Listeriosis, a model infection to study host-pathogen interactions in vivo

2022 ◽  
Vol 66 ◽  
pp. 11-20
Author(s):  
Hana Kammoun ◽  
Minhee Kim ◽  
Lukas Hafner ◽  
Julien Gaillard ◽  
Olivier Disson ◽  
...  
Keyword(s):  
Host Pathogen Interactions ◽  
Host Pathogen

scholarly journals Porcine small intestinal organoids as a model to explore ETEC–host interactions in the gut

2021 ◽  
Vol 52 (1) ◽  
Author(s):  
Bjarne Vermeire ◽  
Liara M. Gonzalez ◽  
Robert J. J. Jansens ◽  
Eric Cox ◽  
Bert Devriendt
Keyword(s):  
Intestinal Epithelial ◽  
Gut Health ◽  
Functional Responses ◽  
Host Pathogen Interactions ◽  
Epithelial Surface ◽  
Host Interactions ◽  
Intestinal Organoids ◽  
Host Pathogen ◽  
Small Intestinal

AbstractSmall intestinal organoids, or enteroids, represent a valuable model to study host–pathogen interactions at the intestinal epithelial surface. Much research has been done on murine and human enteroids, however only a handful studies evaluated the development of enteroids in other species. Porcine enteroid cultures have been described, but little is known about their functional responses to specific pathogens or their associated virulence factors. Here, we report that porcine enteroids respond in a similar manner as in vivo gut tissues to enterotoxins derived from enterotoxigenic Escherichia coli, an enteric pathogen causing postweaning diarrhoea in piglets. Upon enterotoxin stimulation, these enteroids not only display a dysregulated electrolyte and water balance as shown by their swelling, but also secrete inflammation markers. Porcine enteroids grown as a 2D-monolayer supported the adhesion of an F4+ ETEC strain. Hence, these enteroids closely mimic in vivo intestinal epithelial responses to gut pathogens and are a promising model to study host–pathogen interactions in the pig gut. Insights obtained with this model might accelerate the design of veterinary therapeutics aimed at improving gut health.

scholarly journals A bilayer tissue culture model of the bovine alveolus

F1000Research ◽  
2019 ◽  
Vol 8 ◽  
pp. 357
Author(s):  
Diane Lee ◽  
Mark Chambers
Keyword(s):  
Epithelial Cells ◽  
Alveolar Type ◽  
Type Ii Cells ◽  
Type Ii ◽  
Host Pathogen Interactions ◽  
Alveolar Type Ii Cells ◽  
Permeable Membrane ◽  
Host Pathogen

The epithelial lining of the lung is often the first point of interaction between the host and inhaled pathogens, allergens and medications. Epithelial cells are therefore the main focus of studies which aim to shed light on host-pathogen interactions, to dissect the mechanisms of local host immunity and study toxicology. If these studies are not to be conducted exclusively in vivo, it is imperative that in vitro models are developed with a high in vitro-in vivo correlation. We describe here a co-culture bilayer model of the bovine alveolus, designed to overcome some of the limitations encountered with mono-culture and live animal models. Our system includes bovine pulmonary arterial endothelial cells (BPAECs) seeded onto a permeable membrane in 24 well Transwell format. The BPAECs are overlaid with immortalised bovine alveolar type II epithelial cells and the bilayer cultured at air-liquid interface for 14 days before use; in our case to study host-mycobacterial interactions. Characterisation of novel cell lines and the bilayer model have provided compelling evidence that immortalised bovine alveolar type II cells are an authentic substitute for primary alveolar type II cells and their culture as a bilayer in conjunction with BPAECs provides a physiologically relevant in vitro model of the bovine alveolus.   The bilayer model may be used to study dynamic intracellular and extracellular host-pathogen interactions, using proteomics, genomics, live cell imaging, in-cell ELISA and confocal microscopy. The model presented in this article enables other researchers to establish an in vitro model of the bovine alveolus that is easy to set up, malleable and serves as a comparable alternative to in vivo models, whilst allowing study of early host-pathogen interactions, currently not feasible in vivo. The model therefore achieves one of the 3Rs objectives in that it replaces the use of animals in research of bovine respiratory diseases.

scholarly journals Proximity Labeling to Map Host-Pathogen Interactions at the Membrane of a Bacteria Containing Vacuole inChlamydia trachomatisInfected Human Cells

2019 ◽  
Author(s):  
Macy G. Olson ◽  
Ray E. Widner ◽  
Lisa M. Jorgenson ◽  
Alyssa Lawrence ◽  
Dragana Lagundzin ◽  
...  
Keyword(s):  
Protein Interactions ◽  
Protein Protein Interactions ◽  
Host Pathogen Interactions ◽  
Inclusion Membrane ◽  
Eukaryotic Proteins ◽  
Host Pathogen ◽  
Membrane Bound ◽  
Labeling System ◽  
Chlamydial Inclusion

AbstractAs an obligate intracellular pathogenic bacterium,C. trachomatisdevelops within a membrane-bound vacuole, termed the inclusion. The inclusion membrane is modified by chlamydial inclusion membrane proteins (Incs), which act as the mediators of host-pathogen interactions. Anin vivounderstanding of Inc-Inc and Inc-eukaryotic protein interactions and how these contribute to overall host-chlamydial interactions at this unique membrane is lacking. Previous bacterial two-hybrid studies established that certain Incs have the propensity to bind other Incs while others have limited Inc-Inc interactions. We hypothesize some Incs organize the inclusion membrane whereas other Incs bind eukaryotic proteins to promote chlamydial-host interactions. To test this hypothesis, we used the ascorbate peroxidase proximity labeling system (APEX2), which labels proximal proteins with biotinin vivo, and chose to analyze Inc proteins with varying Inc-binding propensities. We inducibly expressed these Incs fused to APEX2 inChlamydia trachomatisL2, verified their localization and labeling activities by transmission electron microscopy, and used affinity purification-mass spectrometry to identify biotinylated proteins. To analyze our mass spectrometry results for statistical significance, we used Significance Analysis of INTeractome (SAINT), which demonstrated that our Inc-APEX2 constructs labeled Inc proteins as well as known and previously unreported eukaryotic proteins that localize to the inclusion. Our results broadly support two types of Inc interactions: Inc-Inc versus Inc-host. One eukaryotic protein, LRRFIP1 (LRRF1) was found in all of our Inc-APEX2 datasets, which is consistent with previously published AP-MS datasets. For the first time, we demonstrate by confocal and super-resolution microscopy that endogenous LRRF1 localizes to the chlamydial inclusion. We also used bacterial two-hybrid studies and pulldown assays to determine if LRRF1 was identified as a true interacting protein or was proximal to our Inc-APEX2 constructs. Combined, our data highlight the utility of APEX2 to capture the complexin vivoprotein-protein interactions at the chlamydial inclusion.Author summaryMany intracellular bacteria, including the obligate intracellular pathogenChlamydia trachomatis, grow within a membrane-bound “bacteria containing vacuole” (BCV) that, in most cases, prevents association with the lysosome. Secreted cytosolic effectors modulate host activity, but an understanding of the host-pathogen interactions that occur at the BCV membrane is limited by the difficulty in purifying membrane fractions from infected host cells. Here, we used the ascorbate peroxidase proximity labeling system (APEX2), which labels proximal proteins with biotinin vivo, to study the interactions that occur at the chlamydial vacuolar, or inclusion, membrane. The inclusion membrane is modified by chlamydial type III secreted inclusion membrane proteins (Incs), which act as the mediators of host-pathogen interactions. Our results broadly support two types of Inc interactions: Inc-Inc versus Inc-host. Our data highlight the utility of APEX2 to capture the complex protein-protein interactions at a membrane sitein vivoin the context of infection.

scholarly journals Dual RNA-Seq of Mtb-Infected Macrophages In Vivo Reveals Ontologically Distinct Host-Pathogen Interactions

Cell Reports ◽  
2020 ◽  
Vol 30 (2) ◽  
pp. 335-350.e4 ◽  
Author(s):  
Davide Pisu ◽  
Lu Huang ◽  
Jennifer K. Grenier ◽  
David G. Russell
Keyword(s):  
Rna Seq ◽  
Host Pathogen Interactions ◽  
Host Pathogen

scholarly journals Illuminating Macrophage Contributions to Host-Pathogen Interactions In Vivo: the Power of Zebrafish

2020 ◽  
Vol 88 (7) ◽  
Author(s):  
Emily E. Rosowski
Keyword(s):  
Cell Culture ◽  
Innate Immunity ◽  
Imaging Techniques ◽  
Innate Immune ◽  
Host Pathogen Interactions ◽  
Culture Studies ◽  
Larval Zebrafish ◽  
Host Pathogen

ABSTRACT Macrophages are a key cell type in innate immunity. Years of in vitro cell culture studies have unraveled myriad macrophage pathways that combat pathogens and demonstrated how pathogen effectors subvert these mechanisms. However, in vitro cell culture studies may not accurately reflect how macrophages fit into the context of an innate immune response in whole animals with multiple cell types and tissues. Larval zebrafish have emerged as an intermediate model of innate immunity and host-pathogen interactions to bridge the gap between cell culture studies and mammalian models. These organisms possess an innate immune system largely conserved with that of humans and allow state-of-the-art genetic and imaging techniques, all in the context of an intact organism. Using larval zebrafish, researchers are elucidating the function of macrophages in response to many different infections, including both bacterial and fungal pathogens. The goal of this review is to highlight studies in zebrafish that utilized live-imaging techniques to analyze macrophage activities in response to pathogens. Recent studies have explored the roles of specific pathways and mechanisms in macrophage killing ability, explored how pathogens subvert these responses, identified subsets of macrophages with differential microbicidal activities, and implicated macrophages as an intracellular niche for pathogen survival and trafficking. Research using this model continues to advance our understanding of how macrophages, and specific pathways inside these cells, fit into complex multicellular innate immune responses in vivo, providing important information on how pathogens evade these pathways and how we can exploit them for development of treatments against microbial infections.

scholarly journals mSphere of Influence: Clearing a Path for High-Resolution Visualization of Host-Pathogen Interactions In Vivo

mSphere ◽  
2019 ◽  
Vol 4 (4) ◽  
Author(s):  
Shumin Tan
Keyword(s):  
Single Cell ◽  
Dimensional Space ◽  
Whole Body ◽  
Host Pathogen Interactions ◽  
Content Type ◽  
3 Dimensional ◽  
Host Interactions ◽  
Host Pathogen ◽  
Tissue Optical Clearing

ABSTRACT Shumin Tan works in the field of Mycobacterium tuberculosis-host interactions. In this mSphere of Influence article, she reflects on how the paper “Single-cell phenotyping within transparent intact tissue through whole-body clearing” by B. Yang et al. (Cell 158:945–958, 2014, https://doi.org/10.1016/j.cell.2014.07.017) impacted her ideas on approaches to visualize and understand heterogeneous host-pathogen interactions in vivo in 3-dimensional space at the single-cell level, through the tractable and broadly compatible tissue optical clearing methods developed.

scholarly journals Dynamics ofMycobacterium tuberculosisAg85B revealed by sensitive ELISA

2019 ◽  
Author(s):  
Joel D. Ernst ◽  
Amber Cornelius ◽  
Miriam Bolz
Keyword(s):  
Protein Secretion ◽  
Bacterial Infections ◽  
Bacterial Population ◽  
Bacterial Protein ◽  
Host Pathogen Interactions ◽  
Gram Positive ◽  
Gram Negative ◽  
Bacterial Proteins ◽  
Host Pathogen

AbstractSecretion of specific proteins contributes to pathogenesis and immune responses in tuberculosis and other bacterial infections, yet the kinetics of protein secretion and fate of secreted proteins in vivo are poorly understood. We generated new monoclonal antibodies that recognize theM. tuberculosissecreted protein, Ag85B, and used them to establish and characterize a sensitive ELISA to quantitate Ag85B in samples generated in vitro and in vivo. We found that nutritional or culture conditions had little impact on secretion of Ag85B, and that there is considerable variation in Ag85B secretion by distinct strains in theM. tuberculosiscomplex: compared with the commonly-used H37Rv strain (Lineage 4),M. africanum(Lineage 6) secretes less, and two strains from Lineage 2 secrete more Ag85B. We also used the ELISA to determine that the rate of secretion of Ag85B is 10-to 100-fold lower than that of proteins secreted by gram-negative and gram-positive bacteria, respectively. ELISA quantitation of Ag85B in lung homogenates ofM. tuberculosisH37Rv-infected mice revealed that although Ag85B accumulates in the lungs as the bacterial population expands, the amount of Ag85B per bacterium decreases nearly 10,000-fold at later stages of infection, coincident with development of T cell responses and arrest of bacterial population growth. These results indicate that bacterial protein secretion in vivo is dynamic and regulated, and quantitation of secreted bacterial proteins can contribute to understanding pathogenesis and immunity in tuberculosis and other infections.ImportanceBacterial protein secretion contributes to host-pathogen interactions, yet the process and consequences of bacterial protein secretion during infection are poorly understood. We developed a sensitive ELISA to quantitate a protein (termed Ag85B) secreted byM. tuberculosisand used it to find that Ag85B secretion occurs with slower kinetics than for proteins secreted by gram positive and gram negative bacteria, and that accumulation of Ag85B in the lungs is markedly regulated as a function of the bacterial population density. Our results demonstrate that quantitation of bacterial proteins during infection can reveal novel insights into host-pathogen interactions.

scholarly journals Host Airway Proteins Interact with Staphylococcus aureus during Early Pneumonia

2008 ◽  
Vol 76 (3) ◽  
pp. 888-898 ◽  
Author(s):  
Christy L. Ventura ◽  
Roger Higdon ◽  
Eugene Kolker ◽  
Shawn J. Skerrett ◽  
Craig E. Rubens
Keyword(s):  
Staphylococcus Aureus ◽  
Shotgun Proteomics ◽  
Community Acquired Pneumonia ◽  
Host Proteins ◽  
Host Pathogen Interactions ◽  
Complement Components ◽  
Host Pathogen ◽  
Hospital Acquired

ABSTRACT Staphylococcus aureus is a major cause of hospital-acquired pneumonia and is emerging as an important etiological agent of community-acquired pneumonia. Little is known about the specific host-pathogen interactions that occur when S. aureus first enters the airway. A shotgun proteomics approach was utilized to identify the airway proteins associated with S. aureus during the first 6 h of infection. Host proteins eluted from bacteria recovered from the airways of mice 30 min or 6 h following intranasal inoculation under anesthesia were subjected to liquid chromatography and tandem mass spectrometry. A total of 513 host proteins were associated with S. aureus 30 min and/or 6 h postinoculation. A majority of the identified proteins were host cytosolic proteins, suggesting that S. aureus was rapidly internalized by phagocytes in the airway and that significant host cell lysis occurred during early infection. In addition, extracellular matrix and secreted proteins, including fibronectin, antimicrobial peptides, and complement components, were associated with S. aureus at both time points. The interaction of 12 host proteins shown to bind to S. aureus in vitro was demonstrated in vivo for the first time. The association of hemoglobin, which is thought to be the primary staphylococcal iron source during infection, with S. aureus in the airway was validated by immunoblotting. Thus, we used our recently developed S. aureus pneumonia model and shotgun proteomics to validate previous in vitro findings and to identify nearly 500 other proteins that interact with S. aureus in vivo. The data presented here provide novel insights into the host-pathogen interactions that occur when S. aureus enters the airway.

scholarly journals Activity-Based Protein Profiling Identifies ATG4B as a Key Host Factor for Enterovirus 71 Proliferation

2019 ◽  
Vol 93 (24) ◽  
Author(s):  
Yang Sun ◽  
Qizhen Zheng ◽  
Yaxin Wang ◽  
Zhengyuan Pang ◽  
Jingwei Liu ◽  
...  
Keyword(s):  
Life Cycle ◽  
Cysteine Protease ◽  
Enterovirus 71 ◽  
Foot And Mouth Disease ◽  
Protein Profiling ◽  
Human Hand ◽  
Host Pathogen Interactions ◽  
Host Pathogen ◽  
Viral Proliferation

ABSTRACT Virus-encoded proteases play diverse roles in the efficient replication of enterovirus 71 (EV71), which is the causative agent of human hand, foot, and mouth disease (HFMD). However, it is unclear how host proteases affect viral proliferation. Here, we designed activity-based probes (ABPs) based on an inhibitor of the main EV71 protease (3Cpro), which is responsible for the hydrolysis of the EV71 polyprotein, and successfully identified host candidates that bind to the ABPs. Among the candidates, the host cysteine protease autophagy-related protein 4 homolog B (ATG4B), a key component of the autophagy machinery, was demonstrated to hydrolytically process the substrate of EV71 3Cpro and had activity comparable to that of the viral protease. Genetic disruption of ATG4B confirmed that the enzyme is indispensable for viral proliferation in vivo. Our results not only further the understanding of host-virus interactions in EV71 biology but also provide a sample for the usage of activity-based proteomics to reveal host-pathogen interactions. IMPORTANCE Enterovirus 71 (EV71), one of the major pathogens of human HFMD, has caused outbreaks worldwide. How EV71 efficiently assesses its life cycle with elaborate interactions with multiple host factors remains to be elucidated. In this work, we deconvoluted that the host ATG4B protein processes the viral polyprotein with its cysteine protease activity and helps EV71 replicate through a chemical biology strategy. Our results not only further the understanding of the EV71 life cycle but also provide a sample for the usage of activity-based proteomics to reveal host-pathogen interactions.

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