Pathology ◽

Dimorphic Fungi ◽

Fungal Virulence ◽

Paracoccidioides Spp ◽

Cell Growth And Viability ◽

Etiological Agents

Abstract Background The thermo-dimorphic fungi Paracoccidioides spp. are the etiological agents of paracoccidioidomycosis. Although poorly studied, paracoccin (PCN) from P. brasiliensis has been shown to harbor lectinic, enzymatic, and immunomodulatory properties that impact disease development. Methods Mutants of P. brasiliensis overexpressing PCN (ov-PCN) were constructed by Agrobacterium tumefaciens-mediated transformation. Ov-PCN strains were analyzed and inoculated intranasally or intravenously to mice. Fungal burden, lung pathology, and survival were monitored to evaluate virulence. Electron microscopy was used to evaluate the size of chito-oligomer particles released by ov-PCN or wild-type strains to growth media. Results ov-PCN strains revealed no differences in cell growth and viability, although PCN overexpression favored cell separation, chitin processing that results in the release of smaller chito-oligomer particles, and enhanced virulence. Our data show that PCN triggers a critical effect in the cell wall biogenesis through the chitinase activity resulting from overexpression of PCN. As such, PCN overexpression aggravates the disease caused by P. brasiliensis. Conclusions Our data is consistent with a model in which PCN modulates the cell wall architecture via its chitinase activity. These findings highlight the potential for exploiting PCN function in future therapeutic approaches.

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Updates in Paracoccidioides Biology and Genetic Advances in Fungus Manipulation

Journal of Fungi ◽

10.3390/jof7020116 ◽

2021 ◽

Vol 7 (2) ◽

pp. 116

Author(s):

Alison Felipe Alencar Chaves ◽

Marina Valente Navarro ◽

Yasmin Nascimento de Barros ◽

Rafael Souza Silva ◽

Patricia Xander ◽

...

Keyword(s):

Rural Areas ◽

Public Health System ◽

Genetic Manipulation ◽

Oxygen Species ◽

Dimorphic Fungi ◽

Pathogenic Yeast ◽

Temperature Changes ◽

The Public ◽

Causative Agents ◽

Paracoccidioides Spp

The dimorphic fungi of the Paracoccidioides genus are the causative agents of paracoccidioidomycosis (PCM). This disease is endemic in Latin America and primarily affects workers in rural areas. PCM is considered a neglected disease, despite being a disabling disease that has a notable impact on the public health system. Paracoccidioides spp. are thermally dimorphic fungi that present infective mycelia at 25 °C and differentiate into pathogenic yeast forms at 37 °C. This transition involves a series of morphological, structural, and metabolic changes which are essential for their survival inside hosts. As a pathogen, the fungus is subjected to several varieties of stress conditions, including the host immune response, which involves the production of reactive nitrogen and oxygen species, thermal stress due to temperature changes during the transition, pH alterations within phagolysosomes, and hypoxia inside granulomas. Over the years, studies focusing on understanding the establishment and development of PCM have been conducted with several limitations due to the low effectiveness of strategies for the genetic manipulation of Paracoccidioides spp. This review describes the most relevant biological features of Paracoccidioides spp., including aspects of the phylogeny, ecology, stress response, infection, and evasion mechanisms of the fungus. We also discuss the genetic aspects and difficulties of fungal manipulation, and, finally, describe the advances in molecular biology that may be employed in molecular research on this fungus in the future.

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Interacting with Hemoglobin: Paracoccidioides spp. Recruits hsp30 on Its Cell Surface for Enhanced Ability to Use This Iron Source

Journal of Fungi ◽

10.3390/jof7010021 ◽

2021 ◽

Vol 7 (1) ◽

pp. 21

Author(s):

Aparecido Ferreira de Souza ◽

Mariana Vieira Tomazett ◽

Kleber Santiago Freitas e Silva ◽

Juliana Santana de Curcio ◽

Christie Ataides Pereira ◽

...

Keyword(s):

Cell Wall ◽

Protein Expression ◽

Paracoccidioides Brasiliensis ◽

Severe Illness ◽

Primary Role ◽

Dimorphic Fungi ◽

Proteomic Data ◽

Hemoglobin Binding ◽

Paracoccidioides Spp ◽

Essential Nutrient

Paracoccidioides spp. are thermally dimorphic fungi that cause paracoccidioidomycosis and can affect both immunocompetent and immunocompromised individuals. The infection can lead to moderate or severe illness and death. Paracoccidioides spp. undergo micronutrients deprivation within the host, including iron. To overcome such cellular stress, this genus of fungi responds in multiple ways, such as the utilization of hemoglobin. A glycosylphosphatidylinositol (GPI)-anchored fungal receptor, Rbt5, has the primary role of acquiring the essential nutrient iron from hemoglobin. Conversely, it is not clear if additional proteins participate in the process of using hemoglobin by the fungus. Therefore, in order to investigate changes in the proteomic level of P. lutzii cell wall, we deprived the fungus of iron and then treated those cells with hemoglobin. Deprived iron cells were used as control. Next, we performed cell wall fractionation and the obtained proteins were submitted to nanoUPLC-MSE. Protein expression levels of the cell wall F1 fraction of cells exposed to hemoglobin were compared with the protein expression of the cell wall F1 fraction of iron-deprived cells. Our results showed that P. lutzii exposure to hemoglobin increased the level of adhesins expression by the fungus, according to the proteomic data. We confirmed that the exposure of the fungus to hemoglobin increased its ability to adhere to macrophages by flow cytometry. In addition, we found that HSP30 of P. lutzii is a novel hemoglobin-binding protein and a possible heme oxygenase. In order to investigate the importance of HSP30 in the Paracoccidioides genus, we developed a Paracoccidioides brasiliensis knockdown strain of HSP30 via Agrobacterium tumefaciens-mediated transformation and demonstrated that silencing this gene decreases the ability of P. brasiliensis to use hemoglobin as a nutrient source. Additional studies are needed to establish HSP30 as a virulence factor, which can support the development of new therapeutic and/or diagnostic approaches.

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In silico identification of GPI-anchored proteins in Paracoccidioides

10.1101/347351 ◽

2018 ◽

Author(s):

L.R. Basso ◽

R.A. Gonçales ◽

E.J.R Vasconcelos ◽

T.F. Reis ◽

P. C. Ruy ◽

...

Keyword(s):

Cell Wall ◽

In Silico ◽

Glycoside Hydrolases ◽

Cellular Mechanisms ◽

Dimorphic Fungi ◽

A Genome ◽

Paracoccidioides Spp ◽

Gpi Proteins ◽

Eukaryotic Organisms ◽

Prediction Approach

ABSTRACTGlycosylphosphatidylinositol-anchored proteins (GPI-proteins) are widely found in eukaryotic organisms. In fungi, GPI-proteins are thought to be involved in diverse cellular mechanisms such as cell wall biosynthesis and cell wall remodeling, adhesion, antigenicity, and virulence. The conserved structural domains of GPI-protein allow the utilization ofin silicoprediction approach to identify this class of proteins using a genome-wide analysis. We used different previously characterized algorithms to search for genes that encode predicted GPI-proteins in the genome ofP. brasiliensis and P. lutzii, thermal dimorphic fungi that causes paracoccidioidomycosis (PCM). By using these methods, 98 GPI-proteins were found inP. brasiliensiswith orthologs inP. lutzii. A series of 28 GPI-proteins were classified in functional categories (such as glycoside hydrolases, chitin-processing proteins, and proteins involved in the biogenesis of the cell wall). Furthermore, 70 GPI-proteins exhibited homology with hypothetical conserved proteins of unknown function. These data will be an important resource for the future analysis of GPI-proteins inParacoccidioides spp.

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Electron Microscopy in Molecular Biology

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100060027 ◽

1967 ◽

Vol 25 ◽

pp. 2-3

Author(s):

Cecil E. Hall

Keyword(s):

Electron Microscopy ◽

Molecular Biology ◽

Noise Level ◽

Molecular Structures ◽

Material Structure ◽

The Arts ◽

Shadow Casting ◽

Electron Image ◽

High Degree ◽

Very High

The visualization of organic macromolecules such as proteins, nucleic acids, viruses and virus components has reached its high degree of effectiveness owing to refinements and reliability of instruments and to the invention of methods for enhancing the structure of these materials within the electron image. The latter techniques have been most important because what can be seen depends upon the molecular and atomic character of the object as modified which is rarely evident in the pristine material. Structure may thus be displayed by the arts of positive and negative staining, shadow casting, replication and other techniques. Enhancement of contrast, which delineates bounds of isolated macromolecules has been effected progressively over the years as illustrated in Figs. 1, 2, 3 and 4 by these methods. We now look to the future wondering what other visions are waiting to be seen. The instrument designers will need to exact from the arts of fabrication the performance that theory has prescribed as well as methods for phase and interference contrast with explorations of the potentialities of very high and very low voltages. Chemistry must play an increasingly important part in future progress by providing specific stain molecules of high visibility, substrates of vanishing “noise” level and means for preservation of molecular structures that usually exist in a solvated condition.

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Molecular Biology and Biotechnology

10.1039/9781849730211 ◽

2009 ◽

Cited By ~ 4

Keyword(s):

Molecular Biology

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An introduction to the themed issue on RNA biology in China

Essays in Biochemistry ◽

10.1042/ebc20200031 ◽

2020 ◽

Vol 64 (6) ◽

pp. 863-866

Author(s):

Zhe Wu

Keyword(s):

Molecular Biology ◽

Chinese Society ◽

Fortieth Anniversary ◽

Rna Biology ◽

Research And Education ◽

Education In China

Abstract The year 2019 marked the fortieth anniversary of the Chinese Society of Biochemistry and Molecular Biology (CSBMB), whose mission is to promote biomolecular research and education in China. The last 40 years have witnessed tremendous growth and achievements in biomolecular research by Chinese scientists and Essays in Biochemistry is delighted to publish this themed issue that focuses on exciting areas within RNA biology, with each review contributed by key experts from China.

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