scholarly journals Mucosal Immunization with a Staphylococcus aureus IsdA-Cholera Toxin A2/B Chimera Induces Antigen-Specific Th2-Type Responses in Mice

2011 ◽  
Vol 18 (9) ◽  
pp. 1543-1551 ◽  
Author(s):  
Britni M. Arlian ◽  
Juliette K. Tinker
Keyword(s):  
Staphylococcus Aureus ◽  
Epithelial Cells ◽  
Cholera Toxin ◽  
Cellular Proliferation ◽  
Mucosal Vaccine ◽  
Enzyme Linked Immunosorbent Assay ◽  
Mucosal Immunization ◽  
Content Type ◽  
Human Nasal Epithelial Cells

ABSTRACTStaphylococcus aureusis a leading cause of opportunistic infection worldwide and a significant public health threat. The iron-regulated surface determinant A (IsdA) adhesin is essential forS. aureuscolonization on human nasal epithelial cells and plays an important role in iron acquisition and resistance to human skin defenses. Here we investigated the murine immune response to intranasal administration of a cholera toxin A2/B (CTA2/B) chimera containing IsdA. Plasmids were constructed to express the IsdA-CTA2/B chimera and control proteins inEscherichia coli. Proper construction of the chimera was verified by SDS-PAGE, Western blotting, GM1 enzyme-linked immunosorbent assay (ELISA), and confocal microscopy. Groups of female BALB/c mice were mock immunized or immunized with IsdA-CTA2/B, IsdA mixed with CTA2/B, or IsdA alone, followed by one booster immunization at 10 days postpriming. Analysis of serum IgG and nasal, intestinal, and vaginal IgA suggested that mucosal immunization with IsdA-CTA2/B induces significant IsdA-specific humoral immunity. Functionalin vitroassays revealed that immune serum significantly blocks the adherence ofS. aureusto human epithelial cells. Splenocytes from mice immunized with IsdA-CTA2/B showed specific cellular proliferation and production of interleukin-4 (IL-4) afterin vitrostimulation. Immunization with IsdA-CTA2/B drove isotype switching to IgG1, indicative of a Th2-type response. Our results suggest that the immunogenicity of theS. aureusIsdA-CTA2/B chimera merits further investigation as a potential mucosal vaccine candidate.

scholarly journals Effect of Coadministration of Vancomycin and BMP-2 on Cocultured Staphylococcus aureus and W-20-17 Mouse Bone Marrow Stromal CellsIn Vitro

2012 ◽  
Vol 56 (7) ◽  
pp. 3776-3784 ◽  
Author(s):  
A. H. Nguyen ◽  
S. Kim ◽  
W. J. Maloney ◽  
J. C. Wenke ◽  
Y. Yang
Keyword(s):  
Staphylococcus Aureus ◽  
Bone Marrow ◽  
Growth Factors ◽  
Bone Cell ◽  
Enzyme Linked Immunosorbent Assay ◽  
Mouse Bone Marrow ◽  
Content Type ◽  
Proliferation And Differentiation ◽  
Dual Administration

ABSTRACTIn this study, we aimed to establish anin vitrobacterium/bone cell coculture model system and to use this model for dose dependence studies of dual administration of antibiotics and growth factorsin vitro. We examined the effect of single or dual administration of the antibiotic vancomycin (VAN) at 0 to 16 μg/ml and bone morphogenetic protein-2 (BMP-2) at 0 or 100 ng/ml on both methicillin-sensitiveStaphylococcus aureusand mouse bone marrow stromal cells (W-20-17) under both mono- and coculture conditions. Cell metabolic activity, Live/Dead staining, double-stranded DNA (dsDNA) amounts, and alkaline phosphatase activity were measured to assess cell viability, proliferation, and differentiation. An interleukin-6 (IL-6) enzyme-linked immunosorbent assay (ELISA) kit was used to test the bone cell inflammation response in the presence of bacteria. Our results suggest that, when delivered together in coculture, VAN and BMP-2 maintain their primary functions as an antibiotic and a growth factor, respectively. Most interestingly, this dual-delivery type of approach has shown itself to be effective at lower concentrations of VAN than those required for an approach relying strictly on the antibiotic. It may be that BMP-2 enhances cell proliferation and differentiation before the cells become infected. In coculture, a dosage of VAN higher than that used for treatment in monoculture may be necessary to effectively inhibit growth ofStaphylococcus aureus. This could mean that the coculture environment may be limiting the efficacy of VAN, possibly by way of bacterial invasion of the bone cells. This report of a coculture study demonstrates a potential beneficial effect of the coadministration of antibiotics and growth factors compared to treatment with antibiotic alone.

scholarly journals Wall Teichoic Acid Glycosylation Governs Staphylococcus aureus Nasal Colonization

mBio ◽  
2015 ◽  
Vol 6 (4) ◽  
Author(s):  
Volker Winstel ◽  
Petra Kühner ◽  
Ferdinand Salomon ◽  
Jesper Larsen ◽  
Robert Skov ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Risk Factor ◽  
Epithelial Cells ◽  
Teichoic Acid ◽  
Nasal Colonization ◽  
Human Pathogen ◽  
Wall Teichoic Acid ◽  
Content Type ◽  
Human Nasal Epithelial Cells ◽  
Key Factor

ABSTRACT Nasal colonization by the human pathogen Staphylococcus aureus is a major risk factor for hospital- and community-acquired infections. A key factor required for nasal colonization is a cell surface-exposed zwitterionic glycopolymer, termed wall teichoic acid (WTA). However, the precise mechanisms that govern WTA-mediated nasal colonization have remained elusive. Here, we report that WTA GlcNAcylation is a pivotal requirement for WTA-dependent attachment of community-acquired methicillin-resistant S. aureus (MRSA) and emerging livestock-associated MRSA to human nasal epithelial cells, even under conditions simulating the nutrient composition and dynamic flow of nasal secretions. Depending on the S. aureus strain, WTA O-GlcNAcylation occurs in either α or β configuration, which have similar capacities to mediate attachment to human nasal epithelial cells, suggesting that many S. aureus strains maintain redundant pathways to ensure appropriate WTA glycosylation. Strikingly, a lack of WTA glycosylation significantly abrogated the ability of MRSA to colonize cotton rat nares in vivo. These results indicate that WTA glycosylation modulates S. aureus nasal colonization and may help to develop new strategies for eradicating S. aureus nasal colonization in the future. IMPORTANCE Nasal colonization by the major human pathogen Staphylococcus aureus is a risk factor for severe endogenous infections and contributes to the spread of this microbe in hospitals and the community. Here, we show that wall teichoic acid (WTA) O-GlcNAcylation is a key factor required for S. aureus nasal colonization. These data provide a mechanistic explanation for the capacity of WTA to modulate S. aureus nasal colonization and may stimulate research activities to establish valuable strategies to eradicate S. aureus nasal colonization in high-risk hospitalized patients and in the general community.

scholarly journals In VivoEfficacy of Ceftaroline Fosamil in a Methicillin-Resistant Panton-Valentine Leukocidin-Producing Staphylococcus aureus Rabbit Pneumonia Model

2014 ◽  
Vol 58 (4) ◽  
pp. 1855-1861 ◽  
Author(s):  
Delphine Croisier-Bertin ◽  
Davy Hayez ◽  
Sonia Da Silva ◽  
Delphine Labrousse ◽  
Donald Biek ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Enzyme Linked Immunosorbent Assay ◽  
Pulmonary Tissue ◽  
Ceftaroline Fosamil ◽  
Methicillin Resistant ◽  
Content Type ◽  
Panton Valentine Leukocidin ◽  
Valentine Leukocidin

ABSTRACTCeftaroline, the active metabolite of the prodrug ceftaroline fosamil, is a cephalosporin with broad-spectrumin vitroactivity against Gram-positive organisms, including methicillin-resistantStaphylococcus aureus(MRSA) and multidrug-resistantStreptococcus pneumoniae(MDRSP), and common Gram-negative pathogens. This study investigated thein vivoactivity of ceftaroline fosamil compared with clindamycin, linezolid, and vancomycin in a severe pneumonia model due to MRSA-producing Panton-Valentine leukocidin (PVL). A USA300 PVL-positive clone was used to induce pneumonia in rabbits. Infected rabbits were randomly assigned to no treatment or simulated human-equivalent dosing with ceftaroline fosamil, clindamycin, linezolid, or vancomycin. Residual bacterial concentrations in the lungs and spleen were assessed after 48 h of treatment. PVL expression was measured using a specific enzyme-linked immunosorbent assay (ELISA). Ceftaroline, clindamycin, and linezolid considerably reduced mortality rates compared with the control, whereas vancomycin did not. Pulmonary and splenic bacterial titers and PVL concentrations were greatly reduced by ceftaroline, clindamycin, and linezolid. Ceftaroline, clindamycin, and linezolid were associated with reduced pulmonary tissue damage based on significantly lower macroscopic scores. Ceftaroline fosamil, clindamycin, and, to a lesser extent, linezolid were efficient in reducing bacterial titers in both the lungs and spleen and decreasing macroscopic scores and PVL production compared with the control.

scholarly journals Cathelicidins Mitigate Staphylococcus aureus Mastitis and Reduce Bacterial Invasion in Murine Mammary Epithelium

2020 ◽  
Vol 88 (7) ◽  
Author(s):  
Paloma Araujo Cavalcante ◽  
Cameron G. Knight ◽  
Yi-Lin Tan ◽  
Ana Paula Alves Monteiro ◽  
Herman W. Barkema ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Epithelial Cells ◽  
Mammary Epithelial Cells ◽  
Mammary Epithelium ◽  
Immune Defense ◽  
Host Cells ◽  
Bacterial Invasion ◽  
Content Type ◽  
Development Of Resistance

ABSTRACT Staphylococcus aureus, an important cause of mastitis in mammals, is becoming increasingly problematic due to the development of resistance to conventional antibiotics. The ability of S. aureus to invade host cells is key to its propensity to evade immune defense and antibiotics. This study focuses on the functions of cathelicidins, small cationic peptides secreted by epithelial cells and leukocytes, in the pathogenesis of S. aureus mastitis in mice. We determined that endogenous murine cathelicidin (CRAMP; Camp) was important in controlling S. aureus infection, as cathelicidin knockout mice (Camp−/−) intramammarily challenged with S. aureus had higher bacterial burdens and more severe mastitis than did wild-type mice. The exogenous administration of both a synthetic human cathelicidin (LL-37) and a synthetic murine cathelicidin (CRAMP) (8 μM) reduced the invasion of S. aureus into the murine mammary epithelium. Additionally, this exogenous LL-37 was internalized into cultured mammary epithelial cells and impaired S. aureus growth in vitro. We conclude that cathelicidins may be potential therapeutic agents against mastitis; both endogenous and exogenous cathelicidins conferred protection against S. aureus infection by reducing bacterial internalization and potentially by directly killing this pathogen.

scholarly journals Characterization of Alpha-ToxinhlaGene Variants, Alpha-Toxin Expression Levels, and Levels of Antibody to Alpha-Toxin in Hemodialysis and Postsurgical Patients with Staphylococcus aureus Bacteremia

2014 ◽  
Vol 53 (1) ◽  
pp. 227-236 ◽  
Author(s):  
Batu K. Sharma-Kuinkel ◽  
Yuling Wu ◽  
David E. Tabor ◽  
Hoyin Mok ◽  
Bret R. Sellman ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Monoclonal Antibody ◽  
Clinical Outcome ◽  
Neutralizing Antibodies ◽  
Neutralizing Antibody ◽  
Enzyme Linked Immunosorbent Assay ◽  
Alpha Toxin ◽  
Content Type ◽  
Antibody Levels

Alpha-toxin is a majorStaphylococcus aureusvirulence factor. This study evaluated potential relationships betweenin vitroalpha-toxin expression ofS. aureusbloodstream isolates, anti-alpha-toxin antibody in serum of patients withS. aureusbacteremia (SAB), and clinical outcomes in 100 hemodialysis and 100 postsurgical SAB patients. Isolates underwentspatyping andhlasequencing. Serum anti-alpha-toxin IgG and neutralizing antibody levels were measured by using an enzyme-linked immunosorbent assay and a red blood cell (RBC)-based hemolysis neutralization assay. Neutralization of alpha-toxin by an anti-alpha-toxin monoclonal antibody (MAb MEDI4893) was tested in an RBC-based lysis assay. Most isolates encodedhla(197/200; 98.5%) and expressed alpha-toxin (173/200; 86.5%).In vitroalpha-toxin levels were inversely associated with survival (cure, 2.19 μg/ml, versus failure, 1.09 μg/ml;P< 0.01). Both neutralizing (hemodialysis, 1.26 IU/ml, versus postsurgical, 0.95;P< 0.05) and IgG (hemodialysis, 1.94 IU/ml, versus postsurgical, 1.27;P< 0.05) antibody levels were higher in the hemodialysis population. Antibody levels were also significantly higher in patients infected with alpha-toxin-expressingS. aureusisolates (P< 0.05). Levels of both neutralizing antibodies and IgG were similar among patients who were cured and those not cured (failures). Sequence analysis ofhlarevealed 12 distincthlagenotypes, and all genotypic variants were susceptible to a neutralizing monoclonal antibody in clinical development (MEDI4893). These data demonstrate that alpha-toxin is highly conserved in clinicalS. aureusisolates. Higherin vitroalpha-toxin levels were associated with a positive clinical outcome. Although patients infected with alpha-toxin-producingS. aureusexhibited higher anti-alpha-toxin antibody levels, these levels were not associated with a better clinical outcome in this study.

scholarly journals Role of 6-Gingerol in Reduction of Cholera Toxin ActivityIn VitroandIn Vivo

2013 ◽  
Vol 57 (9) ◽  
pp. 4373-4380 ◽  
Author(s):  
Pallashri Saha ◽  
Bornita Das ◽  
Keya Chaudhuri
Keyword(s):  
Cholera Toxin ◽  
Diarrheal Disease ◽  
Chemotherapeutic Agents ◽  
Mechanistic Study ◽  
Enzyme Linked Immunosorbent Assay ◽  
Ileal Loop ◽  
Content Type ◽  
Oral Rehydration

ABSTRACTVibrio choleraeis one of the major bacterial pathogens responsible for the devastating diarrheal disease called cholera. Chemotherapy is often used againstV. choleraeinfections; however, the emergence ofV. choleraewith multidrug resistance (MDR) toward the chemotherapeutic agents is a serious clinical problem. This scenario has provided us with the impetus to look into herbal remediation, especially toward blocking the action of cholera toxin (CT). Our studies were undertaken to determine the antidiarrheal potential of 6-gingerol (6G) on the basis of its effect on CT, the virulence factor secreted byV. cholerae. We report here that 6G binds to CT, hindering its interaction with the GM1 receptor present on the intestinal epithelial cells. The 50% inhibitory concentration (IC50) was determined to be 10 μg/ml. The detailed mechanistic study was conducted by enzyme-linked immunosorbent assay (ELISA), fluorescence spectroscopy, and isoelectric focusing. These results were validated within vitrostudies performed with the CHO, HeLa, and HT-29 cell lines, whereas a rabbit ileal loop assay was done to estimate thein vivoaction, which confirms the efficacy of 6G in remediation of the choleragenic effects of CT. Thus, 6G can be an effective adjunctive therapy with oral rehydration solution for severe CT-mediated diarrhea.

scholarly journals Adhesion of Staphylococcus aureus to epithelial cells: an in vitro approach to study interactions within the nasal microbiota

2020 ◽  
Vol 69 (10) ◽  
pp. 1253-1261
Author(s):  
Guillaume Ménard ◽  
Martine Bonnaure-Mallet ◽  
Pierre-Yves Donnio
Keyword(s):  
Staphylococcus Aureus ◽  
Epithelial Cells ◽  
Species Interactions ◽  
Type Species ◽  
Nasal Colonization ◽  
Coagulase Negative Staphylococci ◽  
Content Type ◽  
Link Type ◽  
Nasal Microbiota

Introduction. Staphylococcus aureus is a skin and mucous commensal bacterium of warm-blooded animals. In humans, the nose is the main ecological niche of S. aureus , and nasal carriage is a risk factor for developing an endogenous infection. S. aureus nasal colonization is a multifactorial process, involving inter-species interactions among the nasal microbiota. Aims. The objectives of this study were to characterize the microbiota of carriers and non-carriers of S. aureus and to demonstrate the importance of inter-species relationships in the adhesion of S. aureus , a key step in nasal colonization. Methodology. First, we characterized the nasal microbiota from 30 S. aureus carriers and non-carriers by a culturomic approach. We then evaluated the adhesion of S. aureus , first alone and then along with other bacteria of the nasal microbiota. To do that, we used an in vitro model to measure the interactions among bacteria in the presence of epithelial cells. Results. Analysis of the nasal microbiota of the carriers and non-carriers of S. aureus made it possible to observe that each microbiota has specific features in terms of composition. However, this composition differs significantly between carriers and non-carriers mainly through two bacterial groups: coagulase-negative staphylococci and corynebacteria. In a second part, adhesion of S. aureus to epithelial cells showed competition between S. aureus and these bacteria, suggesting a limitation of nasal colonization by S. aureus . Conclusion. These findings demonstrate the existence of a negative correlation between S. aureus and other species which inhibits adhesion and could limit nasal colonization.

scholarly journals Berberine and Obatoclax Inhibit SARS-Cov-2 Replication in Primary Human Nasal Epithelial Cells In Vitro

Viruses ◽  
2021 ◽  
Vol 13 (2) ◽  
pp. 282
Author(s):  
Finny S. Varghese ◽  
Esther van Woudenbergh ◽  
Gijs J. Overheul ◽  
Marc J. Eleveld ◽  
Lisa Kurver ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Early Stage ◽  
Antiviral Drugs ◽  
Target Cells ◽  
Nasal Epithelial Cells ◽  
Cytokines And Chemokines ◽  
Human Nasal Epithelial Cells ◽  
Apoptotic Protein ◽  
Air Liquid Interface

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) emerged as a new human pathogen in late 2019 and it has infected over 100 million people in less than a year. There is a clear need for effective antiviral drugs to complement current preventive measures, including vaccines. In this study, we demonstrate that berberine and obatoclax, two broad-spectrum antiviral compounds, are effective against multiple isolates of SARS-CoV-2. Berberine, a plant-derived alkaloid, inhibited SARS-CoV-2 at low micromolar concentrations and obatoclax, which was originally developed as an anti-apoptotic protein antagonist, was effective at sub-micromolar concentrations. Time-of-addition studies indicated that berberine acts on the late stage of the viral life cycle. In agreement, berberine mildly affected viral RNA synthesis, but it strongly reduced infectious viral titers, leading to an increase in the particle-to-pfu ratio. In contrast, obatoclax acted at the early stage of the infection, which is in line with its activity to neutralize the acidic environment in endosomes. We assessed infection of primary human nasal epithelial cells that were cultured on an air-liquid interface and found that SARS-CoV-2 infection induced and repressed expression of specific sets of cytokines and chemokines. Moreover, both obatoclax and berberine inhibited SARS-CoV-2 replication in these primary target cells. We propose berberine and obatoclax as potential antiviral drugs against SARS-CoV-2 that could be considered for further efficacy testing.

scholarly journals Rabeprazole inhibits inflammatory reaction by inhibition of cell pyroptosis in gastric epithelial cells

2021 ◽  
Vol 22 (1) ◽  
Author(s):  
Jing Xie ◽  
Long Fan ◽  
Liya Xiong ◽  
Peiyu Chen ◽  
Hongli Wang ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Clinical Sample ◽  
Critical Role ◽  
Enzyme Linked Immunosorbent Assay ◽  
Inflammasome Activation ◽  
Peptic Ulcers ◽  
Gastric Epithelial Cells ◽  
Caspase 1 ◽  
H Pylori

Abstract Background Helicobacter pylori (H. pylori) is a common pathogen in development of peptic ulcers with pyroptosis. Rabeprazole, a critical component of standard triple therapy, has been widely used as the first-line regimen for H. pylori infectious treatment. The aim of this study to explore the function of Rabeprazole on cell pyroptosis in vitro. Methods The clinical sample from patients diagnosed with or without H. pylori-infection were collected to analyze by Immunohistochemistry (IHC). Real-time quantitative PCR (qPCR), western blot (WB) and enzyme linked immunosorbent assay (Elisa) were performed to analyze the effect of Rabeprazole on cell pyroptosis, including LDH, IL-1β and IL-18. Results In this study, we showed that Rabeprazole regulated a phenomenon of cell pyroptosis as confirmed by lactate dehydrogenase (LDH) assay. Further results showed that Rabeprazole inhibited cell pyroptosis in gastric epithelial cells by alleviating GSDMD-executed pyroptosis, leading to decrease IL-1β and IL-18 mature and secretion, which is attributed to NLRP3 inflammasome activation inhibition. Further analysis showed that ASC, NLRP3 and Caspase-1, was significantly repressed in response to Rabeprazole stimulation, resulting in decreasing cleaved-caspase-1 expression. Most important, NLRP3 and GSDMD is significantly increased in gastric tissue of patients with H. pylori infection. Conclusion These findings revealed a critical role of Rabeprazole in cell pyroptosis in patients with H. pylori infection, suggesting that targeting cell pyroptosis is an alternative strategy in improving H. pylori treatment.

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