Bacterial Survival In Synovial Fluid: Is S. aureus in the Knee Joint Persisting Despite Antibiotic Treatment?

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1 Bacterial Survival In Synovial Fluid: Is S. aureus in the Knee Joint Persisting Despite Antibiotic Treatment? Sana Dastgheyb 1, Sommer Hammoud 2, Constantinos Ketonis, MD 3, James Purtill, MD 3, Michael Ciccotti, MD 3, Javad Parvizi, M.D. 3, Noreen J. Hickok, PhD 1. 1 Thomas Jefferson University, Philadelphia, PA, USA, 2 Rothman Institute, Rothman Institute, PA, USA, 3 Rothman Institute, Philadelphia, PA, USA. Disclosures: S. Dastgheyb: None. S. Hammoud: None. C. Ketonis: None. J. Purtill: None. M. Ciccotti: None. J. Parvizi: None. N.J. Hickok: None. Introduction: Infection of joint prosthesis can be a devastating complication of joint arthroplasty, and often results in dramatic revisions, with a high risk of re-infection. Biomaterial surfaces are highly susceptible to colonization by biofilm-forming bacteria in which antibiotic resistance is high, and immune recognition is low. In this study, we report the effect of synovial fluid on biofilm-causing pathogens. Methods: Penetration of pre-operative antibiotics into Synovial Fluid (SF) was first confirmed by Kirby Bauer disc diffusion methods against 6 strains of bacteria (S. aureus ATCC and AH1710, MRSA, S. epidermidis ATCC 35984, and E. coli ATCC and DH5α). Bacterial survival in synovial fluid was assessed by overnight incubation of 107 CFU/mL of S. aureus ATCC in 7 synovial fluid samples obtained from patients prior to invasive knee surgeries. The following day, samples were serially diluted and plated, surviving bacteria were quantified by direct counting. In parallel, Ti90Al6V4 pins were incubated in synovial fluid from patients with pre-operative antibiotics containing 107 CFU/mL of s. aureus. The following day, pins were imaged, or adherent bacteria recovered by sonication in 0.3% Tween-20, which was then serially diluted, plated and quantified by direct counting. Bacterial metabolism in SF was determined by reduction of alamar blue. Bacterial survival was also assessed using incubation of S. aureus Xen-36 luciferase-expressing bacteria in SF. Results: Our results show that despite the presence of antibiotics in SF, the bacteria do not die, nor do they grow in 6 out of 7 SF samples (Fig 1). The addition of titanium pins to the SF showed that adherence to the pin occurred, despite the presence of antibiotics (Fig 2). Alamar blue experiments showed that it is not possible to detect respiration by redox detection in synovial fluid, despite the fact that the bacteria are viable (Fig 3). The Xen-36 strain of bacteria showed that live, luciferase-expressing bacteria were present in synovial fluid. Importantly, confocal imaging of bacteria in TSB (Trypticase Soy Broth), PBS, and SF showed that bacteria readily clump in SF, and even larger clumps are present when antibiotic is present. Discussion: We have shown that bacterial survive in synovial fluid despite the presence of pre-operative antibiotic, and that the phenotype of bacteria is grossly changed in the presence of synovial fluid, which likely confers resistance of the bacteria to antibiotics in the synovial environment. Significance: Despite previous reports that synovial fluid expresses antimicrobial factors, and that antibiotics (both oral and IV) can be found at above minimum inhibitory concentrations (MIC) in synovial fluid, this study describes bacterial survival in synovial fluid. The presence of antibiotics at 20 X MIC, despite being high enough to kill bacteria, did not stop bacterial adherence to titanium pins. This is troubling because it has been reported that only CFU of bacteria are necessary to seed an infection in the presence of a device. Our studies suggest that current antimicrobials used for eradication of bacteria in the synovial environment are not sufficient to eradicate bacterial colonization. Acknowledgments: The authors thank the NIH (National Institute of Arthritis and Musculoskeletal and Skin Diseases training grant T32-AR (SSD), HD (NJH) and DE (NJH). References:

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5 ORS 2014 Annual Meeting Poster No: 1058

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