Fatemeh Askarian

A Staphylococcus aureus TIR domain protein virulence factor blocks TLR2-mediated NF-κB signaling.

Signaling through Toll-like receptors (TLRs), crucial molecules in the induction of host defense responses, requires adaptor proteins that contain a Toll/interleukin-1 receptor (TIR) domain. The pathogen Staphylococcus aureus produces several innate immune-evasion molecules that interfere with the hosts innate immune response. A database search analysis suggested the presence of a gene encoding a homologue of the human TIR domain in S. aureus MSSA476 which was named staphylococcal TIR domain protein (TirS). Ectopic expression of TirS in human embryonic kidney, macrophage and keratinocyte cell lines interfered with signaling through TLR2, including MyD88 and TIRAP, NF-κB and/or mitogen-activated protein kinase pathways. Moreover, the presence of TirS reduced the levels of cytokines MCP-1 and G-CSF secreted in response to S. aureus. The effects on NF-κB pathway were confirmed using S. aureus MSSA476 wild type, an isogenic mutant MSSA476ΔtirS, and complemented MSSA476ΔtirS +pTirS in a Transwell system where bacteria and host cells were physically separated. Finally, in a systematic mouse infection model, TirS promoted bacterial accumulation in several organs 4 days postinfection. The results of this study reveal a new S. aureus virulence factor that can interfere with PAMP-induced innate immune signaling in vitro and bacterial survival in vivo.

Bacterial interference with canonical NFκB signalling

The human body is constantly challenged by a variety of commensal and pathogenic micro-organisms that trigger the immune system. Central in the first line of defence is the pattern-recognition receptor (PRR)-induced stimulation of the NFκB pathway, leading to NFκB activation. The subsequent production of pro-inflammatory cytokines and/or antimicrobial peptides results in recruitment of professional phagocytes and bacterial clearance. To overcome this, bacteria have developed mechanisms for targeted interference in every single step in the PRR–NFκB pathway to dampen host inflammatory responses. This review aims to briefly overview the PRR–NFκB pathway in relation to the immune response and give examples of the diverse bacterial evasion mechanisms including changes in the bacterial surface, decoy production and injection of effector molecules. Targeted regulation of inflammatory responses is needed and bacterial molecules developed for immune evasion could provide future anti-inflammatory agents.

The interaction between Staphylococcus aureus SdrD and desmoglein 1 is important for adhesion to host cells

Staphylococcus aureus is known as a frequent colonizer of the skin and mucosa. Among bacterial factors involved in colonization are adhesins such as the microbial surface components recognizing adhesive matrix molecules (MSCRAMMs). Serine aspartate repeat containing protein D (SdrD) is involved in adhesion to human squamous cells isolated from the nose. Here, we identify Desmoglein 1 (Dsg1) as a novel interaction partner for SdrD. Genetic deletion of sdrD in S. aureus NCTC8325-4 through allelic replacement resulted in decreased bacterial adherence to Dsg1- expressing HaCaT cells in vitro. Complementary gain-of-function was demonstrated by heterologous expression of SdrD in Lactococcus lactis, which increased adherence to HaCaT cells. Also ectopic expression of Dsg1 in HEK293 cells resulted in increased adherence of S. aureus NCTC8325-4 in vitro. Increased adherence of NCTC8325-4, compared to NCTC8325-4ΔsdrD, to the recombinant immobilized Dsg1 demonstrated direct interaction between SdrD and Dsg1. Specificity of SdrD interaction with Dsg1 was further verified using flow cytometry and confirmed binding of recombinant SdrD to HaCaT cells expressing Dsg1 on their surface. These data demonstrate that Dsg1 is a host ligand for SdrD.

Characterisation and identification of enzyme-producing autochthonous bacteria from the gastrointestinal tract of two Indian air-breathing fish

Characterisation and identification of autochthonous enzyme-producing bacteria isolated from the proximal intestine and distal intestine of two species of Indian air-breathing fish, murrel (Channa punctatus) and stinging catfish (Heteropneustes fossilis), were investigated using conventional culture technique. Population levels of proteolytic strains were highest in the digestive tract of stinging catfish. In both species, the viable counts of amylase-producing bacteria were somewhat higher than cellulase-producing bacteria. Among the gut bacteria isolated, 8 strains (4 from murrel and 4 from stinging catfish) were selected as potent enzyme-producers on the basis of quantitative enzyme assays. All these strains were Gram-positive rods, but only four isolates (CPF4, CPH6, CPH7 and HFH4) were capable of forming endospores. The tested bacteria grew in wide range of temperatures and pH. The strains were further identified by 16S rRNA gene sequence analysis. Two strains, CPF3 (isolated from murrel) and HFH4 (isolated from stinging catfish) showed high similarity to Bacillus sp., strain HFH7 (isolated from the stinging catfish) was most closely related to Bacillus subtilis, while five strains belonged to Bacillus licheniformis. Based on the results of the present study, we suggest that incorporation of autochthonous enzyme-producing bacteria in aquafeeds merits further investigations.

Staphylococcus aureus Membrane-Derived Vesicles Promote Bacterial Virulence and Confer Protective Immunity in Murine Infection Models

Staphylococcus aureus produces membrane-derived vesicles (MVs), which share functional properties to outer membrane vesicles. Atomic force microscopy revealed that S. aureus-derived MVs are associated with the bacterial surface or released into the surrounding environment depending on bacterial growth conditions. By using a comparative proteomic approach, a total of 131 and 617 proteins were identified in MVs isolated from S. aureus grown in Luria-Bertani and brain-heart infusion broth, respectively. Purified S. aureus MVs derived from the bacteria grown in either media induced comparable levels of cytotoxicity and neutrophil-activation. Administration of exogenous MVs increased the resistance of S. aureus to killing by whole blood or purified human neutrophils ex vivo and increased S. aureus survival in vivo. Finally, immunization of mice with S. aureus-derived MVs induced production of IgM, total IgG, IgG1, IgG2a, and IgG2b resulting in protection against subcutaneous and systemic S. aureus infection. Collectively, our results suggest S. aureus MVs can influence bacterial–host interactions during systemic infections and provide protective immunity in murine models of infection.

Staphylococcus aureus nasal isolates from healthy individuals cause highly variable host cell responses in vitro

Studies on Staphylococcus aureus populations colonizing the nasal cavity reveal that some bacterial strains are more common, while others are rarely found. This study included five isolates with the most common spa types and five isolates with rare spa types from healthy population. Selected phenotypic traits and genomic content among nasal S.xa0aureus isolates were compared. Besides the rather similar growth rates, our data revealed a high diversity among isolates; that is, in biofilm formation, the ability to attach to and be internalized in keratinocytes as well as ability to induce pro- and anti-inflammatory cytokines. The results showed that S.xa0aureus isolates from healthy hosts are phenotypically diverse and cause highly variable host cell responses. Therefore, generalizing the results from one S.xa0aureus isolate to all is highly questionable.

Serine-aspartate-repeat protein D Increases Staphylococcus aureus Virulence and Survival in Blood

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.

Enterococcus faecium produces membrane vesicles containing virulence factors and antimicrobial resistance related proteins.

Enterococcus faecium is a commensal but also a bacteremia causing pathogen, which is inherently resistant to several antimicrobials and has a great ability to acquire new traits. Bacterial membrane vesicles (MVs) are increasingly recognized as a mode of cell-free communication and a way to deliver virulence factors and/or antimicrobial resistance determinants. These features make MVs interesting research targets in research on critical hospital pathogens. This study describes for the first time that E. faecium strains produce MVs. It presents a morphological as well as a proteomic analysis of MVs isolated from four different, clinically relevant E. faecium strains grown under two different conditions and identifies MV-associated proteins in all of them. Interestingly, 11 virulence factors are found among the MV-associated proteins, including biofilm-promoting proteins and extracellular matrix-binding proteins, which may aid in enterococcal colonization. Additionally, 11 antimicrobial resistance-related proteins were MV-associated. Among those, all proteins encoded by the vanA-cluster of a vancomycin resistant strain were found to be MV-associated. This implies that E. faecium MVs may be utilized by the bacterium to release proteins promoting virulence, pathogenicity and antimicrobial resistance.nnnSIGNIFICANCEnEnterococcal infections, especially bacteremia and endocarditis, are challenging to treat because E. faecium have acquired resistance to multiple classes of antimicrobials, including ampicillin, aminoglycosides, and glycopeptides. Thus, research on different modes of enterococcal pathogenicity is warranted. This study utilized a proteomic approach to identify MV-associated proteins of different nosocomial E. faecium strains representing four clinically relevant sequence types (STs), namely ST17, ST18, ST78, and ST192. The presented data suggest that E. faecium MVs are involved in virulence and antimicrobial resistance.

Staphylococcus aureus modulation of innate immune responses through Toll-like (TLR), (NOD)-like (NLR) and C-type lectin (CLR) receptors

Early recognition of pathogens by the innate immune system is crucial for bacterial clearance. Many pattern recognition receptors (PRRs) such as Toll-like (TLRs) and (NOD)-like (NLRs) receptors have been implicated in initial sensing of bacterial components. The intracellular signaling cascades triggered by these receptors result in transcriptional upregulation of inflammatory pathways. Although this step is crucial for bacterial elimination, it is also associated with the potential for substantial immunopathology, which underscores the need for tight control of inflammatory responses. The leading human bacterial pathogen Staphylococcus aureus expresses over 100 virulence factors that exert numerous effects upon host cells. In this manner, the pathogen seeks to avoid host recognition or perturb PRR-induced innate immune responses to allow optimal survival in the host. These immune system interactions may result in enhanced bacterial proliferation but also provoke systemic cytokine responses associated with sepsis. This review summarizes recent findings on the various mechanisms applied by S. aureus to modulate or interfere with inflammatory responses through PRRs. Detailed understanding of these complex interactions can provide new insights toward future immune-stimulatory therapeutics against infection or immunomodulatory therapeutics to suppress or correct dysregulated inflammation.

Flagella-mediated secretion of a novel Vibrio cholerae cytotoxin affecting both vertebrate and invertebrate hosts

Using Caenorhabditis elegans as anxa0infection host model for Vibrio cholerae predator interactions, we discovered a bacterial cytotoxin, MakA, whose function as a virulence factor relies on secretion via the flagellum channel in a proton motive force-dependent manner. The MakA protein is expressed from the polycistronic makDCBA (motility-associated killing factor) operon. Bacteria expressing makDCBA induced dramatic changes in intestinal morphology leading to a defecation defect, starvation and death in C. elegans. The Mak proteins also promoted V. cholerae colonization of the zebrafish gut causing lethal infection. A structural model of purified MakA at 1.9u2009Å resolution indicated similarities to members of a superfamily of bacterial toxins with unknown biological roles. Our findings reveal an unrecognized role for V. cholerae flagella in cytotoxin export that may contribute both to environmental spread of the bacteria by promoting survival and proliferation in encounters with predators, and to pathophysiological effects during infections.Mitesh Dongre et al. have identified a novel bacterial cytotoxin from Vibrio chloerae, whose virulence depends on flagella-mediated secretion. This suggests that the flagella’s role in cytotoxin export could improve survival and proliferation of bacteria when exposed to environmental stress.