The Candida albicans Histone Acetyltransferase Hat1 Regulates Stress Resistance and Virulence via Distinct Chromatin Assembly Pathways

Human fungal pathogens like Candida albicans respond to host immune surveillance by rapidly adapting their transcriptional programs. Chromatin assembly factors are involved in the regulation of stress genes by modulating the histone density at these loci. Here, we report a novel role for the chromatin assembly-associated histone acetyltransferase complex NuB4 in regulating oxidative stress resistance, antifungal drug tolerance and virulence in C. albicans. Strikingly, depletion of the NuB4 catalytic subunit, the histone acetyltransferase Hat1, markedly increases resistance to oxidative stress and tolerance to azole antifungals. Hydrogen peroxide resistance in cells lacking Hat1 results from higher induction rates of oxidative stress gene expression, accompanied by reduced histone density as well as subsequent increased RNA polymerase recruitment. Furthermore, hat1Delta/Delta cells, despite showing growth defects in vitro, display reduced susceptibility to reactive oxygen-mediated killing by innate immune cells. Thus, clearance from infected mice is delayed although cells lacking Hat1 are severely compromised in killing the host. Interestingly, increased oxidative stress resistance and azole tolerance are phenocopied by the loss of histone chaperone complexes CAF-1 and HIR, respectively, suggesting a central role for NuB4 in the delivery of histones destined for chromatin assembly via distinct pathways. Remarkably, the oxidative stress phenotype of hat1Delta/Delta cells is a species-specific trait only found in C. albicans and members of the CTG clade. The reduced azole susceptibility appears to be conserved in a wider range of fungi. Thus, our work demonstrates how highly conserved chromatin assembly pathways can acquire new functions in pathogenic fungi during coevolution with the host

Novel insights into host-fungal pathogen interactions derived from live-cell imaging

Acknowledgments The authors acknowledge funding from the Wellcome Trust (080088, 086827, 075470 and 099215) including a Wellcome Trust Strategic Award for Medical Mycology and Fungal Immunology 097377 and FP7-2007–2013 grant agreement HEALTH-F2-2010-260338–ALLFUN to NARG.Peer reviewedPublisher PD

High-level production of violacein by the newly isolated Duganella violaceinigra str. NI28 and its impact on Staphylococcus aureus

A violacein-producing bacterial strain was isolated and identified as a relative of Duganella violaceinigra YIM 31327 based upon phylogenetic analyses using the 16S rRNA, gyrB and vioA gene sequences and a fatty acid methyl ester (FAME) analysis. This new strain was designated D. violaceinigra str. NI28. Although these two strains appear related based upon these analyses, the new isolate was phenotypically different from the type strain as it grew 25% faster on nutrient media and produced 45-fold more violacein. When compared with several other violacein producing strains, including Janthinobacterium lividum, D. violaceinigra str. NI28 was the best violacein producer. For instance, the crude violacein yield with D. violaceinigra str. NI28 was 6.0 mg/OD at 24 hours, a value that was more than two-fold higher than all the other strains. Finally, the antibacterial activity of D. violaceinigra str. NI28 crude violacein was assayed using several multidrug resistant Staphylococcus aureus. Addition of 30 mu M crude violacein led to a 96% loss in the initial S. aureus population while the minimum inhibitory concentration was 1.8 mu M. Consequently, this novel isolate represents a phenotypic variant of D. violaceinigra capable of producing much greater quantities of crude violacein, an antibiotic effective against multidrug resistant S. aureusopen

Linked patterns of biological and environmental covariation with brain structure in adolescence: a population-based longitudinal study

Adolescence is a period of major brain reorganization shaped by biologically timed and by environmental factors. We sought to discover linked patterns of covariation between brain structural development and a wide array of these factors by leveraging data from the IMAGEN study, a longitudinal population-based cohort of adolescents. Brain structural measures and a comprehensive array of non-imaging features (relating to demographic, anthropometric, and psychosocial characteristics) were available on 1476 IMAGEN participants aged 14 years and from a subsample reassessed at age 19 years (n = 714). We applied sparse canonical correlation analyses (sCCA) to the cross-sectional and longitudinal data to extract modes with maximum covariation between neuroimaging and non-imaging measures. Separate sCCAs for cortical thickness, cortical surface area and subcortical volumes confirmed that each imaging phenotype was correlated with non-imaging features (sCCA r range: 0.30-0.65, all PFDR < 0.001). Total intracranial volume and global measures of cortical thickness and surface area had the highest canonical cross-loadings (|rho| = 0.31-0.61). Age, physical growth and sex had the highest association with adolescent brain structure (|rho = 0.24-0.62); at baseline, further significant positive associations were noted for cognitive measures while negative associations were observed at both time points for prenatal parental smoking, life events, and negative affect and substance use in youth (|rho| = 0.10-0.23). Sex, physical growth and age are the dominant influences on adolescent brain development. We highlight the persistent negative influences of prenatal parental smoking and youth substance use as they are modifiable and of relevance for public health initiatives

The TolC Protein of Legionella pneumophila Plays a Major Role in Multi-Drug Resistance and the Early Steps of Host Invasion

Pneumonia associated with Iegionnaires's disease is initiated in humans after inhalation of contaminated aerosols. In the environment, Legionella pneumophila is thought to survive and multiply as an intracellular parasite within free-living amoeba. In the genome of L. pneumophila Lens, we identified a unique gene, tolC, encoding a protein that is highly homologous to the outer membrane protein TolC of Escherichia coli. Deletion of tolC by allelic exchange in L. pneumophila caused increased sensitivity to various drugs. The complementation of the tolC mutation in trans restored drug resistance, indicating that TolC is involved in multi-drug efflux machinery. In addition, deletion of tolC caused a significant attenuation of virulence towards both amoebae and macrophages. Thus, the TolC protein appears to play a crucial role in virulence which could be mediated by its involvement in efflux pump mechanisms. These findings will be helpful in unraveling the pathogenic mechanisms of L. pneumophila as well as in developing new therapeutic agents affecting the efflux of toxic compounds

Additive Contributions of Two Manganese-Cored Superoxide Dismutases (MnSODs) to Antioxidation, UV Tolerance and Virulence of Beauveria bassiana

The biocontrol potential of entomopathogenic fungi against arthropod pests depends on not only their virulence to target pests but tolerance to outdoor high temperature and solar UV irradiation. Two Beauveria bassiana superoxide dismutases (SODs), BbSod2 and BbSod3, were characterized as cytosolic and mitochondrial manganese-cored isoenzymes (MnSODs) dominating the total SOD activity of the fungal entomopathogen under normal growth conditions. To probe their effects on the biocontrol potential of B. bassiana, ΔBbSod2, ΔBbSod3, and three hairpin RNA-interfered (RNAi) mutants with the transcripts of both BbSod2 and BbSod3 being suppressed by 91–97% were constructed and assayed for various phenotypic parameters in conjunction with ΔBbSod2/BbSod2, ΔBbSod3/BbSod3 and wild-type (control strains). In normal cultures, the knockout and RNAi mutants showed significant phenotypic alterations, including delayed sporulation, reduced conidial yields, and impaired conidial quality, but little change in colony morphology. Their mycelia or conidia became much more sensitive to menadione or H2O2-induced oxidative stress but had little change in sensitivity to the hyperosmolarity of NaCl and the high temperature of 45°C. Accompanied with the decreased antioxidative capability, conidial tolerances to UV-A and UV-B irradiations were reduced by 16.8% and 45.4% for ΔBbSod2, 18.7% and 44.7% for ΔBbSod3, and ∼33.7% and ∼63.8% for the RNAi mutants, respectively. Their median lethal times (LT50s) against Myzus persicae apterae, which were topically inoculated under a standardized spray, were delayed by 18.8%, 14.5% and 37.1%, respectively. Remarkably, the effects of cytosolic BbSod2 and mitochondrial BbSod3 on the phenotypic parameters important for the fungal bioncontrol potential were additive, well in accordance with the decreased SOD activities and the increased superoxide levels in the knockout and RNAi mutants. Our findings highlight for the first time that the two MnSODs co-contribute to the biocontrol potential of B. bassiana by mediating cellular antioxidative response

Development of an In Vitro Model for the Multi-Parametric Quantification of the Cellular Interactions between Candida Yeasts and Phagocytes

We developed a new in vitro model for a multi-parameter characterization of the time course interaction of Candida fungal cells with J774 murine macrophages and human neutrophils, based on the use of combined microscopy, fluorometry, flow cytometry and viability assays. Using fluorochromes specific to phagocytes and yeasts, we could accurately quantify various parameters simultaneously in a single infection experiment: at the individual cell level, we measured the association of phagocytes to fungal cells and phagocyte survival, and monitored in parallel the overall phagocytosis process by measuring the part of ingested fungal cells among the total fungal biomass that changed over time. Candida albicans, C. glabrata, and C. lusitaniae were used as a proof of concept: they exhibited species-specific differences in their association rate with phagocytes. The fungal biomass uptaken by the phagocytes differed significantly according to the Candida species. The measure of the survival of fungal and immune cells during the interaction showed that C. albicans was the more aggressive yeast in vitro, destroying the vast majority of the phagocytes within five hours. All three species of Candida were able to survive and to escape macrophage phagocytosis either by the intraphagocytic yeast-to-hyphae transition (C. albicans) and the fungal cell multiplication until phagocytes burst (C. glabrata, C. lusitaniae), or by the avoidance of phagocytosis (C. lusitaniae). We demonstrated that our model was sensitive enough to quantify small variations of the parameters of the interaction. The method has been conceived to be amenable to the high-throughput screening of mutants in order to unravel the molecular mechanisms involved in the interaction between yeasts and host phagocytes