RAPID DETECTION OF MRSA FROM CLINICAL SAMPLES USING MAGNETIC SEPARATION AND AK BIOLUMINESCENCE

摘要

Staphylococcus aureus is an opportunistic pathogen that occurs as part of the normal body flora in about a quarter of healthy adults. However, its methicillin-resistant variant (MRSA) has become endemic in hospitals worldwide, causing wound sepsis and other infections, particularly post-operatively. Patients are tested for MRSA by taking swabs, usually from the nose, perineum, armpit or wound. Analysed using conventional microbiological techniques, presumptive negative results are obtained in two days and positive results in two to four days. A prototype test for rapid MRSA detection was developed that used a combination of magnetic separation and adenylate kinase bioluminescence. The technique employed a short culture step (3-4 h) in a methicillin-containing broth to lyse susceptible bacteria prior to capturing resistant Staph. aureus cells using fibrinogen-coated magnetic particles. Staph. aureus has an affinity for fibrinogen that is not shown by other closely related species. Patient swabs were used to inoculate small volumes of broth containing methicillin, which were then incubated for 3 h. Previous experiments had shown that effective cell lysis of susceptible bacterial species could be obtained within this time. The fibrinogen-coated paramagnetic beads were then used to capture intact (i.e. antibiotic-resistant) cells, washed to remove extracellular adenylate kinase (AK) and then assayed using AK bioluminescence. The total assay time was approximately 4 h.