Acidity Is an Excellent Marker of Infection in Hip and Knee Arthroplasty.

BACKGROUND: The diagnosis of joint replacement infection is a difficult clinical challenge that often occurs when the implant cannot be salvaged. We hypothesize that the pH value of synovial fluid could be an important indicator of the inflammatory status of the joint. However, in the literature, there is a lack of data on the pH changes in hip and knee joint replacements and their relation to infection and implant failure. In this study, we aimed to measure the pH levels of synovial fluid in patients with hip and knee joint replacements. We also investigated the potential of pH measurement as a diagnostic tool for joint replacement infection. In this study, we recorded the pH values to be 7.55 and 7.46 in patients where Pseudomonas aeruginosa was identified as the cause of the prosthetic joint infection. We attribute this to the different environments created by this specific bacterium. In other cases where the pH was higher, chronic mitigated infections were diagnosed, caused by strains of Staphylococcus aureus, Streptococcus agalactiase, and coagulase negative staphylococcus. MATERIALS AND METHODS: In our cohort of 155 patients with implanted hip (THA; n = 85) or knee (TKA; n = 70) joint replacements, we conducted a prospective study with a pH measurement. Out of the whole cohort, 44 patients had confirmed joint replacement infection (28.4%) (44/155). In 111 patients, infection was ruled out (71.6%) (111/155). Joint replacement infection was classified according to the criteria of the Musculoskeletal Infection Society (MSIS) from 2018. Based on the measured values, we determined the cut-off level for the probability of ongoing inflammation. We also determined the sensitivity and specificity of the measurement. RESULTS: The group of patients with infection (n = 44) had a significantly lower synovial fluid pH (pH = 6.98 ± 0.48) than the group of patients with no infection (n = 111, pH = 7.82 ± 0.29, p < 0.001). The corresponding median pH values were 7.08 for the patients with infection and 7.83 for the patients with no infection. When we determined the cut-off level of pH 7.4, the sensitivity level of infected replacements was 88.6%, and the specificity level of the measurement was 95.5%. The predictive value of a positive test was 88.6%, and the predictive value of a negative test was 95.5%. CONCLUSIONS: Our results confirm that it is appropriate to include a pH measurement in the diagnostic spectrum of hip and knee replacements. This diagnostic approach has the potential to provide continuous in vivo feedback, facilitated by specialized biosensors. The advantage of this method is the future incorporation of a pH-detecting sensor into intelligent knee and hip replacements that will assess pH levels over time. By integrating these biosensors into intelligent implants, the early detection of joint replacement infections could be achieved, enhancing proactive intervention strategies.

Antimicrobial peptide in polymethylmethacrylate bone cement as a prophylaxis of infectious complications in orthopedics-an experiment in a murine model.

Polymethylmethacrylate (PMMA) bone cement mixed with antibiotics is used in orthopedic surgery to cope with implant-related infections which are typically associated with the formation of bacterial biofilms. Taking into account the growing bacterial resistance to current antibiotics, we examined here the efficacy of a selected antimicrobial peptide (AMP) mixed into the bone cement to inhibit bacterial adhesion and the consequent biofilm formation on its surface. In particular, we followed the formation of bacterial biofilms of methicillin-resistant Staphylococcus aureus (MRSA) on implants made from PMMA bone cement loaded with AMP composed of 12 amino acid residues. This was evaluated by CFU counting of bacteria released by sonication from the biofilms formed on their surfaces after these implants were retrieved from the infected murine femoral canals. The AMP loaded in these model implants prevented adhesion of MRSA and the subsequent formation of MRSA biofilm on the surfaces of more than 80% of these implants, whereas biofilms did form on control implants made from the plain cement. The results of our experiments performed in the murine femoral canal indicate the potential for this murine osteomyelitis model to mimic actual operations in orthopedics.

High-Performance Liquid Chromatography as a Novel Method for the Determination of α-Defensins in Synovial Fluid for Diagnosis of Orthopedic Infections.

The α-defensins (AD) present in synovial fluid have been regarded as constituting the most accurate periprosthetic joint infection (PJI) biomarker. The methods most commonly used for estimating AD as a biomarker are the qualitative Synovasure® PJI tests, based on the technique of lateral flow, and quantitative enzyme-linked immunosorbent assay (ELISA). Here, we propose a novel test based on detecting α-defensins in synovial fluid by high-performance liquid chromatography (HPLC). Synovial fluid was collected from 157 patients diagnosed with PJI, infectious arthritis (IA), arthrosis, reactive arthritis, and rheumatoid arthritis. AD concentrations in the fluid were determined by HPLC, and these same samples were used for additional diagnostic analyses. The results were statistically processed to calculate cutoff concentrations for PJI and IA. HPLC testing showed a sensitivity of 94% and a specificity of 92% for diagnosis of PJI, and a sensitivity of 97% and a specificity of 87% for diagnosis of IA. Using HPLC, we detected in synovial fluid a combination of three α-defensins: human neutrophil peptides HNP1, HNP2, and HNP3. All measured AD concentration values shown in this work refer to the sum of these three individual concentrations. Our study shows that the HPLC method meets the conditions for measuring precise concentrations of the sum of AD and can be recommended as part of a diagnostic array for PJI and IA diagnostics. By this method, we have verified that higher levels of AD in synovial fluid can also be seen in rheumatoid illnesses, crystal arthropathies, and reactive arthritis.

Vancomycin and its crystalline degradation products released from bone grafts and different types of bone cement.

Vancomycin is often used in orthopedic surgery as a local prophylaxis of bacterial infection. The aim of this work was to compare the release of vancomycin and its biologically inactive crystalline degradation products (CDP-1) during in vitro experiments from different types of local antibiotic delivery systems (bone grafts and bone cements). The concentrations of vancomycin and its crystalline degradation products were determined by high-performance liquid chromatography. Each experiment was performed in a phosphate buffer solution over 21 days. Morselized bone grafts, synthetic bone cements Palacos and Copal, and synthetic bone grafts were tested as local carriers of vancomycin. The highest concentration approximately 670 mg/L of vancomycin was released from synthetic bone grafts Actifuse. Even after 21 days, the concentration of vancomycin was still above the minimum inhibitory concentration (MIC). The maximum concentration of vancomycin released in two experiments with human bone grafts exceeded 600 mg/L during the first day and was still above MIC level 21 days later when the experiment was concluded. By comparing the synthetic bone cements Palacos and Copal, Copal had the average maximum concentration of only 32.4 mg/L and Palacos 35.7 mg/L. The concentration of vancomycin fell below the MIC for vancomycin-resistant Staphylococcus aureus (VRSA) on the seventh day with Palacos and the ninth day with Copal. This study showed the insufficient concentration of released vancomycin from synthetic bone cements at the end of the experiment. For improvement of local prophylaxis, it would be beneficial to increase the amount of vancomycin in bone cements.

Antimicrobial peptides prevent bacterial biofilm formation on the surface of polymethylmethacrylate bone cement.

PURPOSE: Antibiotic-loaded polymethylmethacrylate-based bone cement has been implemented in orthopaedics to cope with implant-related infections associated with the formation of bacterial biofilms. In the context of emerging bacterial resistance to current antibiotics, we examined the efficacy of short antimicrobial peptide-loaded bone cement in inhibiting bacterial adhesion and consequent biofilm formation on its surface. METHODOLOGY: The ability of α-helical antimicrobial peptides composed of 12 amino acid residues to prevent bacterial biofilm [methicillin-resistant Staphylococcus aureus (MRSA), Staphylococcus epidermidis, Pseudomonas aeruginosa and Escherichia coli] formation on the surface of model implants made from polymethylmethacrylate-based bone cement was evaluated by colony-forming unit (c.f.u.) counting of bacteria released by sonication from the biofilms formed on their surfaces. The biofilms on model implant surfaces were also visualized by light microscopy after staining with tetrazolium dye (MTT) and by scanning electron microscopy. RESULTS: When incorporated in the implants, these peptides caused a mean reduction in the number of bacterial cells attached to implants' surfaces (by five orders of magnitude), and 88 % of these implants showed no bacterial adhesion after being exposed to growth media containing various bacteria. CONCLUSION: The results showed that the antibiofilm activity of these peptides was comparable to that of the antibiotics, but the peptides exhibited broader specificity than the antibiotics. Given the rapid development of antibiotic resistance, antimicrobial peptides show promise as a substitute for antibiotics for loading into bone cements.

Antimicrobial Peptides for Topical Treatment of Osteomyelitis and Implant-Related Infections: Study in the Spongy Bone.

We examined the benefits of short linear α-helical antimicrobial peptides (AMPs) invented in our laboratory for treating bone infection and preventing microbial biofilm formation on model implants due to causative microorganisms of osteomyelitis. For this purpose, we introduced a model of induced osteomyelitis that utilizes human femur heads obtained from the hospital after their replacement with artificial prostheses. We found that the focus of the infection set up in the spongy part of this bone treated with AMP-loaded calcium phosphate cement was eradicated much more effectively than was the focus treated with antibiotics such as vancomycin or gentamicin loaded into the same cement. This contradicts the minimum inhibitory concentrations (MIC) values of AMPs and antibiotics against some bacterial strains obtained in standard in vitro assays. The formation of microbial biofilm on implants made from poly(methylmethacrylate)-based bone cement loaded with AMP was evaluated after the implants' removal from the infected bone sample. AMPs loaded in such model implants prevented microbial adhesion and subsequent formation of bacterial biofilm on their surface. Biofilms did form, on the other hand, on control implants made from the plain cement when these were implanted into the same infected bone sample. These results of the experiments performed in human bone tissue highlight the clinical potential of antimicrobial peptides for use in treating and preventing osteomyelitis caused by resistant pathogens.

Testing the efficacy of antimicrobial peptides in the topical treatment of induced osteomyelitis in rats.

Joint replacement infections and osteomyelitis are among the most serious complications in orthopaedics and traumatology. The risk factors for these infections are often bacterial resistance to antimicrobials. One of the few solutions available to control bacterial resistance involves antimicrobials, which have a different mechanism of action from traditional antibiotics. Antimicrobial peptides (AMP) appear to be highly promising candidates in the treatment of resistant infections. We have identified several AMP in the venom of various wild bees and designed analogues that show potent antimicrobial activity and low toxicity against eukaryotic cells. The aim of the present study was to test the efficacy of one of those synthetic peptide analogues for the treatment of acute osteomyelitis invoked in laboratory rats. Femoral cavities of 20 laboratory Wistar rats were infected with Staphylococcus aureus. After 1 week, eight rats received an injectable calcium phosphate carrier alone, another eight rats were treated with a calcium phosphate mixed with AMP, and four rats were left without any further treatment. After another week, all rats were euthanized and radiographs were made of both the operated and healthy limbs. The animals with the carrier alone exhibited more severe acute osteomyelitis on radiographs in comparison to the recipients of the calcium phosphate carrier loaded AMP and untreated infected individuals. Based on the results of the above mentioned experiment, it was concluded that when injected directly into the site of femoral acute osteomyelitis, the calcium phosphate carrier mixed with AMP reduced osteomyelitis signs visible on radiographs.