Combined analysis of host immune response, biofilm genes, and 16S rRNA detection in fracture-related infection: an observational cohort study

Abstract. Fracture-related infection (FRI) is a serious complication in orthopaedic trauma that can lead to delayed union, nonunion, and poor clinical outcomes. A better understanding of the host immune response may provide valuable insights into the pathophysiology of FRI and may help identify genomic elements that contribute to the infection. This observational study compared immune responses between patients with FRI and non-infected controls using bone/tissue biopsies and sonication fluid, and it explored the possibility of detecting bacterial and biofilm genes using transcriptome profiling with hybridization technology (nCounter® RNA hybridization technology). A total of 15 infected patients demonstrated significant upregulation of the innate immune pathway, including Toll-like receptor (TLR) signalling and the MyD88 cascade, suggesting an active immune response contributing to both infection control and bone resorption. Among the differentially expressed genes, PTGS2 (COX-2) showed the highest level of upregulation in the infection group. Bone biopsies showed enhanced chemokine (e.g. CXCL1, CXCL2, CCL4/L1/L2) signalling, with higher levels compared to tissue biopsies. Transcriptomic analysis identified bacterial transcripts in cases where conventional culture was negative, revealing potential cases of low-bacterial-load infections causing culture-negativity. Transcriptome profiling exposed distinct immune activation patterns in FRI and enabled the detection of pathogens missed by conventional culture. These findings call for larger, prospective studies to further explore the clinical utility of transcriptomics in understanding and managing FRI.