Experimental Models of Cryptococcosis

Cryptococcosis is a life-threatening fungal disease that infects around one million people each year. Establishment and progression of disease involves a complex interplay between the fungus and a diverse range of host cell types. Over recent years, numerous cellular, tissue, and animal models have been exploited to probe this host-pathogen interaction. Here we review the range of experimental models that are available for cryptococcosis research and compare the relative advantages and limitations of the different systems

Immune Responses Accelerate Ageing: Proof-of-Principle in an Insect Model

The pathology of many of the world's most important infectious diseases is caused by the immune response. Additionally age-related disease is often attributed to inflammatory responses. Consequently a reduction in infections and hence inflammation early in life has been hypothesized to explain the rise in lifespan in industrialized societies. Here we demonstrate experimentally for the first time that eliciting an immune response early in life accelerates ageing. We use the beetle Tenebrio molitor as an inflammation model. We provide a proof of principle for the effects of early infection on morbidity late in life and demonstrate a long-lasting cost of immunopathology. Along with presenting a proof-of-principle study, we discuss a mechanism for the apparently counter-adaptive persistence of immunopathology in natural populations. If immunopathology from early immune response only becomes costly later in life, natural selection on reducing self-harm would be relaxed, which could explain the presence of immune self-harm in nature

Ancient dispersal of the human fungal pathogen Cryptococcus gattii from the Amazon rainforest.

Over the past two decades, several fungal outbreaks have occurred, including the high-profile 'Vancouver Island' and 'Pacific Northwest' outbreaks, caused by Cryptococcus gattii, which has affected hundreds of otherwise healthy humans and animals. Over the same time period, C. gattii was the cause of several additional case clusters at localities outside of the tropical and subtropical climate zones where the species normally occurs. In every case, the causative agent belongs to a previously rare genotype of C. gattii called AFLP6/VGII, but the origin of the outbreak clades remains enigmatic. Here we used phylogenetic and recombination analyses, based on AFLP and multiple MLST datasets, and coalescence gene genealogy to demonstrate that these outbreaks have arisen from a highly-recombining C. gattii population in the native rainforest of Northern Brazil. Thus the modern virulent C. gattii AFLP6/VGII outbreak lineages derived from mating events in South America and then dispersed to temperate regions where they cause serious infections in humans and animals

On sexual dimorphism in immune function

Sexual dimorphism in immune function is a common pattern in vertebrates and also in a number of invertebrates. Most often, females are more ‘immunocompetent’ than males. The underlying causes are explained by either the role of immunosuppressive substances, such as testosterone, or by fundamental differences in male and female life histories. Here, we investigate some of the main predictions of the immunocompetence handicap hypothesis (ICHH) in a comparative framework using mammals. We focus specifically on the prediction that measures of sexual competition across species explain the observed patterns of variation in sex-specific immunocompetence within species. Our results are not consistent with the ICHH, but we do find that female mammals tend to have higher white blood cell counts (WBC), with some further associations between cell counts and longevity in females. We also document positive covariance between sexual dimorphism in immunity, as measured by a subset of WBC, and dimorphism in the duration of effective breeding. This is consistent with the application of ‘Bateman's principle’ to immunity, with females maximizing fitness by lengthening lifespan through greater investment in immune defences. Moreover, we present a meta-analysis of insect immunity, as the lack of testosterone in insects provides a means to investigate Bateman's principle for immunity independently of the ICHH. Here, we also find a systematic female bias in the expression of one of the two components of insect immune function that we investigated (phenoloxidase). From these analyses, we conclude that the mechanistic explanations of the ICHH lack empirical support. Instead, fitness-related differences between the sexes are potentially sufficient to explain many natural patterns in immunocompetence

Intracellular proliferation rates (IPR) illustrate moderate proliferation levels and suggest mitochondrial tubularization is decoupled from IPR in macrophages for <i>C. gattii</i> VGIII isolates.

<p>(A) Intracellular <i>C. gattii</i> proliferation rates in macrophages were similar between environmental and clinical isolates. (B) Percent tubular mitochondria as a result of co-incubation with macrophages (solid bars) in comparison to DMEM controls (hatched bars). Percent tubular mitochondria does not correlate with IPR or virulence. Clinical isolates (VGIIIa dark-purple; VGIIIb light-purple), environmental isolates (VGIIIa dark-green; VGIIIb light-green), and previously reported isolates VGIIIa reference strain B4546 (Black) and VGII (Grey). (C) Ranked IPR of tested <i>C. gattii</i> isolates.</p

Phylogenetic analysis of newly identified <i>C. gattii</i> isolates from California.

<p>Phylogenetic tree obtained by analysis of 7 concatenated MLST loci (<i>IGS1</i>, <i>TEF1</i>, <i>GPD1</i>, <i>LAC1</i>, <i>CAP10</i>, <i>PLB1</i>, and <i>MPD1</i>) using 500 bootstrap replicates and the Maximum Likelihood (MLC) method based on Tamura-Nei model applying Neighbor-Join and BioNL algorithms. The tree with the highest log likelihood is shown. The analysis involved 61 sequences and a total of 4252 positions. Scale bar indicated 0.005 substitutions per nucleotide.</p

Environmental isolates that share an indistinguishable MLST profile with clinical isolates are less virulent in mice.

<p>(A) 11 matched isolates representing four distinct MLST groups, three VGIIIa (G1, G2, G3, two strains each) and one VGIIIb (4 strains, 2 clinical, and 2 environmental), were tested for virulence in the murine animal model. Ten male A/JCr mice per strain were intranasally infected with 10⁶ cells and survival was recorded for 200 days. MLST groups are represented by shared symbol and group number. Clinical isolates (VGIIIa dark-purple; VGIIIb light purple), environmental isolates (VGIIIa dark-green; VGIIIb light green). Median survival CA1232 = 90 days, CA1308 = 100 days, CA1053 = 103 days, CA1508 = 127 days, and BHPP1-S2B = 138 days. (B) Mice were assessed for tissue burden postmortem.</p

<i>C. gattii</i> VGIIIb isolates display higher antifungal susceptibility values to Amphotericin B and flucytosine in contrast to VGIIIa isolates.

<p>(A, E) MIC of VGIIIa (red) versus VGIIIb (blue) for amphotericin B (A), flucytosine (B), fluconazole (C), and ketoconazole (D). MIC of clinical (purple) and environmental (green) isolates of VGIIIa (triangles) and VGIIIb (circles). The MIC of VGIIIa isolates was significantly higher than VGIIIb isolates for amphotericin B (p = 0.0178) and flucytosine (p<0.0001) but not for fluconazole (p = 0.1059) or ketoconazole (p = 0.0685). No significant differences were observed between environmental and clinical isolates of VGIIIa or of VGIIIb in response to amphotericin B (E), flucytosine (F), fluconazole (G), or ketoconazole (H) (<a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1004285#ppat.1004285.s016" target="_blank">Table S10a, S10b and S10c</a>). MIC values, which exceeded the maximum concentration for each Etest, were assigned as the maximum concentration tested for averaging and graphing (<a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1004285#ppat.1004285.s007" target="_blank">Table S1</a>). Maximum concentrations of Etest amphotericin B (32 µg/ml), fluconazole (256 µg/ml), flucytosine (32 µg/ml), and ketoconazole (32 µg/ml).</p