Interspecies Inhibition ofPorphyromonas gingivalisby Yogurt-DerivedLactobacillus delbrueckiiRequires Active Pyruvate Oxidase
ABSTRACTDespite a growing interest in using probiotic microorganisms to prevent disease, the mechanisms by which probiotics exert their action require further investigation.Porphyromonas gingivalisis an important pathogen implicated in the development of periodontitis. We isolated several strains ofLactobacillus delbrueckiifrom dairy products and examined their ability to inhibitP. gingivalisgrowthin vitro. We observed strain-specific inhibition ofP. gingivalisgrowthin vitro. Whole-genome sequencing of inhibitory and noninhibitory strains ofL. delbrueckiirevealed significant genetic differences supporting the strain specificity of the interaction. Extracts of theL. delbrueckiiSTYM1 inhibitory strain contain inhibitory activity that is abolished by treatment with heat, proteinase K, catalase, and sodium sulfite. We purified the inhibitory protein(s) fromL. delbrueckiiSTYM1 extracts using ammonium sulfate precipitation, anion-exchange chromatography, and gel filtration chromatography. Pyruvate oxidase was highly enriched in the purified samples. Lastly, we showed that purified, catalytically active, recombinant pyruvate oxidase is sufficient to inhibitP. gingivalisgrowthin vitrowithout the addition of cofactors. Further, using a saturated transposon library, we isolated transposon mutants ofP. gingivalisin thefeoB2(PG_1294) gene that are resistant to killing by inhibitoryL. delbrueckii, consistent with a mechanism of hydrogen peroxide production by pyruvate oxidase. Our results support the current understanding of the importance of strain selection, not simply species selection, in microbial interactions. SpecificL. delbrueckiistrains or their products may be effective in the treatment and prevention ofP. gingivalis-associated periodontal disease.IMPORTANCEP. gingivalisis implicated in the onset and progression of periodontal disease and associated with some systemic diseases. Probiotic bacteria represent an attractive preventative therapy for periodontal disease. However, the efficacy of probiotic bacteria can be variable between studies. Our data support the known importance of selecting particular strains of bacteria for probiotic use, not simply a single species. Specifically, in the context of probiotic intervention of periodontitis, our data suggest that high-level expression of pyruvate oxidase with hydrogen peroxide production inL. delbrueckiicould be an important characteristic for the design of a probiotic supplement or a microbial therapeutic.