Viral infection triggers interferon-induced expulsion of live Cryptococcus neoformans by macrophages.

Cryptococcus neoformans is an opportunistic human pathogen, which causes serious disease in immunocompromised hosts. Infection with this pathogen is particularly relevant in HIV+ patients, where it leads to around 200,000 deaths per annum. A key feature of cryptococcal pathogenesis is the ability of the fungus to survive and replicate within the phagosome of macrophages, as well as its ability to be expelled from host cells via a novel non-lytic mechanism known as vomocytosis. Here we show that cryptococcal vomocytosis from macrophages is strongly enhanced by viral coinfection, without altering phagocytosis or intracellular proliferation of the fungus. This effect occurs with distinct, unrelated human viral pathogens and is recapitulated when macrophages are stimulated with the anti-viral cytokines interferon alpha or beta (IFNα or IFNß). Importantly, the effect is abrogated when type-I interferon signalling is blocked, thus underscoring the importance of type-I interferons in this phenomenon. Lastly, our data help resolve previous, contradictory animal studies on the impact of type I interferons on cryptococcal pathogenesis and suggest that secondary viral stimuli may alter patterns of cryptococcal dissemination in the host.

Correction: Engineering microbial physiology with synthetic polymers: cationic polymers induce biofilm formation in Vibrio cholerae and downregulate the expression of virulence genes.

[This corrects the article DOI: 10.1039/C7SC00615B.].

Characterizing the Mechanisms of Nonopsonic Uptake of Cryptococci by Macrophages.

The pathogenic fungus Cryptococcus enters the human host via inhalation into the lung and is able to reside in a niche environment that is serum- (opsonin) limiting. Little is known about the mechanism by which nonopsonic phagocytosis occurs via phagocytes in such situations. Using a combination of soluble inhibitors of phagocytic receptors and macrophages derived from knockout mice and human volunteers, we show that uptake of nonopsonized Cryptococcus neoformans and C. gattii via the mannose receptor is dependent on macrophage activation by cytokines. However, although uptake of C. neoformans is via both dectin-1 and dectin-2, C. gattii uptake occurs largely via dectin-1. Interestingly, dectin inhibitors also blocked phagocytosis of unopsonized Cryptococci in wax moth (Galleria mellonella) larvae and partially protected the larvae from infection by both fungi, supporting a key role for host phagocytes in augmenting early disease establishment. Finally, we demonstrated that internalization of nonopsonized Cryptococci is not accompanied by the nuclear translocation of NF-κB or its concomitant production of proinflammatory cytokines such as TNF-α. Thus, nonopsonized Cryptococci are recognized by mammalian phagocytes in a manner that minimizes proinflammatory cytokine production and potentially facilitates fungal pathogenesis.

Quantifying donor-to-donor variation in macrophage responses to the human fungal pathogen Cryptococcus neoformans.

Cryptococcosis remains the leading cause of fungal meningitis worldwide, caused primarily by the pathogen Cryptococcus neoformans. Symptomatic cryptococcal infections typically affect immunocompromised patients. However, environmental exposure to cryptococcal spores is ubiquitous and most healthy individuals are thought to harbor infections from early childhood onwards that are either resolved, or become latent. Since macrophages are a key host cell for cryptococcal infection, we sought to quantify the extent of individual variation in this early phagocyte response within a small cohort of healthy volunteers with no reported immunocompromising conditions. We show that rates of both intracellular fungal proliferation and non-lytic expulsion (vomocytosis) are remarkably variable between individuals. However, we demonstrate that neither gender, in vitro host inflammatory cytokine profiles, nor polymorphisms in several key immune genes are responsible for this variation. Thus the data we present serve to quantify the natural variation in macrophage responses to this important human pathogen and will hopefully provide a useful "benchmark" for the research community.

Engineering microbial physiology with synthetic polymers: cationic polymers induce biofilm formation in Vibrio cholerae and downregulate the expression of virulence genes.

Here we report the first application of non-bactericidal synthetic polymers to modulate the physiology of a bacterial pathogen. Poly(N-[3-(dimethylamino)propyl] methacrylamide) (P1) and poly(N-(3-aminopropyl)methacrylamide) (P2), cationic polymers that bind to the surface of V. cholerae, the infectious agent causing cholera disease, can sequester the pathogen into clusters. Upon clustering, V. cholerae transitions to a sessile lifestyle, characterised by increased biofilm production and the repression of key virulence factors such as the cholera toxin (CTX). Moreover, clustering the pathogen results in the minimisation of adherence and toxicity to intestinal epithelial cells. Our results suggest that the reduction in toxicity is associated with the reduction to the number of free bacteria, but also the downregulation of toxin production. Finally we demonstrate that these polymers can reduce colonisation of zebrafish larvae upon ingestion of water contaminated with V. cholerae. Overall, our results suggest that the physiology of this pathogen can be modulated without the need to genetically manipulate the microorganism and that this modulation is an off-target effect that results from the intrinsic ability of the pathogen to sense and adapt to its environment. We believe these findings pave the way towards a better understanding of the interactions between pathogenic bacteria and polymeric materials and will underpin the development of novel antimicrobial polymers.

Cryptococcus-Epithelial Interactions.

The fungal pathogen, Cryptococcus neoformans, causes devastating levels of morbidity and mortality. Infections with this fungus tend to be predominantly in immunocompromised individuals, such as those with HIV. Infections initiate with inhalation of cryptococcal cells and entry of the pathogen into the lungs. The bronchial epithelial cells of the upper airway and the alveolar epithelial cells of the lower airway are likely to be the first host cells that Cryptococcus engage with. Thus the interaction of cryptococci and the respiratory epithelia will be the focus of this review. C. neoformans has been shown to adhere to respiratory epithelial cells, although if the role of the capsule is in aiding or hindering this adhesion is debatable. The epithelia are also able to react to cryptococci with the release of cytokines and chemokines to start the immune response to this invading pathogen. The activity of surfactant components that line this mucosal barrier towards Cryptococcus and the metabolic and transcriptional reaction of cryptococci when encountering epithelial cells will also be discussed.

New weapons in the Cryptococcus infection toolkit.

The global burden of fungal infections is unacceptably high. The human fungal pathogen Cryptococcus neoformans causes cryptococcosis and accounts for a significant proportion of this burden. Cryptococci undergo a number of elaborate interactions with their hosts, including survival and proliferation within phagocytes as well as dissemination to the central nervous system and other tissues. In this review we highlight a number of exciting recent advances in the field of cryptococcal biology. In particular we discuss new insights into cryptococcal morphology and its impact on virulence, as well as describing novel findings revealing how cryptoccoci may 'talk' to each other.