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Staphylococcus aureus Impairs the Function of and Kills Human Dendritic Cells via the LukAB Toxin
Berends, Evelien T M; Zheng, Xuhui; Zwack, Erin E; Ménager, Mickaël M; Cammer, Michael; Shopsin, Bo; Torres, Victor J
Staphylococcus aureus is a human pathogen responsible for high morbidity and mortality worldwide. Recurrent infections with this bacterium are common, suggesting that S. aureus thwarts the development of sterilizing immunity. S. aureus strains that cause disease in humans produce up to five different bicomponent toxins (leukocidins) that target and lyse neutrophils, innate immune cells that represent the first line of defense against S. aureus infections. However, little is known about the role of leukocidins in blunting adaptive immunity. Here, we explored the effects of leukocidins on human dendritic cells (DCs), antigen-presenting cells required for the development of adaptive immunity. Using an ex vivo infection model of primary human monocyte-derived dendritic cells, we found that S. aureus, including strains from different clonal complexes and drug resistance profiles, effectively kills DCs despite efficient phagocytosis. Although all purified leukocidins could kill DCs, infections with live bacteria revealed that S. aureus targets and kills DCs primarily via the activity of leukocidin LukAB. Moreover, using coculture experiments performed with DCs and autologous CD4+ T lymphocytes, we found that LukAB inhibits DC-mediated activation and proliferation of primary human T cells. Taken together, the data determined in the study reveal a novel immunosuppressive strategy of S. aureus whereby the bacterium blunts the development of adaptive immunity via LukAB-mediated injury of DCs.IMPORTANCE Antigen-presenting cells such as dendritic cells (DCs) fulfill an indispensable role in the development of adaptive immunity by producing proinflammatory cytokines and presenting microbial antigens to lymphocytes to trigger a faster, specific, and long-lasting immune response. Here, we studied the effect of Staphylococcus aureus toxins on human DCs. We discovered that the leukocidin LukAB hinders the development of adaptive immunity by targeting human DCs. The ability of S. aureus to blunt the function of DCs could help explain the high frequency of recurrent S. aureus infections. Taken together, the results from this study suggest that therapeutically targeting the S. aureus leukocidins may boost effective innate and adaptive immune responses by protecting innate leukocytes, enabling proper antigen presentation and T cell activation.
PMID: 30602580
ISSN: 2150-7511
CID: 3562862
Skin Associated Staphylococcus Aureus Contributes to Disease Progression in CTCL [Meeting Abstract]
Tegla, Cosmin A.; Herrera, Alberto M.; Seffens, Angelina M.; Fanok, Melania H.; Dean, George; Kawaoka, John; Laird, Mary E.; Fulmer, Yi; Willerslev-Olsen, Andreas; Hymes, Kenneth B.; Latkowski, Jo-Ann; Odum, Niels; Feske, Stefan; Shopsin, Bo; Torres, Victor; Kadin, Marshall E.; Geskin, Larisa J.; Koralov, Sergei B.
ISI:000518218500534
ISSN: 0006-4971
CID: 4505432
Genome plasticity of agr-defective Staphylococcus aureus during clinical infection
Altman, Deena R; Sullivan, Mitchell J; Chacko, Kieran I; Balasubramanian, Divya; Pak, Theodore R; Sause, William E; Kumar, Krishan; Sebra, Robert; Deikus, Gintaras; Attie, Oliver; Rose, Hannah; Lewis, Martha; Fulmer, Yi; Bashir, Ali; Kasarskis, Andrew; Schadt, Eric E; Richardson, Anthony R; Torres, Victor J; Shopsin, Bo; van Bakel, Harm
Therapy for bacteremia caused by Staphylococcus aureus is often ineffective, even when treatment conditions are optimal according to experimental protocols. Adapted subclones, such as those bearing mutations that attenuate agr-mediated virulence activation, are associated with persistent infection and patient mortality. To identify additional alterations in agr-defective mutants, we sequenced and assembled the complete genomes of clone pairs from colonizing and infected sites of several patients in whom S. aureus demonstrated a within-host loss of agr function. We report that events associated with agr inactivation result in agr-defective blood and nares strain pairs that are enriched in mutations compared to pairs from wild-type controls. The random distribution of mutations between colonizing and infecting strains from the same patient, and between strains from different patients, suggests that much of the genetic complexity of agr-defective strains result from prolonged infection or therapy-induced stress. However, in one of the agr-defective infecting strains, multiple genetic changes resulted in increased virulence in a murine model of bloodstream infection, bypassing the mutation of agr and raising the possibility that some changes were selected. Expression profiling correlated the elevated virulence of this agr-defective mutant to restored expression of the agr-regulated ESAT6-like Type VII secretion system, a known virulence factor. Thus, additional mutations outside the agr locus can contribute to diversification and adaptation during infection by S. aureusagr mutants associated with poor patient outcome.
PMID: 30061376
ISSN: 1098-5522
CID: 3217342
Hierarchy of human IgG recognition within the Staphylococcus aureus immunome
Radke, Emily E; Brown, Stuart M; Pelzek, Adam J; Fulmer, Yi; Hernandez, David N; Torres, Victor J; Thomsen, Isaac P; Chiang, William K; Miller, Andy O; Shopsin, Bo; Silverman, Gregg J
Staphylococcus aureus is an opportunistic pathogen that causes a range of serious infections associated with significant morbidity, by strains increasingly resistant to antibiotics. However, to date all candidate vaccines have failed to induce protective immune responses in humans. We need a more comprehensive understanding of the antigenic targets important in the context of human infection. To investigate infection-associated immune responses, patients were sampled at initial presentation and during convalescence from three types of clinical infection; skin and soft tissue infection (SSTI), prosthetic joint infection (PJI) and pediatric hematogenous osteomyelitis (PHO). Reactivity of serum IgG was tested with an array of recombinant proteins, representing over 2,652 in-vitro-translated open reading frames (ORFs) from a community-acquired methicillin-resistant S. aureus USA300 strain. High-level reactivity was demonstrated for 104 proteins with serum IgG in all patient samples. Overall, high-level IgG-reactivity was most commonly directed against a subset of secreted proteins. Although based on limited surveys, we found subsets of S. aureus proteins with differential reactivity with serum samples from patients with different clinical syndromes. Together, our studies have revealed a hierarchy within the diverse proteins of the S. aureus "immunome", which will help to advance efforts to develop protective immunotherapeutic agents.
PMCID:6125462
PMID: 30185867
ISSN: 2045-2322
CID: 3271732
Staphylococcus aureus biofilms release leukocidins to elicit extracellular trap formation and evade neutrophil-mediated killing
Bhattacharya, Mohini; Berends, Evelien T M; Chan, Rita; Schwab, Elizabeth; Roy, Sashwati; Sen, Chandan K; Torres, Victor J; Wozniak, Daniel J
Bacterial biofilms efficiently evade immune defenses, greatly complicating the prognosis of chronic infections. How methicillin-resistant Staphylococcus aureus (MRSA) biofilms evade host immune defenses is largely unknown. This study describes some of the major mechanisms required for S. aureus biofilms to evade the innate immune response and provides evidence of key virulence factors required for survival and persistence of bacteria during chronic infections. Neutrophils are the most abundant white blood cells in circulation, playing crucial roles in the control and elimination of bacterial pathogens. Specifically, here we show that, unlike single-celled populations, S. aureus biofilms rapidly skew neutrophils toward neutrophil extracellular trap (NET) formation through the combined activity of leukocidins Panton-Valentine leukocidin and γ-hemolysin AB. By eliciting this response, S. aureus was able to persist, as the antimicrobial activity of released NETs was ineffective at clearing biofilm bacteria. Indeed, these studies suggest that NETs could inadvertently potentiate biofilm infections. Last, chronic infection in a porcine burn wound model clearly demonstrated that leukocidins are required for "NETosis" and facilitate bacterial survival in vivo.
PMCID:6048508
PMID: 29941565
ISSN: 1091-6490
CID: 3192162
Genomic and Geographic Context for the Evolution of High-Risk Carbapenem-Resistant Enterobacter cloacae Complex Clones ST171 and ST78
Gomez-Simmonds, Angela; Annavajhala, Medini K; Wang, Zheng; Macesic, Nenad; Hu, Yue; Giddins, Marla J; O'Malley, Aidan; Toussaint, Nora C; Whittier, Susan; Torres, Victor J; Uhlemann, Anne-Catrin
Recent reports have established the escalating threat of carbapenem-resistant Enterobacter cloacae complex (CREC). Here, we demonstrate that CREC has evolved as a highly antibiotic-resistant rather than highly virulent nosocomial pathogen. Applying genomics and Bayesian phylogenetic analyses to a 7-year collection of CREC isolates from a northern Manhattan hospital system and to a large set of publicly available, geographically diverse genomes, we demonstrate clonal spread of a single clone, ST171. We estimate that two major clades of epidemic ST171 diverged prior to 1962, subsequently spreading in parallel from the Northeastern to the Mid-Atlantic and Midwestern United States and demonstrating links to international sites. Acquisition of carbapenem and fluoroquinolone resistance determinants by both clades preceded widespread use of these drugs in the mid-1980s, suggesting that antibiotic pressure contributed substantially to its spread. Despite a unique mobile repertoire, ST171 isolates showed decreased virulence in vitro While a second clone, ST78, substantially contributed to the emergence of CREC, it encompasses diverse carbapenemase-harboring plasmids, including a potentially hypertransmissible IncN plasmid, also present in other sequence types. Rather than heightened virulence, CREC demonstrates lineage-specific, multifactorial adaptations to nosocomial environments coupled with a unique potential to acquire and disseminate carbapenem resistance genes. These findings indicate a need for robust surveillance efforts that are attentive to the potential for local and international spread of high-risk CREC clones.IMPORTANCE Carbapenem-resistant Enterobacter cloacae complex (CREC) has emerged as a formidable nosocomial pathogen. While sporadic acquisition of plasmid-encoded carbapenemases has been implicated as a major driver of CREC, ST171 and ST78 clones demonstrate epidemic potential. However, a lack of reliable genomic references and rigorous statistical analyses has left many gaps in knowledge regarding the phylogenetic context and evolutionary pathways of successful CREC. Our reconstruction of recent ST171 and ST78 evolution represents a significant addition to current understanding of CREC and the directionality of its spread from the Eastern United States to the northern Midwestern United States with links to international collections. Our results indicate that the remarkable ability of E. cloacae to acquire and disseminate cross-class antibiotic resistance rather than virulence determinants, coupled with its ability to adapt under conditions of antibiotic pressure, likely led to the wide dissemination of CREC.
PMCID:5974468
PMID: 29844109
ISSN: 2150-7511
CID: 3658972
Nutritional regulation of the Sae two-component system by CodY in Staphylococcus aureus
Mlynek, Kevin D; Sause, William E; Moormeier, Derek E; Sadykov, Marat R; Hill, Kurt R; Torres, Victor J; Bayles, Kenneth W; Brinsmade, Shaun R
Staphylococcus aureus subverts innate defenses during infection in part by killing host immune cells to exacerbate disease. This human pathogen intercepts host cues and activates a transcriptional response via the S. aureus exoprotein expression (SaeR/S) two-component system to secrete virulence factors critical for pathogenesis. We recently showed that the transcriptional repressor CodY adjusts nuclease (nuc) gene expression via SaeR/S but the mechanism remained unknown. Herein, we identified two CodY binding motifs upstream of the sae P1 promoter, which suggested direct regulation by this global regulator. We show that CodY shares a binding site with the positive activator SaeR and alleviating direct CodY repression at this site is sufficient to abrogate stochastic expression, suggesting that CodY represses sae expression by blocking SaeR binding. Epistasis experiments support a model that CodY also controls sae indirectly through Agr and Rot-mediated repression of the sae P1 promoter. We also demonstrate that CodY repression of sae restrains production of secreted cytotoxins that kill human neutrophils. We conclude that CodY plays a previously unrecognized role in controlling virulence gene expression via SaeR/S, and suggest a mechanism by which CodY acts as a master regulator of pathogenesis by tying nutrient availability to virulence gene expression.Importance Bacterial mechanisms that mediate the switch from a commensal to pathogenic lifestyle is one of the biggest unanswered questions in infectious disease. Since the expression of most virulence genes is often correlated with nutrient depletion, this implies that virulence is a response to the lack of nourishment in host tissues, and that pathogens like S. aureus produce virulence factors in order to gain access to nutrients in the host. Herein, we show that specific nutrient depletion signals appear to be funneled to the SaeR/S system through the global regulator CodY. Our findings reveal a strategy by which S. aureus delays the production of immune evasion and immune cell killing proteins until key nutrients are depleted.
PMCID:5869476
PMID: 29378891
ISSN: 1098-5530
CID: 2933732
Human Memory B Cells TargetingStaphylococcus aureusExotoxins Are Prevalent with Skin and Soft Tissue Infection
Pelzek, Adam J; Shopsin, Bo; Radke, Emily E; Tam, Kayan; Ueberheide, Beatrix M; Fenyo, David; Brown, Stuart M; Li, Qianhao; Rubin, Ada; Fulmer, Yi; Chiang, William K; Hernandez, David N; El Bannoudi, Hanane; Sause, William E; Sommerfield, Alexis; Thomsen, Isaac P; Miller, Andy O; Torres, Victor J; Silverman, Gregg J
Staphylococcus aureus
PMCID:5850327
PMID: 29535203
ISSN: 2150-7511
CID: 2992702
Roles of Staphylococcus aureus Mnh1 and Mnh2 antiporters in salt tolerance, alkali tolerance and pathogenesis
Vaish, Manisha; Price-Whelan, Alexa; Reyes-Robles, Tamara; Liu, Jun; Jereen, Amyeo; Christie, Stephanie; Alonzo, Francis; Benson, Meredith A; Torres, Victor J; Krulwich, Terry A
Staphylococcus aureus has three types of cation/proton antiporters. The type three family includes two multi-subunit Na+/H+ (Mnh) antiporters, Mnh1 and Mnh2. These antiporters are clusters of seven hydrophobic membrane bound protein subunits. Mnh antiporters play important roles in maintaining cytoplasmic pH in prokaryotes, enabling their survival under extreme environmental stress. In this study, we investigated the physiological roles and catalytic properties of Mnh1 and Mnh2 in S. aureus Both Mnh1 and Mnh2 were cloned separately into a pGEM3Z+ vector in the antiporter deficient KNabc E. coli strain. The catalytic properties of the antiporters were measured in everted vesicles (inside out). The Mnh1 antiporter exhibited significant exchange of Na+/H+ cations at pH 7.5. Mnh2 showed significant exchange of both Na+/H+ and K+/H+ cations, especially at pH 8.5. Under elevated salt conditions, deletion of the mnhA1 gene resulted in a significant reduction of the growth rate of S. aureus in the range of pH 7.5 to 9. Deletion of mnhA2 had similar effects but mainly in the range of pH 8.5 -- 9.5. Double deletion of mnhA1 and mnhA2 led to severe reduction of the S. aureus growth rate mainly at pH values above 8.5. The effects of functional losses of both antiporters in S. aureus were also assessed via their support of virulence in the mouse in vivo infection model. Deletion of the mnhA1 gene led to a major loss of S. aureus virulence in mice, while deletion of mnh2 led to no change in virulence.Importance This study focuses on the catalytic properties and physiological roles of Mnh1 and Mnh2 cation/proton antiporters in S. aureus and their contributions under different stress conditions. The Mnh1 antiporter was found to have catalytic activity for Na+/H+ antiport and it plays a significant role in maintaining halotolerance at pH 7.5 while, the Mnh2 antiporter has catalytic activities for Na+/H+ as well as K+/H+ antiport that have roles in both osmotolerance and halotolerance in S. aureus Study of S. aureus with a single deletion of either mnhA1 or mnhA2 was assessed in an infection model of mice. The result shows that mnhA1 but not mnhA2, plays a major role in S. aureus virulence.
PMCID:5809693
PMID: 29263099
ISSN: 1098-5530
CID: 2894002
After the deluge: mining Staphylococcus aureus genomic data for clinical associations and host-pathogen interactions
Copin, Richard; Shopsin, Bo; Torres, Victor J
The genome of Staphylococcus aureus has rapidly become one the most frequently sequenced among bacteria, with more than 40000 genome sequences uploaded to public databases. Computational resources required for analysis and quality assessment have lagged behind accumulation of sequence data. Improved analytic pipelines, in combination with the development of customized S. aureus reference databases, can be used to inform S. aureus biology and potentially predict clinical outcome. Here, we review the currently available data about S. aureus genome in public databases, and discuss their potential utility for understanding S. aureus evolution. Also discussed are ways to overcome challenges to the application of whole-genome sequencing data for prevention and management of S. aureus disease.
PMCID:5862737
PMID: 29197673
ISSN: 1879-0364
CID: 2861662