Equine or porcine synovial fluid as a novel ex vivo model for the study of bacterial free-floating biofilms that form in human joint infections

Bacterial invasion of synovial joints, as in infectious or septic arthritis, can be difficult to treat in both veterinary and human clinical practice. Biofilms, in the form of free-floating clumps or aggregates, are involved with the pathogenesis of infectious arthritis and periprosthetic joint infection (PJI). Infection of a joint containing an orthopedic implant can additionally complicate these infections due to the presence of adherent biofilms. Because of these biofilm phenotypes, bacteria within these infected joints show increased antimicrobial tolerance even at high antibiotic concentrations. To date, animal models of PJI or infectious arthritis have been limited to small animals such as rodents or rabbits. Small animal models, however, yield limited quantities of synovial fluid making them impractical for in vitro research. Herein, we describe the use of ex vivo equine and porcine models for the study of synovial fluid induced biofilm aggregate formation and antimicrobial tolerance. We observed Staphylococcus aureus and other bacterial pathogens adapt the same biofilm aggregate phenotype with significant antimicrobial tolerance in both equine and porcine synovial fluid, analogous to human synovial fluid. We also demonstrate that enzymatic dispersal of synovial fluid aggregates restores the activity of antimicrobials. Future studies investigating the interaction of bacterial cell surface proteins with host synovial fluid proteins can be readily carried out in equine or porcine ex vivo models to identify novel drug targets for treatment of prevention of these difficult to treat infectious diseases

Dogs can detect an odor profile associated with Staphylococcus aureus biofilms in cultures and biological samples

IntroductionThe study investigated the utilization of odor detection dogs to identify the odor profile of Staphylococcus aureus (S. aureus) biofilms in pure in vitro samples and in in vivo biosamples from animals and humans with S. aureus periprosthetic joint infection (PJI). Biofilms form when bacterial communities aggregate on orthopedic implants leading to recalcitrant infections that are difficult to treat. Identifying PJI biofilm infections is challenging, and traditional microbiological cultures may yield negative results even in the presence of clinical signs.MethodsDogs were trained on pure in vitro S. aureus biofilms and tested on lacrimal fluid samples from an in vivo animal model (rabbits) and human patients with confirmed S. aureus PJI.ResultsThe results demonstrated that dogs achieved a high degree of sensitivity and specificity in detecting the odor profile associated with S. aureus biofilms in rabbit samples. Preliminary results suggest that dogs can recognize S. aureus volatile organic compounds (VOCs) in human lacrimal fluid samples.DiscussionTraining odor detection dogs on in vitro S. aureus, may provide an alternative to obtaining clinical samples for training and mitigates biosecurity hazards. The findings hold promise for culture-independent diagnostics, enabling early disease detection, and improved antimicrobial stewardship. In conclusion, this research demonstrates that dogs trained on in vitro S. aureus samples can identify the consistent VOC profile of PJI S. aureus biofilm infections. The study opens avenues for further investigations into a retained VOC profile of S. aureus biofilm infection. These advancements could revolutionize infectious disease diagnosis and treatment, leading to better patient outcomes and addressing the global challenge of antimicrobial resistance

Hydrogen Peroxide, Povidone-Iodine and Chlorhexidine Fail to Eradicate Staphylococcus aureus Biofilm from Infected Implant Materials

Hydrogen peroxide, povidone-iodine, and chlorhexidine are antiseptics that are commonly added to irrigants to either prevent or treat infection. There are little clinical data available that demonstrate efficacy of adding antiseptics to irrigants in the treatment of periprosthetic joint infection after biofilm establishment. The objective of the study was to assess the bactericidal activity of the antiseptics on S. aureus planktonic and biofilm. For planktonic irrigation, S. aureus was exposed to different concentrations of antiseptics. S. aureus biofilm was developed by submerging a Kirschner wire into normalized bacteria and allowing it to grow for forty-eight hours. The Kirschner wire was then treated with irrigation solutions and plated for CFU analysis. Hydrogen peroxide, povidone-iodine, and chlorhexidine were bactericidal against planktonic bacteria with over a 3 log reduction (p < 0.0001). Unlike cefazolin, the antiseptics were not bactericidal (less than 3 log reduction) against biofilm bacteria but did have a statistical reduction in biofilm as compared to the initial time point (p < 0.0001). As compared to cefazolin treatment alone, the addition of hydrogen peroxide or povidone-iodine to cefazolin treatment only additionally reduced the biofilm burden by less than 1 log. The antiseptics demonstrated bactericidal properties with planktonic S. aureus; however, when used to irrigate S. aureus biofilms, these antiseptics were unable to decrease biofilm mass below a 3 log reduction, suggesting that S. aureus biofilm has a tolerance to antiseptics. This information should be considered when considering antibiotic tolerance in established S. aureus biofilm treatment

Microbubble Cavitation Restores Staphylococcus Aureus Antibiotic Susceptibility in Vitro and in a Septic Arthritis Model

Treatment failure in joint infections is associated with fibrinous, antibiotic-resistant, floating and tissue-associated Staphylococcus aureus aggregates formed in synovial fluid (SynF). We explore whether antibiotic activity could be increased against Staphylococcus aureus aggregates using ultrasound-triggered microbubble destruction (UTMD), in vitro and in a porcine model of septic arthritis. In vitro, when bacterially laden SynF is diluted, akin to the dilution achieved clinically with lavage and local injection of antibiotics, amikacin and ultrasound application result in increased bacterial metabolism, aggregate permeabilization, and a 4-5 log decrease in colony forming units, independent of microbubble destruction. Without SynF dilution, amikacin + UTMD does not increase antibiotic activity. Importantly, in the porcine model of septic arthritis, no bacteria are recovered from the SynF after treatment with amikacin and UTMD-ultrasound without UTMD is insufficient. Our data suggest that UTMD + antibiotics may serve as an important adjunct for the treatment of septic arthritis

5-Benzylidene-4-Oxazolidinones are Synergistic with Antibiotics for the Treatment of Staphylococcus Aureus Biofilms

The failure of frontline antibiotics in the clinic is one of the most serious threats to human health and requires a multitude of novel therapeutics and innovative treatment approaches to curtail the growing crisis. In addition to traditional resistance mechanisms resulting in the lack of efficacy of many antibiotics, most chronic and recurring infections are further made tolerant to antibiotic action by the presence of biofilms. Herein, we report an expanded set of 5-benzylidene-4-oxazolidinones that are able to inhibit the formation of Staphylococcus aureus biofilms, disperse preformed biofilms and in combination with common antibiotics are able to significantly reduce the bacterial load in a robust collagen-matrix model of biofilm infection.</div

Effect of BIO-PLYTM, a Platelet-Rich Plasma Derived Biologic on PRRSV-2-Infected Macrophages

Porcine Reproductive and Respiratory Syndrome (PRRS) is the one of the most devastating diseases impacting the swine industry worldwide. Control and prevention methods rely on biosafety measures and vaccination. As an RNA virus with a high rate of mutation, vaccines are only partially effective against circulating and newly emerging strains. To reduce the burden of this disease, research on alternative control methods is needed. Here, we assess the in vitro antiviral effect of a novel platelet-rich plasma-derived biologic termed BIO-PLYTM (for the BIOactive fraction of Platelet-rich plasma LYsate) from both swine and equine origin. Our results show that BIO-PLYTM significantly reduces the amount of PRRSV viral load determined by RT-qPCR and the number of infectious viral particles measured by TCID50 in infected porcine alveolar and parenchymal macrophages. This study also showed limited toxicity of BIO-PLYTM in vitro and aspects of its immunomodulatory capacity evaluating the regulation of reactive oxygen species and cytokines production in infected cells. Finally, this study presents promising data on the effect of BIO-PLYTM on other RNA viruses such as human A influenza viruses and coronavirus

Hydrogen Peroxide, Povidone-Iodine and Chlorhexidine Fail to Eradicate <i>Staphylococcus aureus</i> Biofilm from Infected Implant Materials

Hydrogen peroxide, povidone-iodine, and chlorhexidine are antiseptics that are commonly added to irrigants to either prevent or treat infection. There are little clinical data available that demonstrate efficacy of adding antiseptics to irrigants in the treatment of periprosthetic joint infection after biofilm establishment. The objective of the study was to assess the bactericidal activity of the antiseptics on S. aureus planktonic and biofilm. For planktonic irrigation, S. aureus was exposed to different concentrations of antiseptics. S. aureus biofilm was developed by submerging a Kirschner wire into normalized bacteria and allowing it to grow for forty-eight hours. The Kirschner wire was then treated with irrigation solutions and plated for CFU analysis. Hydrogen peroxide, povidone-iodine, and chlorhexidine were bactericidal against planktonic bacteria with over a 3 log reduction (p p S. aureus; however, when used to irrigate S. aureus biofilms, these antiseptics were unable to decrease biofilm mass below a 3 log reduction, suggesting that S. aureus biofilm has a tolerance to antiseptics. This information should be considered when considering antibiotic tolerance in established S. aureus biofilm treatment