Effect of Biofilms on Recalcitrance of Staphylococcal Joint Infection to Antibiotic Treatment

Joint infection (septic arthritis) is a serious condition that if left untreated can lead to rapid, complete destruction of the joint. It may also spread to other sites in the human body and cause life-threatening secondary site infections. When a prosthetic device is present in the joint, the probability of infection increases, with infection rates for total knee or hip arthroplasty reaching 1%–2% [1, 2]. Staphylococcus aureus is the most common cause of septic arthritis in many countries [2]. Together with coagulase-negative staphylococci, it is also the leading pathogen involved in prosthetic joint infection (PJI) [1]. As with other staphylococcal infections, methicillin-resistant S. aureus (MRSA) joint infections are associated with increased risk of treatment failure and cost [3, 4]. In US hospitals, the community-associated (CA) MRSA strain USA300 has been reported to replace the usual hospital-associated (HA) strains in many types of nosocomial infections [5], including PJI [6]. According to data from a 2005 study in an Atlanta, Georgia, hospital, this strain already has emerged as the predominant pathogen involved in PJI in the United States [6]. The pathogenesis of joint infection is poorly understood. It has been speculated that biofilm formation plays a key role [7], a notion that is predominantly based on the isolation of biofilmlike agglomerations from cases of joint infection [8]. Furthermore, the synovial fluid (SF) present in joint cavities has been attributed a function in inhibiting killing of S. aureus by neutrophils [9], while in a somewhat contradicting manner it has also been described as antimicrobial [10]. Finally, microbiological evaluations of joint infections are often complicated by the frequency of false-negative cultures [11]. All these fairly unexplained observations call for a systematic study to investigate the underlying bacterial phenotypes and mechanisms. The in vitro investigation of bacterial biofilm formation is limited by the chosen growth medium, representing a highly artificial environment with disputable value for the interpretation of biofilm-associated infections [12]. On the other hand, it is difficult to investigate the contribution of biofilm formation to the pathogenesis of joint infections using animal models, because commonly used laboratory animals, such as mice or rats, are too small for that purpose. Notably, despite the frequent use of an S. aureus septic arthritis model in mice [13], biofilm formation has never been linked to septic arthritis based on results from that model. Furthermore, this model is problematic, because bacteria are injected intravenously into the tail and the observed differential degrees of joint infection may be secondary to previous systemic effects. Therefore, in this study we used an ex vivo approach to systematically investigate bacterial behavior in SF during joint infection. We isolated SF from human individuals to monitor bacterial phenotypes in the environment that the pathogen encounters during infection of the joints. We demonstrate extreme biofilm formation and aggregation of S. aureus in SF and show that aggregation in SF dramatically affects resistance to antibiotics. Elucidation of the molecular basis of the observed aggregation pointed to fibrin- and fibronectin-binding proteins, on which ground we tested plasmin to prevent aggregation. Our findings indicate that plasmin treatment efficiently increases the sensitivity of S. aureus to antibiotics in clinical use for joint infections, suggesting a novel pathway for their treatment.