Exploiting the Biofilm Microenvironment to Target Cariogenic Bacteria Using Nanocatalysts

Objectives: To investigate whether a catalytic approach using glucose oxidase (GOx) to enhance H₂O₂ production can boost commensals (Streptococcus oralis) ability to fight Streptococcus mutans in sugar-rich conditions, together with iron oxide nanoparticles (IONPs) that catalyze H₂O₂ to liberate free radicals capable of killing the pathogen and degrading the exopolysaccharides (EPS) matrix, disrupting cariogenic biofilms.
Methods: Dextran coated IONPs were synthesized by co-precipitation of Fe(III) and Fe(II) in the presence of dextran, then GOx was conjugated to dextran aldehyde on the surface of IONPs. The sequential catalysis of IONPs-GOx to yield H₂O₂ followed by free radicals production was tested using the 3,3’,5,5’-tetramethylbenzidine (TMB) assay. S. mutans-S. oralis mixed-species biofilms were formed on saliva-coated hydroxyapatite discs. The cytotoxicity, biofilm uptake and anti-biofilm activity of IONPs-GOx were characterized using biochemical and microbiological methods. The dynamics of biofilm disruption after treatments with IONPs-GOx were analyzed via time-lapse confocal microscopy.
Results: IONPs-GOx have high catalytic efficiency at acidic pH 4.5, but low activity at pH 6.5, resulting in potent S. mutans killing (〉 than 8 log killing vs control, p 〈 0.01), while sparing S. oralis cell viability within biofilms. Furthermore, the EPS was efficiently degraded (73.4% reduction vs control, p 〈 0.05). High-resolution confocal images further confirmed specific inhibition of S. mutans and degradation of EPS-matrix after treatment with IONPs-GOx, altering the biofilm 3D architecture (vs. control). Further mechanistic studies revealed that IONPs-GOx bound preferentially to S. mutans cells compared to S. oralis (60% higher vs S. oralis), which may explain in part of the cause of enhanced pathogen killing vs the commensals.
Conclusions: Our results indicate that a nanocatalytic approach via GOx conjugated onto IONPs increased localized H₂O₂ and free radical production in a pH-dependent manner, which targeted and suppressed cariogenic bacteria accumulation under sugar-rich conditions, enhancing S. oralis fitness within mixed-biofilms.