Pyrrolidonyl Arylamidase (PYR) test is a rapid test which is used for the presumptive identification of group A beta-hemolytic Streptococci and Enterococci. PYR test is available in different format.
Original PYR test used to take 16-20 hours but now 4-hour broth assay is available, which is described in this post. Other formats include rapid (10-15 minutes) tests, in which PYR reagent impregnated filter paper disks or strips are inoculated with the organisms to be tested.
The substrate for the PYR test is L-naphthylamide-β-naphthylamide which is hydrolyzed by a specific bacterial aminopeptidase enzyme.
Hydrolysis of the substrate by this enzyme releases free β-naphthylamide, which is detected by the addition of N, N-dimethylaminocinnamaldehyde. This detection reagent couples with the naphthylamide to form a red Schiff base.
Procedure of PYR test
- With a sterile bacteriologic loop, pick up the growth of two to three morphologically similar colonies and emulsify them in the small volume of PYR broth
- Incubate the tube at 35oC for 4 hours
- Add one drop of PYR reagent and observe for color change.
- The reaction should be read and recorded 1 minute after the addition of reagent.
Results and interpretations
- Positive: the development of a deep cherry red color within a minute of addition of the reagent
- Negative: A yellow or orange color
- Positive control: Enterococcus faecalis, Streptococcus pyogenes
- Negative control: Streptococcus agalactiae
Note: It is essential that testing be performed before the PYR test to determine that the organism is a streptococcus (i.e. gram positive cocci, catalase-negative).
Other organisms (e.g., some aerococci, staphylococci, nutritionally variant streptococci, Arcanobacterium haemolyticum) may also be PYR positive.