Establishment of a clinically relevant beagle model for periprosthetic joint infection with 3D-printed prostheses and multimodal evaluation

Abstract

Objective: Periprosthetic joint infection (PJI) poses significant challenges to arthroplasty outcomes, necessitating translational animal models for pathogenesis studies and therapeutic development. This study aimed to establish a standardized Beagle PJI model by integrating species-specific 3D-printed femoral prostheses with quantitative bacterial inoculation, while evaluating the dose-dependent effects of Staphylococcus aureus (S. aureus) on infection progression.

Methods: Two titanium alloy prostheses were designed using CT-based anatomical data: BFP-C (canine-optimized) and BFP-H (human-derived). Prostheses underwent mechanical compression tests, finite element analysis (FEA) simulating postoperative and osseointegration phases, and in vivo validation in Beagles. The optimized BFP-C was selected for PJI model construction via hemi-hip arthroplasty (HHA), with intraoperative inoculation of S. aureus ranging from 250 to 10^8 colony-forming units (CFU). Longitudinal evaluation included radiography (X-ray/CT), mechanical pull-out tests, histopathology (H&E/Masson/Giemsa staining), bacterial cultures, and mobility assessments using open-field behavioural tracking.

Results: BFP-C exhibited superior biomechanical compatibility, with 12.3-fold higher yield strength (6836 ± 157 N vs. 553 ± 49 N) and 97 % reduction in bone strain (0.71 % vs. 20.32 %) compared to BFP-H. All inoculated groups developed PJI with dose-dependent severity: ultra-high-dose (10^8 CFU) groups displayed severe osteolysis (pull-out strength: 24 ± 8 N vs. 924 ± 45 N in controls), biofilm formation, and mobility impairment (74 % reduction in distance travelled, 2003 ± 276 cm vs. 7976 ± 333 cm in controls), whereas low-dose (250 CFU) groups established PJI with milder manifestations, evidenced by sinus tract formation, 55.1 % reduction in pull-out strength (406 ± 15 N vs. 924 ± 45 N in controls), and concordant radiological/histopathological signs of infection. Imaging examinations revealed differential osteolytic patterns corresponding to bacterial loads. Combined wound evaluation and microbiological analyses confirmed consistent infection establishment across all replicates.

Conclusion: This Beagle PJI model successfully recapitulates clinical infection dynamics, emphasizing the critical role of species-specific prosthesis design and standardized bacterial quantification. The integrated multimodal evaluation system (imaging, biomechanical, and behavioural analyses) demonstrated both the reliability of the model and its sensitivity in detecting infection progression. Its modular design supports customization for studying biofilm-resistant implants or antibiotic delivery systems. These findings not only provide a critical tool for mechanistic PJI research but also establish a theoretical foundation for clinical translation, with the quantitative multimodal framework directly informing diagnostic and therapeutic strategies.

Translational potential: Beyond serving as a preclinical platform for anti-infective therapies, the model provides actionable insights into optimizing human prosthetic biomechanics, such as reducing stress shielding through FEA-informed design principles. The 3D printing workflow further demonstrates rapid prototyping capabilities for patient-specific orthopaedic implants.

Keywords: 3D printed prosthesis; Beagle model; Periprosthetic joint infection (PJI); Translational biomechanics.

Graphical abstract

Scheme 1

(A) Design, 3D printing of prosthesis and construction of the Beagle model of hip PJI. A femoral head prosthesis was designed and 3D-printed to match the Beagle hip anatomy. Subsequently, Beagles underwent HHA with S. aureus injected into the medullary cavity prior to femoral prosthesis implantation. (B) Postoperative infection assessment. Infection was assessed by monitoring postoperative body temperature (POT) and body weight (POW), evaluating wound healing, performing the open field test, conducting X-ray and CT imaging, and analyzing bacterial load, mechanical pull-out strength, and histological staining to determine the severity of hip PJI.

Fig. 1

Design, 3D printing, and implantation of BHP-C & BFP-H. (A) Design and 3D printing of BFP-C and BFP-H. (i-vi) i and iv, Femoral modeling and prosthesis fitting simulation; ii and v, BFP schema showing key parameters: D (femoral head diameter), L (neck length), H (stem height), W (neck width), and CDA (cervical-diaphyseal angle); iii and vi, 3D-printed custom-designed BFP prosthesis. (B) BFP stiffness. (C) BFP yield strength. (D) Hemi-hip arthroplasty procedure (HHA): (1) Disinfect and drape the surgical site; (2) Expose the femoral head; (3) Resect the femoral head; (4) Broach the femoral canal; (5) Implant BFP-C or BFP-H prosthesis; (6) Secure the prosthesis and confirm alignment; (7) Reduce the hip joint; (8) Close the surgical site. Data are presented as mean ± SD (∗P < 0.05, ∗∗P < 0.01, ∗∗∗P < 0.001; n=3).

Fig. 2

Performance comparison of two prostheses ex vivo & in vivo. (A) Finite element analysis of BFP-C and BFP-H. (i–v) i, Mesh generation; ii-iii, short-term deformation for BFP-C and BFP-H; iv-v, long-term deformation for BFP-C and BFP-H. (B) Postoperative imaging data. (i-vi) i, BFP-C at POD 0, with proper prosthesis positioning; ii, BFP-H at POD 0, femoral head dislocation (yellow arrow); iii, BFP-C at POD 28, with proper prosthesis positioning; iv, BFP-H at POD 28, showing BFP-H distal deformation (white arrow); v-vi, CT imaging of BFP-C and BFP-H at POD 28, prostheses within the femoral canal. (C) Biomechanical pull-out test. Pull-out test images, with distant and close-up views. (D) Maximum pull-out forces at POD 0 and POD 28. Data are presented as mean ± SD (∗P < 0.05, ∗∗P < 0.01, ∗∗∗P < 0.001; n = 3).

Fig. 3

Beagle PJI model: postoperative temperature, body weight, and open field test results. (A) Schematic of the Beagle PJI model establishment. (B) Representative trajectory images from the open field test. (C) Quantitative analysis of trajectory data, including (i) distance travelled, (ii) average speed, (iii) maximum acceleration, (iv) duration of movement, (v) activity level, and (vi) rotation frequency. (D) POT and (E) POW in different days. Data are presented as mean ± SD (∗P < 0.05, ∗∗P < 0.01, ∗∗∗P < 0.001; n = 3).

Fig. 4

Postoperative imaging evaluation and maximum pull-out test. (A) X-ray images of PJI model groups on POD 28, with red arrows indicating changes in medullary bone structure. (B) CT images of PJI model groups on POD 28, with red arrows indicating bone destruction. (C) X-ray image scoring on POD 28. (D) CT image scoring on POD 28. (E) Maximum pull-out forces on POD 28 after infection. Data are presented as mean ± SD (∗P < 0.05, ∗∗P < 0.01, ∗∗∗P < 0.001; n = 3).

Fig. 5

Wound evaluation and bacterial detection. (A) Appearance of the surgical wound on POD 28. (B) Subcutaneous exudate of the wound. (C) Representative CFU counts of S. aureus on the BFP-C surface. (D) Wound scoring. (E) Semi-quantification of CFU counts on the BFP-C surface (n = 3). (F) Semi-quantification of CFU counts in periarticular tissue (n = 5). Data are presented as mean ± SD (∗P < 0.05, ∗∗P < 0.01, ∗∗∗P < 0.001).

Fig. 6

Histological evaluation of bone infection. (A) H&E staining of Beagle femurs, yellow arrows indicate inflammatory cells. (B) Masson's trichrome staining of Beagle femurs. (C) Giemsa staining of Beagle femurs, red arrows indicate S. aureus and biofilms. (D) Inflammatory cell count per high-power field. (E) Masson staining scores. (F) S. aureus count per high-power field. Scale bar, 100 μm. Data are presented as mean ± SD (∗P < 0.05, ∗∗P < 0.01, ∗∗∗P < 0.001; n = 6).