Antifungal activity of microbial secondary metabolites (original) (raw)
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Induction of Gliotoxin Secretion in Aspergillus fumigatus by Bacteria-Associated Molecules
PLoS ONE, 2014
Aspergillus fumigatus is the most common causative agent of mold diseases in humans, giving rise to life-threatening infections in immunocompromised individuals. One of its secreted metabolites is gliotoxin, a toxic antimicrobial agent. The aim of this study was to determine whether the presence of pathogen-associated molecular patterns in broth cultures of A. fumigatus could induce gliotoxin production. Gliotoxin levels were analyzed by ultra-performance liquid chromatography and mass spectrometry. The presence of a bacteria-derived lipopolysaccharide, peptidoglycan, or lipoteichoic acid in the growth media at a concentration of 5 mg/ml increased the gliotoxin concentration in the media by 37%, 65%, and 35%, respectively. The findings reveal a correlation between the concentrations of pathogen-associated molecular patterns and gliotoxin secretion. This shows that there is a yet uncharacterized detection system for such compounds within fungi. Inducing secondary metabolite production by such means in fungi is potentially relevant for drug discovery research. Our results also give a possible explanation for the increased virulence of A. fumigatus during bacterial co-infection, one that is important for the transition from colonization to invasiveness in this pulmonary disease.
2019
ABSTRACTAspergillus fumigatus is a major opportunistic human pathogen. Multiple traits contribute to A. fumigatus pathogenicity, including its ability to produce specific secondary metabolites, such as gliotoxin. Gliotoxin is known to inhibit the host immune response, and genetic mutants that inactivate gliotoxin biosynthesis (or secondary metabolism in general) attenuate A. fumigatus virulence. The genome of A. fischeri, a very close non-pathogenic relative of A. fumigatus, contains a biosynthetic gene cluster that exhibits high sequence similarity to the A. fumigatus gliotoxin cluster. However, A. fischeri is not known to produce gliotoxin. To gain further insight into the similarities and differences between the major pathogen A. fumigatus and the non-pathogen A. fischeri, we examined whether A. fischeri strain NRRL 181 biosynthesizes gliotoxin and whether its production, and of secondary metabolites more generally, influence its virulence profile. We found that A. fischeri biosy...
Molecular Microbiology, 2006
Gliotoxin is a secondary metabolite produced by several fungi including the opportunistic human pathogen Aspergillus fumigatus. As gliotoxin exerts immunosuppressive effects in vitro and in vivo, a role as a virulence determinant in invasive aspergillosis has been discussed for a long time but evidence has not been provided until now. Here, by the use of different selection marker genes A. fumigatus knock-out strains were generated that are deficient for the nonribosomal peptide synthetase GliP, the putative key enzyme of the gliotoxin biosynthesis. Deletion of the gliP gene resulted in loss of gliotoxin production, as analysed by high performance liquid chromatography and tandem mass spectrometry. No differences in morphology or growth kinetics between wild-type and gliP-deletion strains were observed. In vitro, the culture supernatant of the gliP-deficient strains showed a reduced cytotoxic effect on both macrophage-like cells and T cell lines. In a low-dose murine infection model of invasive aspergillosis, gliotoxin was detected in the lung and absent when mice were infected with the gliP deletion strain. However, gliP deletion strains showed no difference in virulence compared with the corresponding wildtype strains. Taken together, the non-ribosomal peptide synthetase GliP is essential for gliotoxin production in A. fumigatus. Gliotoxin is not required for pathogenicity of the fungus in immunocompromised mice, despite the fact that a reduced cytotoxicity of the culture supernatant of gliP deletion strains was demonstrated.
Eukaryotic Cell, 2006
The fungal secondary metabolite gliotoxin produced by Aspergillus fumigatus has been hypothesized to be important in the development of invasive aspergillosis. In this study, we addressed this hypothesis by disrupting a nonribosomal peptide synthetase (NRPS) (encoded by gliP) predicted to be involved in gliotoxin production. Mutants with a disrupted gliP locus failed to produce gliotoxin, which confirmed the role of the NRPS encoded by gliP in gliotoxin biosynthesis. We found no morphological, developmental, or physiological defects in ⌬gliP mutant strains. In addition, disruption of gliP resulted in down regulation of gene expression in the gliotoxin biosynthesis gene cluster, which was restored with addition of exogenous gliotoxin. This interesting result suggests a role for gliotoxin in regulating its own production. Culture filtrates from the ⌬gliP mutant were unable to inhibit ionomycin-dependent degranulation of mast cells, suggesting a role for gliotoxin in suppressing mast cell degranulation and possibly in disease development. However, the ⌬gliP mutant did not have an impact on survival or tissue burden in a murine inhalational model of invasive aspergillosis. This result suggests that gliotoxin is not required for virulence in an immunosuppressed host with an invasive pulmonary infection.
PLoS Pathogens, 2010
Gliotoxin, and other related molecules, are encoded by multi-gene clusters and biosynthesized by fungi using nonribosomal biosynthetic mechanisms. Almost universally described in terms of its toxicity towards mammalian cells, gliotoxin has come to be considered as a component of the virulence arsenal of Aspergillus fumigatus. Here we show that deletion of a single gene, gliT, in the gliotoxin biosynthetic cluster of two A. fumigatus strains, rendered the organism highly sensitive to exogenous gliotoxin and completely disrupted gliotoxin secretion. Addition of glutathione to both A. fumigatus DgliT strains relieved gliotoxin inhibition. Moreover, expression of gliT appears to be independently regulated compared to all other cluster components and is up-regulated by exogenous gliotoxin presence, at both the transcript and protein level. Upon gliotoxin exposure, gliT is also expressed in A. fumigatus DgliZ, which cannot express any other genes in the gliotoxin biosynthetic cluster, indicating that gliT is primarily responsible for protecting this strain against exogenous gliotoxin. GliT exhibits a gliotoxin reductase activity up to 9 mM gliotoxin and appears to prevent irreversible depletion of intracellular glutathione stores by reduction of the oxidized form of gliotoxin. Cross-species resistance to exogenous gliotoxin is acquired by A. nidulans and Saccharomyces cerevisiae, respectively, when transformed with gliT. We hypothesise that the primary role of gliotoxin may be as an antioxidant and that in addition to GliT functionality, gliotoxin secretion may be a component of an auto-protective mechanism, deployed by A. fumigatus to protect itself against this potent biomolecule.
Gliotoxin production by clinical and environmental Aspergillus fumigatus strains
International Journal of Medical Microbiology, 2008
The mycotoxin gliotoxin is produced by fungi of the genus Aspergillus, including the important human pathogen Aspergillus fumigatus. Gliotoxin exerts a broad spectrum of immunosuppressive effects in vitro and is detectable in the sera of patients suffering from invasive aspergillosis. In order to correlate the pathogenic potential of A. fumigatus with the ability to produce gliotoxin and to investigate the taxonomic distribution of gliotoxin-producing Aspergillus strains among clinical isolates, a total of 158 Aspergillus isolates comprising four different species (A. fumigatus, n ¼ 100; A. terreus, n ¼ 27; A. niger, n ¼ 16; A. flavus, n ¼ 15) were collected from different medical centers (some originating from probable cases of aspergillosis) and from environmental samples in Germany and Austria. Remarkably, gliotoxin was detected in most culture filtrates of A. fumigatus of both clinical (98%) and environmental (96%) origin. The toxin was also detected, with decreasing frequency, in culture filtrates of A. niger (56%), A. terreus (37%), and A. flavus (13%). The highest gliotoxin concentrations were detected in A. fumigatus strains of clinical (max. 21.35 mg/ml, mean 5.75 mg/ml) and environmental (max. 26.25 mg/ml, mean 5.27 mg/ml) origin. Gliotoxin productivity of other Aspergillus species was significantly lower. Culture supernatants of A. fumigatus strains lacking gliotoxin production showed a significantly lower cytotoxicity on macrophage-like cells and T-cells in vitro. In contrast, lack of gliotoxin production in the other Aspergillus species tested had no significant influence on the cytotoxic effect of culture supernatant on these immune cells.
Improving Conditions for Gliotoxin Production by Local Isolates of Aspergillus fumigatus
Journal of Biotechnology Research Center, 2017
Thirty two isolates of Aspergillus fumigatus were obtained from a total of 44 samples of sputum, nose swab and tracheal aspirate from suspected patient with aspergillosis were also collected from February 2014 to June 2014. A morphological examination of A. fumigatus was first made with naked eye and at low magnification power of microscope after that detailed examination was done by measuring the dimensions of the microscopic structures, photographing the microscopic structures and using relevant literature. Results appeared conical-shaped terminal vesicles, uniseriate row of phialides on the upper two thirds of the vesicle, conidiophore stipes were short, phialides arrange uniseriate upper vesicle conidia and parallel to axis of conidiophore, produced in chains of spore basipetally from phialides, the chains of spore were borne directly in the absence of metulae and represented by septet and branching hyphae. The ability of A. fumigatus for GT production was investigated using t...
Regulation of gliotoxin biosynthesis and protection in Aspergillus species
2021
Aspergillus fumigatus causes a range of human and animal diseases collectively known as aspergillosis. A. fumigatus possesses and expresses a range of genetic determinants of virulence, which facilitate colonisation and disease progression, including the secretion of mycotoxins. Gliotoxin (GT) is the best studied A. fumigatus mycotoxin with a wide range of known toxic effects that impair human immune cell function. GT is also highly toxic to A. fumigatus and this fungus has evolved self-protection mechanisms that include (i) the GT efflux pump GliA, (ii) the GT neutralising enzyme GliT, and (iii) the negative regulation of GT biosynthesis by the bis-thiomethyltransferase GtmA. The transcription factor (TF) RglT is the main regulator of GliT and this GT protection mechanism also occurs in the non-GT producing fungus A. nidulans. However, the A. nidulans genome does not encode GtmA and GliA. This work aimed at analysing the transcriptional response to exogenous GT in A. fumigatus and ...
Influence of media and temperature on gliotoxin production in Aspergillus fumigatus strains
Arhiv za higijenu rada i toksikologiju, 2005
Gliotoxin is a secondary metabolite of the epipolythiodioxopiperazine family with biologically active internal disulfide bridge. It is produced by many fungal species, including Aspergillus fumigatus and A. terreus. A. fumigatus, which produces gliotoxin and more than twenty other secondary metabolites, is the leading cause of invasive aspergillosis. Gliotoxin production in situ influence the development of aspergillosis. This study investigated the in vitro production of gliotoxin in nine A. fumigatus isolates from the upper respiratory tract of immunocompromised patients. The effects of media composition and incubation temperature were studied. Gliotoxin was extracted from biomass and its concentration was semi-quantitatively analysed using thin-layer chromatography. Gliotoxin production was higher in the yeast-extract liquid medium (YES) than in the synthetic Czapek-Dox liquid medium (CZA). Incubation at 37 degrees C resulted in higher gliotoxin production than at 25 degrees C, p...
In Vitro and In Vivo Effects of Gliotoxin, a Fungal Metabolite
Digestive diseases and sciences, 2000
Gliotoxin is a fungal metabolite that has immunosuppressive properties. First, we determined if gliotoxin could inhibit cytokine production from macrophage and colonic epithelial cell lines, as well as whether it inhibited nuclear factor-kappa B in these same cell types. ...