scholarly journals Induction of Salmonella pathogenicity island 1 under different growth conditions can affect Salmonella–host cell interactions in vitro

Microbiology ◽  
2010 ◽  
Vol 156 (4) ◽  
pp. 1120-1133 ◽  
Author(s):  
J. Antonio Ibarra ◽  
Leigh A. Knodler ◽  
Daniel E. Sturdevant ◽  
Kimmo Virtaneva ◽  
Aaron B. Carmody ◽  
...  
Keyword(s):  
Host Cell ◽  
Single Cell Analysis ◽  
Pathogenicity Island ◽  
Growth Conditions ◽  
Culture Conditions ◽  
Effector Proteins ◽  
Host Cells ◽  
Phagocytic Cells ◽  
Synthetic Media

Salmonella invade non-phagocytic cells by inducing massive actin rearrangements, resulting in membrane ruffle formation and phagocytosis of the bacteria. This process is mediated by a cohort of effector proteins translocated into the host cell by type III secretion system 1, which is encoded by genes in the Salmonella pathogenicity island (SPI) 1 regulon. This network is precisely regulated and must be induced outside of host cells. In vitro invasive Salmonella are prepared by growth in synthetic media although the details vary. Here, we show that culture conditions affect the frequency, and therefore invasion efficiency, of SPI1-induced bacteria and also can affect the ability of Salmonella to adapt to its intracellular niche following invasion. Aerobically grown late-exponential-phase bacteria were more invasive and this was associated with a greater frequency of SPI1-induced, motile bacteria, as revealed by single-cell analysis of gene expression. Culture conditions also affected the ability of Salmonella to adapt to the intracellular environment, since they caused marked differences in intracellular replication. These findings show that induction of SPI1 under different pre-invasion growth conditions can affect the ability of Salmonella to interact with eukaryotic host cells.

scholarly journals Differences in Host Cell Invasion and Salmonella Pathogenicity Island 1 Expression between Salmonella enterica Serovar Paratyphi A and NontyphoidalS. Typhimurium

2016 ◽  
Vol 84 (4) ◽  
pp. 1150-1165 ◽  
Author(s):  
Dana Elhadad ◽  
Prerak Desai ◽  
Guntram A. Grassl ◽  
Michael McClelland ◽  
Galia Rahav ◽  
...  
Keyword(s):  
Host Cell ◽  
Cell Invasion ◽  
Salmonella Enterica ◽  
Intestinal Inflammation ◽  
Pathogenicity Island ◽  
Effector Proteins ◽  
Host Cells ◽  
Host Cell Invasion ◽  
Content Type ◽  
Microaerobic Conditions

Active invasion into nonphagocytic host cells is central toSalmonella entericapathogenicity and dependent on multiple genes withinSalmonellapathogenicity island 1 (SPI-1). Here, we explored the invasion phenotype and the expression of SPI-1 in the typhoidal serovarS. Paratyphi A compared to that of the nontyphoidal serovarS. Typhimurium. We demonstrate that whileS. Typhimurium is equally invasive under both aerobic and microaerobic conditions,S. Paratyphi A invades only following growth under microaerobic conditions. Transcriptome sequencing (RNA-Seq), reverse transcription-PCR (RT-PCR), Western blot, and secretome analyses established thatS. Paratyphi A expresses much lower levels of SPI-1 genes and secretes lesser amounts of SPI-1 effector proteins thanS. Typhimurium, especially under aerobic growth. Bypassing the native SPI-1 regulation by inducible expression of the SPI-1 activator, HilA, considerably elevated SPI-1 gene expression, host cell invasion, disruption of epithelial integrity, and induction of proinflammatory cytokine secretion byS. Paratyphi A but not byS. Typhimurium, suggesting that SPI-1 expression is naturally downregulated inS. Paratyphi A. Using streptomycin-treated mice, we were able to establish substantial intestinal colonization byS. Paratyphi A and showed moderately higher pathology and intestinal inflammation in mice infected withS. Paratyphi A overexpressinghilA. Collectively, our results reveal unexpected differences in SPI-1 expression betweenS. Paratyphi A andS. Typhimurium, indicate thatS. Paratyphi A host cell invasion is suppressed under aerobic conditions, and suggest that lower invasion in aerobic sites and suppressed expression of immunogenic SPI-1 components contributes to the restrained inflammatory infection elicited byS. Paratyphi A.

scholarly journals The Secreted Effector Protein EspZ Is Essential for Virulence of Rabbit Enteropathogenic Escherichia coli

2015 ◽  
Vol 83 (3) ◽  
pp. 1139-1149 ◽  
Author(s):  
John Scott Wilbur ◽  
Wyatt Byrd ◽  
Shylaja Ramamurthy ◽  
Hannah E. Ledvina ◽  
Khaldoon Khirfan ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Cell Death ◽  
Epithelial Cell ◽  
Host Cell ◽  
Bacterial Colonization ◽  
Clinical Signs ◽  
Effector Proteins ◽  
Host Cells ◽  
Tunel Staining

Attaching and effacing (A/E) pathogens adhere intimately to intestinal enterocytes and efface brush border microvilli. A key virulence strategy of A/E pathogens is the type III secretion system (T3SS)-mediated delivery of effector proteins into host cells. The secreted protein EspZ is postulated to promote enterocyte survival by regulating the T3SS and/or by modulating epithelial signaling pathways. To explore the role of EspZ in A/E pathogen virulence, we generated an isogenicespZdeletion strain (ΔespZ) and correspondingcis-complemented derivatives of rabbit enteropathogenicEscherichia coliand compared their abilities to regulate the T3SS and influence host cell survivalin vitro. For virulence studies, rabbits infected with these strains were monitored for bacterial colonization, clinical signs, and intestinal tissue alterations. Consistent with data from previous reports,espZ-transfected epithelial cells were refractory to infection-dependent effector translocation. Also, the ΔespZstrain induced greater host cell death than did the parent and complemented strains. In rabbit infections, fecal ΔespZstrain levels were 10-fold lower than those of the parent strain at 1 day postinfection, while the complemented strain was recovered at intermediate levels. In contrast to the parent and complemented mutants, ΔespZmutant fecal carriage progressively decreased on subsequent days. ΔespZmutant-infected animals gained weight steadily over the infection period, failed to show characteristic disease symptoms, and displayed minimal infection-induced histological alterations. Terminal deoxynucleotidyltransferase-mediated dUTP-biotin nick end labeling (TUNEL) staining of intestinal sections revealed increased epithelial cell apoptosis on day 1 after infection with the ΔespZstrain compared to animals infected with the parent or complemented strains. Thus, EspZ-dependent host cell cytoprotection likely prevents epithelial cell death and sloughing and thereby promotes bacterial colonization.

scholarly journals Functional Transfer of Salmonella Pathogenicity Island 2 to Salmonella bongori and Escherichia coli

2004 ◽  
Vol 72 (5) ◽  
pp. 2879-2888 ◽  
Author(s):  
Imke Hansen-Wester ◽  
Dipshikha Chakravortty ◽  
Michael Hensel
Keyword(s):  
Escherichia Coli ◽  
Host Cell ◽  
Pathogenicity Island ◽  
Culture Conditions ◽  
Effector Proteins ◽  
E Coli ◽  
Cell Functions ◽  
A Cell ◽  
Model Transfer ◽  
Systemic Infections

ABSTRACT The type III secretion system (T3SS) encoded by the Salmonella pathogenicity island 2 (SPI2) has a central role in systemic infections by Salmonella enterica and for the intracellular phenotype. Intracellular S. enterica uses the SPI2-encoded T3SS to translocate a set of effector proteins into the host cell, which modify host cell functions, enabling intracellular survival and replication of the bacteria. We sought to determine whether specific functions of the SPI2-encoded T3SS can be transferred to heterologous hosts Salmonella bongori and Escherichia coli Mutaflor, species that lack the SPI2 locus and loci encoding effector proteins. The SPI2 virulence locus was cloned and functionally expressed in S. bongori and E. coli. Here, we demonstrate that S. bongori harboring the SPI2 locus is capable of secretion of SPI2 substrate proteins under culture conditions, as well as of translocation of effector proteins under intracellular conditions. An SPI2-mediated cellular phenotype was induced by S. bongori harboring the SPI2 if the sifA locus was cotransferred. An interference with the host cell microtubule cytoskeleton, a novel SPI2-dependent phenotype, was observed in epithelial cells infected with S. bongori harboring SPI2 without additional effector genes. S. bongori harboring SPI2 showed increased intracellular persistence in a cell culture model, but SPI2 transfer was not sufficient to confer to S. bongori systemic pathogenicity in a murine model. Transfer of SPI2 to heterologous hosts offers a new tool for the study of SPI2 functions and the phenotypes of individual effectors.

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 Perturbation of host autophagy by the Irish potato famine pathogen

2014 ◽  
Vol 70 (a1) ◽  
pp. C826-C826
Author(s):  
Abbas Maqbool ◽  
Richard Richard ◽  
Tolga Bozkurt ◽  
Yasin Dagdas ◽  
Khaoula Belhai ◽  
...  
Keyword(s):  
Host Cell ◽  
Plant Cells ◽  
Irish Potato ◽  
Effector Proteins ◽  
Host Cells ◽  
Cell Physiology ◽  
Biochemical Basis ◽  
The Impact ◽  
Potato Famine ◽  
Irish Potato Famine

Autophagy is a catabolic process involving degradation of dysfunctional cytoplasmic components to ensure cellular survival under starvation conditions. The process involves formation of double-membrane vesicles called autophagosomes and delivery of the inner constituents to lytic compartments. It can also target invading pathogens, such as intracellular bacteria, for destruction and is thus implicated in innate immune pathways [1]. In response, certain mammalian pathogens deliver effector proteins into host cells that inhibit autophagy and contribute to enabling parasitic infection [2]. Pyhtophthora infestans, the Irish potato famine pathogen, is a causative agent of late blight disease in potato and tomato crops. It delivers a plethora of modular effector proteins into plant cells to promote infection. Once inside the cell, RXLR-type effector proteins engage with host cell proteins, to manipulate host cell physiology for the benefit of the pathogen. As plants lack an adaptive immune system, this provides a robust mechanism for pathogens to circumvent host defense. PexRD54 is an intracellular RXLR-type effector protein produced by P. infestans. PexRD54 interacts with potato homologues of autophagy protein ATG8 in plant cells. We have been investigating the structural and biochemical basis of the PexRD54/ATG8 interaction in vitro. We have purified PexRD54 and ATG8 independently and in complex from E. coli. Using protein/protein interaction studies we have shown that PexRD54 binds ATG8 with sub-micromolar affinity. We have also determined the structure of PexRD54 in the presence of ATG8. This crystal structure provides key insights into how the previously reported WY-fold of oomycete RXLR-type effectors [3] can be organized in multiple repeats. The structural data also provides insights into the interaction between PexRD54 and ATG8, suggesting further experiments to understand the impact of this interaction on host cell physiology and how this benefits the pathogen.

Cellular and immunological basis of the host-parasite relationship during infection withNeospora caninum

Parasitology ◽  
2006 ◽  
Vol 133 (3) ◽  
pp. 261-278 ◽  
Author(s):  
A. HEMPHILL ◽  
N. VONLAUFEN ◽  
A. NAGULESWARAN
Keyword(s):  
Host Cell ◽  
Cell Biology ◽  
Neospora Caninum ◽  
Host Cells ◽  
Term Survival ◽  
Prevention Of Infection ◽  
Parasite Relationship ◽  
Potential Applications

Neospora caninumis an apicomplexan parasite that is closely related toToxoplasma gondii, the causative agent of toxoplasmosis in humans and domestic animals. However, in contrast toT. gondii, N. caninumrepresents a major cause of abortion in cattle, pointing towards distinct differences in the biology of these two species. There are 3 distinct key features that represent potential targets for prevention of infection or intervention against disease caused byN. caninum. Firstly, tachyzoites are capable of infecting a large variety of host cellsin vitroandin vivo. Secondly, the parasite exploits its ability to respond to alterations in living conditions by converting into another stage (tachyzoite-to-bradyzoite orvice versa). Thirdly, by analogy withT. gondii, this parasite has evolved mechanisms that modulate its host cells according to its own requirements, and these must, especially in the case of the bradyzoite stage, involve mechanisms that ensure long-term survival of not only the parasite but also of the host cell. In order to elucidate the molecular and cellular bases of these important features ofN. caninum, cell culture-based approaches and laboratory animal models are being exploited. In this review, we will summarize the current achievements related to host cell and parasite cell biology, and will discuss potential applications for prevention of infection and/or disease by reviewing corresponding work performed in murine laboratory infection models and in cattle.

scholarly journals 1,3,4-Thiadiazoles Effectively Inhibit Proliferation of Toxoplasma gondii

Cells ◽  
2021 ◽  
Vol 10 (5) ◽  
pp. 1053
Author(s):  
Lidia Węglińska ◽  
Adrian Bekier ◽  
Katarzyna Dzitko ◽  
Barbara Pacholczyk-Sienicka ◽  
Łukasz Albrecht ◽  
...  
Keyword(s):  
Toxoplasma Gondii ◽  
Host Cell ◽  
Cell Invasion ◽  
Direct Action ◽  
Human Fibroblasts ◽  
Imidazole Ring ◽  
Host Cells ◽  
In Vitro Assays ◽  
Host Cell Invasion

Congenital and acquired toxoplasmosis caused by the food- and water-born parasite Toxoplasma gondii (T. gondii) is one of the most prevalent zoonotic infection of global importance. T. gondii is an obligate intracellular parasite with limited capacity for extracellular survival, thus a successful, efficient and robust host cell invasion process is crucial for its survival, proliferation and transmission. In this study, we screened a series of novel 1,3,4-thiadiazole-2-halophenylamines functionalized at the C5 position with the imidazole ring (1b–12b) for their effects on T. gondii host cell invasion and proliferation. To achieve this goal, these compounds were initially subjected to in vitro assays to assess their cytotoxicity on human fibroblasts and then antiparasitic efficacy. Results showed that all of them compare favorably to control drugs sulfadiazine and trimethoprim in terms of T. gondii growth inhibition (IC50) and selectivity toward the parasite, expressed as selectivity index (SI). Subsequently, the most potent of them with meta-fluoro 2b, meta-chloro 5b, meta-bromo 8b, meta-iodo 11b and para-iodo 12b substitution were tested for their efficacy in inhibition of tachyzoites invasion and subsequent proliferation by direct action on established intracellular infection. All the compounds significantly inhibited the parasite invasion and intracellular proliferation via direct action on both tachyzoites and parasitophorous vacuoles formation. The most effective was para-iodo derivative 12b that caused reduction in the percentage of infected host cells by 44% and number of tachyzoites per vacuole by 93% compared to non-treated host cells. Collectively, these studies indicate that 1,3,4-thiadiazoles 1b–12b, especially 12b with IC50 of 4.70 µg/mL and SI of 20.89, could be considered as early hit compounds for future design and synthesis of anti-Toxoplasma agents that effectively and selectively block the invasion and subsequent proliferation of T. gondii into host cells.

scholarly journals BCG Substrains Change Their Outermost Surface as a Function of Growth Media

Vaccines ◽  
2021 ◽  
Vol 10 (1) ◽  
pp. 40
Author(s):  
Sandra Guallar-Garrido ◽  
Farners Almiñana-Rapún ◽  
Víctor Campo-Pérez ◽  
Eduard Torrents ◽  
Marina Luquin ◽  
...  
Keyword(s):  
Culture Media ◽  
Chemical Properties ◽  
Growth Conditions ◽  
Neutral Red ◽  
Culture Conditions ◽  
Host Cells ◽  
Growth Media ◽  
Phenotypic Characteristics ◽  
Outermost Surface ◽  
Red Dye

Mycobacterium bovis bacillus Calmette-Guérin (BCG) efficacy as an immunotherapy tool can be influenced by the genetic background or immune status of the treated population and by the BCG substrain used. BCG comprises several substrains with genetic differences that elicit diverse phenotypic characteristics. Moreover, modifications of phenotypic characteristics can be influenced by culture conditions. However, several culture media formulations are used worldwide to produce BCG. To elucidate the influence of growth conditions on BCG characteristics, five different substrains were grown on two culture media, and the lipidic profile and physico-chemical properties were evaluated. Our results show that each BCG substrain displays a variety of lipidic profiles on the outermost surface depending on the growth conditions. These modifications lead to a breadth of hydrophobicity patterns and a different ability to reduce neutral red dye within the same BCG substrain, suggesting the influence of BCG growth conditions on the interaction between BCG cells and host cells.

scholarly journals Immune activation of the host cell induces drug tolerance in Mycobacterium tuberculosis both in vitro and in vivo

2016 ◽  
Vol 213 (5) ◽  
pp. 809-825 ◽  
Author(s):  
Yancheng Liu ◽  
Shumin Tan ◽  
Lu Huang ◽  
Robert B. Abramovitch ◽  
Kyle H. Rohde ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Host Cell ◽  
Stress Responses ◽  
Drug Action ◽  
Host Cells ◽  
Transcriptional Analysis ◽  
Drug Pressure ◽  
Cytokine Activation ◽  
The Impact

Successful chemotherapy against Mycobacterium tuberculosis (Mtb) must eradicate the bacterium within the context of its host cell. However, our understanding of the impact of this environment on antimycobacterial drug action remains incomplete. Intriguingly, we find that Mtb in myeloid cells isolated from the lungs of experimentally infected mice exhibit tolerance to both isoniazid and rifampin to a degree proportional to the activation status of the host cells. These data are confirmed by in vitro infections of resting versus activated macrophages where cytokine-mediated activation renders Mtb tolerant to four frontline drugs. Transcriptional analysis of intracellular Mtb exposed to drugs identified a set of genes common to all four drugs. The data imply a causal linkage between a loss of fitness caused by drug action and Mtb’s sensitivity to host-derived stresses. Interestingly, the environmental context exerts a more dominant impact on Mtb gene expression than the pressure on the drugs’ primary targets. Mtb’s stress responses to drugs resemble those mobilized after cytokine activation of the host cell. Although host-derived stresses are antimicrobial in nature, they negatively affect drug efficacy. Together, our findings demonstrate that the macrophage environment dominates Mtb’s response to drug pressure and suggest novel routes for future drug discovery programs.

scholarly journals Contrasting Function of Structured N-Terminal and Unstructured C-Terminal Segments of Mycobacterium tuberculosis PPE37 Protein

mBio ◽  
2018 ◽  
Vol 9 (1) ◽  
Author(s):  
Javeed Ahmad ◽  
Aisha Farhana ◽  
Rita Pancsa ◽  
Simran Kaur Arora ◽  
Alagiri Srinivasan ◽  
...  
Keyword(s):  
Dendritic Cells ◽  
Mycobacterium Tuberculosis ◽  
Host Cell ◽  
Terminal Segment ◽  
Host Protein ◽  
Host Cells ◽  
Productive Infection ◽  
Disordered Proteins ◽  
Intrinsically Disordered

ABSTRACT Pathogens frequently employ eukaryotic linear motif (ELM)-rich intrinsically disordered proteins (IDPs) to perturb and hijack host cell networks for a productive infection. Mycobacterium tuberculosis has a relatively high percentage of IDPs in its proteome, the significance of which is not known. The Mycobacterium-specific PE-PPE protein family has several members with unusually high levels of structural disorder and disorder-promoting Ala/Gly residues. PPE37 protein, a member of this family, carries an N-terminal PPE domain capable of iron binding, two transmembrane domains, and a disordered C-terminal segment harboring ELMs and a eukaryotic nuclear localization signal (NLS). PPE37, expressed as a function of low iron stress, was cleaved by M. tuberculosis protease into N- and C-terminal segments. A recombinant N-terminal segment (P37N) caused proliferation and differentiation of monocytic THP-1 cells, into CD11c, DC-SIGN (dendritic cell-specific intercellular adhesion molecule-3-grabbing nonintegrin)-positive semimature dendritic cells exhibiting high interleukin-10 (IL-10) but negligible IL-12 and also low tumor necrosis factor alpha (TNF-α) secretion—an environment suitable for maintaining tolerogenic immune cells. The C-terminal segment entered the macrophage nucleus and induced caspase-3-dependent apoptosis of host cells. Mice immunized with recombinant PPE37FL and PPE37N evoked strong anti-inflammatory response, validating the in vitro immunostimulatory effect. Analysis of the IgG response of PPE37FL and PPE37N revealed significant immunoreactivities in different categories of TB patients, viz. pulmonary TB (PTB) and extrapulmonary TB (EPTB), vis-a-vis healthy controls. These results support the role of IDPs in performing contrasting activities to modulate the host processes, possibly through molecular mimicry and cross talk in two spatially distinct host environments which may likely aid M. tuberculosis survival and pathogenesis. IMPORTANCE To hijack the human host cell machinery to enable survival inside macrophages, the pathogen Mycobacterium tuberculosis requires a repertoire of proteins that can mimic host protein function and modulate host cell machinery. Here, we have shown how a single protein can play multiple functions and hijack the host cell for the benefit of the pathogen. Full-length membrane-anchored PPE37 protein is cleaved into N- and C-terminal domains under iron-depleted conditions. The N-terminal domain facilitates the propathogen semimature tolerogenic state of dendritic cells, whereas the C-terminal segment is localized into host cell nucleus and induces apoptosis. The immune implications of these in vitro observations were assessed and validated in mice and also human TB patients. This study presents novel mechanistic insight adopted by M. tuberculosis to survive inside host cells.

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