Differences in sialic acid density in pathogenic and non-pathogenic Aspergillus species

Aspergillus fumigatus is a ubiquitous soil fungus that causes invasive lung disease in the immunocompromised host. The structure of the conidial wall has not keen well characterized although it is thought that adhesins presen t on the surface are involved in attachment of the conidia to host lung cel ls and proteins, which is a prerequisite for the establishment of infection . Negatively charged carbohydrates on the conidial surface have been previo usly identified as the molecules responsible for attachment of conidia to e xtracellular matrix proteins. The aim of this research was to identify carb ohydrates on the conidial surface that contribute to its negative charge. D irect chemical analysis and indirect binding assays have demonstrated that A. fumigatus possesses sialic acids on the conidial surface. Pre-treatment of A. fumigatus conidia with sialidase decreased binding of a sialic acid-s pecific lectin, Limax flavus agglutinin (LFA), to the conidial surface and decreased adhesion of conidia to the positively charged polymer poly L-lysi ne. Two other sialic acid-specific lectins, Maackia amurensis agglutinin an d Sambucus nigra agglutinin, exhibited negligible binding to A. fumigatus c onidia indicating that 2,3-alpha- and 2,6-alpha -linked sialic acids are no t the major structures found on the conidial surface. Mild acid hydrolysis and purification of conidial wall carbohydrates yielded a product that had the same R-F as the Neu5Ac standard when analysed by high-performance thin- layer chromatography. A density of 6.7 x 10(5) sialic acid residues per con idium was estimated using a colorimetric assay. Conidia grown on a minimal medium lacking sialic acid also reacted with LFA, indicating that sialic ac id biosynthesis occurs de novo. Sialic acid biosynthesis was shown to be re gulated by nutrient composition: the density of sialic acids on the surface of conidia grown in minimal media was tower than that observed when conidi a were grown on rich, complex media. It has previously been shown that path ogenic Aspergillus species adhere to basal lamina proteins to a greater ext ent than non-pathogenic Aspergillus species. To determine whether the expre ssion of sialic acid on the conidial surface was correlated with adhesion t o basal lamina, conidia from other non-pathogenic Aspergillus species were tested for their reactivity towards LFA. Flow cytometric analysis demonstra ted that A. fumigatus had a significantly greater sialic acid density than three non-pathogenic Aspergillus species. Sialic acids on the conidial wall may be involved in adhesion to fibronectin, a component of the basal lamin a, as binding of A. fumigatus conidia to fibronectin was strongly inhibited in the presence of a sialylated glycoprotein.