Interaction of outer membrane vesicles of the oral pathogen Porphyromonas gingivalis with human red blood cells disclosed by proteome analyses

Porphyromonas gingivalis is a keystone oral pathogen implicated in periodontitis and several other pro-inflammatory or degenerative diseases. Previous studies showed that outer membrane vesicles (OMVs) produced by this Gram-negative bacterium are loaded with virulence factors, including the P. gingivalis peptidyl-arginine deiminase (PPAD). However, our understanding of strain-specific OMV features and the impact of PPAD on OMV cargo composition was so far limited. Therefore, the present study was aimed at comparing OMV proteomes of six different P. gingivalis strains, including PPAD-proficient and -deficient variants of the type strains W83 and ATCC 33277, and two clinical isolates. Per strain, 100–203 OMV-borne proteins were identified, originating mostly from the bacterial periplasm and outer membrane. The OMV core proteome included 79 orthogroups representing 11 biological pathways. Interestingly, 23 identified OMV cargo proteins were related to the type IX secretion system (T9SS) for export of virulence factors, invariably including all four components of the so-called T9SS attachment complex. PPAD-deficiency altered the OMV protein content by ∼18% compared to OMVs from the respective PPAD-proficient strains. Lastly, the investigated OMVs invariably carried multiple proteins involved in iron and heme acquisition. Accordingly, we demonstrate for the first time that P. gingivalis OMVs are highly potent inducers of hemagglutination and to lesser extent hemolysis of human erythrocytes. In conclusion, our present proteome analyses disclose interactions of P. gingivalis OMVs with human erythrocytes related to heme and iron acquisition, and they suggest that attachment to erythrocytes could represent a plausible mechanism for OMV dissemination throughout the human body.