6-Acetylbisdethiobis(methylthio)gliotoxin

Gliotoxin is a fungal secondary metabolite with impact on health and agriculture since it might act as virulence factor and contaminate human and animal food. Homologous gliotoxin (GT) gene clusters are spread across a number of fungal species although if they produce GT or other related epipolythiodioxopiperazines (ETPs) remains obscure. Using bioinformatic tools, we have identified homologous gli gene clusters similar to the A. fumigatus GT gene cluster in several fungal species. In silico study led to in vitro confirmation of GT and Bisdethiobis(methylthio)gliotoxin (bmGT) production in fungal strain cultures by HPLC detection. Despite we selected most similar homologous gli gene cluster in 20 different species, GT and bmGT were only detected in section Fumigati species and in a Trichoderma virens Q strain. Our results suggest that in silico gli homology analyses in different fungal strains to predict GT production might be only informative when accompanied by analysis about mycotoxin production in cell cultures.

As recently described, fungal secondary metabolism activates during infection in response to a hostile host environment. Gliotoxin and bis(methylthio)gliotoxin are two recognized secondary metabolites produced by Aspergillus fumigatus with differential cytotoxicity and involved in virulence. We sought to describe the temporal dynamics of gliotoxin and bis(methylthio)gliotoxin during A. fumigatus progression to further explore their role in the infection. First, we optimized the production of the mycotoxins under different in vitro growth conditions and then specifically measured them using an UHPLC/PDA method. The analytical conditions were selected after testing different parameters such as extraction procedures, column type, and mobile phase composition. We found that gliotoxin and bis(methylthio)gliotoxin are differentially excreted to the extracellular media during the course of A. fumigatus infection regardless of the growth format tested. Dynamic profiles show an early production of gliotoxin, which, after reaching a maximum, decreases coinciding with the increase in the production of the inactive derivative bis(methylthio)gliotoxin. Presence of gliotoxin may indicate an early phase of fungal development, whereas detection of bis(methylthio)gliotoxin may correspond to a more advanced stage of infection. Our chromatographic method successfully characterizes these secondary metabolites. Thus, it may potentially be used to further understand Aspergillus infection. Lay summary: Aspergillus fumigatus secondary metabolites may contribute to fungal survival. A new chromatographic method was applied to simultaneously characterize two relevant metabolites. Presence of toxic gliotoxin may indicate an early phase of development, whereas the detection of the inactive derivate may represent an advanced infection stage.

Background: Invasive pulmonary aspergillosis (IPA) remains a life-threatening opportunistic infection, but can be difficult to diagnose. New biomarkers are therefore needed. Gliotoxin (GT), a secondary metabolite of Aspergillus fumigatus, and bis(methylthio)gliotoxin (bmGT), a degradation product of GT, have been proposed as potential biomarkers. However, these findings have yet to be confirmed. Objectives: To identify the diagnostic potential of GT and bmGT in serum and bronchoalveolar lavage fluid (BALf) in haematology patients compared to galactomannan (GM). Materials and methods: We prospectively collected culture supernatant, serum and BALf from patients with culture-positive IPA and measured GT and bmGT concentrations using ultra high-performance liquid chromatography-quadrupole time of flight mass spectrometry. Galactomannan was detected using a commercially available enzyme immunoassay. Results: We included 18 patients with proven (n = 6) and probable (n = 12) IPA, all with positive cultures for Aspergillus fumigatus. BmGT was only detected in serum from one patient (5.6%), whereas GM was positive (optical density ≥ 0.5) in 11/18 patients (61.1%, P = 0.002). We could not find GT in any serum sample. In BALf, bmGT was detected in 8/18 patients (44.4%) and GT in 9/18 patients (50%), compared to GM (optical density ≥ 1.0) in all patients (100%). Conclusions: Gliotoxin and bis(methylthio)gliotoxin had a very poor performance for diagnosing IPA. As other biomarkers are more sensitive and easier to detect, we would not recommend serum or BALf GT/bmGT to be used in the diagnosis of IPA.