Comparison of chelating agent, soap, and antibiotic irrigation in removing adherent bacteria from orthopaedic implants

Introduction: Adherent bacteria on orthopaedic implants are difficult to eradicate despite irrigation and treatment with IV antibiotics. This has led to the use of other irrigation solutions including surfactants (castile soap), antiseptics (povidine, chlorhexidine, hydrogen peroxide) and antibiotics (bacitracin, neomycin, and polymyxin). The mechanism of bacteria adhesion offers another yet unexplored strategy for irrigation. Bacterial adhesion ligands require calcium. Calcium removal has been shown to interfere with adhesion to surfaces(1). Thus an agent that can bind calcium ions may disrupt bacterial adhesion. Such agents include chelators, organic compounds that bind to and inactivate metal ions. Ethlyenediaminetetraacetic acid (EDTA) is a chelator that binds calcium and iron ions. EDTA has multiple uses that indicate its relative nontoxicity: administered by IV it binds lead for treatment of lead poisoning, for irrigation of infected root canals in dentistry, and in ophthalmic solutions. Non-orthopedic literature suggests EDTA may work well as irrigant in infections involving hardware. Currently, no studies have investigated EDTA as a potential surgical irrigation solution. This study compares the efficacy of EDTA, soap, antibiotics and normal saline as irrigation solution in removing adherent bacteria from metal hardware. Materials and Methods: Stainless steel AO cortical screws of 3.5 mm diameter and 20 mm length were incubated with staphylococcus epidermidis (s. epi) strain RP62A (ATCC 35984). This strain is a high-slime producing strain that has been extensively characterized and used in other studies of bacterial adhesion. The incubation was performed with a standardized inoculum of s. epi in a total volume of 5 ml trypticase soy broth (TSB), resulting in a 2 x 10^7 CFU inoculum for each screw. Five irrigation solutions were tested: 1% EDTA (Fisher scientific), 5% EDTA, 1% soap solution (Huntington Episoft, contains sodium olefin sulfonate (SOS)), 0.5% neomycin and normal saline. 1% soap was prepared by injecting 10ml of Huntington Episoft hand soap into 1 L of sterile normal saline. All irrigation solutions had normal saline as base. Screws in TSB were incubated at 35°C for 18 hours in ambient air. After overnight incubation, TSB was removed and each screw was placed into a new 50 mL, sterile, polypropylene conical tube and irrigated with each solution using low pressure pulsatile lavage (Surgilav plus, Stryker). After irrigation, each screw was placed into 5 mL sterile normal saline and sonicated at 35 kHz to remove residual bacteria(Gen probe, Lab Line Instruments Inc, Melrose, Illinois) for 10 minutes at 37°C, and then vortexed for 10 seconds. Serial 10-fold dilutions were made from the saline and 25 μl of each dilution was plated onto Columbia agar with 5% sheep blood for colony counts. Results: All samples in the soap group had no residual bacterial growth while all other groups had CFU/mL in the thousands. ANOVA on the logs of the results shows a difference among groups with p<0.000002. Analysis using Tukey’s shows significant difference between soap and every other treatment group. However, no difference existed among saline, EDTA or neomycin. Also, there was no difference between the different concentrations of EDTA. Residual bacterial count (Colony forming units per mL) Differences among treatment groups