Elevated Cholesterol in the <italic toggle="yes">Coxiella burnetii</italic> Intracellular Niche Is Bacteriolytic

ABSTRACT Coxiella burnetii is an intracellular bacterial pathogen and a significant cause of culture-negative endocarditis in the United States. Upon infection, the nascent Coxiella phagosome fuses with the host endocytic pathway to form a large lysosome-like vacuole called the parasitophorous vacuo...

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Autores principales:	Minal Mulye, Dhritiman Samanta, Seth Winfree, Robert A. Heinzen, Stacey D. Gilk
Formato:	article
Lenguaje:	EN
Publicado:	American Society for Microbiology 2017
Materias:	Microbiology QR1-502
Acceso en línea:	https://doaj.org/article/be01b1fbc33648889d32486b6313c022
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spelling	oai:doaj.org-article:be01b1fbc33648889d32486b6313c0222021-11-15T15:51:07ZElevated Cholesterol in the <italic toggle="yes">Coxiella burnetii</italic> Intracellular Niche Is Bacteriolytic10.1128/mBio.02313-162150-7511https://doaj.org/article/be01b1fbc33648889d32486b6313c0222017-03-01T00:00:00Zhttps://journals.asm.org/doi/10.1128/mBio.02313-16https://doaj.org/toc/2150-7511ABSTRACT Coxiella burnetii is an intracellular bacterial pathogen and a significant cause of culture-negative endocarditis in the United States. Upon infection, the nascent Coxiella phagosome fuses with the host endocytic pathway to form a large lysosome-like vacuole called the parasitophorous vacuole (PV). The PV membrane is rich in sterols, and drugs perturbing host cell cholesterol homeostasis inhibit PV formation and bacterial growth. Using cholesterol supplementation of a cholesterol-free cell model system, we found smaller PVs and reduced Coxiella growth as cellular cholesterol concentration increased. Further, we observed in cells with cholesterol a significant number of nonfusogenic PVs that contained degraded bacteria, a phenotype not observed in cholesterol-free cells. Cholesterol had no effect on axenic Coxiella cultures, indicating that only intracellular bacteria are sensitive to cholesterol. Live-cell microscopy revealed that both plasma membrane-derived cholesterol and the exogenous cholesterol carrier protein low-density lipoprotein (LDL) traffic to the PV. To test the possibility that increasing PV cholesterol levels affects bacterial survival, infected cells were treated with U18666A, a drug that traps cholesterol in lysosomes and PVs. U18666A treatment led to PVs containing degraded bacteria and a significant loss in bacterial viability. The PV pH was significantly more acidic in cells with cholesterol or cells treated with U18666A, and the vacuolar ATPase inhibitor bafilomycin blocked cholesterol-induced PV acidification and bacterial death. Additionally, treatment of infected HeLa cells with several FDA-approved cholesterol-altering drugs led to a loss of bacterial viability, a phenotype also rescued by bafilomycin. Collectively, these data suggest that increasing PV cholesterol further acidifies the PV, leading to Coxiella death. IMPORTANCE The intracellular Gram-negative bacterium Coxiella burnetii is a significant cause of culture-negative infectious endocarditis, which can be fatal if untreated. The existing treatment strategy requires prolonged antibiotic treatment, with a 10-year mortality rate of 19% in treated patients. Therefore, new clinical therapies are needed and can be achieved by better understanding C. burnetii pathogenesis. Upon infection of host cells, C. burnetii grows within a specialized replication niche, the parasitophorous vacuole (PV). Recent data have linked cholesterol to intracellular C. burnetii growth and PV formation, leading us to further decipher the role of cholesterol during C. burnetii-host interaction. We observed that increasing PV cholesterol concentration leads to increased acidification of the PV and bacterial death. Further, treatment with FDA-approved drugs that alter host cholesterol homeostasis also killed C. burnetii through PV acidification. Our findings suggest that targeting host cholesterol metabolism might prove clinically efficacious in controlling C. burnetii infection.Minal MulyeDhritiman SamantaSeth WinfreeRobert A. HeinzenStacey D. GilkAmerican Society for MicrobiologyarticleMicrobiologyQR1-502ENmBio, Vol 8, Iss 1 (2017)
institution	DOAJ
collection	DOAJ
language	EN
topic	Microbiology QR1-502
spellingShingle	Microbiology QR1-502 Minal Mulye Dhritiman Samanta Seth Winfree Robert A. Heinzen Stacey D. Gilk Elevated Cholesterol in the <italic toggle="yes">Coxiella burnetii</italic> Intracellular Niche Is Bacteriolytic
description	ABSTRACT Coxiella burnetii is an intracellular bacterial pathogen and a significant cause of culture-negative endocarditis in the United States. Upon infection, the nascent Coxiella phagosome fuses with the host endocytic pathway to form a large lysosome-like vacuole called the parasitophorous vacuole (PV). The PV membrane is rich in sterols, and drugs perturbing host cell cholesterol homeostasis inhibit PV formation and bacterial growth. Using cholesterol supplementation of a cholesterol-free cell model system, we found smaller PVs and reduced Coxiella growth as cellular cholesterol concentration increased. Further, we observed in cells with cholesterol a significant number of nonfusogenic PVs that contained degraded bacteria, a phenotype not observed in cholesterol-free cells. Cholesterol had no effect on axenic Coxiella cultures, indicating that only intracellular bacteria are sensitive to cholesterol. Live-cell microscopy revealed that both plasma membrane-derived cholesterol and the exogenous cholesterol carrier protein low-density lipoprotein (LDL) traffic to the PV. To test the possibility that increasing PV cholesterol levels affects bacterial survival, infected cells were treated with U18666A, a drug that traps cholesterol in lysosomes and PVs. U18666A treatment led to PVs containing degraded bacteria and a significant loss in bacterial viability. The PV pH was significantly more acidic in cells with cholesterol or cells treated with U18666A, and the vacuolar ATPase inhibitor bafilomycin blocked cholesterol-induced PV acidification and bacterial death. Additionally, treatment of infected HeLa cells with several FDA-approved cholesterol-altering drugs led to a loss of bacterial viability, a phenotype also rescued by bafilomycin. Collectively, these data suggest that increasing PV cholesterol further acidifies the PV, leading to Coxiella death. IMPORTANCE The intracellular Gram-negative bacterium Coxiella burnetii is a significant cause of culture-negative infectious endocarditis, which can be fatal if untreated. The existing treatment strategy requires prolonged antibiotic treatment, with a 10-year mortality rate of 19% in treated patients. Therefore, new clinical therapies are needed and can be achieved by better understanding C. burnetii pathogenesis. Upon infection of host cells, C. burnetii grows within a specialized replication niche, the parasitophorous vacuole (PV). Recent data have linked cholesterol to intracellular C. burnetii growth and PV formation, leading us to further decipher the role of cholesterol during C. burnetii-host interaction. We observed that increasing PV cholesterol concentration leads to increased acidification of the PV and bacterial death. Further, treatment with FDA-approved drugs that alter host cholesterol homeostasis also killed C. burnetii through PV acidification. Our findings suggest that targeting host cholesterol metabolism might prove clinically efficacious in controlling C. burnetii infection.
format	article
author	Minal Mulye Dhritiman Samanta Seth Winfree Robert A. Heinzen Stacey D. Gilk
author_facet	Minal Mulye Dhritiman Samanta Seth Winfree Robert A. Heinzen Stacey D. Gilk
author_sort	Minal Mulye
title	Elevated Cholesterol in the <italic toggle="yes">Coxiella burnetii</italic> Intracellular Niche Is Bacteriolytic
title_short	Elevated Cholesterol in the <italic toggle="yes">Coxiella burnetii</italic> Intracellular Niche Is Bacteriolytic
title_full	Elevated Cholesterol in the <italic toggle="yes">Coxiella burnetii</italic> Intracellular Niche Is Bacteriolytic
title_fullStr	Elevated Cholesterol in the <italic toggle="yes">Coxiella burnetii</italic> Intracellular Niche Is Bacteriolytic
title_full_unstemmed	Elevated Cholesterol in the <italic toggle="yes">Coxiella burnetii</italic> Intracellular Niche Is Bacteriolytic
title_sort	elevated cholesterol in the <italic toggle="yes">coxiella burnetii</italic> intracellular niche is bacteriolytic
publisher	American Society for Microbiology
publishDate	2017
url	https://doaj.org/article/be01b1fbc33648889d32486b6313c022
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Elevated Cholesterol in the <italic toggle="yes">Coxiella burnetii</italic> Intracellular Niche Is Bacteriolytic

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