# In vitro evaluation of biofilm formation by Streptococcus mutans and Candida albicans in orthodontic aligners

## TL;DR

This study shows that orthodontic aligners can accumulate harmful biofilms that may lead to tooth decay.

## Contribution

The study experimentally demonstrates biofilm formation on orthodontic aligners by S. mutans and C. albicans.

## Key findings

- Monospecies and mixed biofilms of S. mutans and C. albicans formed robustly on Invisalign® aligners.
- Biofilm accumulation could increase the risk of carious lesions in users.
- Proper cleaning of aligners is essential to prevent microbial buildup.

## Abstract

Aligners have been used by the orthodontic community for approximately 20 years, but little research has been carried out on the accumulation of biofilm on the surface of these aligners, as well as their possible impact on the oral ecosystem.

Ten hemi-arches of Invisalign® brand orthodontic aligners were used. The hemi-arches were placed inside sterile flasks containing 25 mL of Gibbons and Nygaard broth, with standardized suspensions of the two microorganisms on the 0.5 MacFarland scale and incubated in aerophily (C. albicans) and microaerophily (S. mutans and mixed biofilm) at 37°C for 72h. The biofilm formed was removed by the multiple rinses method to quantify the microorganisms in the biofilms in CFU/mL. A qualitative analysis with scanning electron microscopy was performed to observe the structure of the formed biofilms.

It was observed the accumulation of a monospecies biofilm (S. mutans - 2.55 x 106 and C. albicans - 1.62 x 107) and mixed biofilm (S. mutans - 2.21 x 105 and C. albicans - 1.06 x 107) very robust on the surface of orthodontic aligners.

According to the results obtained, one can conclude that Invisalign® brand orthodontic aligners are susceptible to the accumulation of monospecies and mixed biofilms of S. mutans and C. albicans. Therefore, it is necessary to consider the possibility of Invisalign® users developing carious lesions associated with the biofilm formed by these two microbial species, and to guide the patient on the correct cleaning of the device during treatment.