Effects of magnitude and duration of cathodic voltage-controlled electrical stimulation of titanium for prevention of biofilm infections

Abstract

Event Abstract Back to Event Effects of magnitude and duration of cathodic voltage-controlled electrical stimulation of titanium for prevention of biofilm infections Mary Canty1*, Anthony Campagnari2* and Mark Ehrensberger1, 3* 1 SUNY at Buffalo, Biomedical Engineering, United States 2 SUNY at Buffalo, Microbiology and Immunology, United States 3 SUNY at Buffalo, Orthopaedics Surgery, United States Introduction: Periprosthetic joint infection (PJI) is a devastating outcome of total joint arthroplasty (TJA). Gram-positive methicillin-resistant Staphylococcus aureus (NRS70) and gram-negative Acinetobacter baumanni (Ab307) develop biofilms on the surface of titanium implants. This study evaluated the effects of applying cathodic voltage-controlled electrical stimulations (CVCES) of -1.5V and -1.8V for 2, 4, and 8hrs to commercially pure titanium (cpTi) incubated in NRS70 and Ab307 as a method of preventing bacterial attachment to the surface. Materials and Methods: cpTi coupons were wet sanded to a 600-grit finish, sonicated in deionized water for 10mins, and sterilized under ultraviolet light for 30mins. Experiments were conducted with NRS70 (respiratory isolate) and Ab307 (blood isolate). Fresh cultures of NRS70 were incubated in 5mL of tryptic soy broth with 0.25% glucose (TSBG). This was repeated for Ab307 in Muller Hinton (MH) broth. Solutions were adjusted to an OD600 of 0.1 (~107 CFU/mL) and placed in a shaking water bath at 180rpm and 37°C. Cultures were grown to mid-log, diluted to an OD600 of 0.1, and then 2.75mL of this solution was added to a sterile cpTi coupon in a custom stimulation chamber[1]. The agar chamber uses a standard three-electrode system with a cpTi working electrode, a graphite counter electrode and an Ag/AgCl reference electrode. With respect to Ag/AgCl, voltages of -1.8V for 2, 4 and 8hrs, and -1.5V for 4 and 8hrs were applied to the working electrode at room temperature using a potentiostat (Interface 1000, Gamry Instruments). Control experiments were conducted at open circuit potential (OCP). After incubation, cpTi coupons were washed in sterile PBS, sonicated and dilution plated for colony forming units (CFU). The media was collected for pH measurement and dilution plated for CFU. Potentiostatic EIS was performed on cpTi in sterile media for each CVCES condition. EIS Impedance results were fit to a modified Randles circuit (EIS 300 software, Gamry Instruments) with a complex-non-linear-least-squares method. The media pH was assessed following EIS. Four independent samples were evaluated in all conditions. For 4 and 8hrs, a one-way ANOVA followed by Tukey post-hoc compared log-transformed CFU, Rp, and C across CVCES conditions. If the Levene’s test was significant, a Welch’s test followed by a Games-Howell post-hoc test was performed. Student’s t-tests compared the log-transformed CFU, Rp, and C for 2hr CVCES. Results and Discussion: As compared to OCP, CVCES for 4hrs at -1.8V significantly reduced coupon-associated NRS70 and Ab307 CFU by 99.9% (p=0.029 and p<0.001). It also completely eradicated all planktonic CFU in media for NRS70 and Ab307 (p<0.001). CVCES at -1.8V for 8hrs completely eradicated both coupon-associated and planktonic CFU in media for NRS70 and Ab307. CVCES at -1.5V for 4 and 8hrs, and -1.8V for 2hrs, did not result in clinically relevant CFU reductions compared to OCP. For 4 and 8hr CVCES with and without bacteria, the current density was significantly higher at -1.8V than -1.5V (6.56x10-4 vs. 5.60x10-5 A/cm2) and resulted in increases in pH. Conclusions: CVCES of cpTi implants may provide effective prevention of PJI. Stimulation increases the pH and reduces viable CFU. This is directly related to the rates of water and oxygen reduction on the cpTi. The voltage-dependent properties of cpTi are a suspected factor in the observed antimicrobial effects of CVCES.