Staphylococcus aureus remains a common cause of nosocomial bacterial infections and are often antibiotic resistant. The role of NK cells and IL-15 and their relationship in host defense against extracellular bacterial pathogens including S. aureus remain unclear. We have undertaken several approaches to address this issue using wild type (WT), IL-15 gene knock-out (KO), and NK cell-depleted mouse models. Upon pulmonary staphylococcal infection WT mice had markedly increased activated NK cells, but not NKT or γδ T cells, in the airway lumen that correlated with IL-15 production in the airway and with alveolar macrophages. In vitro exposure to staphylococcal products and/or coculture with lung macrophages directly activated NK cells. In contrast, lung macrophages better phagocytosed S. aureus in the presence of NK cells. In sharp contrast to WT controls, IL-15 KO mice deficient in NK cells were found to be highly susceptible to pulmonary staphylococcal infection despite markedly increased neutrophils and macrophages in the lung. In further support of these findings, WT mice depleted of NK cells were similarly susceptible to staphylococcal infection while they remained fully capable of IL-15 production in the lung at levels similar to those of NK-competent WT hosts. Our study thus identifies a critical role for NK cells in host defense against pulmonary extracellular bacterial infection and suggests that IL-15 is involved in this process via its indispensable effect on NK cells, but not other innate cells. These findings hold implication for the development of therapeutics in treating antibiotic-resistant S. aureus infection.

Lung immunopathology is the main cause of influenzamediated morbidity and death, and much of its molecular mechanisms remain unclear. Whereas tumor necrosis factor-␣ (TNF-␣) is traditionally considered a proinflammatory cytokine, its role in influenza immunopathology is unresolved. We have investigated this issue by using a model of acute H1N1 influenza infection established in wild-type and TNF-␣-deficient mice and evaluated lung viral clearance, inflammatory responses, and immunopathology. Whereas TNF-␣ was up-regulated in the lung after influenza infection, it was not required for normal influenza viral clearance. However, TNF-␣ deficiency led not only to a greater extent of illness but also to heightened lung immunopathology and tissue remodeling. The severe lung immunopathology was associated with increased inflammatory cell infiltration, anti-influenza adaptive immune responses, and expression of cytokines such as monocyte chemoattractant protein-1 (MCP-1) and fibrotic growth factor, TGF-␤1. Thus, in vivo neutralization of MCP-1 markedly attenuated lung immunopathology and blunted TGF-␤1 production following influenza infection in these hosts. On the other hand, in vivo transgenic expression of MCP-1 worsened lung immunopathology following influenza infection in wild-type hosts. Thus, TNF-␣ is dispensable for influenza clearance; however, different from the traditional belief, this cytokine is critically required for negatively regulating the extent of lung immunopathology during acute influenza infection.

Vaccine-inducible airway luminal antituberculosis memory CD8 T cells are self-renewable in an antigen-dependent manner, and can be maintained independent of peripheral T cell supply.

Protection by parenteral immunization with plasmid DNA vaccines against pulmonary tuberculosis (TB) is very modest. In this study, we have investigated the underlying mechanisms for the poor mucosal protective efficacy and the avenues and mechanisms to improve the efficacy of a single i.m. immunization with a monogenic plasmid DNA TB vaccine in a murine model. We show that i.m. DNA immunization fails to elicit accumulation of Ag-specific T cells in the airway lumen despite robust T cell responses in the spleen. Such systemically activated T cells cannot be rapidly mobilized into the airway lumen upon Mycobacterium tuberculosis exposure. However, airway deposition of low doses of soluble mycobacterial Ags in previously immunized mice effectively mobilizes the systemically activated T cells into the airway lumen. A fraction of such airway luminal T cells can persist in the airway lumen, undergo quick, robust expansion and activation and provide marked immune protection upon airway M. tuberculosis exposure. Airway mucosal deposition of soluble mycobacterial Ags was found to create a tissue microenvironment rich in proinflammatory molecules including chemokines and hence conducive to T cell recruitment. Thus, in vivo neutralization of MIP-1α or IFN-inducible protein-10 markedly inhibited the accumulation of Ag-specific T cells in the airway lumen. Our data suggest that immunoprotective efficacy on the mucosal surface by i.m. plasmid DNA immunization could be substantially improved by simple mucosal soluble Ag inoculation and restoration of mucosal luminal T cells. Our study holds implication for the future design of DNA vaccination strategies against intracellular infections.

Pulmonary tuberculosis (TB) remains a serious health problem worldwide. Effective vaccination strategies are needed. We report the development of a novel TB vaccine using vesicular stomatitis virus (VSV) as a viral vector system to express Ag85A. VSVAg85A was shown to be immunogenic when given to mice by either an intranasal or an intramuscular (i.m.) route. Although distinct T-cell profiles resulted from both routes of immunization, only intranasal delivery generated a mucosal T-cell response that was protective upon pulmonary Mycobacterium tuberculosis (M.tb) challenge. While this protection manifested at an early time-point after immunization, it was not sustained. The potential of VSVAg85A to be used as a mucosal booster for parenteral priming by an adenoviral TB vaccine expressing Ag85A (AdAg85A) was investigated. VSVAg85A immunization markedly boosted antigen-specific T-cell responses in the airway lumen while also augmenting immune activation in the systemic compartment, after AdAg85A priming. This translated into significantly better protective efficacy against pulmonary challenge with M.tb than either vaccine used alone. Our study therefore suggests that VSV as a vector system is a promising candidate to be used in a heterologous viral prime-boost immunization regimen against intracellular bacterial infection.