Staphyloccocus aureus and Staphyloccocus epidermidis: Biofilm development on different cobalt-chrome alloys for implant materials in orthopedy

Objectives Peri-prosthetic infections (PIs), caused by the adhesion of bacteria to medical implant materials, are among the most serious complications which can occur following joint replacement surgeries. The bacteria form biofilms on the surfaces, triggering inflammatory reactions in the tissue surrounding the periprosthetic area. Those infectious reactions occur mostly within the first two prosthetic years and can results in the rejection of the implant or, in the worst case, in amputations. The most common pathogens in this formation are coagulase-negative Staphylococcus spp. (up to 43%), Staphylococcus aureus (up to 23%), Streptococcus spp. (9-10%), as well as gram-negative rods or Enterococci. The basic aim of the here presented work was to understand which casting alloys have the worst properties for the adhesion of infectious microorganisms - specifically S. aureus and S. epidermidis. For this purpose, six different cobalt-chromium-molybdenum (CoCrMo) alloys were tested: CoCrMo, “CoCrMo TiN (titanium nitride)”, “CoCrMo porous coated”, “CoCrMo cpTi (commercially pure titanium)”, “CoCrMo cpTi + TCP (tricalcium phosphate) and “CoCrMo highly-polished.”

Methods For biofilm preparation, 24 well plates were prepared by filling them with four platelets per alloy type. After adding 1.5 mL of the respective cell suspension (S. aureus or S. epidermidis in a cell density of McFarland 0.5) into each well, the well plates were sealed with a breathable membrane and incubated on the shaker at 37°C and 90 RPM for 48 hours. After incubation the biofilms were collected by scraping them of the platelet surfaces. The protein and polysaccharide contents were measured using the Pierce BCA protein kit (Thermo Fisher Scientific) and the phenol-sulfuric-aid method, respectively. The LIVE/DEAD BacLight Bacterial Viability Kit (Thermo Fisher Scientific) was used in a flow cytometric assay for the determination of cell number as well as cell viability. R studio was used for statistical analysis.

Results and Conclusion The smooth surfaces (TiN and polished) showed the lowest values in both protein and polysaccharide measurements. Interestingly CoCrMo displayed an increased quantity for both proteins and polysaccharides compared to the modified, smooth plates. In the protein measurement, the rough plates showed twice the µg value per mL after just 48 hours, TCP as much as 2.5 times. The polysaccharides did not show any surprises either, with the highest average value in cpTi. Flow cytometry data let to the conclusion that within the 48 hours growth time most bacteria survived on the surfaces and that the cell numbers go hand in hand with the protein and polysaccharide contents of the biofilms.

In conclusion smoother alloy coatings might be the best choice when considering material for an implant when only looking at bacterial infections. Genetic analysis of the biofilms will be performed in a follow up study and should give more insight in the developmental stage of the biofilm on the different alloy types.

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Publication History

Article published online:
15 October 2020