Th17-stimulating Protein Vaccines Confer Protection against Pseudomonas aeruginosa Pneumonia

Abstract

Rationale: New vaccine approaches are needed for Pseudomonas aeruginosa, which continues to be a major cause of serious pulmonary infections. Although Th17 cells can protect against gram-negative pathogens at mucosal surfaces, including the lung, the bacterial proteins recognized by Th17 cells are largely unknown and could be potential new vaccine candidates. Objectives: We describe a strategy to identify Th17-stimulating protein antigens of Pseudomonas aeruginosa to assess their efficacy as vaccines against pneumonia.Methods: Using a library of in vitro transcribed and translated P. aeruginosa proteins, we screened for Th17-stimulating antigens by coculturing the library proteins with splenocytes from mice immunized with a live-attenuated P. aeruginosa vaccine that is protective via Th17-based immunity. We measured antibody and Th17 responses after intranasal immunization of mice with the purified proteins mixed with the Th17 adjuvant curdlan, and we tested the protective efficacy of vaccination in a murine model of acute pneumonia. Measurements and Main Results: The proteins PopB, FpvA, FptA, OprL, and PilQ elicited strong IL-17 secretion in the screen, and purified versions of PopB, FpvA, and OprL stimulated high IL-17 production from immune splenocytes. Immunization with PopB, which is a highly conserved component of the type III secretion system and a known virulence factor, elicited Th17 responses and also enhanced clearance of P. aeruginosa from the lung and spleen after challenge. PopB-immunized mice were protected from lethal pneumonia in an antibody-independent, IL-17-dependent manner. Conclusions: Screening for Th17-stimulating protein antigens identified PopB as a novel and promising vaccine candidate for P. aeruginosa.Keywords: vaccine; pneumonia; Th17; IL-17; Pseudomonas aeruginosa Pseudomonas aeruginosa is a major cause of serious, often antibiotic-resistant, lung infections in humans, particularly in patients receiving mechanical ventilation and those with cystic fibrosis (1, 2). Most P. aeruginosa vaccines developed to date, including those based on the LPS O antigen (3), the outer membrane proteins F and I (4, 5), or the type III secretion system component PcrV (6), have relied on conventional protective mechanisms-namely, antibody-mediated opsonophagocytic killing and/or antibody-mediated toxin inhibition. Although LPS O antigen-based vaccines can mediate high levels of immunity to P. aeruginosa, the protection is limited to strains having the same LPS serogroup (3). The failure of the Federal Hyperimmune Immunoglobulin Trial (7), which found no benefit in critically ill adults passively immunized with P. aeruginosa LPS O antigenspecific IgG, perhaps best illustrates that antibody-mediated protective mechanisms are not sufficient.Th17 cells have recently been shown to mediate antibodyindependent host defense against Klebsiella pneumoniae (8), although the bacterial proteins recognized by the Th17 cells in those studies were not fully characterized. In our own evaluations of live-attenuated P. ...

Linocin and OmpW Are Involved in Attachment of the Cystic Fibrosis-Associated Pathogen Burkholderia cepacia Complex to Lung Epithelial Cells and Protect Mice against Infection

“…While the cell-mediated responses required to protect against Bcc have not been studied to date, the stimulation of IL-17A by both antigens is likely to contribute to protection against Bcc infection by these antigens, which have both an extracellular and an intracellular component to their lifestyle (27,(56)(57)(58)(59)(60). IL-17-stimulating antigens were recently shown to confer protection against P. aeruginosa in immunization studies (61). Although further studies, such as immunization and protection of another animal species, toxicity studies, and clinical trials, on the potential of these two antigens as vaccine candidates to protect or treat CF patients and other susceptible populations against Bcc infection need to be carried out, the current data indicate that the two antigens OmpW and linocin show promise as protective antigens.…”

PrtR Homeostasis Contributes to Pseudomonas aeruginosa Pathogenesis and Resistance against Ciprofloxacin

“…Female BALB/c mice (6 to 8 weeks old) were anesthetized with an intraperitoneal injection of 7.5% chloral hydrate (100 l per mouse). Anesthetized mice were intranasally inoculated with 10 l of bacterial suspension in each nostril, giving a total infection volume of 20 l. Bacterial colonization in the lung was determined as previously described (16). Briefly, 16 h postinfection, mice were sacrificed by inhalation of CO 2 .…”

“…To examine bacterial gene expression levels during infection, mice were sacrificed by CO 2 at various time points postinfection. Bronchoalveolar lavage fluid (BALF) was obtained by cannulation of the trachea followed by two instillations of 1 ml sterile PBS with 0.5 mM EDTA (16). Two hundred microliters of the BALF was used for bacterial counting, while the remaining BALF was centrifuged and the pellets were immediately resuspended in 200 l TRIzol reagent (Invitrogen).…”

Microbiota-Driven Immune Cellular Maturation Is Essential for Antibody-Mediated Adaptive Immunity to Staphylococcus aureus Infection in the Eye

“…It has become increasingly apparent that endogenous innate immune factors can also be important components for efficacy of acquired immune effectors such as antibodies (28,29), notably as cofactors in vaccine-induced immunity in the lung (30)(31)(32). We therefore evaluated the role of T cells and the IL-17/IL-22 pathways in antibody-mediated reductions in corneal pathology and bacterial levels in S. aureus-infected eyes.…”