[A chromogenic method for rapid identification of Staphylococcus aureus]

Staphylococcus aureus is responsible for septicaemia and serious nosocomial infections. A rapid and specific identification of this species is of great importance in clinical microbiology. Current methods for S. aureus identification require a 18 to 24 h-incubation. We describe a two hour-identification method based on the detection of the staphylocoagulase, using human prothrombin and a chromogenic substrate. 242 staphylococcal strains (160 S. aureus, 82 coagulase-negative staphylococci (CNS)) were collected from 4 French hospitals. They have been identified by the following methods: (i) clotting of citrated rabbit plasma, which is considered as reference method; (ii) biochemical tests (Rapidec Staph and Api Staph or ID 32 Staph); (iii) and agglutination test (Pastorex Staph or Pastorex Staph-plus). A strain of S. intermedius was provided by the Collection of the Pasteur Institute (Paris). An adapted culture medium is inoculated with staphylococci and adjusted to 2 Mac Farland unities. This medium is then mixed to an equal volume with a human prothrombin solution and the chromogenic substrate. After 1 to 2 hours incubation at 37 degrees C, the strength of the yellow colour of the mixture is observed to the naked eye, or measured at 405 nm with a spectrophotometer. Fifteen chromogenic tripeptides having a thrombin-like affinity and paranitroanilin as leaving group were compared. With the substrate which has the higher hydrolysis velocity and enzymatic affinity (SQ149), all S. aureus strains gave a positive result: 94.7% of the methicillin-susceptible S. aureus were detected after 1 hour incubation, but only 52.3% of the methicillin-resistant S. aureus. 98.4% of the methicillin-resistant S. aureus were detected after 2 hours. No false positive result was observed for the 82 CNS strains. The chromogenic method shows good within-run and day-to-day precision tests. It doesn't need any complementary test. The sensitivity and the specificity are 99.4% and 100% respectively.