Protocols for processing of tissue from arthroplasty infections vary and might affect the recovery of bacteria. We compared homogenization, bead beating and enzymatic disruption for recovery of live bacteria from tissue samples.

Suspensions of Staphylococcus aureus and Escherichia coli were prepared as controls. Three samples were taken from each and the first was bead beaten, the second homogenized, and Proteinase K was added for 10 and 30 minutes to the third sample before culturing. In addition, artificially inoculated pork tissue and known infected human tissue samples were processed by either homogenization or bead beating prior to cultures and results were compared.

Number of cycles of bead beating and homogenization and duration of Proteinase K treatment had significant effects. Bead beating for 2 and 4 cycles reduced the yield of S.aureus to 52% and 20% of control, and E.coli to 33% and 8%. Homogenization for 2 and 4 cycles reduced S.aureus to 86% and 65% of control, and E.coli to 90% and 87%. Proteinase K for 10 minutes and 30 minutes reduced the yield of S.aureus to 75% and 33% of control, and E.coli to 91% and 49% respectively. Inoculated Pork tissue showed a reduction in S.aureus recovery of 90% for bead beating compared to homogenization, and 80% in the case of E.coli. Bead beating of infected human tissue samples reduced the yield by 58% compared to homogenization.

Bead-beating is a common recommended method of processing tissue from arthroplasty cases. However, even though it produces a homogeneous sample, it does so at the cost of significant loss of viable bacteria. Homogenization and 10 minutes of Proteinase K incubation are almost equivalent, but the homogenizer is preferred being more controllable and cheaper. This should help to define guidelines for diagnosing infections using tissue samples.

Metal instrumentation (rods and screws) is used to stabilise the spine after trauma, malignancy or deformity. Approx 3% become infected often necessitating removal of metal. At surgery tissue samples and metal are removed for culture, but many clinical laboratories are not equipped to process metal or use simple culture methods. The causative bacteria exist as biofilms on the metal and they are often anaerobic and slow-growing, so conventional culture methods often fail to detect them. Also, they are common contaminants leading to diagnostic uncertainty. We have established a laboratory protocol to overcome these problems.

Removed metalwork was sonicated and the sonicate centrifuged and the supernatant discarded. Quantitative aerobic and anaerobic culture of the resuspended pellet for 14 days and microscopy were carried out.

Metalwork from 11 suspected infected cases was culture-positive (median 2857, 60–5000cfu/mL). Microscopy revealed an infection due to Candida albicans that would not have been detected otherwise. Bacteria were isolated from 8 of 10 non-infected cases (median 15, 0–35 cfu/mL). Conventionally processed samples failed to grow in 4 infected cases. (cfu/mL infected vs noninfected cases p=0.0093)

Micro-organisms on spinal metalwork grow as biofilms and they require sonication to dislodge them. The causative bacteria are slow-growing and P acnes is anaerobic and requires prolonged incubation. S epidermidis and P acnes are common contaminants and quantitative culture helps to distinguish pathogens from contaminants, removing the diagnostic uncertainty that conventional methods give. Microscopy of the sonicate can reveal micro-organisms that fail to grow on culture. We recommend that sonication of metalwork, prolonged anaerobic incubation and quantitative culture be adopted to improve diagnostic clarity for spinal instrumentation infections.

Background

Antibiotic loaded bone cement spacers are used as an adjunct to treatment in 2-stage arthroplasty revisions. If release of the correct choice of antimicrobials is optimised, systemic therapy might be curtailed and emergence of resistance minimised. Aims: To determine the elution period of antimicrobials from bone cement with and without a copolymer, polyvinylpyrrolidone (PVP) and to limit resistance development by the use of two or more antimicrobials.

Background

Deep infection rates of 1 - 2% following primary hip and knee arthroplasty are mainly due to endogenous contamination of the surgical site from bacteria within the patient's own skin. However surgical skin preparation removes only bacteria from the surface of the skin, leaving viable bacteria in the deeper layers of the skin within hair follicles and sweat and sebaceous glands. The aim of our study was to test the hypothesis that surface skin swabs taken after skin preparation with alcoholic povidone iodine would not grow bacteria, whereas full thickness biopsies taken from the line of surgical incision would grow bacteria.

Introduction

Open fractures occur with an annual incidence of 11.5 per 100,000 (6900 pa in UK). Infection rates, even with intravenous broad-spectrum antibiotics, remain as high as 22%. For this reason necessary bone grafting is usually delayed until soft-tissue cover of the bone injury is achieved. A biodegradable bone graft that released sustained high concentrations of antibiotics and encouraged osteogenesis, that could be implanted safely on the day of injury would reduce infection rates and avoid reoperation and secondary grafting. The non –union rate (approx 350 pa in UK) should also be reduced. Such a graft, consisting of a PLA/PGA co –polymer and containing antibiotics, is under development and here we report assessment of spectrum and duration of antimicrobial activity and effect of addition of antibiotics on mechanical properties.

Infection of orthopaedic implants is a significant problem, with increased antibiotic resistance of adherent ‘biofilm’ bacteria causing difficulties in treatment. We have investigated the in vitro effect of a pulsed electromagnetic field (PEMF) on the efficacy of antibiotics in the treatment of infection of implants.

Five-day biofilms of Staphylococcus epidermidis were grown on the tips of stainless-steel pegs. They were exposed for 12 hours to varying concentrations of gentamicin or vancomycin in microtitre trays at 37°C and 5% CO₂. The test group were exposed to a PEMF. The control tray was not exposed to a PEMF. After exposure to antibiotic the pegs were incubated overnight, before standard plating onto blood agar for colony counting.

Exposure to a PEMF increased the effectiveness of gentamicin against the five-day biofilms of Staphylococcus epidermidis. In three of five experiments there was reduction of at least 50% in the minimum biofilm inhibitory concentration. In a fourth experiment there was a two-log difference in colony count at 160 mg/l of gentamicin. Analysis of variance (ANOVA) confirmed an effect by a PEMF on the efficacy of gentamicin which was significant at p < 0.05. There was no significant effect with vancomycin.