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Whole-genome and sex-related transcriptomics of the pathogenic fungus Pneumocystis jirovecii within human lungs

English title Whole-genome and sex-related transcriptomics of the pathogenic fungus Pneumocystis jirovecii within human lungs
Applicant Hauser Philippe
Number 165825
Funding scheme Project funding
Research institution Institut de Microbiologie - CHUV Faculté de Biologie et Médecine Université de Lausanne
Institution of higher education University of Lausanne - LA
Main discipline Experimental Microbiology
Start/End 01.05.2016 - 30.04.2020
Approved amount 600'000.00
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All Disciplines (2)

Discipline
Experimental Microbiology
Medical Microbiology

Keywords (3)

Transcriptomics; whole-genome transcriptional profiles; Pneumocystis jirovecii

Lay Summary (French)

Lead
Les patients immunocompromis sont à risques d’infections par Pneumocystis jirovecii, un champignon qui cause des pneumonies sévères, parfois fatales. Le séquençage récent du génome de ce pathogène permet d’analyser l’expression de ce génome dans diverses conditions. Nous allons comparer cette expression entre les patients avec pneumonie et des individus qui ne sont que colonisés par une faible charge du champignon. Les buts sont d’identifier et étudier des facteurs de virulence, ainsi que la reproduction sexuée qui a probablement lieu de façon obligatoire dans les poumons humains.
Lay summary

Les gènes de P. jirovecii exprimés chez des patients avec pneumonie ainsi que chez des individus colonisés seront déterminés et comparés.

Partie 1 : Les éventuels facteurs de virulence surexprimés lors de la pneumonie seront investigués.

Partie 2 : Les protéines de surface sont un facteur de virulence reconnu impliqués la variation antigénique et l’adhésion aux cellules humaines. Leur expression et leur diversité seront comparées entre pneumonie et colonisation.

Partie 3 : L’expression des gènes de P. jirovecii impliqués dans sa reproduction sexuée sera comparée entre pneumonie et colonisation.

 

Valeur attendue du projet : l’étude de l’expression des gènes des pathogènes dans les humains infectés est particulièrement relevante car les modèles animaux ne reproduisent pas complètement les infections naturelles. L’identification de facteurs de virulence de P. jirovecii et la caractérisation de sa reproduction sexuée pourrait aider à combattre la maladie grâce à l’identification de nouvelle cibles thérapeutiques. Les résultats pourraient aussi aboutir à de nouvelles connaissances concernant le mode de vie opportuniste des champignons, ce qui pourrait avoir des applications importantes pour combattre d’autres champignons pathogènes pour l’humain.

Direct link to Lay Summary Last update: 07.04.2016

Responsible applicant and co-applicants

Employees

Publications

Publication
Reply to Nevez and Le Gal, “Caspofungin and Pneumocystis Pneumonia: It Is Time To Go Ahead”
Luraschi Amanda, Richard Sophie, Hauser Philippe Marcel (2019), Reply to Nevez and Le Gal, “Caspofungin and Pneumocystis Pneumonia: It Is Time To Go Ahead”, in Antimicrobial Agents and Chemotherapy, 63(10), 01320.
Expression and Immunostaining Analyses Suggest that Pneumocystis Primary Homothallism Involves Trophic Cells Displaying Both Plus and Minus Pheromone Receptors
Luraschi A., Richard S., Almeida J. M. G. C. F., Pagni M., Cushion M. T., Hauser P. M. (2019), Expression and Immunostaining Analyses Suggest that Pneumocystis Primary Homothallism Involves Trophic Cells Displaying Both Plus and Minus Pheromone Receptors, in mBio, 10(4), e01145.
Site-Directed Mutagenesis of the 1,3-β-Glucan Synthase Catalytic Subunit of Pneumocystis jirovecii and Susceptibility Assays Suggest Its Sensitivity to Caspofungin
Luraschi A., Richard S., Hauser P. M. (2018), Site-Directed Mutagenesis of the 1,3-β-Glucan Synthase Catalytic Subunit of Pneumocystis jirovecii and Susceptibility Assays Suggest Its Sensitivity to Caspofungin, in Antimicrobial Agents and Chemotherapy, 62(12), 01159.
Functional and Expression Analyses of the Pneumocystis MAT Genes Suggest Obligate Sexuality through Primary Homothallism within Host Lungs
Richard S., Almeida J. M. G. C. F., Cissé O. H., Luraschi A., Nielsen O., Pagni M., Hauser P. M. (2018), Functional and Expression Analyses of the Pneumocystis MAT Genes Suggest Obligate Sexuality through Primary Homothallism within Host Lungs, in mBio, 9(1), 00221.
Identification and Functional Ascertainment of the Pneumocystis jirovecii Potential Drug Targets Gsc1 and Kre6 Involved in Glucan Synthesis
Luraschi Amanda, Cissé Ousmane H., Pagni Marco, Hauser Philippe M. (2017), Identification and Functional Ascertainment of the Pneumocystis jirovecii Potential Drug Targets Gsc1 and Kre6 Involved in Glucan Synthesis, in Journal of Eukaryotic Microbiology, 64(4), 481-490.

Collaboration

Group / person Country
Types of collaboration
University of Copenhagen, Prof O. Nielsen Denmark (Europe)
- in-depth/constructive exchanges on approaches, methods or results
- Publication
Swiss Bioinformatics Institute, Vital-it, Dr M. Pagni Switzerland (Europe)
- in-depth/constructive exchanges on approaches, methods or results
- Publication
- Research Infrastructure
University of Cincinnati, Prof MT Cushion United States of America (North America)
- in-depth/constructive exchanges on approaches, methods or results
- Publication
University of Lisbon, Dr J Almeida Portugal (Europe)
- in-depth/constructive exchanges on approaches, methods or results
- Publication

Associated projects

Number Title Start Funding scheme
146135 Characterization of the telomeres and comparative genomics of the human pathogenic fungus Pneumocystis jirovecii 01.05.2013 Project funding
124998 De novo sequencing of the genome of the human pathogenic fungus Pneumocystis jirovecii and study of Pneumocystis pneumonia lung microbiome 01.10.2009 Project funding
192802 Expression and functional analyses of the major surface antigens of the human pathogenic fungus Pneumocystis jirovecii 01.05.2020 Project funding

Abstract

Background: Pneumocystis jirovecii is a fungus which causes life-threatening pneumonia in patients who are immuno-compromised because of AIDS, organ transplantation, or cancer. The absence of in vitro culture method for this pathogen slowed down the understanding of its biology and pathogenicity. We recently succeeded to sequence the P. jirovecii genome from the microbiome present in a single bronchoalveolar lavage (BAL) specimen of a single patient with Pneumocystis pneumonia (PcP). The availability of this genome allows new analyses to be performed, such as transcriptomics. The reservoir of P. jirovecii includes patients without recognized PcP who are colonized with a low fungal load. The fungus is probably in a different physiological state (slow growth) during colonization than during active PcP (rapid growth). Consequently, the fungal genome is probably expressed differentially, virulence factors being over-expressed during PcP. Comparative whole-genome transcriptional analyses can be used to identify these virulence factors. A recognized virulence factor of P. jirovecii is the gene family encoding isoforms of the major surface glycoprotein (MSG) involved in antigenic variation and adhesion to target host lung cells. Only one MSG isoform localized downstream of a conserved expression site present at a single copy per genome is believed to be expressed at a time in a given cell. We hypothesize that the diversity of the expressed MSGs within P. jirovecii populations is smaller during colonization than during PcP because of an increased pressure by the human immune system. Another crucial feature of P. jirovecii is its probable obligate sexual reproduction within human lungs. We recently identified the mating type (MAT) genes responsible for sexual differentiation as well as other sex-related genes in the genome of P. jirovecii. The structure of the MAT locus and the identity of the genes present suggest self-fertility through primary homothallism. Transcriptional analyses of these genes within human lungs are required to further characterize this sexual reproduction. Specific aims: - to identify the virulence factors of P. jirovecii (Part 1).- to study the expression and diversity of P. jirovecii MSGs (Part 2).- to characterize the occurrence and mechanisms of the sexual reproduction of P. jirovecii (Part 3).Experimental design :Part 1.: Identification of the virulence factors of P. jirovecii P. jirovecii transcriptomes will be determined using illumina sequencing of the enriched cDNAs and the differential expression of the whole genome between the two conditions will be analyzed. Potential virulence factors over-expressed during PcP will be investigated. Part 2.: Study of the expression and diversity of P. jirovecii MSGsThe differential expression of the MSGs between colonization and PcP will be analyzed in the whole-genome transcriptional profiles obtained in part 1 using the expression site as surrogate marker. The diversity of the expressed MSGs will be determined by the analysis of the MSGs linked downstream to the expression site within the transcriptional raw reads. Part 3.: Characterization of the sexual reproduction of P. jirovecii 3.1. Characterization of the expression of MAT and sex-related genes within human lungsThe differential expression of the P. jirovecii MAT and other sex-related genes between colonization and PcP will be analyzed in the whole-genome transcriptional profiles obtained in part 1. These analyses will allow characterizing the system of P. jirovecii sexual reproduction, as well as assessing its occurrence during colonization and PcP. 3.2. Detection of receptors to mating pheromones on the surface of trophic formsThe receptors to pheromones of mating types Minus and Plus will be stained on the surface of P. jirovecii trophic cells using fluorescent antibodies. Controls will consist in staining Saccharomyces cerevisiae cells expressing the P. jirovecii receptors. These experiments will allow further assessing P. jirovecii sexuality, and determining if each cell expresses both mating types or only one.Importance and impact:In vivo transcriptomics during human infections are particularly relevant because, as shown for other fungal pathogens, animal and in vitro models do not reproduce fully the natural infections. The identification of the virulence factors of P. jirovecii and the characterization of its sexual reproduction may help fighting the disease by the identification of potential new targets for therapeutic intervention. The results may lead to new knowledge associated with the fungal opportunistic life style which may have broad applications to other fungi pathogenic to humans.
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