scholarly journals Virulence Vs. Immunomodulation: Roles of the Paracoccin Chitinase and Carbohydrate-Binding Sites in Paracoccidioides brasiliensis Infection

2021 ◽  
Vol 8 ◽  
Author(s):  
Nayla de Souza Pitangui ◽  
Fabrício Freitas Fernandes ◽  
Relber Aguiar Gonçales ◽  
Maria Cristina Roque-Barreira
Keyword(s):  
Cell Wall ◽  
Host Response ◽  
Paracoccidioides Brasiliensis ◽  
Severe Disease ◽  
Chitinase Activity ◽  
Carbohydrate Binding ◽  
Toll Like Receptor ◽  
Dimorphic Fungus ◽  
Fungal Virulence ◽  
Yeast Infection

Paracoccin (PCN) is a bifunctional protein primarily present in the cell wall of Paracoccidioides brasiliensis, a human pathogenic dimorphic fungus. PCN has one chitinase region and four potential lectin sites and acts as both a fungal virulence factor and an immunomodulator of the host response. The PCN activity on fungal virulence, mediated by the chitinase site, was discovered by infecting mice with yeast overexpressing PCN (PCN-ov). PCN-ov are characterized by increased chitin hydrolysis, a narrow cell wall, and augmented resistance to phagocytes' fungicidal activity. Compared to wild-type (wt) yeast, infection with PCN-ov yeast causes a more severe disease, which is attributed to the increased PCN chitinase activity. In turn, immunomodulation of the host response was demonstrated by injecting, subcutaneously, recombinant PCN in mice infected with wt-P. brasiliensis. Through its carbohydrate binding site, the injected recombinant PCN interacts with Toll-like receptor 2 (TLR2) and Toll-like receptor 4 (TLR4) N-glycans on macrophages, triggers M1 polarization, and stimulates protective Th1 immunity against the fungus. The PCN-treatment of wt yeast-infected mice results in mild paracoccidioidomycosis. Therefore, PCN paradoxically influences the course of murine paracoccidioidomycosis. The disease is severe when caused by yeast that overexpress endogenous PCN, which exerts a robust local chitinase activity, followed by architectural changes of the cell wall and release of low size chito-oligomers. However, the disease is mild when exogenous PCN is injected, which recognizes N-glycans on systemic macrophages resulting in immunomodulation.

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

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.

scholarly journals Drk1, a Dimorphism Histidine Kinase, Contributes to Morphology, Virulence, and Stress Adaptation in Paracoccidioides brasiliensis

2021 ◽  
Vol 7 (10) ◽  
pp. 852
Author(s):  
Caroline Maria Marcos ◽  
Haroldo Cesar de Oliveira ◽  
Patrícia Akemi Assato ◽  
Rafael Fernando Castelli ◽  
Ana Marisa Fusco-Almeida ◽  
...  
Keyword(s):  
Cell Wall ◽  
Latin American ◽  
Histidine Kinase ◽  
Paracoccidioides Brasiliensis ◽  
Yeast Cells ◽  
Lower Amount ◽  
Stress Adaptation ◽  
Infection Model ◽  
Dimorphic Fungus ◽  
Virulence Attenuation

P. brasiliensis is a thermally dimorphic fungus belonging to Paracoccidioides complex, causative of a systemic, endemic mycosis limited to Latin American countries. Signal transduction pathways related to important aspects as surviving, proliferation according to the biological niches are linked to the fungal pathogenicity in many species, but its elucidation in P. brasiliensis remains poorly explored. As Drk1, a hybrid histidine kinase, plays regulators functions in other dimorphic fungi species, mainly in dimorphism and virulence, here we investigated its importance in P. brasilensis. We, therefore generated the respective recombinant protein, anti-PbDrk1 polyclonal antibody and a silenced strain. The Drk1 protein shows a random distribution including cell wall location that change its pattern during osmotic stress condition; moreover the P. brasiliensis treatment with anti-PbDrk1 antibody, which does not modify the fungus’s viability, resulted in decreased virulence in G. mellonella model and reduced interaction with pneumocytes. Down-regulating PbDRK1 yielded phenotypic alterations such as yeast cells with more elongated morphology, virulence attenuation in G. mellonella infection model, lower amount of chitin content, increased resistance to osmotic and cell wall stresses, and also caspofungin, and finally increased sensitivity to itraconazole. These observations highlight the importance of PbDrk1 to P. brasiliensis virulence, stress adaptation, morphology, and cell wall organization, and therefore it an interesting target that could help develop new antifungals.

scholarly journals Paracoccin overexpression in Paracoccidioides brasiliensis reveals the influence of chitin hydrolysis on fungal virulence and host immune response

2019 ◽  
Author(s):  
Relber Aguiar Gonçales ◽  
Vanessa Cristina Silva Vieira ◽  
Rafael Ricci-Azevedo ◽  
Fabrício Freitas Fernandes ◽  
Sandra Maria de Oliveira Thomaz ◽  
...  
Keyword(s):  
Immune Response ◽  
Host Response ◽  
Paracoccidioides Brasiliensis ◽  
In Vitro Assays ◽  
Significant Advance ◽  
Fungal Virulence ◽  
Tnf Α ◽  
Chitin Hydrolysis ◽  
Yeast Genes

ABSTRACTParacoccidioides brasiliensis and P. lutzii, etiological agents of paracoccidioidomycosis (PCM), develop as mycelia at 25-30 °C and as yeast at 35-37 °C. Only a few Paracoccidioides spp. proteins are well characterized. Thus, we studied paracoccin (PCN) from P. brasiliensis, its role in the fungus biology, and its relationship with the host innate immune cells. Cloning and heterologous expression analysis revealed its lectin, enzymatic, and immunomodulatory properties. Recently, we employed a system based on Agrobacterium tumefaciens-mediated transformation to manipulate P. brasiliensis yeast genes to obtain clones knocked-down for PCN, which after all, are unable to transit from yeast to mycelium forms, causing a mild pulmonary disease. Herein, we generate P. brasiliensis overexpressing PCN (ov-PCN). To date, it was not explored the overexpressing of endogenous components in Paracoccidioides spp. Therefore, we investigate the role of PCN in fungal biology and pathogenesis. Augmented levels of PCN mRNA and protein, and N-acetylglucosaminidase activity confirmed PCN overexpression in ov-PCN of P. brasiliensis yeasts. Interestingly, PCN overexpression did not affect the yeasts’ growth or viability and favored cell separation. The ov-PCN clones transitioned faster to the mycelium form than the wt-PCN yeasts. Concerning infection, while most of mice infected with the wt-yeasts (90%) survive at least until the 70th day, all mice infected with ov-PCN yeasts were already died at the 35th day post-infection. In vitro assays showed that ov-PCN were more susceptible to phagocytosis by macrophages. Finally, it was verified that the chitin particles isolated from the ov-PCN cells were smaller than those obtained from the wt-PCN yeasts. Macrophages stimulated with the chitin isolated from ov-PCN produce IL-10, whereas the particles with a wider size range harvested from wt-PCN yeasts induced TNF-α and IL-1β secretion. The anti-inflammatory microenvironment from macrophage stimulation with small chitin particles hampers the development of a protective immune response against the fungus. We postulated that the high grade of chitin cleavage, as the results of augmented PCN expression, favors pathogenesis following P. brasiliensis infection. Thus, PCN is a relevant virulence fungal factor.AUTHOR SUMMARYParacoccidioides spp. are pathogenic fungi that cause paracoccidioidomycosis (PCM) in humans, the main deep mycosis of Latin America. Recently, by knocking down the paracoccin gene, our group showed that this lectin is necessary for the morphological transition from yeast to hyphae, and that this decrease results in low P. brasiliensis virulence. Here, after overexpress PCN, we revealed the importance of the yeast chitin hydrolysis to the host response. Infection of mice with ov-PCN yeasts causes severe lung disease compared to moderate disease caused by wt-PCN yeasts. The release of smaller chitin particles was as a result of an accelerated chitin hydrolysis provided by ov-PCN yeasts. Interestingly, these smallest chitin particles are able to modulate host response by increasing IL-10 in the meantime that decrease TNF-α secretion, thus hampering Th1 immune response that is crucial in the fight against this fungi. These findings represent a significant advance in the knowledge about the role of PCN chitinase in P. brasiliensis.

scholarly journals De-Coding the Contributions of the Viral RNAs to Alphaviral Pathogenesis

Pathogens ◽  
2021 ◽  
Vol 10 (6) ◽  
pp. 771
Author(s):  
Autumn T. LaPointe ◽  
Kevin J. Sokoloski
Keyword(s):  
Host Response ◽  
Severe Disease ◽  
Review Article ◽  
Structural Proteins ◽  
Virulence Determinants ◽  
Healthy Individuals ◽  
Viral Rnas ◽  
Positive Sense ◽  
Identification And Characterization

Alphaviruses are positive-sense RNA arboviruses that are capable of causing severe disease in otherwise healthy individuals. There are many aspects of viral infection that determine pathogenesis and major efforts regarding the identification and characterization of virulence determinants have largely focused on the roles of the nonstructural and structural proteins. Nonetheless, the viral RNAs of the alphaviruses themselves play important roles in regard to virulence and pathogenesis. In particular, many sequences and secondary structures within the viral RNAs play an important part in the development of disease and may be considered important determinants of virulence. In this review article, we summarize the known RNA-based virulence traits and host:RNA interactions that influence alphaviral pathogenesis for each of the viral RNA species produced during infection. Overall, the viral RNAs produced during infection are important contributors to alphaviral pathogenesis and more research is needed to fully understand how each RNA species impacts the host response to infection as well as the development of disease.

scholarly journals Genome and Secretome Analysis of Staphylotrichum longicolleum DSM105789 Cultured on Agro-Residual and Chitinous Biomass

2021 ◽  
Vol 9 (8) ◽  
pp. 1581
Author(s):  
Arslan Ali ◽  
Bernhard Ellinger ◽  
Sophie C. Brandt ◽  
Christian Betzel ◽  
Martin Rühl ◽  
...  
Keyword(s):  
Sugarcane Bagasse ◽  
Chitinase Activity ◽  
Growth Conditions ◽  
Glycoside Hydrolases ◽  
Carbohydrate Binding ◽  
Waste Products ◽  
Enzyme Mixture ◽  
Secretome Analysis ◽  
Polysaccharide Lyases ◽  
Carbohydrate Esterases

Staphylotrichum longicolleum FW57 (DSM105789) is a prolific chitinolytic fungus isolated from wood, with a chitinase activity of 0.11 ± 0.01 U/mg. We selected this strain for genome sequencing and annotation, and compiled its growth characteristics on four different chitinous substrates as well as two agro-industrial waste products. We found that the enzymatic mixture secreted by FW57 was not only able to digest pre-treated sugarcane bagasse, but also untreated sugarcane bagasse and maize leaves. The efficiency was comparable to a commercial enzymatic cocktail, highlighting the potential of the S. longicolleum enzyme mixture as an alternative pretreatment method. To further characterize the enzymes, which efficiently digested polymers such as cellulose, hemicellulose, pectin, starch, and lignin, we performed in-depth mass spectrometry-based secretome analysis using tryptic peptides from in-gel and in-solution digestions. Depending on the growth conditions, we were able to detect from 442 to 1092 proteins, which were annotated to identify from 134 to 224 putative carbohydrate-active enzymes (CAZymes) in five different families: glycoside hydrolases, auxiliary activities, carbohydrate esterases, polysaccharide lyases, glycosyl transferases, and proteins containing a carbohydrate-binding module, as well as combinations thereof. The FW57 enzyme mixture could be used to replace commercial enzyme cocktails for the digestion of agro-residual substrates.

scholarly journals Membrane-Associated Proteins of a Lipopolysaccharide-Deficient Mutant of Neisseria meningitidis Activate the Inflammatory Response through Toll-Like Receptor 2

2001 ◽  
Vol 69 (4) ◽  
pp. 2230-2236 ◽  
Author(s):  
Robin R. Ingalls ◽  
Egil Lien ◽  
Douglas T. Golenbock
Keyword(s):  
Cell Wall ◽  
Inflammatory Response ◽  
Neisseria Meningitidis ◽  
Parental Strain ◽  
Host Responses ◽  
Deficient Mutant ◽  
Toll Like Receptor ◽  
Gram Negative ◽  
Negative Cell ◽  
Toll Like Receptor 2

ABSTRACT The recent isolation of a lipopolysaccharide (LPS)-deficient mutant of Neisseria meningitidis has allowed us to explore the roles of other gram-negative cell wall components in the host response to infection. The experiments in this study were designed to examine the ability of this mutant strain to activate cells. Although it was clearly less potent than the parental strain, we found the LPS-deficient mutant to be a capable inducer of the inflammatory response in monocytic cells, inducing a response similar to that seen with Staphylococcus aureus. Cellular activation by the LPS mutant was related to expression of CD14, a high-affinity receptor for LPS and other microbial products, as well as Toll-like receptor 2, a member of the Toll family of receptors recently implicated in host responses to gram-positive bacteria. In contrast to the parental strain, the synthetic LPS antagonist E5564 did not inhibit the LPS-deficient mutant. We conclude that even in the absence of LPS, the gram-negative cell wall remains a potent inflammatory stimulant, utilizing signaling pathways independent of those involved in LPS signaling.

Host response to cytoadherence in Plasmodium falciparum

2008 ◽  
Vol 36 (2) ◽  
pp. 221-228 ◽  
Author(s):  
Srabasti J. Chakravorty ◽  
Katie R. Hughes ◽  
Alister G. Craig
Keyword(s):  
Plasmodium Falciparum ◽  
Tumour Necrosis ◽  
Host Response ◽  
Severe Disease ◽  
Signalling Pathways ◽  
Malarial Infection ◽  
Extensive Information ◽  
Adhesive Interactions ◽  
Necrosis Factor ◽  
Tnf Tumour Necrosis Factor

Cytoadherence of PRBCs (Plasmodium falciparum-infected red blood cells) to host endothelium has been associated with pathology in severe malaria, but, despite extensive information on the primary processes involved in the adhesive interactions, the mechanisms underlying the disease are poorly understood. Endothelial cells have the ability to mobilize immune and pro-adhesive responses when exposed to both PRBCs and TNF (tumour necrosis factor). In addition, there is also an up-regulation by PRBCs and TNF and a concurrent down-regulation of a range of genes involved in inflammation and cell death, by PRBCs and TNF. We propose that the balance between positive and negative regulation will contribute to endothelial pathology during malarial infection. Apposition of PRBCs has been shown by a number of groups to activate signalling pathways. This is dependent, at least in part, on the cytoadherence characteristics of the invading isolate, such that the avidity of the PRBC for the receptor on host endothelium is proportional to the level of activation of the signalling pathways. An understanding of the post-adhesive processes produced by cytoadherence may help us to understand the variable pathology seen in malaria and to design appropriate therapies to alleviate severe disease.

scholarly journals Had1 Is Required for Cell Wall Integrity and Fungal Virulence inCryptococcus neoformans

2017 ◽  
Vol 8 (2) ◽  
pp. 643-652 ◽  
Author(s):  
Won-Hee Jung ◽  
Ye-Eun Son ◽  
Sang-Hun Oh ◽  
Ci Fu ◽  
Hye Shin Kim ◽  
...  
Keyword(s):  
Cell Wall ◽  
Cell Wall Integrity ◽  
Fungal Virulence

scholarly journals OPTIMALISASI PRODUKSI ENZIM KITINASE PADA ISOLAT JAMUR KITINOLITIK DARI SAMPEL TANAH RIZOSFER

2018 ◽  
Vol 3 (01) ◽  
pp. 62-69
Author(s):  
Eka Corneliyawati ◽  
Massora Massora ◽  
Khikmah Khikmah ◽  
As’ad Syamsul Arifin
Keyword(s):  
Biological Control ◽  
Cell Wall ◽  
Rhizosphere Soil ◽  
Plant Root ◽  
Optimal Growth ◽  
Chitinase Activity ◽  
Fungal Species ◽  
Plant Roots ◽  
Chitinase Production

The rhizosphere is the zone of soil surrounding a plant root where plant roots, soil and the soil biota interact with each other. Chitinolytic fungi has been effectively used in biological control agens. The chitinase activity causes lysis of the fungi cell wall pathogen. The aim of the research was to find optimization of activity chitinase enzyme from rhizosphere soil was conducted in vitro. Optimal growth chitinase production for TKR3 fungi isolate were concentration of chitin 0,2% (b/v), pH 5,5, temperature 30ºC, agitation 150 rpm and incubation time at four days. The optimum yield of chitinase production is influenced by fungal species and environmental conditions.

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