Background. Cryptococcal meningitis (CM) is a leading cause of HIV-associated mortality globally. High fungal burden in cerebrospinal fluid (CSF) at diagnosis and poor fungal clearance during treatment are recognized adverse prognostic markers; however, the underlying pathogenic factors that drive these clinical manifestations are incompletely understood. We profiled a large set of clinical isolates for established cryptococcal virulence traits to evaluate the contribution of C. neoformans phenotypic diversity to clinical presentation and outcome in human cryptococcosis.Methods. Sixty-five C. neoformans isolates from clinical trial patients with matched clinical data were assayed in vitro to determine murine macrophage uptake, intracellular proliferation rate (IPR), capsule induction, and laccase activity. Analysis of the correlation between prognostic clinical and host immune parameters and fungal phenotypes was performed using Spearman's r, while the fungal-dependent impact on long-term survival was determined by Cox regression analysis.Results. High levels of fungal uptake by macrophages in vitro, but not the IPR, were associated with CSF fungal burden (r = 0.38, P = 0.002) and long-term patient survival (hazard ratio [HR] 2.6, 95% CI 1.2-5.5, P = 0.012). High-uptake strains were hypocapsular (r = -0.28, P = 0.05) and exhibited enhanced laccase activity (r = 0.36, P = 0.003). Fungal isolates with greater laccase activity exhibited heightened survival ex vivo in purified CSF (r = 0.49, P < 0.0001) and resistance to clearance following patient antifungal treatment (r = 0.39, P = 0.003).Conclusion. These findings underscore the contribution of cryptococcal-phagocyte interactions and laccasedependent melanin pathways to human clinical presentation and outcome. Furthermore, characterization of fungal-specific pathways that drive clinical manifestation provide potential targets for the development of therapeutics and the management of CM.

ERK5 regulates nonlytic expulsion of live pathogens from phagocytes to limit dissemination of infections.

The innate immune system is a critical line of defense against pathogenic fungi. Macrophages act at an early stage of infection, detecting and phagocytizing infectious propagules. To avoid killing at this stage, fungal pathogens use diverse strategies ranging from evasion of uptake to intracellular parasitism. This article will discuss five of the most important human fungal pathogens (Candida albicans, Aspergillus fumigatus, Cryptococcus neoformans, Coccidiodes immitis, and Histoplasma capsulatum) and consider the strategies and virulence factors adopted by each to survive and replicate within macrophages. Macrophages are phagocytic immune cells, derived from monocyte differentiation, and are involved in the first line of defense during microbial invasion. They recognize, engulf, and destroy foreign bodies, such as pathogenic organisms before then presenting antigen to coordinate subsequent adaptive immunity (Fig. 1). Macrophages are found in almost all tissues and are particularly abundant at mucosal surfaces in which pathogen exposure is naturally higher, such as the alveoli. They are able to engulf particles ranging from 0.5 mm (such as bacteria) to .5 mm (such as yeast), although an upper size limit does exist (Chen et al. 1997;Krombach et al. 1997;Kinchen and Ravichandran 2008).Macrophages express a wide range of different receptors able to detect nonself particles. These receptors can be split into two groups: opsonic and nonopsonic. Opsonic receptors, such as the Fc receptor or complement receptor families (Flannagan et al. 2012), are able to recognize particles coated (opsonized) in antibody or complement (Johnston and May 2013). In contrast, nonopsonic receptors are pathogen recognition receptors (PRRs) that directly detect pathogen-associated molecular patterns (PAMPs) on the surface of microorganisms. Macrophages are heterogeneous in the receptors they express, and can change this repertoire when differentiating into different subtypes. For instance, the nonopsonic Dectin-1 receptor Once engulfment has been achieved, the macrophage must then digest the pathogen, now segregated into a phagosome (digesting vesicle). To complete the digestion, the phagosome must mature via the fusion of early and late stage endosomes and ultimately fuse with the lysosome, generating a phagolysosome (Kinchen and Ravichandran 2008). The phagolysosome uses vacuolar ATPases to pump H þ ions into the phagolysosome, reducing the pH. Once the pH is reduced sufficiently, acid-dependent proteases, such as cathepsin D, are activated to degrade the pathogen (Fig. 1) (Kinchen and Ravichandran 2008).Many human fungal pathogens have developed strategies to resist phagocytic attack, thus facilitating pathogenicity. The field of fungal research has exploded over the last three decades because of the increase in fungal-related illnesses that appears to correlate with the AIDS pandemic and more effective immunosuppressive medicines . Macrophages have been shown to play a role in resistance to disseminated candidiasis, although they are ...

Footrot causes 70–90% of lameness in sheep in Great Britain. With approximately 5% of 18 million adult sheep lame at any one time, it costs the UK sheep industry £24–84 million per year. The Gram-negative anaerobe Dichelobacter nodosus is the causative agent, with disease severity influenced by bacterial load, virulence, and climate. The aim of the current study was to characterize strains of D. nodosus isolated by culture of swabs from healthy and diseased feet of 99 ewes kept as a closed flock over a 10-month period and investigate persistence and transmission of strains within feet, sheep, and the flock. Overall 268 isolates were characterized into strains by serogroup, proline–glycine repeat (pgr) status, and multi-locus variable number tandem repeat analysis (MLVA). The culture collection contained 87 unique MLVA profiles and two major MLVA complexes that persisted over time. A subset of 189 isolates tested for the virulence marker aprV2 were all positive. The two MLVA complexes (76 and 114) comprised 62 and 22 MLVA types and 237 and 28 isolates, respectively. Serogroups B, and I, and pgrB were associated with MLVA complex 76, whereas serogroups D and H were associated with MLVA complex 114. We conclude that within-flock D. nodosus evolution appeared to be driven by clonal diversification. There was no association (P > 0.05) between serogroup, pgr, or MLVA type and disease state of feet. Strains of D. nodosus clustered within sheep and were transmitted between ewes over time. D. nodosus was isolated at more than one time point from 21 feet, including 5 feet where the same strain was isolated on two occasions at an interval of 1–33 weeks. Collectively, our results indicate that D. nodosus strains persisted in the flock, spread between sheep, and possibly persisted on feet over time.