title: In vitro Paracoccidioides brasiliensis biofilm and gene expression of adhesins and hydrolytic enzymes.
abstract: Paracoccidioides species are dimorphic fungi that initially infect the lungs but can also spread throughout the body. The spreading infection is most likely due to the formation of a biofilm that makes it difficult for the host to eliminate the infection. Biofilm formation is crucial for the development of infections and confines the pathogen to an extracellular matrix. Its presence is associated with antimicrobial resistance and avoidance of host defenses. This current study provides the first description of biofilm formation by Paracoccidioides brasiliensis (Pb18) and an analysis of gene expression, using real-time PCR, associated with 3 adhesins and 2 hydrolytic enzymes that could be associated with the virulence profile. Biofilm formation was analyzed using fluorescence microscopy, scanning electron microscopy (SEM) and confocal laser scanning microscopy (CLSM). Metabolic activity was determined using the XTT reduction assay. P. brasiliensis was able to form mature biofilm in 144 h with a thickness of 100 μm. The presence of a biofilm was found to be associated with an increase in the expression of adhesins and enzymes. GP43, enolase, GAPDH and aspartyl proteinase genes were over-expressed, whereas phospholipase was down-regulated in biofilm. The characterization of biofilm formed by P. brasiliensis may contribute to a better understanding of the pathogenesis of paracoccidioidomycosis as well as the search for new therapeutic alternatives; while improving the effectiveness of treatment.
authors.affiliation: Departamento de Anáalises Clínicas; Laboratório de Micologia Clínica; Faculdade de Ciências Farmacêuticas; UNESP - Univ. Estadual Paulista; Araraquara, Brasil
authors.forename: Janaina de Cássia Orlandi
authors.initials: Jde C
authors.lastname: Sardi
authors.affiliation: Departamento de Anáalises Clínicas; Laboratório de Micologia Clínica; Faculdade de Ciências Farmacêuticas; UNESP - Univ. Estadual Paulista; Araraquara, Brasil
authors.forename: Nayla de Souza
authors.initials: Nde S
authors.lastname: Pitangui
authors.affiliation: Departamento de Anáalises Clínicas; Laboratório de Micologia Clínica; Faculdade de Ciências Farmacêuticas; UNESP - Univ. Estadual Paulista; Araraquara, Brasil
authors.forename: Aline Raquel
authors.initials: AR
authors.lastname: Voltan
authors.affiliation: Departamento de Anáalises Clínicas; Laboratório de Micologia Clínica; Faculdade de Ciências Farmacêuticas; UNESP - Univ. Estadual Paulista; Araraquara, Brasil
authors.forename: Jaqueline Derissi
authors.initials: JD
authors.lastname: Braz
authors.affiliation: Departamento de Patologia ; Instituto Oswaldo Cruz ; Rio de Janeiro , Brasil.
authors.forename: Marcelo Pelajo
authors.initials: MP
authors.lastname: Machado
authors.affiliation: Departamento de Anáalises Clínicas; Laboratório de Micologia Clínica; Faculdade de Ciências Farmacêuticas; UNESP - Univ. Estadual Paulista; Araraquara, Brasil
authors.forename: Ana Marisa
authors.initials: AM
authors.lastname: Fusco Almeida
authors.affiliation: Departamento de Anáalises Clínicas; Laboratório de Micologia Clínica; Faculdade de Ciências Farmacêuticas; UNESP - Univ. Estadual Paulista; Araraquara, Brasil
authors.forename: Maria Jose Soares
authors.initials: MJ
authors.lastname: Mendes Giannini
mesh_terms: D018441:Biofilms / Q000254:growth & development*; D002478:Cells, Cultured; D005562:Formazans / Q000032:analysis; D020869:Gene Expression Profiling; D006801:Humans; D006867:Hydrolases / Q000032:analysis*; D018613:Microscopy, Confocal; D008855:Microscopy, Electron, Scanning; D008856:Microscopy, Fluorescence; D010084:Oxidation-Reduction; D010228:Paracoccidioides / Q000201:enzymology* / Q000378:metabolism / Q000502:physiology*; D060888:Real-Time Polymerase Chain Reaction; D013778:Tetrazolium Salts / Q000378:metabolism; D037521:Virulence Factors / Q000032:analysis*
chemical_list: D005562:Formazans; D013778:Tetrazolium Salts; D037521:Virulence Factors; C059087:2,3-bis(2-methoxy-4-nitro-5-sulfophenyl)-5-((phenylamino)carbonyl)-2H-tetrazolium hydroxide; C059088:1,5-bis(2-methoxy-4-nitro-5-sulfophenyl)-3-((phenylamino)carbonyl)formazan; D006867:Hydrolases
keywords: Paracoccidioides brasiliensis; adhesins; biofilms; gene expression; real-time PCR; virulence factors