Association of oral and non-oral microorganisms in a multispecies biofilm model formed on orthodontic appliances: an in vitro study.

Orthodontic devices can promote dental biofilm formation and increase the risk of oral diseases when hygiene is inadequate. Invisible aligners, although removable and easier to clean, are frequently exposed to external environments, which may allow contamination by non-oral microorganisms. This study aimed to evaluate the association and viability of oral and non-oral microorganisms in mono- and multispecies biofilm models formed on orthodontic appliances.

Methods

Stimulated human saliva was used to form the acquired pellicle on specimens of invisible aligners, conventional brackets, and self-ligating brackets. The samples were exposed to Streptococcus mutans, Candida albicans, Staphylococcus aureus, and Pseudomonas aeruginosa, and incubated for 48 h under mono- and multispecies conditions. After ultrasonic detachment, viable cells were quantified by CFU/mL enumeration, and extracellular polymeric substance (EPS) content was determined. Statistical analysis was performed using one-way ANOVA followed by Tukey's post hoc test for monospecies biofilms, and two-way ANOVA followed by Bonferroni's post hoc test for multispecies biofilms (p < 0.05). Higher CFU/mL values were observed for S. aureus and P. aeruginosa compared to C. albicans and S. mutans (p < 0.05). In multispecies biofilms, C. albicans showed higher values than S. mutans (p < 0.05). CFU/mL increased over time, with no differences among orthodontic devices (p > 0.05). EPS content was higher in conventional brackets, intermediate in self-ligating brackets, and lower in aligners (p < 0.05). Oral and non-oral microorganisms remained viable and were recovered from biofilm models formed on orthodontic appliances. These findings suggest that such devices may act as substrates for the persistence of non-oral microorganisms under biofilm-forming conditions, highlighting the importance of appropriate hygiene and storage practices.