Community-associated methicillin-resistant Staphylococcus aureus (CA-MRSA), especially the pandemic USA300 clone, is associated with severe infections and high mortality rates, particularly in patients with pneumonia. USA300 produces Panton-Valentine leukocidin (PVL), a potent exotoxin that lyses polymorphonuclear leukocytes (PMNs). It has been generally assumed that PVL accounts for hypervirulence of USA300 and has a key role in pathogenesis of necrotizing pneumonia, but data from rodent infection models are conflicting. Rodent PMNs are less susceptible than human PMNs to PVL cytotoxicity, which may account for the inconclusive results. Rabbit and human PMNs are similar in their susceptibility to PVL. Postulating that PMNs play a role in the tissue destructive process (and therefore susceptibility of this target cell to PVL effects matters), we developed a rabbit model of pneumonia. In this model, an isogenic PVL-negative USA300 mutant was attenuated in its capacity to cause lethal pneumonia compared to the PVL-positive parent;virulence was fully restored in the PVL-complemented mutant. Infection with the PVL-positive strain was remarkably similar to the human disease. Airspace instillation of purified PVL alone induced rapid onset of acute lung injury and lung inflammation. The overall objective of this project is to determine the mechanisms of PVL-induced acute lung injury, lung inflammation and lethal necrotizing pneumonia.
Four specific aims are proposed.
Aim 1. To determine the role of host inflammatory response in PVL-induced necrotizing pneumonia. We hypothesize that PVL-induced acute lung injury is mediated by PMNs. To test this hypothesis, we will determine whether blocking interleukin 8, the principal chemokine that recruits PMNs into the lung, attenuates PVL-induced acute lung injury and lung inflammation.
Aim 2. To determine whether PVL and related bicomponent toxins, leukocidin (LukED) and -haemolysin (HlgABC), individually or synergistically induce acute lung injury and lung inflammation. We will investigate whether deletion of the lukED and hlgABC genes attenuates virulence;whether gene complementation restores virulence;and determine the levels of LukED and HlgABC expression during infection. Potential synergism among PVL, LukED and HlgABC in inducing acute lung injury and lung inflammation also will be investigated as this information has important implications for designing therapeutic interventions.
Aim 3. To determine the relative effects of PVL, -hemolysin (HLA), and -type phenol-soluble modulins (PSM-) on acute lung injury and necrotizing pneumonia in the rabbit model. These experiments will determine whether other exotoxins similar in their activities to PVL contribute to virulence in the pneumonia model.
Aim 4. To determine whether passive and active immunization protects against PVL-induced lung injury and death in the rabbit pneumonia model. Our proposed experiments will increase knowledge of the role of PVL and related toxins in pathogenesis of lung injury and establish a basis for the development of new therapeutic approaches.

Public Health Relevance

Because of the high mortality rate associated with necrotizing hemorrhagic pneumonia caused by Panton Valentine leukocidin (PVL)-producing Staphylococcus aureus, there is an urgent need to understand better the mechanisms of this toxin-mediated disease and develop new therapeutic approaches. We propose to use a rabbit model to investigate the mechanisms of PVL and related toxins in causing acute lung injury and lung inflammation. Specific treatment strategies to block the toxigenic effects of PVL will be evaluated in the rabbit model.

National Institute of Health (NIH)
National Institute of Allergy and Infectious Diseases (NIAID)
Research Project (R01)
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Host Interactions with Bacterial Pathogens Study Section (HIBP)
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Huntley, Clayton C
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University of California San Francisco
Internal Medicine/Medicine
Schools of Medicine
San Francisco
United States
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Diep, Binh An; Hilliard, Jamese J; Le, Vien T M et al. (2017) Targeting Alpha Toxin To Mitigate Its Lethal Toxicity in Ferret and Rabbit Models of Staphylococcus aureus Necrotizing Pneumonia. Antimicrob Agents Chemother 61:
Diep, Binh An; Le, Vien T M; Badiou, Cedric et al. (2016) IVIG-mediated protection against necrotizing pneumonia caused by MRSA. Sci Transl Med 8:357ra124
Le, Vien T M; Tkaczyk, Christine; Chau, Sally et al. (2016) Critical Role of Alpha-Toxin and Protective Effects of Its Neutralization by a Human Antibody in Acute Bacterial Skin and Skin Structure Infections. Antimicrob Agents Chemother 60:5640-8
Diep, Binh An; Le, Vien T M; Visram, Zehra C et al. (2016) Improved Protection in a Rabbit Model of Community-Associated Methicillin-Resistant Staphylococcus aureus Necrotizing Pneumonia upon Neutralization of Leukocidins in Addition to Alpha-Hemolysin. Antimicrob Agents Chemother 60:6333-40
Cohen, Taylor S; Hilliard, Jamese J; Jones-Nelson, Omari et al. (2016) Staphylococcus aureus ? toxin potentiates opportunistic bacterial lung infections. Sci Transl Med 8:329ra31
Diep, Binh An; Phung, Qui; Date, Shailesh et al. (2014) Identifying potential therapeutic targets of methicillin-resistant Staphylococcus aureus through in vivo proteomic analysis. J Infect Dis 209:1533-41
Diep, Binh An; Afasizheva, Anna; Le, Hoan N et al. (2013) Effects of linezolid on suppressing in vivo production of staphylococcal toxins and improving survival outcomes in a rabbit model of methicillin-resistant Staphylococcus aureus necrotizing pneumonia. J Infect Dis 208:75-82
Le, Vien Thi Minh; Diep, Binh An (2013) Selected insights from application of whole-genome sequencing for outbreak investigations. Curr Opin Crit Care 19:432-9
Tattevin, Pierre; Schwartz, Brian S; Graber, Christopher J et al. (2012) Concurrent epidemics of skin and soft tissue infection and bloodstream infection due to community-associated methicillin-resistant Staphylococcus aureus. Clin Infect Dis 55:781-8
Perret, Magali; Badiou, Cedric; Lina, Gerard et al. (2012) Cross-talk between Staphylococcus aureus leukocidins-intoxicated macrophages and lung epithelial cells triggers chemokine secretion in an inflammasome-dependent manner. Cell Microbiol 14:1019-36

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