Purification and characterization of factors produced by Aspergillus fumigatus which affect human ciliated respiratory epithelium

Abstract

The mechanisms by which Aspergillus fumigatus colonizes the respiratory mucosa are unknown. Culture filtrates of eight of nine clinical isolates of A. fumigatus slowed ciliary beat frequency and damaged human respiratory epithelium in vitro. These changes appeared to occur concurrently. Culture filtrates of two clinical isolates of Candida albicans had no effect on ciliated epithelium. We have purified and characterized cilioinhibitory factors of a clinical isolate of A. fumigatus. The cilioinhibitory activity was heat labile, reduced by dialysis, and partially extractable into chloroform. The activity was associated with both high-and lowmolecular-weight factors, as determined by gel filtration on Sephadex G-50. A low-molecular-weight cilioinhibitory factor was further purified by reverse-phase high-performance liquid chromatography and shown by mass spectrometry to be gliotoxin, a known metabolite of A. fumigatus. Gliotoxin significantly slowed ciliary beat frequency in association with epithelial damage at concentrations above 0.2 g/ml; other Aspergillus toxins, i.e., fumagillin and helvolic acid, were also cilioinhibitory but at much higher concentrations. Highmolecular-weight (>35,000 and 25,000) cilioinhibitory materials had neither elastolytic nor proteolytic activity and remain to be identified. Thus, A. fumigatus produces a number of biologically active substances which slow ciliary beating and damage epithelium and which may influence colonization of the airways.

Redundant synthesis of a conidial polyketide by two distinct secondary metabolite clusters in Aspergillus fumigatus

“…This species is notorious for producing a plethora of SMs (e.g. gliotoxin, helvolic acid, hexadehydroastechrome, trypacidin, endocrocin, neosartoricin, and fumagillin) shown to exhibit immunomodulatory and/or cytotoxic properties that are thought to facilitate the pathogenicity of this fungus (Amitani et al, 1995;Bok et al, 2005;Spikes et al, 2008;Lodeiro et al, 2009;Gauthier et al, 2012;Berthier et al, 2013;Chooi et al, 2013;Yin et al, 2013), although their ecological role may relate to their antimicrobial activity (Carberry et al, 2012;Kang et al, 2013) and UV protective capacity (Allam and Abd El-Zaher, 2012).…”

“…Secondary metabolism is spatially and temporally regulated in fungi with metabolites produced in different compartments of the fungal thallus Kistler and Broz, 2015). Mycelial SMs such as gliotoxin and helvolic acid have been extensively studied over the last decades (Amitani et al, 1995;Bok et al, 2006;Spikes et al, 2008;Kwon-Chung and Sugui, 2009). However, with the exception of dihydroxynaphthalene (DHN) melanin (Jahn et al, 1997;Sugui et al, 2007;Bayry et al, 2014), the spore-borne SMs are relatively understudied, despite spores being crucial to the fungus for dispersal and colonization of new substrates.…”

Detection of early phase specific surface appendages during germination of Aspergillus fumigatus conidia

“…The common route of infection is inhalation of conidia and the major di¡erence between A. fumigatus and less pathogenic aspergilli is that A. fumigatus conidia reach the lower parts of the lung where they start to multiply. A reason for this di¡erence might be the ability of A. fumigatus conidia to inhibit the ciliary activity of the mucous epithelium in the respiratory tract [5]. In the lung of healthy individuals A. fumigatus cells are attacked and killed by alveolar macrophages and neutrophils [6].…”

Excursions in biomedical mass spectrometry

“…purification), two ADP-ribosyl DEA-BAG anomers were is metabolised by reductive methylation to bis-dethiobis-(thiomethyl)-gliotoxin which is has no biological observed in the h.p.l.c.-MS profile (Figure 4 ); analysis of the electrospray mass spectrum of peaks I and II indicated effect on CBF. Neither gliotoxin nor its metabolite are amenable to GC-MS analysis, however, they were that the major ion species was m/z 775 corresponding to the protonated molecular ion (M+H+) of the anomers; readily identified in culture filtrates of A. fumigatus by microbore h.p.l.c.-electrospray MS (Figure 3 ) [33]. We these peaks were absent in the control incubations with no cells.…”