Vishnu Chaturvedi

Characterization of Cryptococcus neoformans variety gattii SOD2 reveals distinct roles of the two superoxide dismutases in fungal biology and virulence

We studied superoxide dismutases (SODs) in the encapsulated yeast Cryptococcus neoformans (Cn) variety gattii to analyse the role of mitochondrial MnSOD (SOD2) in fungal biology and virulence. SOD2 was cloned from a Cn cosmid library, sod2 mutant and sod2 + SOD2 reconstituted strains were constructed by homologous recombination, and two sod1sod2 double mutants were constructed by replacing SOD2 in the sod1 mutant with the sod2::HYG allele. The SOD2 protein (SOD2p) encoded 225 amino acids, with 36–66% identity with other fungal SOD2ps. SOD2 deletion rendered Cn highly growth‐defective at 37°C in 19–20% oxygen (normal air), and this defect was reversed by limiting oxygen to 1.3% as well in the presence of antioxidant, ascorbic acid. The sod2 mutant accumulated significantly more reactive oxygen species (ROS) at 37°C as well at 30°C in the presence of antimycin A, suggesting that SOD2p is the primary defence of Cn against the superoxide anion (O2• −) in the mitochondria. The sod2 was also highly susceptible to redox‐cycling agents, high salt and nutrient limitations. The sod2 mutant was avirulent in intranasally infected mice and markedly attenuated in its virulence in intravenously infected mice. The virulence defect of sod2 mutant appeared related to its growth defects in high oxygen environment, but not resulting from increased sensitivity to oxidative killing by phagocytes. The sod1sod2 double mutants were avirulent in mice. Additionally, sod1sod2 double mutants showed a marked reduction in the activities of other known Cn virulence factors; and they were more susceptible to PMN killing than was the sod2 single mutant. Previously, we reported that the attenuation of sod1 mutant in mice was resulting from enhanced susceptibility to phagocyte killing, combined with a reduction in the activities of a number of virulence factors. Thus, SOD1p and SOD2p play distinct roles in the biology and virulence of Cn var. gattii via independent modes of action.

Role of Ess1 in Growth, Morphogenetic Switching, and RNA Polymerase II Transcription in Candida albicans

Candida albicans is a fungal pathogen that causes potentially fatal infections among immune-compromised individuals. The emergence of drug resistant C. albicans strains makes it important to identify new antifungal drug targets. Among potential targets are enzymes known as peptidyl-prolyl cis/trans isomerases (PPIases) that catalyze isomerization of peptide bonds preceding proline. We are investigating a PPIase called Ess1, which is conserved in all major human pathogenic fungi. Previously, we reported that C. albicans Ess1 is essential for growth and morphogenetic switching. In the present study, we re-evaluated these findings using more rigorous genetic analyses, including the use of additional CaESS1 mutant alleles, distinct marker genes, and the engineering of suitably-matched isogenic control strains. The results confirm that CaEss1 is essential for growth in C. albicans, but show that reduction of CaESS1 gene dosage by half (δ/+) does not interfere with morphogenetic switching. However, further reduction of CaEss1 levels using a conditional allele does reduce morphogenetic switching. We also examine the role of the linker α-helix that distinguishes C. albicans Ess1 from the human Pin1 enzyme, and present results of a genome-wide transcriptome analysis. The latter analysis indicates that CaEss1 has a conserved role in regulation of RNA polymerase II function, and is required for efficient termination of small nucleolar RNAs and repression of cryptic transcription in C. albicans.

The presence of 3-hydroxy oxylipins in pathogenic microbes.

There is a sufficient body of work documenting the distribution of 3-hydroxy oxylipins in microbes. However, there is limited information on the role of these compounds in microbial pathogenesis. When derived from mammalian cells, these compounds regulate patho-biological processes, thus an understanding of 3-hydroxy oxylipin function and metabolism could prove important in shedding light on how these compounds mediate cellular pathology and physiology. This could present 3-hydroxy oxylipin biosynthetic pathways as targets for drug development. In this minireview, we interrogate the relevant yeast and bacterial 3-hydroxy oxylipin literature in order to appreciate how these compounds may influence the inflammatory response leading to disease development.

Cryptococcus and Cryptococcosis in the Twenty-First Century

Cryptococcus neoformans has been recognized as a human pathogen for more than a century. During this period, the fungus has continued to intrigue physicians and laboratory scientists with its unusual forms, widespread occurrence in specialized niches and varied manifestations of its disease in healthy and immunodeficient individuals. The 1937 inaugural issue of Mycopathologia had a rather interesting take on this pathogen wherein Lodder argued that Cryptoccocus represents a ‘‘nomen dubium’’ and a ‘‘nomen confusum’’ and proposed that this generic name should be avoided in yeast taxonomy [9]. Much has changed in the intervening years as taxonomy of Cryptococcus, and especially C. neoformans was placed on firm footing and cryptococcosis came to be recognized as an important fungal disease globally. The pace of progress on these topics has been especially noticeable in past 25 years. Thus, it is fitting that this special thematic issue of Mycopathologia carries a number of articles commemorating the 8th International Conference on Cryptococcus and Cryptococcosis (ICCC), which was held in Charleston, SC from May 1–5, 2011. In their inaugural article, Kwon-Chung, Perfect and Levitz [8] chronicle the impetus for ICCC. These three leading experts had played crucial roles since the first ICCC was held in 1989, and their perspective on the evolution of this field of study is a fitting start to the contents of this special issue. A broad overview of the meeting in Charleston along with a contemporary perspective on the fungal virulence, outbreaks, and host factors is summarized by members of the Heitman laboratory [19]. The next perspective article by Del Poeta and Casadevall [7] lays out future challenges facing the scientific community in its quest for better understanding of the pathogen and for the control of this deadly disease. M. Del Poeta (&) Department of Biochemistry and Molecular Biology, Medical University of South Carolina, 173 Ashley Avenue, BSB 512A, Charleston, SC 29425, USA e-mail: delpoeta@musc.edu