c.i.-fluorescent-brightening-agent-28 and galactomannan

Owing to the essential role in protection of the Aspergillus fumigatus cell against human defense reactions, its cell wall has long been taken as a promising antifungal target. The cell wall of A. fumigatus composed of chitin, glucan and galactomannan and mannoproteins. Although galactomannan has been used as a diagnostic target for a long time, its biosynthesis remains unknown in A. fumigatus. In this study, a putative α1,6-mannosyltransferase gene mnn9 was identified in A. fumigatus. Deletion of the mnn9 gene resulted in an increased sensitivity to calcofluor white, Congo red, or hygromycin B as well as in reduced cell wall components and abnormal polarity. Although there was no major effect on N-glycan synthesis, covalently-linked cell wall mannoprotein Mp1 was significantly reduced in the mutant. Based on our results, we propose that Mnn9p is a mannosyltransferase responsible for the formation of the α-mannan in cell wall mannoproteins, potentially via elongation of O-linked mannose chains.

Topics: Aspergillus fumigatus; Benzenesulfonates; Cell Wall; Congo Red; Galactose; Gene Deletion; Hygromycin B; Mannans; Mannosyltransferases; Membrane Glycoproteins

Echinocandins inhibit β-1,3-glucan synthesis and are one of the few antimycotic drug classes effective against Aspergillus spp. In this study, we characterized the β-1,3-glucan synthase Fks1 of Aspergillus fumigatus, the putative target of echinocandins. Data obtained with a conditional mutant suggest that fks1 is not essential. In agreement, we successfully constructed a viable Δfks1 deletion mutant. Lack of Fks1 results in characteristic growth phenotypes similar to wild type treated with echinocandins and an increased susceptibility to calcofluor white and sodium dodecyl sulfate. In agreement with Fks1 being the only β-1,3-glucan synthase in A. fumigatus, the cell wall is devoid of β-1,3-glucan. This is accompanied by a compensatory increase of chitin and galactosaminogalactan and a significant decrease in cell wall galactomannan due to a massively enhanced galactomannan shedding. Our data furthermore suggest that inhibition of hyphal septation can overcome the limitations of echinocandin therapy. Compounds inhibiting septum formation boosted the antifungal activity of caspofungin. Thus, development of clinically applicable inhibitors of septum formation is a promising strategy to improve existing antifungal therapy.

Topics: Antifungal Agents; Aspergillosis; Aspergillus fumigatus; Benzenesulfonates; beta-Glucans; Caspofungin; Cell Wall; Chitin; Echinocandins; Galactose; Glucosyltransferases; Hyphae; Lipopeptides; Mannans; Mutation; Phenotype; Polysaccharides

We assessed Calcofluor white staining, Aspergillus polymerase chain reaction, and a galactomannan enzyme immunoassay for diagnosis of fungal infection with use of computed tomography-guided percutaneous lung biopsy specimens obtained from 61 patients. The sensitivity and specificity of computerized tomography, Aspergillus polymerase chain reaction, and galactomannan enzyme immunoassay were 100% and 50%, 100% and 86%, and 88% and 94%, respectively.

Topics: Adult; Aspergillosis; Aspergillus; Benzenesulfonates; Biopsy; Contrast Media; False Positive Reactions; Female; Galactose; Humans; Immunocompromised Host; Immunoenzyme Techniques; Lung Diseases, Fungal; Male; Mannans; Middle Aged; Polymerase Chain Reaction; Prospective Studies; Sensitivity and Specificity; Tomography, X-Ray Computed