Cryptococcus: from environmental saprophyte to global pathogen

Abstract

Cryptococcosis is a globally distributed invasive fungal infection that is caused by species within the genus Cryptococcus which presents substantial therapeutic challenges. Although natural human-to-human transmission has never been observed, recent work has identified multiple virulence mechanisms that enable cryptococci to infect, disseminate within and ultimately kill their human host. In this Review, we describe these recent discoveries that illustrate the intricacy of host-pathogen interactions and reveal new details about the host immune responses that either help to protect against disease or increase host susceptibility. In addition, we discuss how this improved understanding of both the host and the pathogen informs potential new avenues for therapeutic development.

Figure 1. Inflammatory signalling in response to cryptococcal infection

Cryptococci, which can assume a titan cell morphology, inevitably shed microbial molecules that contain pathogen-associated molecular patterns (PAMPs). Such fungal molecules are typically components of the cell wall or capsule such as β-glucan, chitin or glucuronoxylomannan (GXM), which are detected by immune sentinel cells, most notably dendritic cells (DCs). DC activation then summons T cells, inducing CD4⁺ T cells to secrete cytokines that activate a T helper cell 1 (T_H1) response (including the secretion of interleukin-12 (IL-12) and IL-23). T_H1 cells produce pro-inflammatory cytokines (such as interferon-γ (IFNγ)), which ultimately control fungal infection. However, some fungal PAMPs can influence DC activation, including modulating the levels of major histocompatibility complex class II (MHC II) or of nuclear factor-κB (NF-κB) signalling. This activates a T_H2 response (mediated by the production of cytokines such as IL-4 and IL-33); this anti-inflammatory environment affects macrophage activation (M1 classic activation and M2 alternative activation) and the ability of macrophages to mediate fungal clearance.

Figure 2. Infection establishment and dissemination with in the human host

Cryptococcal cells typically enter the human host through the lung (bottom). There, they are recognized by patrolling phagocytes but can avoid uptake either by growing into very large titan cells, or by relying on the antiphagocytic properties of the fungal capsule. If uptake occurs, however, cryptococci can survive and persist in phagocytes. For most strains, a failure in host immune function is then required to allow intracellular proliferation. However, the unusual Pacific Northwest outbreak (PNO) strains of Cryptococcus gattii can proliferate in immunocompetent host cells by exploiting a poorly characterized ‘division of labour’ mechanism: in response to reactive oxygen species generated by the phagocyte, some cryptococcal cells acquire an unusual morphology characterized by extensive tubularization of their mitochondria, which increases survival of neighbouring cells (via a mechanism that remains unclear). Cryptococcus spp. proliferation within phagocytes ultimately leads either to host cell lysis or to a nonlytic escape mechanism termed vomocytosis. Upon replication in the lung, cryptococci are able to disseminate to other tissues, including the central nervous system (CNS). Entry into the CNS can occur in three ways: by squeezing between host endothelial cells (paracytosis), which involves the fungal protease Mpr1 and the enzyme urease (which probably weakens the endothelial vessel wall to facilitate entry); by moving directly through endothelial cells (transcytosis), in a process that is mediated by hyaluronic acid in the fungal capsule and the host receptor CD44; or by ‘hitching a ride’ within migrating phagocytes, through what is known as the ‘Trojan horse’ hypothesis. BBB, blood–brain barrier.

Figure 3. Current and future therapies for cryptococcosis

Schematic representation of a cryptococcal cell, showing key current and potential therapeutic targets and examples of antifungal drugs acting at each site. Drugs in current clinical use are shown in red, new drugs in blue and repurposed drugs in green. The three classes of antifungal agents currently used to treat cryptococcosis are polyenes (such as amphotericin B), azoles (such as fluconazole) and the pyrimidine analogue flucytosine (5-FC). Amphotericin B deoxycholate (AmBd) acts by binding to ergosterol in the cryptococcal cell wall, generating pores in the cell membrane, and by inducing cell death by oxidative damage. 5-FC is deaminated by the fungal enzyme cytosine deaminase into 5-fluorouracil (5-FU), which then inhibits thymidylate synthetase and blocks DNA synthesis, or is converted into 5-fluorouridine triphosphate, which is incorporated into RNA and disrupts protein synthesis. Fluconazole inhibits the fungal cytochrome P450 enzyme 14α-demethylase, which is required for conversion of lanosterol to ergosterol, an essential component of the fungal cell membrane. E1210 inhibits the synthesis of the cell wall component glycosylphosphatidylinositol (GPI)-anchored mannoproteins. VT-1129 blocks the activity of CYP51, an essential enzyme in the pathway to produce ergosterol. The arlyamidine T-2307 targets the fungal mitochondrial membrane. Tamoxifen (an oestrogen antagonist that is used in the treatment of breast cancer) targets calmodulin, and the antidepressant sertraline seems to target fungal protein synthesis through an unknown mechanism.