scholarly journals Non-Invasive Luciferase Imaging of Type I Interferon Induction in a Transgenic Mouse Model of Biomaterial Associated Bacterial Infections: Microbial Specificity and Inter-Bacterial Species Interactions

2020 ◽  
Vol 8 (10) ◽  
pp. 1624
Author(s):  
Muhammad Imran Rahim ◽  
Andreas Winkel ◽  
Stefan Lienenklaus ◽  
Nico S. Stumpp ◽  
Szymon P. Szafrański ◽  
...  
Keyword(s):  
Mouse Model ◽  
Species Interactions ◽  
Bacterial Infections ◽  
Bacterial Species ◽  
Inflammatory Cells ◽  
Single Species ◽  
Oral Bacteria ◽  
Type I ◽  
Non Invasive

The performance of biomaterials is often compromised by bacterial infections and subsequent inflammation. So far, the conventional analysis of inflammatory processes in vivo involves time-consuming histology and biochemical assays. The present study employed a mouse model where interferon beta (IFN-β) is monitored as a marker for non-invasive rapid detection of inflammation in implant-related infections. The mouse model comprises subcutaneous implantation of morphologically modified titanium, followed by experimental infections with four taxonomically diverse oral bacteria: Streptococcus oralis, Aggregatibacter actinomycetemcomitans, Porphyromonas gingivalis and Treponema denticola (as mono culture or selected mixed-culture). IFN-β expression increased upon infections depending on the type of pathogen and was prolonged by the presence of the implant. IFN-β expression kinetics reduced with two mixed species infections when compared with the single species. Histological and confocal microscopy confirmed pathogen-specific infiltration of inflammatory cells at the implant-tissue interface. This was observed mainly in the vicinity of infected implants and was, in contrast to interferon expression, higher in infections with dual species. In summary, this non-invasive mouse model can be used to quantify longitudinally host inflammation in real time and suggests that the polymicrobial character of infection, highly relevant to clinical situations, has complex effects on host immunity.

A novel mouse model based on intersectional genetics enables unambiguous in vivo discrimination between plasmacytoid and other dendritic cells and their comparative characterization

2022 ◽  
Author(s):  
Michael Valente ◽  
Nils Collinet ◽  
Thien-Phong Vu Manh ◽  
Karima Naciri ◽  
Gilles Bessou ◽  
...  
Keyword(s):  
Dendritic Cells ◽  
Steady State ◽  
Mouse Model ◽  
Single Cell ◽  
Ex Vivo ◽  
High Specificity ◽  
Type I ◽  
Specific Expression ◽  
Physiological Functions

Plasmacytoid dendritic cells (pDC) were identified about 20 years ago, based on their unique ability to rapidly produce copious amounts of all subsets of type I and type III interferon (IFN-I/III) upon virus sensing, while being refractory to infection. Yet, the identity and physiological functions of pDC are still a matter of debate, in a large part due to their lack of specific expression of any single cell surface marker or gene that would allow to track them in tissues and to target them in vivo with high specificity and penetrance. Indeed, recent studies showed that previous methods that were used to identify or deplete pDC also targeted other cell types, including pDC-like cells and transitional DC (tDC) that were proposed to be responsible for all the antigen presentation ability previously attributed to steady state pDC. Hence, improving our understanding of the nature and in vivo choreography of pDC physiological functions requires the development of novel tools to unambiguously identify and track these cells, including in comparison to pDC-like cells and tDC. Here, we report successful generation of a pDC-reporter mouse model, by using an intersectional genetic strategy based on the unique co-expression of Siglech and Pacsin1 in pDC. This pDC-Tomato mouse strain allows specific ex vivo and in situ detection of pDC. Breeding them with Zbtb46GFP mice allowed side-by-side purification and transcriptional profiling by single cell RNA sequencing of bona fide pDC, pDC-like cells and tDC, in comparison to type 1 and 2 conventional DC (cDC1 and cDC2), both at steady state and during a viral infection, revealing diverging activation patterns of pDC-like cells and tDC. Finally, by breeding pDC-Tomato mice with Ifnb1EYFP mice, we determined the choreography of pDC recruitment to the micro-anatomical sites of viral replication in the spleen, with initially similar but later divergent behaviors of the pDC that engaged or not into IFN-I production. Our novel pDC-Tomato mouse model, and newly identified gene modules specific to combinations of DC types and activations states, will constitute valuable resources for a deeper understanding of the functional division of labor between DC types and its molecular regulation at homeostasis and during viral infections.

scholarly journals Common adaptive strategies underlie within-host evolution of bacterial pathogens

2020 ◽  
Author(s):  
Yair E Gatt ◽  
Hanah Margalit
Keyword(s):  
Bacterial Infections ◽  
Large Scale ◽  
Bacterial Species ◽  
Adaptive Strategies ◽  
Single Species ◽  
Adaptive Strategy ◽  
Host Immune System ◽  
Glycerol Phosphate ◽  
Genomic Changes ◽  
Virulence Attenuation

Abstract Within-host adaptation is a hallmark of chronic bacterial infections, involving substantial genomic changes. Recent large-scale genomic data from prolonged infections allow the examination of adaptive strategies employed by different pathogens and open the door to investigate whether they converge towards similar strategies. Here, we compiled extensive data of whole-genome sequences of bacterial isolates belonging to miscellaneous species sampled at sequential time points during clinical infections. Analysis of these data revealed that different species share some common adaptive strategies, achieved by mutating various genes. While the same genes were often mutated in several strains within a species, different genes related to the same pathway, structure or function were changed in other species utilizing the same adaptive strategy (e.g. mutating flagellar genes). Strategies exploited by various bacterial species were often predicted to be driven by the host immune system, a powerful selective pressure that is not species-specific. Remarkably, we find adaptive strategies identified previously within single species to be ubiquitous. Two striking examples are shifts from siderophore-based to heme-based iron scavenging (previously shown for Pseudomonas aeruginosa), and changes in glycerol-phosphate metabolism (previously shown to decrease sensitivity to antibiotics in Mycobacterium tuberculosis). Virulence factors were often adaptively affected in different species, indicating shifts from acute to chronic virulence and virulence attenuation during infection. Our study presents a global view on common within-host adaptive strategies employed by different bacterial species and provides a rich resource for further studying these processes.

Enhanced in vivo efficacy of a long-life type I Interferon superagonist in a mouse model of multiple sclerosis

2014 ◽  
Vol 275 (1-2) ◽  
pp. 219
Author(s):  
Daniel Harari ◽  
Nadine Kallweit ◽  
Renne Abramovich ◽  
Keren Sasson ◽  
Alla Zozulya ◽  
...  
Keyword(s):  
Multiple Sclerosis ◽  
Mouse Model ◽  
Type I Interferon ◽  
Type I ◽  
In Vivo Efficacy ◽  
Long Life ◽  
Life Type

scholarly journals New Approaches for Isolation of Previously Uncultivated Oral Bacteria

2011 ◽  
Vol 78 (1) ◽  
pp. 194-203 ◽  
Author(s):  
M. V. Sizova ◽  
T. Hohmann ◽  
A. Hazen ◽  
B. J. Paster ◽  
S. R. Halem ◽  
...  
Keyword(s):  
Oral Cavity ◽  
Single Cell ◽  
Single Species ◽  
Oral Bacteria ◽  
Cell Cultivation ◽  
Human Oral Cavity ◽  
Successful Recovery ◽  
Number Of Microorganisms ◽  
Enrichment Techniques

ABSTRACTA significant number of microorganisms from the human oral cavity remain uncultivated. This is a major impediment to the study of human health since some of the uncultivated species may be involved in a variety of systemic diseases. We used a range of innovations previously developed to cultivate microorganisms from the human oral cavity, focusing on anaerobic species. These innovations include (i)in vivocultivation to specifically enrich for species actively growing in the oral cavity (the “minitrap” method), (ii) single-cell long-term cultivation to minimize the effect of fast-growing microorganisms, and (iii) modifications of conventional enrichment techniques, using media that did not contain sugar, including glucose. To enable cultivation of obligate anaerobes, we maintained strict anaerobic conditions in most of our cultivation experiments. We report that, on a per cell basis, the most successful recovery was achieved using minitrap enrichment (11%), followed by single-cell cultivation (3%) and conventional plating (1%). Taxonomically, the richest collection was obtained using the single-cell cultivation method, followed by minitrap and conventional enrichment, comprising representatives of 13, 9, and 4 genera, respectively. Interestingly, no single species was isolated by all three methods, indicating method complementarity. An important result is the isolation and maintenance in pure culture of 10 strains previously only known by their molecular signatures, as well as representatives of what are likely to be three new microbial genera. We conclude that the ensemble of new methods we introduced will likely help close the gap between cultivated and uncultivated species from the human oral cavity.

scholarly journals Molecular Basis for Lytic Bacteriophage Resistance in Enterococci

mBio ◽  
2016 ◽  
Vol 7 (4) ◽  
Author(s):  
Breck A. Duerkop ◽  
Wenwen Huo ◽  
Pooja Bhardwaj ◽  
Kelli L. Palmer ◽  
Lora V. Hooper
Keyword(s):  
Cell Wall ◽  
Mouse Model ◽  
Enterococcus Faecalis ◽  
Molecular Basis ◽  
Bacterial Infections ◽  
Hypervariable Region ◽  
Cell Wall Protein ◽  
Phage Resistance ◽  
Human Intestine

ABSTRACT The human intestine harbors diverse communities of bacteria and bacteriophages. Given the specificity of phages for their bacterial hosts, there is growing interest in using phage therapies to combat the rising incidence of multidrug-resistant bacterial infections. A significant barrier to such therapies is the rapid development of phage-resistant bacteria, highlighting the need to understand how bacteria acquire phage resistance in vivo . Here we identify novel lytic phages in municipal raw sewage that kill Enterococcus faecalis , a Gram-positive opportunistic pathogen that resides in the human intestine. We show that phage infection of E. faecalis requires a predicted integral membrane protein that we have named PIP EF (for phage infection protein from E. faecalis ). We find that PIP EF is conserved in E. faecalis and harbors a 160-amino-acid hypervariable region that determines phage tropism for distinct enterococcal strains. Finally, we use a gnotobiotic mouse model of in vivo phage predation to show that the sewage phages temporarily reduce E. faecalis colonization of the intestine but that E. faecalis acquires phage resistance through mutations in PIP EF . Our findings define the molecular basis for an evolutionary arms race between E. faecalis and the lytic phages that prey on them. They also suggest approaches for engineering E. faecalis phages that have altered host specificity and that can subvert phage resistance in the host bacteria. IMPORTANCE Bacteriophage therapy has received renewed attention as a potential solution to the rise in antibiotic-resistant bacterial infections. However, bacteria can acquire phage resistance, posing a major barrier to phage therapy. To overcome this problem, it is necessary to understand phage resistance mechanisms in bacteria. We have unraveled one such resistance mechanism in Enterococcus faecalis , a Gram-positive natural resident of the human intestine that has acquired antibiotic resistance and can cause opportunistic infections. We have identified a cell wall protein hypervariable region that specifies phage tropism in E. faecalis . Using a gnotobiotic mouse model of in vivo phage predation, we show that E. faecalis acquires phage resistance through mutations in this cell wall protein. Our findings define the molecular basis for lytic phage resistance in E. faecalis . They also suggest opportunities for engineering E. faecalis phages that circumvent the problem of bacterial phage resistance.

scholarly journals Phage Therapy as a Focused Management Strategy in Aquaculture

2021 ◽  
Vol 22 (19) ◽  
pp. 10436
Author(s):  
José Ramos-Vivas ◽  
Joshua Superio ◽  
Jorge Galindo-Villegas ◽  
Félix Acosta
Keyword(s):  
Bacterial Infections ◽  
Therapeutic Potential ◽  
Current Knowledge ◽  
Control Strategies ◽  
Bacterial Species ◽  
Lytic Enzymes ◽  
Wide Range ◽  
Cultured Fish

Therapeutic bacteriophages, commonly called as phages, are a promising potential alternative to antibiotics in the management of bacterial infections of a wide range of organisms including cultured fish. Their natural immunogenicity often induces the modulation of a variated collection of immune responses within several types of immunocytes while promoting specific mechanisms of bacterial clearance. However, to achieve standardized treatments at the practical level and avoid possible side effects in cultivated fish, several improvements in the understanding of their biology and the associated genomes are required. Interestingly, a particular feature with therapeutic potential among all phages is the production of lytic enzymes. The use of such enzymes against human and livestock pathogens has already provided in vitro and in vivo promissory results. So far, the best-understood phages utilized to fight against either Gram-negative or Gram-positive bacterial species in fish culture are mainly restricted to the Myoviridae and Podoviridae, and the Siphoviridae, respectively. However, the current functional use of phages against bacterial pathogens of cultured fish is still in its infancy. Based on the available data, in this review, we summarize the current knowledge about phage, identify gaps, and provide insights into the possible bacterial control strategies they might represent for managing aquaculture-related bacterial diseases.

scholarly journals Intrinsic Abnormalities of Cystic Fibrosis Airway Connective Tissue Revealed by an In Vitro 3D Stromal Model

Cells ◽  
2020 ◽  
Vol 9 (6) ◽  
pp. 1371
Author(s):  
Claudia Mazio ◽  
Laura S. Scognamiglio ◽  
Rossella De Cegli ◽  
Luis J. V. Galietta ◽  
Diego Di Bernardo ◽  
...  
Keyword(s):  
Cystic Fibrosis ◽  
Inflammatory Response ◽  
Bacterial Infections ◽  
Inflammatory Cells ◽  
Lung Model ◽  
Lung Dysfunction ◽  
Novel Therapeutic Strategies ◽  
And Function

Cystic fibrosis is characterized by lung dysfunction involving mucus hypersecretion, bacterial infections, and inflammatory response. Inflammation triggers pro-fibrotic signals that compromise lung structure and function. At present, several in vitro cystic fibrosis models have been developed to study epithelial dysfunction but none of these focuses on stromal alterations. Here we show a new cystic fibrosis 3D stromal lung model made up of primary fibroblasts embedded in their own extracellular matrix and investigate its morphological and transcriptomic features. Cystic fibrosis fibroblasts showed a high proliferation rate and produced an abundant and chaotic matrix with increased protein content and elastic modulus. More interesting, they had enhanced pro-fibrotic markers and genes involved in epithelial function and inflammatory response. In conclusion, our study reveals that cystic fibrosis fibroblasts maintain in vitro an activated pro-fibrotic state. This abnormality may play in vivo a role in the modulation of epithelial and inflammatory cell behavior and lung remodeling. We argue that the proposed bioengineered model may provide new insights on epithelial/stromal/inflammatory cells crosstalk in cystic fibrosis, paving the way for novel therapeutic strategies.

Abstract 17280: A New in vivo Imaging Protocol for Non-Invasive Detection of Right Ventricular Dysfunction in a Pulmonary Hypertension Mouse Model

Circulation ◽  
2018 ◽  
Vol 138 (Suppl_1) ◽  
Author(s):  
Thomas Hansen ◽  
Kristen Bubb ◽  
Gemma Figtree
Keyword(s):  
Pulmonary Hypertension ◽  
Mouse Model ◽  
Right Ventricular ◽  
Systolic Pressure ◽  
Saline Control ◽  
Novel Therapies ◽  
Imaging Protocol ◽  
Non Invasive ◽  
Rv Function

Introduction: Accurate measurements of right ventricular (RV) function are critical for studying novel therapies impacting the heart and pulmonary circulation. Until now, assessment in mouse models has relied on invasive measures. Improvements in mouse echocardiography may facilitate application of measures recently validated in humans, including tricuspid annular plane systolic excursion (TAPSE) and RV-S’ (systolic excursion velocity), to allow non-invasive assessment of RV function. Aims: To apply and validate TAPSE and RV-S’ using high-resolution echocardiography for the measurement of RV function in a mouse model of pulmonary hypertension (PH). Methods: Echocardiography was performed on mice 3 weeks after induction of PH using inhaled bleomycin or saline control. PAT, TAPSE and RV-S’ were recorded in mice using a 55-mHz transducer (Visualsonics, Vevo3100). Invasive measurements of right ventricular systolic pressure (RVSP) were obtained via catheterisation of the internal jugular vein, prior to culling. Results: RVSP was significantly elevated in bleomycin-treated mice ( 33.41±0.8mmHg n=10) compared to controls ( 25.66±0.9mmHg n=11; p<0.0001). Similarly, RV hypertrophy was observed in bleomycin mice [RV:body weight 1.156±0.03g/kg n=11] compared with control ( 0.968±0.02g/kg n=12; p=0.0002). TAPSE was sensitive to these differences, being significantly reduced in bleomycin mice ( 0.5739±0.020mm n=8) compared with control ( 0.7387±0.033mm n=10; p=0.0012), and correlated significantly with invasive RVSP (r 2 =0.7218; p<0.0001). RV-S’ was also reduced in bleomycin mice (18.14±0.98mm/s n=7) compared with control (25.38±1.24mm/s n=8; p=0.0006) and correlated strongly with RVSP (r 2 =0.6378; p=0.0011). The correlation of both TAPSE and RV-S’ with RVSP compared favourably to the previously used surrogate measure of RVSP in mice, PAT (r 2 =0.5278; p=0.0002). Conclusions: TAPSE and RV-S’ can be applied in mouse echocardiography, and are sensitive, non-invasive measures of PH and RV dysfunction, comparing well with gold-standard invasive right ventricular systolic pressures. This may benefit the power of future preclinical studies of novel therapies in pulmonary hypertension and RV dysfunction.

Experimental induced wound cicatrisation after highly diluted products treatment

2021 ◽  
Vol 11 (40) ◽  
pp. 211-212
Author(s):  
Fernando Fortunato Jeronimo ◽  
Jenifer Pendiuk Gonçalves ◽  
Katia Fialho Do Nascimento ◽  
Simone Martins De Oliveira ◽  
Carolina Camargo De Oliveira ◽  
...  
Keyword(s):  
Connective Tissue ◽  
Inflammatory Cells ◽  
Control Group ◽  
Type I ◽  
Chelidonium Majus ◽  
Conium Maculatum ◽  
The Matrix ◽  
Aconitum Napellus

Introduction: Skin is an attractive target to study extracellular matrix, due to abundance in Connective tissue. In cases of injuries the first step is an inflammatory reaction and subsequent the healing that involves several changes in the matrix. These changes are fundamental to inflammatory cells activities allowing healing. Highly diluted products were shown to facilitate inflammatory mediators and to activate immune cells in vivo and in vitro, thus it can be effective to wound healing. Aims: This study aims to evaluate highly diluted products effects on inflammation and cicatrization process. Methodology: Three compounds (M8 (Aconitum napellus 20dH, Arsenicum album 18dH, Asa foetida 20dH, Calcarea carbonica 16dH, Conium maculatum 17dH, Ipecacuanha 13dH, Phosphorus 20dH, Rhus toxicodendron 17dH, Silicea 20dH, Sulphur 24dH, Thuja occidentalis 19dH), M1 (Chelidonium majus 20dH, Cinnamon 20dH, Echinaceae purpurea 20dH, Gelsemium sempervirens 20dH plus all M8 compounds) and Curcuma cH30 – simple product), were manipulated as a gel and applied on mice dorsal flank after incision and suture (approximately 1 cm and three points), for 3 consecutive days. After the treatments the scars were evaluated macroscopically, the animals were killed, the skin samples collected, fixed and processed for Hematoxilin-Eosin (HE) and Masson Tricromic (to observe the collagen fibers type I). The slices were analyzed and images collected by a light microscope Olympus BX51 with camera attached Olympus DP72. Results: It was observed a higher and faster rate of tissue epithelization in the treated groups after three days of gel-product application. This could be observed in lower rates in the control group (no treatment) - Figure 1 and 2). Regeneration and organization of connective tissue were proportional to epithelization the treated groups. We also observed evidences of changes in amount of neutrophils and fibroblasts, resulting in changes in the healing period. Analyses for these confirmations are in progress.

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