Biofilm Formation on Dental Implant Biomaterials by Staphylococcus aureus Strains Isolated from Patients with Cystic Fibrosis

Abstract

Implants made of ceramic and metallic elements, which are used in dentistry, may either promote or hinder the colonization and adhesion of bacteria to the surface of the biomaterial to varying degrees. The increased interest in the use of dental implants, especially in patients with chronic systemic diseases such as cystic fibrosis (CF), is caused by an increase in disease complications. In this study, we evaluated the differences in the in vitro biofilm formation on the surface of biomaterials commonly used in dentistry (Ti-6Al-4V, cobalt-chromium alloy (CoCr), and zirconia) by Staphylococcus aureus isolated from patients with CF. We demonstrated that S. aureus adherence and growth depends on the type of material used and its surface topography. Weaker bacterial biofilm formation was observed on zirconia surfaces compared to titanium and cobalt-chromium alloy surfaces. Moreover, scanning electron microscopy showed clear differences in bacterial aggregation, depending on the type of biomaterial used. Over the past several decades, S. aureus strains have developed several mechanisms of resistance, especially in patients on chronic antibiotic treatment such as CF. Therefore, the selection of an appropriate implant biomaterial with limited microorganism adhesion characteristics can affect the occurrence and progression of oral cavity infections, particularly in patients with chronic systemic diseases.

Keywords: AFM; CoCr alloy; SEM; Staphylococcus aureus; Ti-6Al-4V; biofilm; biomaterials; cystic fibrosis; zirconia.

Figure 1

Biofilm-forming capacity of S. aureus clinical strains. The strains of S. aureus were assessed for their ability to form a biofilm and accordingly classified as strong, moderate, weak, and non-producers. Results are expressed as the mean value ± standard deviations of the mean of at least 5 independent experiments performed in triplicate. To classify the strains into groups that showed significant differences, statistical analysis was performed using Student’s t-test (* p < 0.05). The isolate number 3 that formed high levels of biofilm is indicated (◆).

Figure 2

Biofilm formation (in colony-forming units (log CFU/mL-1)) on the surfaces of different biomaterials. Error bars represent the pooled standard deviations of the mean (n = 5). The level of significance was preset at * p = 0.05. The mean and standard deviation are shown.

Figure 3

(A) Scanning electron microscopy (SEM) micrographs. Representative SEM images of the different biomaterial specimens. In all cases, the examined disks generally exhibited a smooth surface topography, with some fine polishing marks homogeneously distributed over the surface. (a), Ti-6Al-4V (grade 5 titanium); (b) zirconium dioxide (yttria-stabilized tetragonal zirconia polycrystals-3Y-TZP); (c) cobalt-chromium (CoCr) alloy (Duceralloy C). Original magnification ×1000 (Scale bar = 10 μm). (B) AFM micrographs show the surface topography of the tested biomaterials (a), Ti-6Al-4V; (b) zirconium dioxide; (c) cobalt-chromium (CoCr) alloy. Calibrated to 225 μm2 sample surface. (C) 3D atomic force microscopy (AFM) images for (a) Ti-6Al-4V; (b) zirconium dioxide; (c) cobalt-chromium (CoCr) alloy.

Figure 4

(A) Scanning electron microscopy images (a–c) of biofilms on different biomaterials. (a), Ti-6Al-4V (grade 5 titanium); (b) zirconium dioxide; (c) cobalt-chromium (CoCr) alloy (Duceralloy C). (B) Fluorescence microscopy images (a–c), after staining with a LIVE/DEAD BacLight kit on different biomaterials. (a) Ti-6Al-4V; (b) zirconium dioxide; (c) CoCr alloy (Duceralloy C). Original magnification ×10,000 (Scale bar = 1 μm).