Differential metabolism between biofilm and suspended Pseudomonas aeruginosa cultures in bovine synovial fluid by 2D NMR-based metabolomics

Abstract Total joint arthroplasty is a common surgical procedure resulting in improved quality of life; however, a leading cause of surgery failure is periprosthetic joint infection. Periprosthetic infection often involves biofilms, making treatment challenging. Periprosthetic joint infections are difficult to diagnose by traditional culturing methods and there are no clinical biomarkers for the presence of biofilms. Further, the metabolic state of pathogens in the joint space is difficult to diagnose, the mechanism of their tolerance to antibiotics and host defenses is not well understood, and their culturing in the laboratory is challenging. Thus, there is a critical need for improved pathogen- and phenotype-specific diagnosis as well as improved treatment strategies toward better patient outcomes. Here, we present a quantitative, untargeted NMR-based metabolomics strategy for Pseudomonas aeruginosa suspended culture and biofilm phenotypes grown in bovine synovial fluid. We identified 21 unique metabolites as potential markers of P. aeruginosa and one unique marker of the biofilm phenotype in synovial fluid. Significant differences in metabolic pathways were found between the suspended culture and biofilm phenotypes including creatine, glutathione, alanine, and choline metabolism and the tricarboxylic acid cycle. These unique metabolite and pathway differences have the potential to serve as targets for P. aeruginosa and specifically biofilm diagnosis and biofilm control in synovial fluid. Author Summary Joint replacement surgery is a common procedure frequently required in later stages of life due to damage in the joint. Over one million joint replacement surgeries are performed annually with rates increasing every year. A devastating complication associated with joint replacement is the development of infection around the implant device in the joint space, known as a periprosthetic joint infection. Bacteria in the joint space can form a biofilm, which is a gel-like matrix encasing the cells that increases resistance to treatment and exacerbates chronic infections. A particular challenge for the diagnosis of biofilm-mediated periprosthetic joint infections is the slowly growing nature of biofilm-mediated phenotypes, resulting in frequent failure to detect these bacteria by clinical microbiological culturing methods. Small molecule metabolites are uniquely produced by strains of bacteria in the biofilm versus planktonic or suspended culture phenotype. Identification of metabolites as specific markers of infection and biofilm could allow a new culture-free diagnostic approach to diagnose infection by biofilm. Furthermore, knowledge of metabolic pathway populations in biofilm in joint fluid could point to specific targets to prevent biofilm formation in the joint space.