scholarly journals Leukocidins and the Nuclease Nuc Prevent Neutrophil-Mediated Killing of Staphylococcus aureus Biofilms

2020 ◽  
Vol 88 (10) ◽  
Author(s):  
Mohini Bhattacharya ◽  
Evelien T. M. Berends ◽  
Xuhui Zheng ◽  
Preston J. Hill ◽  
Rita Chan ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Innate Immune ◽  
Human Neutrophils ◽  
Chronic Infections ◽  
Content Type ◽  
Persistent Infections ◽  
Biofilm Bacteria ◽  
Key Aspects ◽  
Neutrophil Response ◽  
Insight Into

ABSTRACT Bacterial biofilms are linked with chronic infections and have properties distinct from those of planktonic, single-celled bacteria. The virulence mechanisms associated with Staphylococcus aureus biofilms are becoming better understood. Human neutrophils are critical for the innate immune response to S. aureus infection. Here, we describe two virulence strategies that converge to promote the ability of S. aureus biofilms to evade killing by neutrophils. Specifically, we show that while neutrophils exposed to S. aureus biofilms produce extracellular traps (NETs) and phagocytose bacteria, both mechanisms are inefficient in clearance of the biofilm biomass. This is attributed to the leukocidin LukAB, which promotes S. aureus survival during phagocytosis. We also show that the persistence of biofilm bacteria trapped in NETs is facilitated by S. aureus nuclease (Nuc)-mediated degradation of NET DNA. This study describes key aspects of the interaction between primary human neutrophils and S. aureus biofilms and provides insight into how S. aureus evades the neutrophil response to cause persistent infections.

scholarly journals d-Amino Acids Enhance the Activity of Antimicrobials against Biofilms of Clinical Wound Isolates of Staphylococcus aureus and Pseudomonas aeruginosa

2014 ◽  
Vol 58 (8) ◽  
pp. 4353-4361 ◽  
Author(s):  
Carlos J. Sanchez ◽  
Kevin S. Akers ◽  
Desiree R. Romano ◽  
Ronald L. Woodbury ◽  
Sharanda K. Hardy ◽  
...  
Keyword(s):  
Amino Acids ◽  
Staphylococcus Aureus ◽  
Pseudomonas Aeruginosa ◽  
Clinical Isolates ◽  
Chronic Infections ◽  
Content Type ◽  
Log Reduction ◽  
Viable Bacteria ◽  
Biofilm Bacteria

ABSTRACTWithin wounds, microorganisms predominantly exist as biofilms. Biofilms are associated with chronic infections and represent a tremendous clinical challenge. As antibiotics are often ineffective against biofilms, use of dispersal agents as adjunctive, topical therapies for the treatment of wound infections involving biofilms has gained interest. We evaluatedin vitrothe dispersive activity ofd-amino acids (d-AAs) on biofilms from clinical wound isolates ofStaphylococcus aureusandPseudomonas aeruginosa; moreover, we determined whether combinations ofd-AAs and antibiotics (clindamycin, cefazolin, oxacillin, rifampin, and vancomycin forS. aureusand amikacin, colistin, ciprofloxacin, imipenem, and ceftazidime forP. aeruginosa) enhance activity against biofilms.d-Met,d-Phe, andd-Trp at concentrations of ≥5 mM effectively dispersed preformed biofilms ofS. aureusandP. aeruginosaclinical isolates, an effect that was enhanced when they were combined as an equimolar mixture (d-Met/d-Phe/d-Trp). When combined withd-AAs, the activity of rifampin was significantly enhanced against biofilms of clinical isolates ofS. aureus, as indicated by a reduction in the minimum biofilm inhibitory concentration (MBIC) (from 32 to 8 μg/ml) and a >2-log reduction of viable biofilm bacteria compared to treatment with antibiotic alone. The addition ofd-AAs was also observed to enhance the activity of colistin and ciprofloxacin against biofilms ofP. aeruginosa, reducing the observed MBIC and the number of viable bacteria by >2 logs and 1 log at 64 and 32 μg/ml in contrast to antibiotics alone. These findings indicate that the biofilm dispersal activity ofd-AAs may represent an effective strategy, in combination with antimicrobials, to release bacteria from biofilms, subsequently enhancing antimicrobial activity.

scholarly journals Serine-Aspartate Repeat Protein D Increases Staphylococcus aureus Virulence and Survival in Blood

2016 ◽  
Vol 85 (1) ◽  
Author(s):  
Fatemeh Askarian ◽  
Satoshi Uchiyama ◽  
J. Andrés Valderrama ◽  
Clement Ajayi ◽  
Johanna U. E. Sollid ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Ex Vivo ◽  
Innate Immune ◽  
Human Neutrophils ◽  
Infection Model ◽  
Bacterial Survival ◽  
Bacterial Killing ◽  
Content Type ◽  
Repeat Protein

ABSTRACT Staphylococcus aureus expresses a panel of cell wall-anchored adhesins, including proteins belonging to the microbial surface components recognizing adhesive matrix molecule (MSCRAMM) family, exemplified by the serine-aspartate repeat protein D (SdrD), which serve key roles in colonization and infection. Deletion of sdrD from S. aureus subsp. aureus strain NCTC8325-4 attenuated bacterial survival in human whole blood ex vivo, which was associated with increased killing by human neutrophils. Remarkably, SdrD was able to inhibit innate immune-mediated bacterial killing independently of other S. aureus proteins, since addition of recombinant SdrD protein and heterologous expression of SdrD in Lactococcus lactis promoted bacterial survival in human blood. SdrD contributes to bacterial virulence in vivo, since fewer S. aureus subsp. aureus NCTC8325-4 ΔsdrD bacteria than bacteria of the parent strain were recovered from blood and several organs using a murine intravenous infection model. Collectively, our findings reveal a new property of SdrD as an important key contributor to S. aureus survival and the ability to escape the innate immune system in blood.

scholarly journals Further Insight into the Mechanism of Human PMN Lysis following Phagocytosis of Staphylococcus aureus

2021 ◽  
Author(s):  
Viktoria Rungelrath ◽  
Adeline R. Porter ◽  
Natalia Malachowa ◽  
Brett A. Freedman ◽  
Jacqueline M. Leung ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Human Neutrophils ◽  
Human Pathogen ◽  
Content Type ◽  
Insight Into

S. aureus strain USA300 has the ability to cause rapid lysis of human neutrophils after phagocytosis. Although this phenomenon likely contributes to the success of USA300 as a human pathogen, our knowledge of the mechanism remains incomplete.

scholarly journals Pseudomonas aeruginosa PA14 Enhances the Efficacy of Norfloxacin against Staphylococcus aureus Newman Biofilms

2020 ◽  
Vol 202 (18) ◽  
Author(s):  
Giulia Orazi ◽  
Fabrice Jean-Pierre ◽  
George A. O’Toole
Keyword(s):  
Cystic Fibrosis ◽  
Staphylococcus Aureus ◽  
Pseudomonas Aeruginosa ◽  
Antimicrobial Agents ◽  
Respiratory Infections ◽  
Multiple Infection ◽  
Bacterial Biofilms ◽  
Interspecies Interactions ◽  
Chronic Infections ◽  
Content Type

ABSTRACT The thick mucus within the airways of individuals with cystic fibrosis (CF) promotes frequent respiratory infections that are often polymicrobial. Pseudomonas aeruginosa and Staphylococcus aureus are two of the most prevalent pathogens that cause CF pulmonary infections, and both are among the most common etiologic agents of chronic wound infections. Furthermore, the ability of P. aeruginosa and S. aureus to form biofilms promotes the establishment of chronic infections that are often difficult to eradicate using antimicrobial agents. In this study, we found that multiple LasR-regulated exoproducts of P. aeruginosa, including 2-heptyl-4-hydroxyquinoline N-oxide (HQNO), siderophores, phenazines, and rhamnolipids, likely contribute to the ability of P. aeruginosa PA14 to shift S. aureus Newman norfloxacin susceptibility profiles. Here, we observe that exposure to P. aeruginosa exoproducts leads to an increase in intracellular norfloxacin accumulation by S. aureus. We previously showed that P. aeruginosa supernatant dissipates the S. aureus membrane potential, and furthermore, depletion of the S. aureus proton motive force recapitulates the effect of the P. aeruginosa PA14 supernatant on shifting norfloxacin sensitivity profiles of biofilm-grown S. aureus Newman. From these results, we hypothesize that exposure to P. aeruginosa PA14 exoproducts leads to increased uptake of the drug and/or an impaired ability of S. aureus Newman to efflux norfloxacin. Surprisingly, the effect observed here of P. aeruginosa PA14 exoproducts on S. aureus Newman susceptibility to norfloxacin seemed to be specific to these strains and this antibiotic. Our results illustrate that microbially derived products can alter the ability of antimicrobial agents to kill bacterial biofilms. IMPORTANCE Pseudomonas aeruginosa and Staphylococcus aureus are frequently coisolated from multiple infection sites, including the lungs of individuals with cystic fibrosis (CF) and nonhealing diabetic foot ulcers. Coinfection with P. aeruginosa and S. aureus has been shown to produce worse outcomes compared to infection with either organism alone. Furthermore, the ability of these pathogens to form biofilms enables them to cause persistent infection and withstand antimicrobial therapy. In this study, we found that P. aeruginosa-secreted products dramatically increase the ability of the antibiotic norfloxacin to kill S. aureus biofilms. Understanding how interspecies interactions alter the antibiotic susceptibility of bacterial biofilms may inform treatment decisions and inspire the development of new therapeutic strategies.

scholarly journals Biofilm/Persister/Stationary Phase Bacteria Cause More Severe Disease Than Log Phase Bacteria – II Infection with Persister Forms of Staphylococcus aureus Causes a Chronic Persistent Skin Infection with More Severe Lesion that Takes Longer to Heal and is not Eradicated by the Current Recommended Treatment in Mice

2018 ◽  
Author(s):  
Rebecca Yee ◽  
Yuting Yuan ◽  
Cory Brayton ◽  
Andreina Tarff Leal ◽  
Jie Feng ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Stationary Phase ◽  
Skin Infection ◽  
Bacterial Load ◽  
Mouse Skin ◽  
Infection Model ◽  
Persister Cells ◽  
Persistent Infections ◽  
Recommended Treatment ◽  
Biofilm Bacteria

AbstractStaphylococcus aureus is an opportunistic pathogen that can cause persistent infections clinically. Treatment for chronic S. aureus infections ranges from at least one week to several months and such infections are prone to relapse likely due to the presence of persistent forms of bacteria such as persister cells. Persister cells, which are bacterial cells that become dormant under stress conditions, can be isolated in vitro but their clinical significance in in vivo infections are largely unclear. Here, we evaluated S. aureus persistent forms using stationary phase cultures and biofilm bacteria (enriched in persisters) in comparison with log phase cultures in terms of their ability to cause disease in a mouse skin infection model. Surprisingly, we found that infection of mice with stationary phase cultures and biofilm bacteria produced a more severe chronic skin infection with more pronounced lesions which took longer to heal than log phase (actively growing) cultures. After two week infection, the bacterial load and skin tissue pathology, as determined by hyperplasia, immune cell infiltration, and crust/lesion formation, of mice infected with the more persistent forms (e.g. stationary phase bacteria and biofilm bacteria) were greater than mice infected with log phase bacteria. Using our persistent infection mouse model, we showed that the clinically recommended treatment for recurrent S. aureus skin infection, doxycycline + rifampin, was not effective in eradicating the bacteria in the treatment study, despite reducing lesion sizes and pathology in infected mice. Analogous findings were also observed in a Caenorhabditis elegans model, where S.aureus stationary phase cultures caused a greater mortality than log phase culture as early as two days post-infection. Thus, we established a new model for chronic persistent infections using persister bacteria that could serve as a relevant model to evaluate therapeutic options for persistent infections in general. Our findings connect persisters with persistent infections, have implications for understanding disease pathogenesis, and are likely to be broadly valid for other pathogens.

scholarly journals In Staphylococcus aureus, the Particulate State of the Cell Envelope Is Required for the Efficient Induction of Host Defense Responses

2019 ◽  
Vol 87 (12) ◽  
Author(s):  
ByungHyun Kim ◽  
TingTing Jiang ◽  
Jun-Hyun Bae ◽  
Hye Su Yun ◽  
Seong Han Jang ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Peritoneal Cavity ◽  
Immune Activation ◽  
Bacterial Cell ◽  
Physical State ◽  
Cell Envelope ◽  
Innate Immune ◽  
Soluble Form ◽  
Content Type ◽  
Bacterial Cell Envelope

ABSTRACT Upon microbial infection, host immune cells recognize bacterial cell envelope components through cognate receptors. Although bacterial cell envelope components function as innate immune molecules, the role of the physical state of the bacterial cell envelope (i.e., particulate versus soluble) in host immune activation has not been clearly defined. Here, using two different forms of the staphylococcal cell envelope of Staphylococcus aureus RN4220 and USA300 LAC strains, we provide biochemical and immunological evidence that the particulate state is required for the effective activation of host innate immune responses. In a murine model of peritoneal infection, the particulate form of the staphylococcal cell envelope (PCE) induced the production of chemokine (C-X-C motif) ligand 1 (CXCL1) and CC chemokine ligand 2 (CCL2), the chemotactic cytokines for neutrophils and monocytes, respectively, resulting in a strong influx of the phagocytes into the peritoneal cavity. In contrast, compared with PCE, the soluble form of cell envelope (SCE), which was derived from PCE by treatment with cell wall-hydrolyzing enzymes, showed minimal activity. PCE also induced the secretion of calprotectin (myeloid-related protein 8/14 [MRP8/14] complex), a phagocyte-derived antimicrobial protein, into the peritoneal cavity at a much higher level than did SCE. The injected PCE particles were phagocytosed by the infiltrated neutrophils and monocytes and then delivered to mediastinal draining lymph nodes. More importantly, intraperitoneally (i.p.) injected PCE efficiently protected mice from S. aureus infection, which was abolished by the depletion of either monocytes/macrophages or neutrophils. This study demonstrated that the physical state of bacterial cells is a critical factor for efficient host immune activation and the protection of hosts from staphylococcal infections.

scholarly journals Three-Dimensional In Vitro Staphylococcus aureus Abscess Communities Display Antibiotic Tolerance and Protection from Neutrophil Clearance

2020 ◽  
Vol 88 (11) ◽  
Author(s):  
Marloes I. Hofstee ◽  
Martijn Riool ◽  
Igors Terjajevs ◽  
Keith Thompson ◽  
Martin J. Stoddart ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Immune Cells ◽  
Early Stage ◽  
Three Dimensional ◽  
Collagen Gel ◽  
Maximum Size ◽  
Human Neutrophils ◽  
Content Type

ABSTRACT Staphylococcus aureus is a prominent human pathogen in bone and soft-tissue infections. Pathophysiology involves abscess formation, which consists of central staphylococcal abscess communities (SACs), surrounded by a fibrin pseudocapsule and infiltrating immune cells. Protection against the ingress of immune cells such as neutrophils, or tolerance to antibiotics, remains largely unknown for SACs and is limited by the lack of availability of in vitro models. We describe a three-dimensional in vitro model of SACs grown in a human plasma-supplemented collagen gel. The in vitro SACs reached their maximum size by 24 h and elaborated a fibrin pseudocapsule, as confirmed by electron and immunofluorescence microscopy. The in vitro SACs tolerated 100× the MIC of gentamicin alone and in combination with rifampin, while planktonic controls and mechanically dispersed SACs were efficiently killed. To simulate a host response, SACs were exposed to differentiated PLB-985 neutrophil-like (dPLB) cells and to primary human neutrophils at an early stage of SAC formation or after maturation at 24 h. Both cell types were unable to clear mature in vitro SACs, but dPLB cells prevented SAC growth upon early exposure before pseudocapsule maturation. Neutrophil exposure after plasmin pretreatment of the SACs resulted in a significant decrease in the number of bacteria within the SACs. The in vitro SAC model mimics key in vivo features, offers a new tool to study host-pathogen interactions and drug efficacy assessment, and has revealed the functionality of the S. aureus pseudocapsule in protecting the bacteria from host phagocytic responses and antibiotics.

scholarly journals Staphylococcus aureus Releases Proinflammatory Membrane Vesicles To Resist Antimicrobial Fatty Acids

mSphere ◽  
2020 ◽  
Vol 5 (5) ◽  
Author(s):  
Arnaud Kengmo Tchoupa ◽  
Andreas Peschel
Keyword(s):  
Fatty Acids ◽  
Staphylococcus Aureus ◽  
Immune Responses ◽  
Membrane Vesicle ◽  
Membrane Vesicles ◽  
Innate Immune ◽  
Toll Like Receptor ◽  
Content Type ◽  
Healthy Humans ◽  
Toll Like Receptor 2

ABSTRACT Staphylococcus aureus is a major pathogen, which colonizes one in three otherwise healthy humans. This significant spread of S. aureus is largely due to its ability to circumvent innate immune responses, including antimicrobial fatty acids (AFAs) on the skin and in nasal secretions. In response to AFAs, S. aureus swiftly induces resistance mechanisms, which have yet to be completely elucidated. Here, we identify membrane vesicle (MV) release as a resistance strategy used by S. aureus to sequester host-specific AFAs. MVs protect S. aureus against a wide array of AFAs. Strikingly, beside MV production, S. aureus modulates MV composition upon exposure to AFAs. MVs purified from bacteria grown in the presence of linoleic acid display a distinct protein content and are enriched in lipoproteins, which strongly activate Toll-like receptor 2 (TLR2). Cumulatively, our findings reveal the protective capacities of MVs against AFAs, which are counteracted by an increased TLR2-mediated innate immune response. IMPORTANCE The nares of one in three humans are colonized by Staphylococcus aureus. In these environments, and arguably on all mucosal surfaces, bacteria encounter fatty acids with antimicrobial properties. Our study uncovers that S. aureus releases membrane vesicles (MVs) that act as decoys to protect the bacterium against antimicrobial fatty acids (AFAs). The AFA-neutralizing effects of MVs were neither strain specific nor restricted to one particular AFA. Hence, MVs may represent “public goods” playing an overlooked role in shaping bacterial communities in AFA-rich environments such as the skin and nose. Intriguingly, in addition to MV biogenesis, S. aureus modulates MV composition in response to exposure to AFAs, including an increased release of lipoproteins. These MVs strongly stimulate the innate immunity via Toll-like receptor 2 (TLR2). TLR2-mediated inflammation, which helps to fight infections, may exacerbate inflammatory disorders like atopic dermatitis. Our study highlights intricate immune responses preventing infections from colonizing bacteria.

scholarly journals OptimizedIn VitroAntibiotic Susceptibility Testing Method for Small-Colony Variant Staphylococcus aureus

2016 ◽  
Vol 60 (3) ◽  
pp. 1725-1735 ◽  
Author(s):  
Mimi R. Precit ◽  
Daniel J. Wolter ◽  
Adam Griffith ◽  
Julia Emerson ◽  
Jane L. Burns ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Susceptibility Testing ◽  
Brain Heart Infusion ◽  
Chronic Infections ◽  
Poor Growth ◽  
Content Type ◽  
Treatment History ◽  
Mueller Hinton ◽  
Mueller Hinton Agar ◽  
Small Colony

Staphylococcus aureussmall-colony variants (SCVs) emerge frequently during chronic infections and are often associated with worse disease outcomes. There are no standardized methods for SCV antibiotic susceptibility testing (AST) due to poor growth and reversion to normal-colony (NC) phenotypes on standard media. We sought to identify reproducible methods for AST ofS. aureusSCVs and to determine whether SCV susceptibilities can be predicted on the basis of treatment history, SCV biochemical type (auxotrophy), or the susceptibilities of isogenic NC coisolates. We tested the growth and stability of SCV isolates on 11 agar media, selecting for AST 2 media that yielded optimal SCV growth and the lowest rates of reversion to NC phenotypes. We then performed disk diffusion AST on 86S. aureusSCVs and 28 isogenic NCs and Etest for a subset of 26 SCVs and 24 isogenic NCs. Growth and reversion were optimal on brain heart infusion agar and Mueller-Hinton agar supplemented with compounds for which most clinical SCVs are auxotrophic: hemin, menadione, and thymidine. SCVs were typically nonsusceptible to either trimethoprim-sulfamethoxazole or aminoglycosides, in accordance with the auxotrophy type. In contrast, SCVs were variably nonsusceptible to fluoroquinolones, macrolides, lincosamides, fusidic acid, and rifampin;mecA-positive SCVs were invariably resistant to cefoxitin. All isolates (both SCVs and NCs) were susceptible to quinupristin-dalfopristin, vancomycin, minocycline, linezolid, chloramphenicol, and tigecycline. Analysis of SCV auxotrophy type, isogenic NC antibiograms, and antibiotic treatment history had limited utility in predicting SCV susceptibilities. With clinical correlation, this AST method and these results may prove useful in directing treatment for SCV infections.

scholarly journals ComparativeIn VitroActivities of Oritavancin, Dalbavancin, and Vancomycin against Methicillin-Resistant Staphylococcus aureus Isolates in a Nondividing State

2016 ◽  
Vol 60 (7) ◽  
pp. 4342-4345 ◽  
Author(s):  
Adam Belley ◽  
David Lalonde Seguin ◽  
Francis Arhin ◽  
Greg Moeck
Keyword(s):  
Staphylococcus Aureus ◽  
Bactericidal Activity ◽  
Antibacterial Agents ◽  
Methicillin Resistant Staphylococcus Aureus ◽  
Methicillin Resistant ◽  
Content Type ◽  
Skin Structure ◽  
Persistent Infections ◽  
Time Kill

ABSTRACTAntibacterial agents that kill nondividing bacteria may be of utility in treating persistent infections. Oritavancin and dalbavancin are bactericidal lipoglycopeptides that are approved for acute bacterial skin and skin structure infections in adults caused by susceptible Gram-positive pathogens. Using time-kill methodology, we demonstrate that oritavancin exerts bactericidal activity against methicillin-resistantStaphylococcus aureus(MRSA) isolates that are maintained in a nondividing statein vitro, whereas dalbavancin and the glycopeptide vancomycin do not.

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