Section 1: Clinical Microbiology Laboratory handbook Trauma and Orthopaedics

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1 Section 1: Clinical Microbiology Laboratory handbook Trauma and Orthopaedics Guidance for use of the microbiology laboratory / specimen taking in patients with suspected infection related to T+O Approved by T. Lewis Page 1 of 7 Date of Issue: January 2012

2 Specimen type: Joint aspirate (native joint, prosthetic joint with acute infection) Patients with suspected septic arthritis, or suspected crystal arthritis. Crystal identification, organism identification and sensitivity testing is important in the diagnosis and subsequent management of an acutely inflamed joint. Sterile, no touch technique using a needle and syringe. Fluid should be sent to the laboratory for microscopy, gram stain, direct culture and enrichment culture in a universal container. Antibiotic effect will be reported. Direct sensitivities will be performed if organisms are seen on the gram stain. Rejection criteria : None. Most organisms in pure growth, but particularly S. aureus, haemolytic streptococci, Strep. pneumoniae, HACEK organisms, coliforms, Pseudomonas aeruginosa, Neisseria gonorrhoea. Phone to discuss if mycobacteria or other unusual pathogens (eg. Fungi) are being considered. Specimens should be transported to the laboratory at room temperature. If there is to be delay in processing, the universal container should be stored at 4 o C. Blood culture bottles should be stored at room temperature. Gram stain and microscopy results will be available within 2 hours of receipt. Preliminary culture results within 24 hours of receipt. Final direct culture results within 48 hours of receipt. Final enrichment culture within 7 days of receipt. 09:00 22:00 Monday Sunday. Will not be routinely processed outside this time period. Monday to Sunday. Plates will be routinely inspected on Sundays and Bank holidays. Positive gram stain, positive culture results with likely / possible pathogens. Approved by T. Lewis Page 2 of 7 Date of Issue: January 2012

3 Specimen type: Joint aspirate (prosthetic joint with chronic infection) Patients with possible infection of prosthetic joint. These are not acutely infected joints (see above for management of this situation). The aim of this test is to inform the likelihood of infection, and ensure that any organisms are covered by operative prophylaxis and cements. Various cut-off values for white cell counts have been proposed, but the probability of infection is increased with counts of more than cells / ml. The probability of infection is also increased if there is a preponderance of neutrophils. Sterile, no touch technique using a needle and syringe. A small amount of fluid should be sent to the laboratory for a cell count using a counting chamber and detection of antimicrobial activity. Remaining fluid should be inoculated directly into a paediatric blood culture bottle. Rejection criteria: None Any, but particularly Coagulase negative Staphylococci. Specimens should be transported to the laboratory at room temperature. If there is to be delay in processing, the universal container should be stored at 4 o C. Blood culture bottles should be stored at room temperature. Microscopy results will be available within 2 hours of receipt. Final enrichment culture (from blood culture bottle) within 5 days of receipt. 09:00-17:00 Monday Friday. Will not be routinely processed outside this time period. Monday to Saturday. Plates will not be routinely inspected on Sundays and Bank holidays. None a comment will be added to reports as to suggested prophylaxis. An will be sent to the responsible consultant. Approved by T. Lewis Page 3 of 7 Date of Issue: January 2012

4 Specimen type: Tissue from infected joint (native joint, prosthetic joint with acute infection), osteomyelitis, or other infected soft tissue structures. Washout or debridement in theatre of acutely inflamed joint or infected bone. Organism identification and sensitivity testing is important in the diagnosis and subsequent management of an acutely inflamed joint or osteomyelitis. Biopsy of tissue from debrided skin may be important in identification of unusual pathogens. Surgical technique, or radiologically guided biopsy. Necrotic tissue should be debrided and discarded before sampling. Necrotic tissue will be discarded by the laboratory. Send vital tissue in a universal tube. It is important that the surgical team dissect out the areas of tissue most likely to be infected this will not be done by the laboratory. If necessary, send multiple specimens in separate universal tubes. Specimens will receive a gram stain and culture. Antibiotic effect will be reported. Direct sensitivities will be performed if organisms are seen on the gram stain. Samples will be inoculated into enrichment culture (brain heart infusion) for 48 hours before subculturing. Rejection criteria: None Most organisms in pure growth, but particularly S. aureus, haemolytic streptococci, Strep. pneumoniae, HACEK organisms, coliforms, Pseudomonas aeruginosa, Neisseria gonorrhoea. Phone to discuss if mycobacteria or other unusual pathogens (eg. Fungi) are being considered. Specimens should be transported to the laboratory and kept at 4 o C pending processing. Microscopy results will be available within 2 hours of receipt. Final enrichment culture within 7 days of receipt. 09:00 22:00 Monday Sunday. Will not be routinely processed outside this time period. Monday to Sunday. Plates will be routinely inspected on Sundays and Bank holidays. Any positive microscopy, any positive culture. Approved by T. Lewis Page 4 of 7 Date of Issue: January 2012

5 Specimen type: Tissue from infected joint (prosthetic joint with chronic infection) Joint revision with possible infection Surgical technique. After debridement of necrotic tissue, at least 5 specimens should be taken from around the joint, including sinus tract, capsule and other possibly infected tissues. Specimens should be inoculated directly into a Robertsons Cooked Meat (RCM) Broth in theatre using a sterile technique. The tops of the bottles should be wiped with an alcohol wipe and allowed to dry before opening. Bottles will be incubated for up to 14 days. Broths will be subcultured if turbid, and will receive a terminal subculture. Rejection criteria: None Most organisms in pure growth, but particularly Staphylococci, Streptococci, and Proprionibacteria. Specimens should be transported to the laboratory at room temperature, and can be stored in a 37 o C incubator. Culture only minimum of 14 day enrichment culture with up to 5 days to identify organisms 09:00 17:00 Monday Friday. Will not be routinely processed outside this time period. Monday to Saturday. Plates will not be routinely inspected on Sundays and Bank holidays. The consultant microbiologist will be informed of any positive culture and will contact the clinical team to advise on management. Approved by T. Lewis Page 5 of 7 Date of Issue: January 2012

6 Specimen type: Wound swab Wound swabs are not predictive of organisms causing underlying deep infection, with the exception of S. aureus. Wound swabs cannot be used to diagnose infection, as all organisms may colonise a wound. Swabs are most useful in ensuring sensitivity to empirical antibiotic choices. Biopsy may be necessary for diagnosis of infection in unusual, or non-resolving cases. Slough and necrotic tissue must be debrided before swabbing the wound with a charcoal swab. Rejection criteria: Inadequately labelled specimens and forms will be rejected (see specimen acceptance polic). Specimens will be rejected if there are inadequate clinical details forms must specify the nature of the suspected infection and intended antibiotics if relevant. Unlabelled forms, or forms labelled?infection or similar, will be rejected. S. aureus, haemolytic streptococci. Pasteurella spp. if history of animal bite. Swabs of leg ulcers will be screened for MRSA only, unless the form specifies cellulitis or an intended antibiotic. Room temperature Culture. Plates are incubated for 48 hours after receipt. 09:00-17:00 Monday Friday. 09: Saturday. Will not be routinely processed outside this time period. Monday to Saturday. None. Approved by T. Lewis Page 6 of 7 Date of Issue: January 2012

7 Summary of Specimen Types by Indication Indication Specimen Tests Septic arthritis (native joint and acute arthritis of prosthetic joint) Joint aspirate into a universal container Microscopy (including crystals) Direct culture Sub-acute Septic arthritis (prosthetic joint) Infected native joint (theatre specimen) Infected prosthetic joint (theatre specimens) 1. Joint aspirate into universal container 2. Joint aspirate into paediatric blood culture bottle Tissue after debridement (discard necrotic / nonviable material) into universal container 1. Minimum of 5 samples of vital tissue after debridement (discard necrotic / non-viable material). See sampling protocol for suspected prosthetic joint infection. Tissues should be placed directly into Robertson s Cooked Meat (RCM) Broth. 2. Joint fluid inoculated directly into paediatric blood culture bottle Antibiotic effect White Cell count Microscopy Direct Culture Antibiotic effect Superficial wound infection Wound swab (charcoal) after debridement of slough / necrotic tissue. Culture Approved by T. Lewis Page 7 of 7 Date of Issue: January 2012

Disc Diffusion Susceptibility Methods

Disc Diffusion Susceptibility Methods Introduction When a filter paper disc impregnated with a chemical is placed on agar the chemical will diffuse from the disc into the agar. This diffusion will place