scholarly journals Coxiella burnetiiType 4B Secretion System-dependent manipulation of endolysosomal maturation is required for bacterial growth

2019 ◽  
Author(s):  
Dhritiman Samanta ◽  
Tatiana M. Clemente ◽  
Stacey D. Gilk
Keyword(s):  
Host Cell ◽  
Secretion System ◽  
Intracellular Bacterium ◽  
Host Cells ◽  
Acidic Ph ◽  
Obligate Intracellular Bacterium ◽  
Cell Infection ◽  
Obligate Intracellular ◽  
Endosomal Acidification ◽  
Infected Cells

AbstractUpon host cell infection, the obligate intracellular bacteriumC. burnetiiresides and multiplies within theCoxiella–ContainingVacuole (CCV). The nascent CCV progresses through the endosomal maturation pathway into a phagolysosome, acquiring lysosomal markers as well as acidic pH and active proteases and hydrolases. Approximately 24-48 hours post infection, heterotypic fusion between the CCV and host endosomes/lysosomes leads to CCV expansion and subsequent bacterial replication in the mature CCV. Initial CCV acidification is required to activateC. burnetiimetabolism and the Type 4B Secretion System (T4BSS), which secretes effector proteins required for CCV maturation. However, we recently found that the mature CCV is less acidic (pH~5.2) than lysosomes (pH~4.8). Further, CCV acidification to pH~4.8 causesC. burnetiilysis, suggestingC. burnetiiactively regulates CCV pH. Because heterotypic fusion with host endosomes/lysosomes may influence CCV pH, we investigated endosomal maturation in cells infected with wildtype (WT) or T4BSS mutant (ΔdotA)C. burnetii. We observed significantly fewer LAMP1-positive lysosomes, along with less acidic “mature” endosomes (pH~5.8), in WT-infected cells, compared to mock or ΔdotA-infected cells. Further, while endosomes progressively acidified from the periphery (pH~5.5) to the perinuclear area (pH~4.7) in both mock and ΔdotA-infected cells, endosomes did not acidify beyond pH~5.2 in WT-infected cells, indicating that theC. burnetiiT4BSS inhibits endosomal maturation. Finally, increasing the number of acidic lysosomes by overexpressing the transcription factor EB inhibitedC. burnetiigrowth, indicating lysosomes are detrimental toC. burnetii. Overall, our data suggest thatC. burnetiiregulates CCV pH, possibly by reducing the number of host lysosomes available for heterotypic fusion.Author summaryThe obligate intracellular bacteriumCoxiella burnetiicauses human Q fever, which manifests as a flu-like illness but can develop into a life-threatening and difficult to treat endocarditis.C. burnetii,in contrast to many other intracellular bacteria, thrives within a lysosome-like vacuole in host cells. However, we previously found that theC. burnetiivacuole is not as acidic as lysosomes and increased acidification kills the bacteria, suggesting thatC. burnetiiregulates the pH of its vacuole. Here, we discovered thatC. burnetiiblocks endosomal maturation and acidification during host cell infection, resulting in fewer lysosomes in the host cell. Moreover, increasing lysosomes in the host cells blockedC. burnetiigrowth. Together, our study suggests thatC. burnetiiregulates vacuole acidity and blocks endosomal acidification in order to produce a permissive intracellular niche.

scholarly journals Developmental differentiation in a cytoplasm-dwelling obligate intracellular bacterium

2020 ◽  
Author(s):  
Suparat Giengkam ◽  
Jantana Wongsantichon ◽  
Sharanjeet Atwal ◽  
Yanin Jaiyen ◽  
Wah Ing Goh ◽  
...  
Keyword(s):  
Host Cell ◽  
Metabolic Activity ◽  
Model System ◽  
Intracellular Bacterium ◽  
Host Cells ◽  
Orientia Tsutsugamushi ◽  
Obligate Intracellular Bacterium ◽  
New Model ◽  
Obligate Intracellular ◽  
Developmental Differentiation

AbstractDevelopmental differentiation has been described for several vacuole-dwelling obligate intracellular bacteria but never for an obligate intracellular bacterium that resides in the cytoplasm. Here, we show that the cytoplasm-dwelling obligate intracellular bacterium Orientia tsutsugamushi (Ot) exists in five distinct subpopulations. We show that Ot differentiates into a distinct, metabolically inactive, extracellular state upon budding from the surface of host cells and that this stage is preceded by a surface-associated maturation stage. We identify proteins that are differentially expressed in intracellular, replicative bacteria and extracellular, metabolically inactive bacteria. Metabolic activity resumes rapidly upon entry into the cytoplasm and is triggered by the host cell reducing environment. This example of developmental differentiation in a species of Rickettsiaceae provides a new model system for studying synchronized differentiation in a bacterium that has a minimal genome and where the interactions between bacterium and host cell are more direct than they are for bacteria separated from the eukaryote cell host by a vacuolar membrane.Author SummaryScrub typhus is a life-threatening human infection that is caused by the bacterium Orientia tsutsugamushi and spread by mites. Although the disease is estimated to affect at least one million people annually and is often fatal, the infectious agent is much less well understood than many other pathogens. O. tsutsugamushi is an intracellular bacterium that can only grow and divide within eukaryotic cells. During infection, it is found primarily in the cells that make up the lining of blood vessels and in certain immune cell types. O. tsutsugamushi bacteria can remain inside a single infected cell for seven days or more before budding out. The ways in which the bacterium itself changes during the course of an intracellular infection cycle have not been studied. In the current work, we used a range of techniques to show that O. tsutsugamushi differentiates into five distinct subpopulations, and that these are associated with measurable differences in metabolic activity, replication and infectivity. This work opens new avenues of research into the regulation and mechanisms of differentiation of O. tsutsugamushi, which could lead to improved diagnosis and treatments. It also provides a new model system for studying fundamental questions about bacterial development.

scholarly journals Role of Bcl-2 Family Members in Caspase-Independent Apoptosis during Chlamydia Infection

2002 ◽  
Vol 70 (1) ◽  
pp. 55-61 ◽  
Author(s):  
Jean-Luc Perfettini ◽  
John C. Reed ◽  
Nicole Israël ◽  
Jean-Claude Martinou ◽  
Alice Dautry-Varsat ◽  
...  
Keyword(s):  
Caspase 3 ◽  
Chlamydia Psittaci ◽  
Host Cells ◽  
Chlamydia Infection ◽  
Caspase Inhibitor ◽  
Obligate Intracellular Bacterium ◽  
Obligate Intracellular ◽  
Infected Cells ◽  
Induced Apoptosis

ABSTRACT Infection with an obligate intracellular bacterium, the Chlamydia trachomatis lymphogranuloma venereum (LGV/L2) strain or the guinea pig inclusion conjunctivitis serovar of Chlamydia psittaci, leads to apoptosis of host cells. The apoptosis is not affected by a broad-spectrum caspase inhibitor, and caspase-3 is not activated in infected cells, suggesting that apoptosis mediated by these two strains of Chlamydia is independent of known caspases. Overexpression of the proapoptotic Bcl-2 family member, Bax, was previously shown to induce caspase-independent apoptosis, and we find that Bax is activated and translocates from the cytosol to the mitochondria in C. psittaci-infected cells. C. psittaci-induced apoptosis is inhibited in host cells overexpressing Bax inhibitor-1 and is inhibited through overexpression of Bcl-2, which blocks both caspase-dependent and -independent apoptosis. As Bax and mitochondria are ideally located to sense stress-related metabolic changes emanating from the interior of an infected cell, it is likely that Bax-dependent apoptosis may also be observed in cells infected with other intracellular pathogens.

scholarly journals Extrusions promote engulfment and Chlamydia survival within macrophages

2016 ◽  
Author(s):  
Meghan Zuck ◽  
Tisha C Ellis ◽  
Anthony Venida ◽  
Kevin Hybiske
Keyword(s):  
Immune Responses ◽  
Host Cell ◽  
Membrane Structure ◽  
Functional Role ◽  
Host Cells ◽  
Obligate Intracellular Bacterium ◽  
Obligate Intracellular ◽  
Primary Bone ◽  
New Hosts ◽  
Primary Bone Marrow

All obligate intracellular pathogens must exit their host cells in order to propagate and survive as a species; the precise strategies they use have a direct impact on their ability to disseminate within a host, transmit to new hosts, and engage or avoid immune responses. The obligate intracellular bacterium Chlamydia trachomatis exits the host cell by two distinct exit strategies, lysis and extrusion. Despite being equally active pathways, lysis and extrusion differ greatly in their mechanisms. The defining characteristics of extrusions, and advantages gained by Chlamydia within this unique double-membrane structure are not well understood. Here, we present data that defines extrusions as being largely devoid of host organelles, comprised mostly of Chlamydia elementary bodies, and containing phosphatidylserine on the outer surface of the extrusion membrane. Towards defining a functional role for extrusions in Chlamydia pathogenesis, we demonstrate that extrusions confer significant infectious advantages for Chlamydia by serving as transient, intracellular-like niches for extracellular Chlamydia, as compared to Chlamydia that would otherwise exit by lysing the host cell. In addition to enhanced survival outside of the host cell, we report the key discovery that chlamydial extrusions can be engulfed by primary bone marrow-derived macrophages, after which they provide a protective microenvironment for Chlamydia. Extrusion-derived Chlamydia were able to stave off macrophage based killing beyond 8 h, and culminated in the release of infectious EB from the macrophage. Based on these findings, we propose a model in which a major outcome of Chlamydia exiting epithelial cells inside extrusions is to hijack macrophages as vehicles for dissemination within the host.

scholarly journals Exploitation of the Endocytic Pathway by Orientia tsutsugamushi in Nonprofessional Phagocytes

2006 ◽  
Vol 74 (7) ◽  
pp. 4246-4253 ◽  
Author(s):  
Hyuk Chu ◽  
Jung-Hee Lee ◽  
Seung-Hoon Han ◽  
Se-Yoon Kim ◽  
Nam-Hyuk Cho ◽  
...  
Keyword(s):  
Host Cell ◽  
Causative Agent ◽  
Scrub Typhus ◽  
Adaptor Protein ◽  
Endocytic Pathway ◽  
Intracellular Bacterium ◽  
Orientia Tsutsugamushi ◽  
Obligate Intracellular Bacterium ◽  
Obligate Intracellular ◽  
Endocytic Vesicles

ABSTRACT Orientia tsutsugamushi, a causative agent of scrub typhus, is an obligate intracellular bacterium that requires the exploitation of the endocytic pathway in the host cell. We observed the localization of O. tsutsugamushi with clathrin or adaptor protein 2 within 30 min after the infection of nonprofessional phagocytes. We have further confirmed that the infectivity of O. tsutsugamushi is significantly reduced by drugs that block clathrin-mediated endocytosis but not by filipin III, an inhibitor that blocks caveola-mediated endocytosis. In the present study, with a confocal microscope, O. tsutsugamushi was sequentially colocalized with the early and late endosomal markers EEA1 and LAMP2, respectively, within 1 h after infection. The colocalization of O. tsutsugamushi organisms with EEA1 and LAMP2 gradually disappeared until 2 h postinfection, and then free O. tsutsugamushi organisms were found in the cytoplasm. When the acidification of endocytic vesicles was blocked by treating the cells with NH4Cl or bafilomycin A, the escape of O. tsutsugamushi organisms from the endocytic pathway was severely impaired, and the infectivity of O. tsutsugamushi was drastically reduced. To our knowledge, this is the first report that the invasion of O. tsutsugamushi is dependent on the clathrin-dependent endocytic pathway and the acidification process of the endocytic vesicles in nonprofessional phagocytes.

COVID 19 – Eye Issues

2020 ◽  
Vol 5 (Special) ◽  
Keyword(s):  
Host Cell ◽  
Target Cell ◽  
Cell Receptor ◽  
Human Tissues ◽  
Type Ii ◽  
Cell Infection ◽  
Host Cell Receptor ◽  
Infected Cells ◽  
Cellular Entry

The coronavirus illness (COVID-19) is caused by a new recombinant SARS-CoV (SARS-CoV) virus (SARS-CoV-2). Target cell infection by SARS-CoV is mediated by the prickly protein of the coronavirus and host cell receptor, enzyme 2 converting angiotensin (ACE2) [3]. Similarly, a recent study suggests that cellular entry by SARS-CoV-2 is dependent on both ACE2 as well as type II transmembrane axial protease (TMPRSS2) [4]. This means that detection of ACE2 and PRSS2 expression in human tissues can predict potential infected cells and their respective effects in COVID-19 patients [1].

scholarly journals Revisiting Ehrlichia ruminantium Replication Cycle Using Proteomics: The Host and the Bacterium Perspectives

2021 ◽  
Vol 9 (6) ◽  
pp. 1144
Author(s):  
Isabel Marcelino ◽  
Philippe Holzmuller ◽  
Ana Coelho ◽  
Gabriel Mazzucchelli ◽  
Bernard Fernandez ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Host Cell ◽  
Cell Metabolism ◽  
Replication Cycle ◽  
Host Cells ◽  
Ehrlichia Ruminantium ◽  
Host Interaction ◽  
Infected Cells ◽  
Related Proteins

The Rickettsiales Ehrlichia ruminantium, the causal agent of the fatal tick-borne disease Heartwater, induces severe damage to the vascular endothelium in ruminants. Nevertheless, E. ruminantium-induced pathobiology remains largely unknown. Our work paves the way for understanding this phenomenon by using quantitative proteomic analyses (2D-DIGE-MS/MS, 1DE-nanoLC-MS/MS and biotin-nanoUPLC-MS/MS) of host bovine aorta endothelial cells (BAE) during the in vitro bacterium intracellular replication cycle. We detect 265 bacterial proteins (including virulence factors), at all time-points of the E. ruminantium replication cycle, highlighting a dynamic bacterium–host interaction. We show that E. ruminantium infection modulates the expression of 433 host proteins: 98 being over-expressed, 161 under-expressed, 140 detected only in infected BAE cells and 34 exclusively detected in non-infected cells. Cystoscape integrated data analysis shows that these proteins lead to major changes in host cell immune responses, host cell metabolism and vesicle trafficking, with a clear involvement of inflammation-related proteins in this process. Our findings led to the first model of E. ruminantium infection in host cells in vitro, and we highlight potential biomarkers of E. ruminantium infection in endothelial cells (such as ROCK1, TMEM16K, Albumin and PTPN1), which may be important to further combat Heartwater, namely by developing non-antibiotic-based strategies.

scholarly journals CXCR2 Blockade Influences Anaplasma phagocytophilum Propagation but Not Histopathology in the Mouse Model of Human Granulocytic Anaplasmosis

2004 ◽  
Vol 11 (5) ◽  
pp. 963-968 ◽  
Author(s):  
Diana G. Scorpio ◽  
Mustafa Akkoyunlu ◽  
Erol Fikrig ◽  
J. Stephen Dumler
Keyword(s):  
Anaplasma Phagocytophilum ◽  
Tissue Injury ◽  
Treated Group ◽  
Obligate Intracellular Bacterium ◽  
Human Granulocytic Anaplasmosis ◽  
Obligate Intracellular ◽  
Liver Histopathology ◽  
Granulocytic Anaplasmosis ◽  
Infected Cells ◽  
And Control

ABSTRACT Anaplasma phagocytophilum is an obligate intracellular bacterium that infects neutrophils and causes human granulocytic anaplasmosis. Infection induces neutrophil secretion of interleukin-8 or murine homologs and perpetuates infection by recruiting susceptible neutrophils. We hypothesized that antibody blockade of CXCR2 would decrease A. phagocytophilum tissue load by interrupting neutrophil recruitment but would not influence murine hepatic pathology. C3H-scid mice were treated with CXCR2 antiserum or control prior to or on day 14 after infection. Quantitative PCR and immunohistochemistry for A. phagocytophilum were performed and severity of liver histopathology was ranked. Control mice had more infected cells in tissues than the anti-CXCR2-treated group. The histopathological rank was not different between treated and control animals. Infected cells of control mice clustered in tissue more than in treated mice. The results support the hypothesis of bacterial propagation through chemokine induction and confirm that tissue injury is unrelated to A. phagocytophilum tissue load.

scholarly journals Chlamydial Lytic Exit from Host Cells Is Plasmid Regulated

mBio ◽  
2015 ◽  
Vol 6 (6) ◽  
Author(s):  
Chunfu Yang ◽  
Tregei Starr ◽  
Lihua Song ◽  
John H. Carlson ◽  
Gail L. Sturdevant ◽  
...  
Keyword(s):  
Type Iii Secretion ◽  
Actin Polymerization ◽  
Type Iii Secretion System ◽  
Secretion System ◽  
Cytoskeletal Proteins ◽  
Host Cells ◽  
Genetic Mechanism ◽  
Developmental Cycle ◽  
Obligate Intracellular ◽  
Type Iii

ABSTRACTChlamydia trachomatisis an obligate intracellular bacterium that is a globally important human pathogen. The chlamydial plasmid is an attenuating virulence factor, but the molecular basis for attenuation is not understood. Chlamydiae replicate within a membrane-bound vacuole termed an inclusion, where they undergo a biphasic developmental growth cycle and differentiate from noninfectious into infectious organisms. Late in the developmental cycle, the fragile chlamydia-laden inclusion retains its integrity by surrounding itself with scaffolds of host cytoskeletal proteins. The ability of chlamydiae to developmentally free themselves from this cytoskeleton network is a fundamental virulence trait of the pathogen. Here, we show that plasmidless chlamydiae are incapable of disrupting their cytoskeletal entrapment and remain intracellular as stable mature inclusions that support high numbers of infectious organisms. By using deletion mutants of the eight plasmid-carried genes (Δpgp1to Δpgp8), we show that Pgp4, a transcriptional regulator of multiple chromosomal genes, is required for exit. Exit of chlamydiae is dependent on protein synthesis and is inhibited by the compound C1, an inhibitor of the type III secretion system (T3S). Exit of plasmid-free and Δpgp4organisms, which failed to lyse infected cells, was rescued by latrunculin B, an inhibitor of actin polymerization. Our findings describe a genetic mechanism of chlamydial exit from host cells that is dependent on an unknownpgp4-regulated chromosomal T3S effector gene.IMPORTANCEChlamydia's obligate intracellular life style requires both entry into and exit from host cells. Virulence factors that function in exiting are unknown. The chlamydial inclusion is stabilized late in the infection cycle by F-actin. A prerequisite of chlamydial exit is its ability to disassemble actin from the inclusion. We show that chlamydial plasmid-free organisms, and also a plasmid gene protein 4 (pgp4) null mutant, do not disassociate actin from the inclusion and fail to exit cells. We further provide evidence that Pgp4-regulated exit is dependent on the chlamydial type III secretion system. This study is the first to define a genetic mechanism that functions in chlamydial lytic exit from host cells. The findings also have practical implications for understanding why plasmid-free chlamydiae are highly attenuated and have the ability to elicit robust protective immune responses.

scholarly journals A streamlined method for transposon mutagenesis ofRickettsia parkeriyields numerous mutations that impact infection

2018 ◽  
Author(s):  
Rebecca L. Lamason ◽  
Natasha M. Kafai ◽  
Matthew D. Welch
Keyword(s):  
Host Cell ◽  
Virulence Factors ◽  
Vascular Endothelial Cells ◽  
Spotted Fever ◽  
Transposon Mutagenesis ◽  
Host Cells ◽  
Spotted Fever Group ◽  
Loss Of Function ◽  
Rickettsia Parkeri ◽  
Obligate Intracellular

AbstractThe rickettsiae are obligate intracellular alphaproteobacteria that exhibit a complex infectious life cycle in both arthropod and mammalian hosts. As obligate intracellular bacteria,Rickettsiaare highly adapted to living inside a variety of host cells, including vascular endothelial cells during mammalian infection. Although it is assumed that the rickettsiae produce numerous virulence factors that usurp or disrupt various host cell pathways, they have been challenging to genetically manipulate to identify the key bacterial factors that contribute to infection. Motivated to overcome this challenge, we sought to expand the repertoire of available rickettsial loss-of-function mutants, using an improvedmariner-based transposon mutagenesis scheme. Here, we present the isolation of over 100 transposon mutants in the spotted fever group speciesRickettsia parkeri. These mutants targeted genes implicated in a variety of pathways, including bacterial replication and metabolism, hypothetical proteins, the type IV secretion system, as well as factors with previously established roles in host cell interactions and pathogenesis. Given the need to identify critical virulence factors, forward genetic screens such as this will provide an excellent platform to more directly investigate rickettsial biology and pathogenesis.

Host range of obligate intracellular bacterium Parachlamydia acanthamoebae

2010 ◽  
Vol 54 (11) ◽  
pp. 707-713 ◽  
Author(s):  
Yasuhiro Hayashi ◽  
Shinji Nakamura ◽  
Junji Matsuo ◽  
Tatsuya Fukumoto ◽  
Mitsutaka Yoshida ◽  
...  
Keyword(s):  
Host Range ◽  
Intracellular Bacterium ◽  
Obligate Intracellular Bacterium ◽  
Obligate Intracellular
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