DOCSLIB.ORG

Stimulation of Innate Immunity Leads to Clearance of C. Neoformans

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
Stimulation of Innate Immunity Leads to Clearance of C. Neoformans STIMULATION OF INNATE IMMUNITY LEADS TO CLEARANCE OF C. NEOFORMANS INFECTION IN ZEBRAFISH ALFRED ALINAFE KAMUYANGO A thesis submitted to the University of Sheffield for fulfillment of the requirements for the degree of Doctor of Philosophy in the Department of Infection, Immunity and Cardiovascular Diseases December 2017 Abstract Cryptococcus neoformans is an opportunistic pathogen and a leading cause of life‑threatening fungal infections in the immunocompromised. Individuals that are at greater risk are those with defective T-cell mediated immunity such as those with HIV/AIDS. Despite treatment with anti-fungal drugs, mortality remains excessively high and patients experience serious side effects hence the need to explore new therapeutic strategies. Activation of macrophages is essential for the control of cryptococcal infection. However, in the absence of T-cell mediated immunity, activation of macrophages is disrupted and clearance of cryptococcal infection is abrogated. Herein I first report a zebrafish-C. neoformans model of infection and show that zebrafish can clear, control or fail to control cryptococcal infection. I then go on to test whether cytokines or PRR ligands are capable of stimulating innate immune resistance to C. neoformans in zebrafish. I demonstrate that concomitant injection of IFNγ with C. neoformans results in reduced fungal burden and increased fungal clearance. IFNγ increases the recruitment of phagocytes to the site of infection and enhances phagocytosis by macrophages. Macrophage deficient larvae fail to clear or suppress cryptococcal infection despite treatment with IFNγ. In addition, infected macrophages display low lysosomal pH and elevated expression of IL-1β in IFNγ-treated larvae. Although neutrophils take up the fungus, their depletion does not alter cryptococcal burden. Secondly, I demonstrate that S. aureus CWP is a potent inducer of innate defences against C. neoformans. Using chemically digested S. aureus CWP to remove wall teichoic acid or mutants that do not produce wall teichoic acid (tarO) or lipoproteins (lgt), I establish that protective effects of S. aureus CWP require wall teichoic acid but not lipoproteins. Protection by S. aureus CWP are associated with increased recruitment of macrophages but not enhanced phagocytosis or TNFα expression. ABSTRACT i This work provides new insights into cellular responses and effector molecules in the context of cryptococcal disease progression while identifying potential immunomodulatory targets. ABSTRACT ii Acknowledgements Not just one or two, but many people have made a positive contribution to this piece of work and I express my sincere appreciation. Firstly, I would like to express my heartfelt appreciation to my supervisor Simon Johnston. Simon, you have provided extraordinary support and guidance throughout my PhD. Your help was beyond expectation. You were so dedicated as if I was the only student you had. You have been very encouraging, for example, whenever I went to your office with my results I thought were not very good, I would come out smiling and feeling encouraged. I could not ask for a better supervisor, your approach is thrilling and I have learned a lot from you. You have been a blessing. I am forever thankful to Jenny Wright for proofreading my thesis. I also thank Josie Gibson who selflessly provided a graph for my thesis. I would also like to thank Josh Hooker, a technician in Simon Foster’s laboratory for making all the S. aureus cell wall preparations used in my project. My thanks also go to Victoriano Mulero at Department of Cell Biology and Histology, Faculty of Biology, University of Murcia, Spain for providing Vibrio anguillarum DNA. I would also like to express my appreciation to my lab colleagues in the Johnston lab. Our socialising and laughter were comforts in my stressful moments, and your hard work and encouragement kept me focused on my work in those tough times. Words cannot articulate how grateful I am. Special thanks go to Robert Evans, Aleksandra Bojarczuk and Kate Pline. I am very grateful to the Wellcome for funding my PhD. Last, with love, extra special thanks go to all my family and friends both in Malawi and in the UK. Many thanks for the conversations, for texts and words of encouragement. I am eternally grateful. Truly, I am thankful to and humbled by all those who have contributed to my success. My prayers are with you always. Above all, I thank God for bringing each and every individual mentioned here to support me. ACKNOWLEDGEMENTS iii Table of Contents Table of Figures Table of Tables CHAPTER 1 INTRODUCTION ....................................................................... 1 1.1 Cryptococcus and cryptococcosis ................................................... 2 1.2 Pathogenic Cryptococcus species ................................................... 2 1.2.1 Cryptococcus neoformans ............................................................. 3 1.2.2 Cryptococcus gattii ........................................................................ 4 1.3 Life cycle of Cryptococcus ................................................................ 8 1.3.1 Infective forms of C. neoformans ................................................. 10 1.3.2 C. neoformans genome ............................................................... 10 1.4 Cryptococcosis ................................................................................. 11 1.4.1 Cause, transmission and affected organs, life cycle .................... 11 1.4.2 Epidemiology ............................................................................... 13 1.4.2.1 In HIV/AIDS patients ............................................................ 13 1.4.2.2 In HIV naïve patients ............................................................ 15 1.4.3 Clinical manifestations ................................................................. 17 1.4.3.1 CNS cryptococcosis ............................................................. 17 1.4.3.2 Pulmonary cryotococcosis .................................................... 17 1.4.3.3 Cryptococcosis in other sites ................................................ 18 1.4.4 Diagnosis ..................................................................................... 19 1.4.4.1 Cryptococcal meningitis ....................................................... 19 1.4.4.2 Pulmonary cryptococcosis .................................................... 20 1.4.5 Management of cryptococcal disease ......................................... 21 1.4.5.1 Management of meningeal cryptococcosis .......................... 21 1.4.5.2 Management of cryptococcosis complications ..................... 22 1.4.5.3 Management of non-meningeal cryptococcosis ................... 23 1.4.6 C. neoformans virulence factors .................................................. 24 1.4.6.1 The Capsule ......................................................................... 24 1.4.6.2 Melanin ................................................................................. 28 1.4.6.3 Anti-phagocytic protein 1 ...................................................... 28 CONTENTS iv 1.4.6.4 Urease .................................................................................. 29 1.4.6.5 Thermotolerance .................................................................. 29 1.4.6.6 Phospholipase ...................................................................... 30 1.4.6.7 Other virulence factors ......................................................... 30 1.4.7 Pathogenesis of C. neoformans .................................................. 30 1.4.7.1 Dissemination to the CNS .................................................... 33 1.5 Host immunity to Cryptococcus neoformans ................................ 35 1.5.1 Innate immune and cellular responses against C. neoformans ... 35 1.5.2 Phagocytes .................................................................................. 39 1.5.2.1 Neutrophils ........................................................................... 39 1.5.2.2 Dendritic cells ....................................................................... 41 1.5.2.3 Macrophages ........................................................................ 44 1.6 Experimental models for cryptococcosis ....................................... 48 1.6.1 In vitro cell models ....................................................................... 48 1.7 Invertebrate models .......................................................................... 49 1.7.1 Galleria mellonella ....................................................................... 49 1.7.2 Drosophila melanogaster and Drosophila S2 cells ...................... 50 1.7.3 Caenorhabditis elegans ............................................................... 50 1.7.4 Dictyostelium discoideum (Social amoebae) ............................... 51 1.7.5 Acanthamoeba castellanii ............................................................ 51 1.8 Vertebrate models ............................................................................ 52 1.8.1 Mouse (Mus musculus) ............................................................... 52 1.8.2 Rat (Rattus rattus) ....................................................................... 52 1.8.3 Rabbit (Oryctolagus cuniculus) .................................................... 53 1.8.4 Guinea pigs (Cavia porcellus) ..................................................... 53 1.8.5 The zebrafish ..............................................................................
Load more

Recommended publications
  • Emanuele Giurisato Curriculum Vitae
    Emanuele Giurisato Curriculum Vitae Present position: Lecturer at University of Siena. Department of Biotechnology, Chemistry and Pharmacy, University of Siena. Via Aldo Moro, 1 53100, Siena, Italy Phone: +39 0577 232125 (office) e-mail: [email protected] Visiting scientist at University of Manchester Faculty of Biology, Medicine and Health Oxford Road, M13 9PT, Manchester, UK e-mail: [email protected] Education and training: 2014-2016 Marie-Curie Fellowship (I-MOVE 2013-1-OUT-24-EXP) University of Manchester 2009-present Lecturer at University of Siena 2008 Post-doctoral fellowship Department of Physiopathology Molecular Medicine and Public Health, University of Siena, Italy 2003-2007 Post-doctoral fellowship Department of Pathology and Immunology, Washington University School of Medicine, St Louis, MO (USA) 1999-2002 Ph.D student in “Molecular Medicine” Department of Physiopathology Molecular Medicine and Public Health, University of Siena, Italy 1996-1999 Research assistant, Department of Biomedical Experimental Sciences, CRIBI (University of Padua, Italy) 1996 BSc, Biological Sciences (specialization in Molecular Biology), Department of Biomedical Experimental Sciences, (University of Padua, Italy) Research experience: Nov.’96 -Sept. ‘99: Research assistant at the University of Padua, Department of Biomedical Experimental Sciences, CRIBI. Advisor: Dr. M. Cantini Telethon projects A101 “Myoblast-selective mitogens released by macrophage as a tool for muscle regeneration and gene therapy” and A095 “FasL engineered myoblasts can escape from death: requisite for immune protection in gene therapy via myoblast”. Oct.1999-Dec 2002: Ph. D. student in “Molecular Medicine” at the University of Siena. Advisor: Prof. A. Benedetti Research project: (i) role of plasma-membrane microdomains (rafts) in cellular responses to stimulatory molecules in T lymphocytes, (ii) Calcium signalling in T cells and (iii) involvement of lipid rafts during the immunological synapse formation.
    [Show full text]
  • Cryptococcus Neoformans Thermotolerance to Avian Body Temperature Is Sufficient for Extracellular Growth but Not Intracellular S
    www.nature.com/scientificreports OPEN Cryptococcus neoformans Thermotolerance to Avian Body Temperature Is Sufficient Received: 30 October 2015 Accepted: 14 January 2016 For Extracellular Growth But Published: 17 February 2016 Not Intracellular Survival In Macrophages Simon A Johnston1,2, Kerstin Voelz3 & Robin C May3,4 Cryptococcus neoformans is a fatal fungal pathogen of humans that efficiently parasitises macrophages. Birds can be colonised by cryptococci and can transmit cryptococcosis to humans via inhalation of inoculated bird excreta. However, colonisation of birds appears to occur in the absence of symptomatic infection. Here, using a pure population of primary bird macrophages, we demonstrate a mechanism for this relationship. We find that bird macrophages are able to suppress the growth of cryptococci seen in mammalian cells despite C. neoformans being able to grow at bird body temperature, and are able to escape from bird macrophages by vomocytosis. A small subset of cryptococci are able to adapt to the inhibitory intracellular environment of bird macrophages, exhibiting a large cell phenotype that rescues growth suppression. Thus, restriction of intracellular growth combined with survival at bird body temperature explains the ability of birds to efficiently spreadC. neoformans in the environment whilst avoiding systemic disease. Cryptococcus neoformans is an environmental fungus that causes fatal human and animal disease. In humans, cryptococcosis causes hundreds of thousands of deaths each year, the vast majority in immunocompromised patients1. As with many significant pathogens, cryptococci are able to parasitise host cells. This potential for an intracellular lifestyle allows C. neoformans the potential to evade additional host immune responses and thus spread within the body, leading to systemic disease2,3.
    [Show full text]
  • Memory in Fungal Pathogens Promotes Immune Evasion, Colonisation And
    Memory in fungal pathogens promotes immune evasion, colonisation and infection Alistair J.P. Brown*, Neil A.R. Gow, Adilia Warris and Gordon D. Brown Medical Research Council Centre for Medical Mycology at the University of Aberdeen, Institute of Medical Sciences, Foresterhill, Aberdeen, AB25 2ZD, United Kingdom. * Correspondence: Alistair J P Brown E: [email protected] T: +44 (0)1224 437482 Key words: Fungal pathogenicity; immune evasion; fungal infection; fungal adaptation; adaptive prediction; fungal memory Abstract By analogy with Pavlov’s dogs, certain pathogens have evolved anticipatory behaviours that exploit specific signals in the human host to prepare themselves against imminent host challenges. This adaptive prediction, a type of history-dependent microbial behaviour, represents a primitive form of microbial memory. For fungal pathogens, adaptive prediction helps them circumvent nutritional immunity, protects them against phagocytic killing, and activates immune evasion strategies. We describe how these anticipatory responses, and the contrasting lifestyles and evolutionary trajectories of fungal pathogens, have influenced the evolution of such adaptive behaviours, and how these behaviours affect host colonisation and infection. 1 Microbial memory Microorganisms often inhabit dynamic niches where they are confronted by continuously fluctuating environmental challenges that are often perceived as stresses. Consequently, to survive, microbes have evolved adaptation mechanisms that mitigate against these challenges. Certain niches impose stresses that recur in a reasonably predictable manner in which one input is often followed by a second input of a certain type. This presents the resident microbes with an opportunity to develop anticipatory behaviours that have the potential to enhance their fitness in this niche. This is somewhat analogous to Pavlov’s dogs, who were entrained to salivate upon hearing a bell in anticipation of being fed [1].
    [Show full text]
  • Intracellular Parasitism of Macrophages by Cryptococcus
    Intracellular parasitism of macrophages by Cryptococcus by Hansong Ma, BSc A thesis submitted to The University of Birmingham for the degree of DOCTOR OF PHILOSOPHY School of Biosciences The University of Birmingham August 2009 University of Birmingham Research Archive e-theses repository This unpublished thesis/dissertation is copyright of the author and/or third parties. The intellectual property rights of the author or third parties in respect of this work are as defined by The Copyright Designs and Patents Act 1988 or as modified by any successor legislation. Any use made of information contained in this thesis/dissertation must be in accordance with that legislation and must be properly acknowledged. Further distribution or reproduction in any format is prohibited without the permission of the copyright holder. Abstract The pathogenic fungi Cryptococcus neoformans and Cryptococcus gattii are two of the main causes of life-threatening meningoencephalitis in immunocompromised and immunocompetent individuals respectively. Following inhalation, cryptococci are engulfed by phagocytic cells in the lung and previous studies by our group and others have demonstrated that they are then able to survive inside these cells (especially macrophages), thus acting as intracellular parasites. This intracellular phase is thought to underlie the ability of the pathogens to remain latent for long periods of time within infected individuals. Here, we demonstrate that cryptococci can also manipulate host macrophages in order to mediate an exquisitely controlled ‘escape’ process. This expulsive process, which we have termed ‘vomocytosis’, can occur either into the extracellular milieu or, remarkably, into neighbouring host cells, thus resulting in direct cell-to-cell transmission (‘lateral transfer’).
    [Show full text]
  • 2018 BMES MOBILE THURSDAY Annual Meeting Platform Sessions Th-1 APP (Thursday 8:00-9:30Am)
    TABLE OF CONTENTS TABLE SCIENTIFIC PROGRAM 2018 BMES MOBILE THURSDAY Annual Meeting Platform Sessions Th-1 APP (Thursday 8:00-9:30am) ....................................... 4–13 Go to the Apple or Android Store and search for: Platform Sessions Th-2 BMES (Thursday 1:30-3:00pm) ....................................14–22 Download the Free App > Select BMES2018 Platform Sessions Th-3 (Thursday 3:45-5:15pm) ....................................23–31 Poster Sessions–Thursday ..............................32–91 Browse the program by date or session type FRIDAY Platform Sessions Fri-1 keywords Search (Friday 8:00-9:30am) ....................................... 93–101 Search Author list Platform Sessions Fri-2 Add presentations to (Friday 1:15-2:45pm) .................................. 102–110 a custom itinerary Platform Sessions Fri-3 Click a link to show (Friday 3:30-5:00pm) .................................... where a presentation 111–119 is on the map of the convention center Poster Sessions–Friday .............................. 120–179 SATURDAY Platform Sessions Sat-1 (Saturday 8:00am-9:30am) .........................180–189 Platform Sessions Sat-2 Don't forget to turn your BMES (Saturday 1:30-3:00pm) ............................... 190–198 BASH ticket in for a wristband at Platform Sessions Sat-3 (Saturday 3:15-4:45pm) ...............................199–205 the information or registration Poster Sessions–Saturday ....................... 206–249 booths before Friday afternoon Omni Hotel Floorplan ..........................................
    [Show full text]
  • Dinoflagellate Symbionts Escape Vomocytosis by Host Cell Immune Suppression
    bioRxiv preprint doi: https://doi.org/10.1101/864579; this version posted December 6, 2019. The copyright holder for this preprint (which was not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission. Title: Dinoflagellate symbionts escape vomocytosis by host cell immune suppression Authors: Marie R. Jacobovitz#, Sebastian Rupp#, Philipp A. Voss, Sebastian G. Gornik, Annika Guse* Affiliation: Centre for Organismal StuDies (COS), Heidelberg University, Germany. #contributeD equally *corresponding author A.G.: [email protected] Abstract Emergence of the symbiotic lifestyle fostereD the immense Diversity of all ecosystems on Earth, but symbiosis plays a particularly remarkable role in marine ecosystems. Photosynthetic dinoflagellate enDosymbionts power reef ecosystems by transferring vital nutrients to their coral hosts. The mechanisms Driving this symbiosis, specifically those which allow hosts to Discriminate between beneficial symbionts anD pathogens, are not well unDerstooD. Here, we uncover that host immune suppression is key for dinoflagellate enDosymbionts to avoiD elimination by the host using a comparative, moDel systems approach. UnexpecteDly, we finD that the clearance of non- symbiotic microalgae occurs by non-lytic expulsion (vomocytosis) anD not intracellular Digestion, the canonical mechanism useD by professional immune cells to Destroy foreign invaDers. We proviDe eviDence that suppression of TLR signalling by targeting the conserveD MyD88 adapter protein has been co-opteD for this enDosymbiotic lifestyle, suggesting that this is an evolutionarily ancient mechanism exploiteD to facilitate symbiotic associations ranging from coral enDosymbiosis to the microbiome of vertebrate guts. Main Text Summary Symbiotic interactions appear in all Domains of life and are key drivers of adaption and evolutionary diversification.
    [Show full text]
  • Investigating the Molecular Mechanisms of Vomocytosis
    Investigating the Molecular Mechanisms of Vomocytosis By Andrew Stephen Gilbert A thesis submitted to the University of Birmingham for the degree of DOCTOR OF PHILOSPHY School of Biosciences Institute of Microbiology and Infection (IMI) University of Birmingham Birmingham, UK September 2016 University of Birmingham Research Archive e-theses repository This unpublished thesis/dissertation is copyright of the author and/or third parties. The intellectual property rights of the author or third parties in respect of this work are as defined by The Copyright Designs and Patents Act 1988 or as modified by any successor legislation. Any use made of information contained in this thesis/dissertation must be in accordance with that legislation and must be properly acknowledged. Further distribution or reproduction in any format is prohibited without the permission of the copyright holder. Abstract The opportunistic fungal pathogen Cryptococcus neoformans is the major etiological agent of the life threatening disease cryptococcosis, which is responsible for over half a million human deaths per annum (Park et al., 2009). Professional phagocytes, such as alveolar macrophages, phagocytose inhaled spores and attempt to destroy the pathogen. However, this process is inefficient in immunocompromised hosts, such as those suffering from HIV/AIDS. In such hosts the macrophage is thought to behave like a “Trojan Horse”, acting as both a cryptococcal dissemination vector and as a protective niche against antifungal agents/cells present in the circulation (Casadevall et al., 2010). Vomocytosis, first discovered in C. neoformans (Ma et al., 2006) (Alvarez and Casadevall 2006), is a non-lytic expulsive mechanism whereby C. neoformans or C. gattii exit the macrophage leaving both pathogen and the host macrophage with a morphologically normal phenotype.
    [Show full text]
  • Viral Infection Enhances Vomocytosis of Intracellular Fungi Via Type I
    bioRxiv preprint doi: https://doi.org/10.1101/512293; this version posted January 4, 2019. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY 4.0 International license. 1 Viral infection enhances vomocytosis of intracellular fungi via Type I 2 interferons 3 Paula I Seoane1, Leanne M. Taylor-Smith1, David Stirling2, Lucy C. K. Bell2, Mahdad 4 Noursadeghi2, Dalan Bailey3, Robin C. May1* 5 6 1Institute of Microbiology & Infection and School of Biosciences, University of Birmingham, 7 Edgbaston, Birmingham, B15 2TT, UK 8 2Division of Infection and Immunity, University College London, Gower Street, London WC1E 9 6BT, UK 10 3The Pirbright Institute, Ash Rd, Surrey, GU24 0NF, UK 11 12 *Correspondence to: [email protected] 13 14 Abstract 15 Cryptococcus neoformans is an opportunistic human pathogen, which causes serious disease 16 in immunocompromised hosts. Infection with this pathogen is particularly relevant in HIV+ 17 patients, where it leads to around 200,000 deaths per annum. A key feature of cryptococcal 18 pathogenesis is the ability of the fungus to survive and replicate within the phagosome of 19 macrophages, as well as its ability to escape via a novel non-lytic mechanism known as 20 vomocytosis. We have been exploring whether viral infection affects the interaction between 21 C. neoformans and macrophages. Here we show that viral infection enhances cryptococcal 22 vomocytosis without altering phagocytosis or intracellular proliferation of the fungus.
    [Show full text]
  • 444. Dendritic Cells Perform Vomocytosis of Cryptococcus
    Dendritic Cells Perform Vomocytosis of Cryptococcus neoformans Noah Pacifici, Melissa Cruz-Acuña, Neeraj Senthil, and Jamal Lewis University of California, Davis, Davis, CA Statement of Purpose Pathogens have evolved over time to evade the host immune system in various ways. For instance, the fungal species Cryptococcus neoformans (CN), following engulfment by phagocytes, has been observed to stay alive within the acidic phagolysosome and escape through a process called vomocytosis1. Using this phenomenon, CN utilizes host immune cells to disseminate infection throughout the body. One condition due to infection of this microbe is cryptococcal meningitis (CM). Primarily affecting immunocompromised individuals, including an estimated 220,000 HIV/AIDS patients, CM causes ~181,000 deaths/ year worldwide2. Greater understanding of vomocytosis could lead to the development of new CM treatments for patients, as well as novel biomaterial Figure 1. (A) An example image of a vomocytosis event of particulate vaccines. Current studies have mainly focused CN escaping from a DC. (B) Measured expulsions and cell-to- on vomocytosis occurrences in macrophages, as alveolar cell transfer events of live and heat-killed CN from DCs. macrophages are the first immune cell to phagocytose CN Timing of vomocytosis events were also measured for live (C) and ultimately expel this fungal pathogen. However, CN and heat-killed (D) groups. (n=1) may have the ability to induce vomocytosis in other phagocytes due to conserved molecular machinery. windows per condition. Expulsion events were counted as Recently, Yang et al.3 observed vomocytosis-like any time one or more CN escaped from a DC (Figure 1A). expulsion of CN from neutrophils, the most abundant phagocyte found in the body.
    [Show full text]
  • Using Flow Cytometry to Analyze Cryptococcus Infection of Macrophages
    This is a repository copy of Using Flow Cytometry to Analyze Cryptococcus Infection of Macrophages.. White Rose Research Online URL for this paper: http://eprints.whiterose.ac.uk/117211/ Version: Accepted Version Book Section: Evans, R.J. orcid.org/0000-0003-0678-6510, Voelz, K., Johnston, S.A. orcid.org/0000-0002-3429-9536 et al. (1 more author) (2017) Using Flow Cytometry to Analyze Cryptococcus Infection of Macrophages. In: Botelho, R., (ed.) Phagocytosis and Phagosomes. Methods in Molecular Biology (1519). Springer New York , pp. 349-357. ISBN 978-1-4939-6579-3 https://doi.org/10.1007/978-1-4939-6581-6_24 Reuse Unless indicated otherwise, fulltext items are protected by copyright with all rights reserved. The copyright exception in section 29 of the Copyright, Designs and Patents Act 1988 allows the making of a single copy solely for the purpose of non-commercial research or private study within the limits of fair dealing. The publisher or other rights-holder may allow further reproduction and re-use of this version - refer to the White Rose Research Online record for this item. Where records identify the publisher as the copyright holder, users can verify any specific terms of use on the publisher’s website. Takedown If you consider content in White Rose Research Online to be in breach of UK law, please notify us by emailing [email protected] including the URL of the record and the reason for the withdrawal request. [email protected] https://eprints.whiterose.ac.uk/ USING FLOW CYTOMETRY TO ANALYSE CRYPTOCOCCUS INFECTION OF MACROPHAGES Robert J.
    [Show full text]
  • The Multiplicity of Infection Does Not Affect Interactions Between Cryptococcus Neoformans and Macrophages
    bioRxiv preprint doi: https://doi.org/10.1101/2020.02.05.936427; this version posted February 7, 2020. The copyright holder for this preprint (which was not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission. The multiplicity of infection does not affect interactions between Cryptococcus neoformans and macrophages Katherine Pline1and Simon Johnston1* 1. Department of Infection, Immunity and Cardiovascular Disease, Bateson Centre and Florey Institute, University of Sheffield, UK *Corresponding author: Simon Johnston [email protected], tel (+44) 0144 222 2301. bioRxiv preprint doi: https://doi.org/10.1101/2020.02.05.936427; this version posted February 7, 2020. The copyright holder for this preprint (which was not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission. Summary Major determinants of the outcome of infection include growth of the pathogen and response of immune cells such as macrophages. Cryptococcus neoformans is a fungal pathogen which may grow within the extracellular environment, or may exploit host macrophages as a niche for replication and dissemination. The clinical outcome of cryptococcal infection varies widely between individuals even when key host and pathogen molecular factors are the same. For a broad range of infections altering pathogen density is known to influence progression and outcome of infection by affecting immune response and pathogen biology (e.g. via innate immune signalling or microbial quorum sensing). Here, using time lapse imaging of murine cell line and human primary macrophages in vitro, we examined the effect of altering pathogen density on the interactions of macrophages with cryptococci.
    [Show full text]
  • Cryptococcus Neoformans from Macrophages Mbio.Asm.Org Occurs in Vivo and Is Influenced by Phagosomal Ph on June 21, 2016 - Published by André Moraes Nicola,A,B Emma J
    Downloaded from RESEARCH ARTICLE Nonlytic Exocytosis of Cryptococcus neoformans from Macrophages mbio.asm.org Occurs In Vivo and Is Influenced by Phagosomal pH on June 21, 2016 - Published by André Moraes Nicola,a,b Emma J. Robertson,a Patrícia Albuquerque,a Lorena da Silveira Derengowski,a,b and Arturo Casadevalla,c Department of Microbiology and Immunology, Albert Einstein College of Medicine of the Yeshiva University, Bronx, New York, USAa; Instituto de Ciências Biológicas, Universidade de Brasília, Campus Universitário Darcy Ribeiro, Brasília, Brazilb; and Division of Infectious Diseases, Department of Medicine, Albert Einstein College of Medicine of the Yeshiva University, Bronx, New York, USAc ABSTRACT A unique aspect of the interaction of the fungus Cryptococcus neoformans with macrophages is the phenomenon of nonlytic exocytosis, also referred to as “vomocytosis” or phagosome extrusion/expulsion, which involves the escape of fungal cells from the phagocyte with the survival of both cell types. This phenomenon has been observed only in vitro using subjective and time-consuming microscopic techniques. In spite of recent advances in our knowledge about its mechanisms, a major ques- tion still remaining is whether this phenomenon also occurs in vivo. In this study, we describe a novel flow cytometric method that resulted in a substantial gain in throughput for studying phagocytosis and nonlytic exocytosis in vitro and used it to explore mbio.asm.org the occurrence of this phenomenon in a mouse model of infection. Furthermore, we tested the hypothesis that host cell phago- somal pH affected nonlytic exocytosis. The addition of the weak bases ammonium chloride and chloroquine resulted in a signifi- cant increase of nonlytic exocytosis events, whereas the vacuolar ATPase inhibitor bafilomycin A1 had the opposite effect.
    [Show full text]