Background Rothia dentocariosa is an opportunistic pathogen found in the oral cavity and is found to be involved in many oral infections as it has the ability to attach to tooth and mucosal surfaces, produce substantial amounts of acids and integrate into dental plaque biofilms. Aim To analyze the proteome of R. dentocariosa by using bioinformatics tools and to investigate the proinflammatory potential of R. dentocariosa. Material and Methods Protein sequences of R. dentocariosa were downloaded from NCBI and various in silico analyses were performed using bioinformatics tools. R. dentocariosa CCUG 35437 was grown on blood agar in 5%CO2 in air at 37 C for 2 days. Biofilms were cultured for 2 days and quantified by crystal violet staining. Human whole blood was stimulated with biofilms, biofilm-supernatants, planktonic cells, and whole cells. Proteome Profiler and ELISA based quantification of cytokines was performed for the samples. Results In silico analysis of the whole genome and proteome of R. dentocariosa revealed a number of proteins predicted to be potentially secreted but also possess virulence properties. R. dentocariosa was able to form only moderate biofilms. The ability of R. dentocariosa to induce different cytokines varied depending on the stimulant being used. Biofilms and planktonic cultures induced specific cytokines that were not induced by whole cells or biofilm-supernatants. While IL-8 was induced at near-similar levels from biofilm and planktonic cells, IL-10 was induced at significantly higher levels (P<0.05) only by the planktonic cultures. The biofilm-supernatant and the whole cell stimulants induced lower levels of cytokines than biofilm and planktonic cultures. Conclusion Identification of potential virulence factors predicted to be secreted extracellularly may suggest a key role for R. dentocariosa in oral and non-oral infections. Different stimulants from R. dentocariosa showed varying potential to induce cytokines from human whole blood. This may suggest differences in the composition/concentration of the bacterial components in the stimulants, with varying abilities to induce cytokine production, may be the reason for the observed differences.