Phenylethyl Alcohol Blood Agar (PEA) is an enriched selective medium for the isolation of obligately anaerobic bacteria from clinical specimens. Phenylethyl Alcohol Blood Agar medium contains phenylethyl alcohol to inhibit the growth of gram-negative, facultatively-anaerobic bacteria and prevent swarming. This medium is based on the Brucella Agar forumulation and will support the growth of most obligate anaerobic bacteria, both gram positive and gram negative. The Phenylethyl Alcohol Blood Agar medium is especially useful in selective isolation of anaerobes from mixed populations that contain rapidly growing gram-negative bacteria such as Proteus species. This medium is prepared, stored and dispensed under oxygen-free conditions to prevent the formation of oxidized products prior to use. 

   Anaerobe Systems PRAS Phenylethyl Alcohol
                   Blood Agar (PEA) AS-113

Formula

Pancreatic Digest of Casein, 10.0 g

Soy Peptone, 3.0 g

Animal Tissue, 10.0 g

Dextrose, 1.0 g

Yeast Extract, 2.0 g

Sodium Chloride, 5.0 g

Sodium Bisulfite, 0.1 g

Hemin (0.1% soln), 5.0 ml

Vitamin K₁ (1% soln), 1.0 ml

L-Cystine, 0.4 g

Agar, 15.0 g

Sheep Blood, 45.5 ml

Phenylethyl Alcohol, 2.7 ml

Distilled Water, 1000.0 ml

 

Final pH 7.1 +/- 0.4 at 25 degrees C.

Final weight 16.0 g +/- 1.6 for monoplate.

Final weight 8.0 g +/- 0.8 for Bi-plate. 

Precautions

For IN VITRO DIAGNOSTIC USE only.  Approved biohazard precautions and aseptic techniques should be observed when using this product.  This product is for use only by properly-trained and qualified personnel.  Sterilize all biohazard waste prior to disposal.

Storage and Shelf Life

Storage:  Upon receipt, store at room temperature (13°C - 27°C) in original container until use.  Avoid overheating or freezing.  Do not use medium if there are signs of deterioration (shrinking, cracking or discoloration) or contamination.  The expiration date applies to the product in its original packaging and stored as directed.  Do not use product past the expiration date shown on the container.

Shelf Life:  90 days from date of manufacture. 

Procedure

Specimen Collection:  Specimens for anaerobic culture should be protected from air (oxygen) during collection, transport and processing.  Consult appropriate references for detailed instructions concerning collection and transport of anaerobes.

Methods for Use:  Phenylethyl Alcohol Blood Agar should be inoculated directly with clinical material or a broth that has been previously inoculated from clinical material.  Inoculated plates should be streaked to obtain isolated colonies, immediately placed in an anaerobic atmosphere and incubated at 35-37^oC for 18-48 hours.  Additional periods of incubation may be necessary to recover some anaerobes.  Detailed instructions for processing anaerobic cultures can be found in the appropriate references.

Materials Required But Not Provided

Standard microbiological supplies and equipment such as loops, saline blanks, slides, microscope, incinerator / autoclave, incubators, anaerobic chamber, other culture media and serological and biochemical reagents.

Limitations

Phenylethyl Alcohol Blood Agar will not provide complete information for the identification of bacterial isolates.  Additional test procedures and media are required for complete identification. It is recommended that a non-selective medium such as Anaerobic Brucella Blood Agar also be inoculated from clinical specimens to assure growth of all species present.  Some strains of facultative organisms, which are normally inhibited, may grow on the Phenylethyl Alcohol Blood Agar medium.  A test for aerotolerance should be used to confirm that each colony type is an obligate anaerobe.  Consult reference material for additional information.

Quality Control

This medium should support good growth of anaerobes found in clinical infections.  In addition, Phenylethyl Alcohol Blood Agar should inhibit the growth of facultatively-anaerobic, gram-negative rods such as Escherichia coli and the swarming of Proteus mirabilis. 

The following organisms are routinely used for quality control performance testing at Anaerobe Systems.

Organism Tested	ATCC #	Results	Time (Hours)
Bacteroides fragilis	25285	Growth	24 h
Prevotella melaninogenica	25845	No Growth
Fusobacterium necrophorum	25286	Growth	24 h
Fusobacterium nucleatum	25586	No Growth
Clostridium perfringens	13124	Growth	24 h
Peptostreptococcus anaerobius	27337	Growth	24 h
Staphylococcus aureus or    Enterococcus faecalis	25923     29212	Growth	24 h
Escherichia coli	25922	Inhibited to No Growth
Proteus mirabilis	12453	Inhibited to No Growth
Propionibacterium acnes  or  Clostridium difficile	6919     9689	Growth	24 – 48 h 24 h

 

 User Quality Control: The final determination to the extent and quantity of user laboratory quality control must be determined by the end user.

If sterility testing is to be performed on this product, the appropriate percentage of the original shipment amount should be incubated anaerobically and aerobically for 48 – 96 hours.

If the nutritive/inhibitory capacity of this media is to be tested for performance, it is recommended that the following ATCC organisms be evaluated for growth/inhibition.

 

Organism	Expected Growth
B. fragilis	24 hrs
P. melaninogenica	NG
F. nucleatum	Inhibited
E. coli	Inhibited

 

Physical Appearance:  Phenylethyl Alcohol Blood Agar should appear opaque burgundy red in color.

 

References

1.        Dowell, V. R., Jr. and T. M. Hawkins. 1974. Laboratory Methods in Anaerobic Bacteriology.  CDC Laboratory Manual. USDHEW C. D. C. Atlanta, GA 30333.

 

2.        Dowell, V. R., Jr. and G. L. Lombard. 1977. Presumptive Identification of Anaerobic Non-sporeforming Gram-negative Bacilli. USDHEW, CDC. Atlanta, GA 30333.

 

3.        Dowell, V. R., Jr., G. L. Lombard, F. S. Thompson and A. Y. Armfiedl. 1977. Media for the Isolation, Characterization, and Identification of Obligately Anaerobic Bacteria. USDHEW, CDC, Atlanta, GA 30333.

 

4.        Holdeman, L. V., F. P. Cato and W. E. C. Moore. 1977. Anaerobe Laboratory Manual.  Virginia Polytechnic Institute and State University. Blacksburg, VA 24061.

 

5.        Jousimies-Somer, H. R., Summanen, P., Citron, D. M., Baron, E. J., Wexler, H. M. and S. M. Finegold. 2002. Wadsworth – KTL Anaerobic Bacteriology Manual. Star Publishing Co., Belmont, CA 94002

 

6.        NCCLS. Quality Control for Commercially Prepared Microbiological Culture Media; Approved Standard- Third Edition. (2004). NCCLS document M22-A3. NCCLS, 940 West Valley Road, Suite 1400, Wayne, PA 19087-1898.