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Cell wall composition in Cryptococcus neoformans is media dependent and alters host response, inducing protective immunity

INTRODUCTION: Cryptococcus neoformans is a basidiomycete fungus that can cause meningoencephalitis, especially in immunocompromised patients. Cryptococcus grows in many different media, although little attention has been paid to the role of growth conditions on the cryptococcal cell wall or on virul...

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Autores principales: Upadhya, Rajendra, Lam, Woei C., Hole, Camaron R., Vasselli, Joseph G., Lodge, Jennifer K.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Frontiers Media S.A. 2023
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10399910/
https://www.ncbi.nlm.nih.gov/pubmed/37538303
http://dx.doi.org/10.3389/ffunb.2023.1183291
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author Upadhya, Rajendra
Lam, Woei C.
Hole, Camaron R.
Vasselli, Joseph G.
Lodge, Jennifer K.
author_facet Upadhya, Rajendra
Lam, Woei C.
Hole, Camaron R.
Vasselli, Joseph G.
Lodge, Jennifer K.
author_sort Upadhya, Rajendra
collection PubMed
description INTRODUCTION: Cryptococcus neoformans is a basidiomycete fungus that can cause meningoencephalitis, especially in immunocompromised patients. Cryptococcus grows in many different media, although little attention has been paid to the role of growth conditions on the cryptococcal cell wall or on virulence. OBJECTIVE: The purpose of this study was to determine how different media influenced the amount of chitin and chitosan in the cell wall, which in turn impacted the cell wall architecture and host response. METHODS: Yeast extract, peptone, and dextrose (YPD) and yeast nitrogen base (YNB) are two commonly used media for growing Cryptococcus before use in in vitro or in vivo experiments. As a result, C. neoformans was grown in either YPD or YNB, which were either left unbuffered or buffered to pH 7 with MOPS. These cells were then labeled with cell wall-specific fluorescent probes to determine the amounts of various cell wall components. In addition, these cells were employed in animal virulence studies using the murine inhalation model of infection. RESULTS: We observed that the growth of wild-type C. neoformans KN99 significantly changes the pH of unbuffered media during growth. It raises the pH to 8.0 when grown in unbuffered YPD but lowers the pH to 2.0 when grown in unbuffered YNB (YNB-U). Importantly, the composition of the cell wall was substantially impacted by growth in different media. Cells grown in YNB-U exhibited a 90% reduction in chitosan, the deacetylated form of chitin, compared with cells grown in YPD. The decrease in pH and chitosan in the YNB-U-grown cells was associated with a significant increase in some pathogen-associated molecular patterns on the surface of cells compared with cells grown in YPD or YNB, pH 7. This altered cell wall architecture resulted in a significant reduction in virulence when tested using a murine model of infection. Furthermore, when heat-killed cells were used as the inoculum, KN99 cells grown in YNB-U caused an aberrant hyper-inflammatory response in the lungs, resulting in rapid animal death. In contrast, heat-killed KN99 cells grown in YNB, pH 7, caused little to no inflammatory response in the host lung, but, when used as a vaccine, they conferred a robust protective response against a subsequent challenge infection with the virulent KN99 cells. CONCLUSION: These findings emphasize the importance of culture media and pH during growth in shaping the content and organization of the C. neoformans cell wall, as well as their impact on fungal virulence and the host response.
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spelling pubmed-103999102023-08-03 Cell wall composition in Cryptococcus neoformans is media dependent and alters host response, inducing protective immunity Upadhya, Rajendra Lam, Woei C. Hole, Camaron R. Vasselli, Joseph G. Lodge, Jennifer K. Front Fungal Biol Fungal Biology INTRODUCTION: Cryptococcus neoformans is a basidiomycete fungus that can cause meningoencephalitis, especially in immunocompromised patients. Cryptococcus grows in many different media, although little attention has been paid to the role of growth conditions on the cryptococcal cell wall or on virulence. OBJECTIVE: The purpose of this study was to determine how different media influenced the amount of chitin and chitosan in the cell wall, which in turn impacted the cell wall architecture and host response. METHODS: Yeast extract, peptone, and dextrose (YPD) and yeast nitrogen base (YNB) are two commonly used media for growing Cryptococcus before use in in vitro or in vivo experiments. As a result, C. neoformans was grown in either YPD or YNB, which were either left unbuffered or buffered to pH 7 with MOPS. These cells were then labeled with cell wall-specific fluorescent probes to determine the amounts of various cell wall components. In addition, these cells were employed in animal virulence studies using the murine inhalation model of infection. RESULTS: We observed that the growth of wild-type C. neoformans KN99 significantly changes the pH of unbuffered media during growth. It raises the pH to 8.0 when grown in unbuffered YPD but lowers the pH to 2.0 when grown in unbuffered YNB (YNB-U). Importantly, the composition of the cell wall was substantially impacted by growth in different media. Cells grown in YNB-U exhibited a 90% reduction in chitosan, the deacetylated form of chitin, compared with cells grown in YPD. The decrease in pH and chitosan in the YNB-U-grown cells was associated with a significant increase in some pathogen-associated molecular patterns on the surface of cells compared with cells grown in YPD or YNB, pH 7. This altered cell wall architecture resulted in a significant reduction in virulence when tested using a murine model of infection. Furthermore, when heat-killed cells were used as the inoculum, KN99 cells grown in YNB-U caused an aberrant hyper-inflammatory response in the lungs, resulting in rapid animal death. In contrast, heat-killed KN99 cells grown in YNB, pH 7, caused little to no inflammatory response in the host lung, but, when used as a vaccine, they conferred a robust protective response against a subsequent challenge infection with the virulent KN99 cells. CONCLUSION: These findings emphasize the importance of culture media and pH during growth in shaping the content and organization of the C. neoformans cell wall, as well as their impact on fungal virulence and the host response. Frontiers Media S.A. 2023-05-12 /pmc/articles/PMC10399910/ /pubmed/37538303 http://dx.doi.org/10.3389/ffunb.2023.1183291 Text en Copyright © 2023 Upadhya, Lam, Hole, Vasselli and Lodge https://creativecommons.org/licenses/by/4.0/This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.
spellingShingle Fungal Biology
Upadhya, Rajendra
Lam, Woei C.
Hole, Camaron R.
Vasselli, Joseph G.
Lodge, Jennifer K.
Cell wall composition in Cryptococcus neoformans is media dependent and alters host response, inducing protective immunity
title Cell wall composition in Cryptococcus neoformans is media dependent and alters host response, inducing protective immunity
title_full Cell wall composition in Cryptococcus neoformans is media dependent and alters host response, inducing protective immunity
title_fullStr Cell wall composition in Cryptococcus neoformans is media dependent and alters host response, inducing protective immunity
title_full_unstemmed Cell wall composition in Cryptococcus neoformans is media dependent and alters host response, inducing protective immunity
title_short Cell wall composition in Cryptococcus neoformans is media dependent and alters host response, inducing protective immunity
title_sort cell wall composition in cryptococcus neoformans is media dependent and alters host response, inducing protective immunity
topic Fungal Biology
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10399910/
https://www.ncbi.nlm.nih.gov/pubmed/37538303
http://dx.doi.org/10.3389/ffunb.2023.1183291
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