Biofilm on Toothbrushes of Children with Cystic Fibrosis: A Potential Source of Lung Re-Infection after Antibiotic Treatment?

Abstract

Frequent recurrent lung infections result in irreversible lung damage in children with cystic fibrosis (CF). This study aimed to determine if toothbrushes contain biofilms of pathogens, and act as potential reservoirs for lung re-infection following antibiotic treatment of acute exacerbations. Toothbrushes were collected from children with CF of lung infection before, during and after antibiotic treatment. Toothbrushes were rinsed with sterile saline and cultured. Bacterial isolates from toothbrushes were identified by 16s rRNA gene sequencing and compared with isolates from a sputum sample of the same patient. Scanning electron microscopy (SEM) was used to visually confirm the presence of bacterial biofilms and confocal laser scanning microscopy (CLSM) combined with Live/Dead stain to confirm bacterial viability. Large numbers of bacteria and biofilms were present on all toothbrushes. SEM confirmed the presence of biofilms and CLSM confirmed bacterial viability on all toothbrushes. Pathogens identified on toothbrushes from children before and during antibiotics treatment were in concordance with the species found in sputum samples. Pseudomonas aeruginosa and Staphylococcus aureus was able to be cultured from children's toothbrushes despite antibiotic treatment. Toothbrushes were shown to be contaminated with viable pathogens and biofilms before and during antibiotic treatment and could be a potential source of lung re-infections.

Keywords: antibiotic treatment; biofilm; cystic fibrosis; recurrent lung infection; toothbrush.

Figure 1

Confocal laser scanning microscopy image of toothbrush bristles with bacteria stained with LIVE/DEAD^® BacLight™ Bacterial Viability Kit. Live bacteria are stained green and dead bacteria are stained red. (a) Toothbrush from a child with CF before antibiotic treatment; (b) toothbrush from a child with CF receiving antibiotic treatment.

Figure 2

Scanning electron microscopy images of toothbrushes obtained from children with active disease before antibiotic treatment showing large numbers of bacteria both attached to and encased in extracellular polymeric substances (EPS) on toothbrush bristles. (a) Low power magnified ×1000; (b) high power magnified ×5000.

Figure 3

Scanning electron microscopy images of toothbrushes obtained from children receiving antibiotics treatment showing thick EPS with little surface bacteria, suggesting that bacteria are completely encased in thick EPS. (a) Low power magnified ×1000; (b) high power magnified ×5000.