Cell Wall Modifications during Conidial Maturation of the Human Pathogenic Fungus Pseudallescheria boydii

Abstract

Progress in extending the life expectancy of cystic fibrosis (CF) patients remains jeopardized by the increasing incidence of fungal respiratory infections. Pseudallescheria boydii (P. boydii), an emerging pathogen of humans, is a filamentous fungus frequently isolated from the respiratory secretions of CF patients. It is commonly believed that infection by this fungus occurs through inhalation of airborne conidia, but the mechanisms allowing the adherence of Pseudallescheria to the host epithelial cells and its escape from the host immune defenses remain largely unknown. Given that the cell wall orchestrates all these processes, we were interested in studying its dynamic changes in conidia as function of the age of cultures. We found that the surface hydrophobicity and electronegative charge of conidia increased with the age of culture. Melanin that can influence the cell surface properties, was extracted from conidia and estimated using UV-visible spectrophotometry. Cells were also directly examined and compared using electron paramagnetic resonance (EPR) that determines the production of free radicals. Consistent with the increased amount of melanin, the EPR signal intensity decreased suggesting polymerization of melanin. These results were confirmed by flow cytometry after studying the effect of melanin polymerization on the surface accessibility of mannose-containing glycoconjugates to fluorescent concanavalin A. In the absence of melanin, conidia showed a marked increase in fluorescence intensity as the age of culture increased. Using atomic force microscopy, we were unable to find rodlet-forming hydrophobins, molecules that can also affect conidial surface properties. In conclusion, the changes in surface properties and biochemical composition of the conidial wall with the age of culture highlight the process of conidial maturation. Mannose-containing glycoconjugates that are involved in immune recognition, are progressively masked by polymerization of melanin, an antioxidant that is commonly thought to allow fungal escape from the host immune defenses.

Production of melanin pigments in saprophytic fungi in vitro and during infection

“…MAb 8D6 also confirmed melanin production in P. boydii , which confirms earlier work using electron paramagnetic resonance (EPR) and is supported by the identification of two genes, PKS type I ( PKSI ) and tetrahydroxynaphtalene reductase ( 4HNR ), involved in the DHN‐melanin biosynthesis pathway .…”

A Multifaceted Study of Scedosporium boydii Cell Wall Changes during Germination and Identification of GPI-Anchored Proteins

“…The fungal strain S . boydii IHEM 15155 (formerly Pseudallescheria boydii ) was used throughout this study [ 29 ]. It was maintained on yeast extract-peptone-dextrose (YPD; 0.5% w/v yeast extract, 2% w/v glucose, 1% w/v peptone, 0.05% w/v chloramphenicol) agar plates.…”

“…Cell surface charge and hydrophobicity were evaluated by microelectrophoresis and two-phase partitioning as described previously [ 29 ]. For evaluation of the cell surface charge, resting or germinating conidia (10 6 cells) were washed and resuspended in Milli-Q water containing 1 mM NaCl, then their electrophoretic mobility was measured at 25°C using Zetasizer Nano ZS (Malvern Instruments Ltd, Malvern, UK).…”

“…We previously demonstrated that the conidial cell wall of S . boydii contains dihydroxynaphtalene (DHN)-melanin and that the cell wall content in melanin and mannose-containing glycoconjugates increases during maturation of conidia along with the cell surface physical properties [ 29 ]. Here, we tracked the cell wall modifications during the germination process using various approaches, including investigation, at the molecular level, of glycosylphosphatidylinositol (GPI)-anchored proteins in conidial and hyphal walls as these integral cell wall proteins (CWPs) play a major role in normal morphology and virulence in other fungal models [ 30 – 32 ].…”

A case of chronic unilateral maxillary sinusitis by Pseudoallescheria boydii

“…The mechanisms of adherence and establishment of an infection by these fungi in the lung are still largely unknown. It is thought that the infection process in the respiratory tract starts by inhalation and adhesion of airborne conidia that differentiate into hyphae, with both processes mediated by the spore cell wall since that acts as the interface between the fungus and lung tissues [3]. Aspergillus fumigatus is the most common species implicated in paranasal sinus infection, with other species being rarely reported.…”