Novel in vivo mouse model of implant related spine infection

Abstract

Post-operative spine infections are a challenge, as hardware must often be retained to prevent destabilization of the spine, and bacteria form biofilm on implants, rendering them inaccessible to antibiotic therapy, and immune cells. A model of posterior-approach spinal surgery was created in which a stainless steel k-wire was transfixed into the L4 spinous process of 12-week-old C57BL/six mice. Mice were then randomized to receive either one of three concentrations (1 × 102, 1 × 103, and 1 × 104 colony forming units (CFU)) of a bioluminescent strain of Staphylococcus aureus or normal saline at surgery. The mice were then longitudinally imaged for bacterial bioluminescence to quantify infection. The 1 × 102 CFU group had a decrease in signal down to control levels by POD 25, while the 1 × 103 and 1 × 104 CFU groups maintained a 10-fold higher signal through POD 35. Bacteria were then harvested from the pin and surrounding tissue for confirmatory CFU counts. All mice in the 1 × 104 CFU group experienced wound breakdown, while no mice in the other groups had this complication. Once an optimal bacterial concentration was determined, mice expressing enhanced green fluorescent protein in their myeloid cells (Lys-EGFP) were utilized to contemporaneously quantify bacterial burden, and immune response. Neutrophil fluorescence peaked for both groups on POD 3, and then declined. The infected group continued to have a response above the control group through POD 35. This study, establishes a noninvasive in vivo mouse model of spine implant infection that can quantify bacterial burden and host inflammation longitudinally in real time without requiring animal sacrifice.

Multimodal imaging guides surgical management in a preclinical spinal implant infection model

“…It should be mentioned that the spinal implant S. aureus infection also resulted in increased systemic inflammation with an elevated SUV signal in the spleen. On POD 42, S. aureus CFUs were enumerated from ex vivo implants and tissue, and CFUs were only detected from implants of infected mice but not sterile mice ( Figure 4, J and K), confirming the in vivo BLI data, as previously described for this model ( Figure 4I) (27,28).…”

“…Mouse model of spinal implant S. aureus infection using bioluminescence imaging. To evaluate the ability of conventional 18 F-FDG PET-CT versus 89 Zr-NIR680-1D9 PET-CT or in vivo FLI to detect an in vivo biofilm infection, an established mouse model of S. aureus spinal implant infection was employed (27). Spinal surgery with implantation of an L-shaped implant and inoculation with bioluminescent S. aureus (Xen36 strain) was performed in conjunction with in vivo bioluminescence imaging (BLI) on postoperative days (POD) 0, 1, 3, 5, and 7 (Figures 2 and 3).…”

“…This strain was derived from the parental strain S. aureus ATCC-49525 (Wright), which is a clinical isolate from a septic patient. This specific bioluminescent derivative has been utilized in prior validated studies (27,47). S. aureus Xen36 uniquely utilizes a luxABCDE operon, which is optimized and integrated into the host's native plasmid (48).…”

“…Preparation of S. aureus for inoculation. Bacterial preparations were performed in accordance with previously published experiments, with minor alterations as needed (27,47). First, S. aureus Xen36 was isolated from potential contaminants using kanamycin plates.…”

“…However, it is unclear so far whether the 1D9 probe could facilitate diagnosis as well as guide treatment of a more complex biofilm-associated S. aureus implant infection. Therefore, we investigated the 1D9 probe (25, 26), dual labeled with 89 Zr and the NIR680 dye (i.e., 89 Zr-NIR680-1D9), as a multimodal noninvasive approach to distinguish septic from aseptic inflammation, diagnose S. aureus infection specifically, and facilitate intraoperative image-guided selective debridement of fluorescently labeled infected tissue in a preclinical S. aureus spinal implant infection model that utilized bioluminescently labeled bacteria to further confirm the specificity and sensitivity of this targeting probe (27).…”