PYR broth is generally used for the isolation and identification of Streptococcus pyogenes .It is highly sensitive test and often replaces bacitracin and salt tolerance tests.
It is a presumptive test for both group A and D enterococcal streptococci which determine the hydrolysis activity of enzyme L-pyrrolidonyl arylamidase (PYR) produced by Streptococcus pyogenes but not by other beta haemolytic streptococci were free b-naphthylamide is then detected by addition of the diazo dye complex, N ,Dimethylaminocinnamaldehyde.A Development of a red colour is indicative of PYR hydrolysis .
Principle of PYR Broth
- Beef heart infusion and peptic digest of animal tissue provide nitrogenous nutrients.
- The sugar dextrose is the carbohydrate serving as an energy source.
- Disodium phosphate serves as buffering agent
- As for sodium chloride,it maintains osmotic balance.
- Chromogenic mixture provides substrate for PYR enzyme.
Composition of PYR Broth
|Beef heart infusion from||500.000|
|Peptic digest of animal tissue||20.000|
|Final pH at temperature at 25°C||7.8±0.2|
Preparation of PYR Broth
- Start by Suspending 37 grams in 1000 ml distilled water
- Heat if necessary to dissolve the medium completely.
- Now mix well and dispense as desired.
- Sterilize by autoclaving at 15 lbs pressure (121°C) for 15 minutes.
Appearance of PYR Broth
- It appears as a Light yellow coloured clear solution
PYR Test Procedure using PYR Broth
- Pick 3-5 colonies of the suspected organisms from a 18-24 hours pure culture plate and Inoculate on PYR broth
- Now incubate the tube aerobically at 35-37°C for 4 hours.
- Add 1-2 drops of PYR reagent directly into the inoculated PYR broth
- Observe for the red color development within 1-2 minutes.
PYR Test Procedure using Disk Method
- Start by wetting the PYR test disc on the strip with 10 µl sterile distilled water or deionized water.
Take Note not to flood the disk
- Using a wire loop,pick up 5-10 colonies of the suspected organism from a 18-24 hours pure culture and make a light smear on the surface of the disc
- Incubate the disc for 1-2 minutes at room temperature.
- After incubation, add 1 drop of PYR reagent (N, N-dimethylaminocinnamaldehyde)
- Observe for red color development within 1-2 minutes.
Interpretation of PYR Test
|Positive||Bright pink or cherry-red color within 1-2 minutes.|
|Negative||No color change or a blue color due to a positive indole reaction|
Cultural Test Response
|Streptococcus pyogenes||positive, red colouration|
|Enterococcus faecalis||positive, red colouration|
- Facklam R. R., Thacker L. G, Fox B., Eriquez L., 1982, J. Clin. Microbiol., 15 (6), a, 987-990.
- MacFaddin J. F., 2000, Biochemical Tests for Identification of Medical Bacteria, 3rd Edition, Lippinocott Williams and Wilkins, N.Y. 407-410.
- Koneman E. W., Allen S. D., Janda W. M., Schreckenberger P. C., Winn W. C. Jr., 1992, Colour Atlas and Textbook of Diagnostic Microbiology, 4th Ed., J. B. Lippinccott Company.
- Murray P. R., Baron J. H., Pfaller M. A., Jorgensen J. H. and Yolken R. H., (Eds.), 2003, Manual of Clinical Microbiology, 8th Ed., American Society for Microbiology, Washington, D.C