Nontypeable Haemophilus influenzae Redox Recycling of Protein Thiols Promotes Resistance to Oxidative Killing and Bacterial Survival in Biofilms in a Smoke-Related Infection Model

Academic Article

Abstract

  • Smoke exposure is a risk factor for community-acquired pneumonia, which is typically caused by host-adapted airway opportunists like nontypeable Haemophilus influenzae (NTHi). Genomic analyses of NTHi revealed homologs of enzymes with predicted roles in reduction of protein thiols, which can have key roles in oxidant resistance. Using a clinical NTHi isolate (NTHi 7P49H1), we generated isogenic mutants in which homologs of glutathione reductase (open reading frame NTHI 0251), thioredoxin-dependent thiol peroxidase (NTHI 0361), thiol peroxidase (NTHI 0907), thioredoxin reductase (NTHI 1327), and glutaredoxin/peroxiredoxin (NTHI 0705) were insertionally inactivated. Bacterial protein analyses revealed that protein oxidation after hydrogen peroxide treatment was elevated in all the mutant strains. Similarly, each of these mutants was less resistant to oxidative killing than the parental strain; these phenotypes were reversed by genetic complementation. Analysis of biofilm communities formed by the parental and mutant strains showed reduction in overall biofilm thickness and density and significant sensitization of bacteria within the biofilm structure to oxidative killing. Experimental respiratory infection of smoke-exposed mice with NTHi 7P49H1 showed significantly increased bacterial counts compared to control mice. Immunofluorescent staining of lung tissues showed NTHi communities on lung mucosae, interspersed with neutrophil extracellular traps; these bacteria had transcript profiles consistent with NTHi biofilms. In contrast, infection with the panel of NTHi mutants showed a significant decrease in bacterial load. Comparable results were observed in bactericidal assays with neutrophil extracellular traps in vitro. Thus, we conclude that thiol-mediated redox homeostasis is a determinant of persistence of NTHi within biofilm communities.
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    Author List

  • Hunt BC; Xu X; Gaggar A; Swords WE
  • Volume

  • 7
  • Issue

  • 1