L-Pyrrolidonyl Arylamidase (PYR) test is a biochemical assay used in microbiology to detect specific bacterial species, specifically streptococci vs enterococci. The test detects the enzyme L-pyrrolidonyl arylamidase.
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L-Pyrrolidonyl Arylamidase (PYR)
L-Pyrrolidonyl Arylamidase (PYR) is a bacterial enzyme that breaks down the substrate L-pyrrolidonyl-β-naphthylamide. The PYR test, a quick and specific biochemical assay used in microbiology to detect some bacteria, mainly streptococci and enterococci, is based on the activity of this enzyme.
Key Features:
L-Pyrrolidonyl Arylamidase
L-Pyrrolidonyl Arylamidase is an enzyme that breaks down L-pyrrolidonyl-β-naphthylamide into β-naphthylamine.
Bacterial Identification: The presence of PYR enzyme activity distinguishes certain bacterial species, most notably Streptococcus pyogenes (Group A Streptococcus) and Enterococcus spp. These species are PYR-positive, implying that they synthesize the enzyme.
Biochemical Assay
The PYR test utilizes a substrate (L-pyrrolidonyl-β-naphthylamide) that is digested by the enzyme to yield β-naphthylamine. When this substance combines with a color developer (such as p-dimethylaminocinnamaldehyde), it produces a red compound, signifying a favorable outcome.
Clinical Relevance
Identifying bacteria that make PYR is critical for identifying infections and directing treatment. Streptococcus pyogenes, for example, causes strep throat and other dangerous infections, whereas Enterococcus spp. are significant in urinary tract infections and other nosocomial.
Principle
The PYR test detects the enzyme L-pyrrolidonyl arylamidase, which converts the substrate L-pyrrolidonyl-β-naphthylamide into β-naphthylamine. This chemical combines with a color developer (often p-dimethylaminocinnamaldehyde) to generate a bright red color, indicating a favorable outcome.
Requirements
- PYR disks or strips contain the substrate L-pyrrolidonyl-β-naphthylamide.
- Sterile water or saline: To moisten the disk or strip.
- To transmit bacterial colonies, use an inoculation loop or wooden applicator sticks.
- PYR reagent (color developer) is typically p-dimethylaminocinnamaldehyde.
- Control organisms: Known positive and negative controls are used to validate the test results.
- Incubator (optional): For incubation at 35-37°C as needed.
Procedure
Preparation:
Moisten a PYR disc or strip with a drop of sterile water or saline.
Inoculation:
Using an inoculating loop or a wooden applicator stick, apply a little amount of the test organism (usually from a pure culture) to the surface of the wet PYR disc or strip.
Incubation:
Allow the infected disc or strip to rest at room temperature for 2-5 minutes to ensure optimal absorption and interaction.
Development:
Apply one or two drops of PYR reagent to the disc or strip.
Observation:
Within a few minutes, a red colour should appear.
Results
Positive results: The appearance of a brilliant red colour within 1-2 minutes confirms the existence of L-pyrrolidonyl arylamidase.
Negative result: The absence of red colour (the disc or strip remains the same colour or becomes yellow) shows the absence of L-pyrrolidonyl arylamidase.
Interpretation
Positive organisms for the PYR test include Streptococcus pyogenes (Group A Streptococcus) and Enterococcus spp.
Negative organisms: Most other streptococci (except Group A) and many other gram-positive and gram-negative bacteria do not produce L-pyrrolidonyl arylamidase, resulting in a negative test.
Quality Control
Positive Control: A PYR-positive bacterium (such as Enterococcus faecalis).
A PYR-negative bacterium, such as Streptococcus agalactiae, serves as a negative control.
The accuracy and reliability of the PYR test are dependent on proper method, ideal incubation conditions, and the use of control organisms to verify the results. This test is a valuable tool for quickly identifying specific bacterial species in clinical microbiology laboratories.
Frequently Asked Question
What is L-Pyrrolidonyl Arylamidase (PYR) ?
L-Pyrrolidonyl Arylamidase (PYR) is a bacterial enzyme that breaks down the substrate L-pyrrolidonyl-β-naphthylamide. The PYR test, a quick and specific biochemical assay used in microbiology to detect some bacteria, mainly streptococci and enterococci, is based on the activity of this enzyme.
What does a positive Pyr test indicate?
A positive PYR test presumptively identifies group A streptococci (Streptococcus pyogenes) and group D enterococci.
What are the requirements of L-Pyrrolidonyl Arylamidase (PYR) ?
The requirements of L-Pyrrolidonyl Arylamidase (PYR) are
1. PYR disks or strips contain the substrate L-pyrrolidonyl-β-naphthylamide.
2. Sterile water or saline: To moisten the disk or strip.
3. To transmit bacterial colonies, use an inoculation loop or wooden applicator sticks.
4. PYR reagent (color developer) is typically p-dimethylaminocinnamaldehyde.
5. Control organisms: Known positive and negative controls are used to validate the test results.
6. Incubator (optional): For incubation at 35-37°C as needed.
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