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. ...
Pseudomonas aeruginosa strain PA14 is a multi-host pathogen that infects plants, nematodes, insects, and vertebrates. Many PA14 factors are required for virulence in more than one of these hosts. Noting that plants have a fundamentally different cellular architecture from animals, we sought to identify PA14 factors that are specifically required for plant pathogenesis. We show that synthesis by PA14 of the disaccharide trehalose is required for pathogenesis in Arabidopsis, but not in nematodes, insects, or mice. In-frame deletion of two closely-linked predicted trehalose biosynthetic operons, treYZ and treS, decreased growth in Arabidopsis leaves about 50 fold. Exogenously co-inoculated trehalose, ammonium, or nitrate, but not glucose, sulfate, or phosphate suppressed the phenotype of the double ΔtreYZΔtreS mutant. Exogenous trehalose or ammonium nitrate does not suppress the growth defect of the double ΔtreYZΔtreS mutant by suppressing the plant defense response. Trehalose also does not function intracellularly in P. aeruginosa to ameliorate a variety of stresses, but most likely functions extracellularly, because wild-type PA14 rescued the in vivo growth defect of the ΔtreYZΔtreS in trans. Surprisingly, the growth defect of the double ΔtreYZΔtreS double mutant was suppressed by various Arabidopsis cell wall mutants that affect xyloglucan synthesis, including an xxt1xxt2 double mutant that completely lacks xyloglucan, even though xyloglucan mutants are not more susceptible to pathogens and respond like wild-type plants to immune elicitors. An explanation of our data is that trehalose functions to promote the acquisition of nitrogen-containing nutrients in a process that involves the xyloglucan component of the plant cell wall, thereby allowing P. aeruginosa to replicate in the intercellular spaces in a leaf. This work shows how P. aeruginosa, a multi-host opportunistic pathogen, has repurposed a highly conserved “house-keeping” anabolic pathway (trehalose biosynthesis) as a potent virulence factor that allows it to replicate in the intercellular environment of a leaf.
Solitary Peutz-Jeghers type polyps are characterized by a hamartomatous polyp of the gastrointestinal (GI) tract in a patient without mucocutaneous pigmentation, family history of Peutz-Jeghers syndrome, or STK11/LKB1 mutations. Histologically identical to the polyps in Peutz-Jeghers syndrome, these sporadic polyps can arise anywhere along the GI tract, with typical arborizing smooth muscles extending from the muscularis mucosa. While the lining mucosa is generally the same as the organ in which it arises, gastric pyloric and osseous metaplasia have been reported in intestinal polyps in Peutz-Jeghers syndrome. Herein, the authors report the first case of a small intestinal solitary Peutz-Jeghers type polyp with gastric antral and fundic gland lining mucosa. A 43-year-old male was admitted for small bowel obstruction. Diagnostic laparoscopy revealed jejuno-jejunal intussusception with an associated polyp measuring 7.2 cm. Histological examination showed a hamartomatous polyp with arborizing smooth muscle bundles extending from the muscularis mucosae. The polyp was lined by non-dysplastic gastric antral and fundic gland mucosa, and was sharply demarcated from the adjacent non-polypoid intestinal mucosa. Colonoscopy, esophagogastroduodenoscopy and small bowel enteroscopy revealed no additional polyps or masses. Thorough investigation of the patient's family history was negative for Peutz-Jeghers syndrome or mucocutaneous pigmentation. Molecular analysis of the lesion was negative for STK11/LKB1 mutations. A diagnosis of solitary Peutz-Jeghers type polyp of the small bowel with gastric antral and fundic gland mucosal lining was rendered.
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