Basic Science Focus Forum

Wed., 10/12/11 BSFF: Biomaterials & Surface Tech, Paper #20, 5:03 pm OTA-2011

Testing for Contrasting Infection Rates and Biofilm Formation Among Biomaterials in an In Vitro Biofilm Infection Model

Brian Stover, BS1; Zalman Vaksman, MS1; Heidi B. Kaplan, PhD1; Catherine G. Ambrose, PhD2; Milan K. Sen, MD2
1Dept. of Microbiology and Molecular Genetics,
2Department of Orthopaedic Surgery, UT Health, Houston, Texas, USA

Purpose: This study presents an in vitro model for biofilm formation on orthopaedic biomaterials. Several materials have been evaluated to determine if Staphylococcus aureus biofilm growth rates vary among the materials.

Methods: An in vitro S. aureus biofilm model adapted from one previously developed in our laboratory was used to study biofilm formation on different biomaterials. Materials tested in this study included stainless steel, TMZF alloy, and grades 2 and 5 titanium treated with type II and III anodization, which were in the form of 10-mm disks that were sterilized and then coated with serum proteins prior to use. A clinical isolate of S. aureus obtained from a human osteomyelitis infection was used to evaluate the rate of biofilm growth on the different materials. Disks of each material were incubated statically for 24 hours at 37°C, with 1 × 106 S. aureus cells in a synthetic interstitial fluid (SIF) consisting of physiologic concentrations of salt, glucose, hyaluronic acid, and fetal bovine serum. On each subsequent day the SIF was carefully removed and replaced with fresh, sterile SIF. For analysis on days 1 through 7 the biofilms on each disk to be examined were stained using a LIVE/DEAD BacLight Bacterial Viability Kit (Invitrogen) and imaged with a Zeiss LSM 510 Meta confocal laser scanning microscope. The data were then analyzed to compare the biofilm growth on the different materials by using the nPHLIP 2.0 software previously developed in our lab. Polymethylmethacrylate (PMMA) disks were tested along with the metals and used as controls. For each time point and material tested, a total of 4 disks were examined in 2 separate experiments.

Results: All results were normalized by dividing the total biovolume on each disk by the total biovolume on the control disks on the day of maximum growth (day 6). On days 2 through 6, there were statistically significant differences between the biovolumes on the disks. For each of these days, the biofilm biovolumes on the stainless steel and TMZF alloy disks were significantly greater than those on the grade 2 and grade 5 titanium and PMMA disks. At no time point was there a significant difference between biofilm growth on the grade 2 and grade 5 titanium disks.

Conclusion: S. aureus biofilms were detected on all materials tested. The biovolume of live cells within the biofilms was significantly greater on the stainless steel and TMZF alloy materials, indicating that the grade 2 and grade 5 titanium materials might be more resistant to implant-associated infections. The type of anodization was not found in this study to result in significantly different biofilm growth rates.

Alphabetical Disclosure Listing (628K PDF)

• The FDA has not cleared this drug and/or medical device for the use described in this presentation   (i.e., the drug or medical device is being discussed for an “off label” use).  ◆FDA information not available at time of printing. Δ OTA Grant.