The capsule of the fungal pathogen Cryptococcus neoformans has been studied extensively in recent decades, and a large body of information is now available to the scientific community. Well-known aspects of the capsule include its structure, antigenic properties and its function as a virulence factor. The capsule is composed primarily of two polysaccharides, glucuronoxylomannan (GXM) and galactoxylomannan (GalXM), in addition to a smaller proportion of mannoproteins (MP). Most of the studies on the composition of the capsule have focused on GXM, which comprises more than 90% of the capsule's polysaccharide mass. It is GalXM, however, that is of particular scientific interest because of its immunological properties. The molecular structure of these polysaccharides is very complex and has not yet been fully elucidated. Both GXM and GalXM are high molecular mass polymers with the mass of GXM equaling roughly 10 times that of GalXM. Recent findings suggest, however, that the actual Mw might be different to what it has traditionally been thought to be. In addition to their structural roles in the polysaccharide capsule, these molecules have been associated with many deleterious effects on the immune response. Capsular components are therefore considered key virulence determinants in Cryptococcus neoformans, which has motivated their use in vaccines and made them targets for monoclonal antibody treatments. In this review we will provide an update on the current knowledge of the C. neoformans capsule, covering aspects related to its structure, synthesis, and particularly, its role as a virulence factor.

The most distinctive feature of the human pathogenic fungus is a polysaccharide capsule that is essential for virulence and is composed primarily of glucuronoxylomannan (GXM) and galactoxylomannan (GalXM). GXM mediates multiple deleterious effects on host immune function, yet relatively little is known about its physical properties. The average mass of Cryptococcus neoformans GXM from four antigenically different strains ranged from 1.7 to 7 ؋ 10 6 daltons as calculated from Zimm plots of light-scattering data. GalXM was significantly smaller than GXM, with an average mass of 1 ؋ 10 5 daltons. These molecular masses imply that GalXM is the most numerous polysaccharide in the capsule on a molar basis. The radius of gyration of the capsular polysaccharides ranged between 68 and 208 nm. Viscosity measurements suggest that neither polysaccharide altered fluid dynamics during infection since GXM behaved in solution as a polyelectrolyte and GalXM did not increase solution viscosity. Immunoblot analysis indicated heterogeneity within GXM. In agreement with this, scanning transmission electron microscopy of GXM preparations revealed a tangled network of two different types of molecules. Mass per length measurements from light scattering and scanning transmission electron microscopy were consistent and suggested GXM molecules self-associate. A mechanism for capsule growth is proposed based on the extracellular release and entanglement of GXM molecules.

ABSTRACTAutophagy is used by eukaryotes in bulk cellular material recycling and in immunity to intracellular pathogens. We evaluated the role of macrophage autophagy in the response toCryptococcus neoformansandCandida albicans, two important opportunistic fungal pathogens. The autophagosome marker LC3 (microtubule-associated protein 1 light chain 3 alpha) was present in most macrophage vacuoles containingC. albicans. In contrast, LC3 was found in only a few vacuoles containingC. neoformanspreviously opsonized with antibody but never after complement-mediated phagocytosis. Disruption of host autophagyin vitroby RNA interference against ATG5 (autophagy-related 5) decreased the phagocytosis ofC. albicansand the fungistatic activity of J774.16 macrophage-like cells against both fungi, independent of the opsonin used. ATG5-knockout bone marrow-derived macrophages (BMMs) also had decreased fungistatic activity againstC. neoformanswhen activated. In contrast, nonactivated ATG5-knockout BMMs actually restrictedC. neoformansgrowth more efficiently, suggesting that macrophage autophagy plays different roles againstC. neoformans, depending on the macrophage type and activation. Interference with autophagy in J774.16 cells also decreased nonlytic exocytosis ofC. neoformans, increased interleukin-6 secretion, and decreased gamma interferon-induced protein 10 secretion. Mice with a conditionally knocked out ATG5 gene in myeloid cells showed increased susceptibility to intravenousC. albicansinfection. In contrast, these mice manifested no increased susceptibility toC. neoformans, as measured by survival, but had fewer alternatively activated macrophages and less inflammation in the lungs after intratracheal infection than control mice. These results demonstrate the complex roles of macrophage autophagy in restricting intracellular parasitism by fungi and reveal connections with nonlytic exocytosis, humoral immunity, and cytokine signaling.

SummaryThe major virulence factor of Cryptococcus neoformans is its polysaccharide capsule composed of glucuronoxylomannan (GXM), galactoxylomannan (GalXM) and mannoproteins. A variety of immunomodulating activities have been described for GXM and mannoproteins but little is known about possible interactions of GalXM with the immune system. In the present article, we investigate the effect of purified soluble GalXM on human T lymphocytes. The results indicate that, GalXM (i) can affect selected immune responses; (ii) causes significant impairment of T cell proliferation and increases interferon-γ γ γ γ and interleukin-10 production; and (iii) induces apoptosis of T lymphocytes through activation of caspase-8 that terminates with fragmentation of DNA. These results are the first to suggest a role for GalXM in C. neoformans virulence by demonstrating that it can target human T cells, and that it may impair the development of an effective specific T cell response.