Formula
Peptone
Yeast Extract Broth Basal Medium:
Pancreatic Digest of Casein, 20.0 g
Yeast Extract, 10.0
g
Resazurin, 0.001
g
Calcium Chloride, anhydrous, 0.008 g
Magnesium Sulfate, anhydrous, 0.008 g
Potassium Phosphate, monobasic, 0.04 g
Potassium Phosphate, dibasic, 0.04 g
Sodium Chloride, 0.08
g
Sodium Bicarbonate, 0.4
g
Hemin (0.1% Soln), 5.0
ml
Vitamin K1 (1% Soln), 0.1ml
L-Cysteine, 0.5
g
Distilled Water, 1000.0
ml
The following components have been added per liter
to the basal medium to form the following products.
PY-Adonitol: Adonitol, 5.0 g
PY-Amygdalin: Amygdalin, 5.0 g
PY-Arabinose: Arabinose, 5.0 g
PY-Arginine: Arginine, 3.0 g
PYG-Bile: Oxgall, 20.0 g
Glucose, 10.0 g
PY-Cellobiose: Cellobiose, 10.0 g
PY-Dulcitol: Dulcitol, 10.0 g
PY-Erythritol: Erythritol, 5.0 g
PY-Esculin: Esculin, 5.0 g
PYG: Glucose, 10.0g
PY-Fructose: Fructose, 10.0 g
PY-Galactose: Galactose, 10.0 g
PYG-Gelatin: Gelatin, 120.0 g
Glucose, 10.0 g
PY-Glycerol: Glycerol, 8.0 ml
PY-Glycogen: Glycogen, 5.0 g
PY-Inositol: Inositol, 10.0 g
PY-Inulin: Inulin, 10.0 g
PY-Lactose: Lactose, 10.0 g
PY-Maltose: Maltose, 10.0 g
PY-Mannitol: Mannitol, 10.0 g
PY-Mannose: Mannose, 10.0 g
PY-Melezitose: Melezitose, 5.0 g
PY-Melibiose: Melibiose, 5.0 g
PY-Raffinose: Raffinose, 10.0 g
PY-Rhamnose: Rhamnose, 10.0 g
PY-Ribose: Ribose, 10.0 g
PY-Salicin: Salicin, 10.0 g
PY-Sorbitol: Sorbitol, 10.0 g
PY-Starch: Starch, 10.0 g
PY-Sucrose: Sucrose, 10.0 g
PY-Trehalose: Trehalose, 5.0 g
PY-Xylan: Xylan, 10.0 g
PY-Xylose: Xylose, 10.0 g
Final pH 7.3+/- 0.5 at 25 degrees C.
Final volume 7.0 ml +/- 0.7.
Final volume 5.0 ml +/- 0.5 for AS-860.
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 packaging until
use. Avoid overheating or freezing. Do not use medium if there are signs of
deterioration (discoloration or evaporation), oxygen exposure, 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: 1 year from date of manufacture.
Procedure
Specimen Collection:
Specimens for anaerobic culture should be protected from air (oxygen) during
collection, transport and processing of specimens. Consult appropriate references for detailed
instructions concerning collection and transport of anaerobes.
Methods for Use: Peptone
Yeast Extract Broth Based Media should be inoculated with pure cultures of an
organism. These media are supplied with
a screw cap tube fitted with a Hungate cap, which allows for direct injection
of inoculum using a syringe.
Decontaminate the rubber septa with alcohol and pierce with the needle;
the inoculum can then be syringed into the medium. Inoculated tubes should be immediately placed
in an anaerobic atmosphere and incubated at 35 – 37oC for 24 – 48
hours. Fermentation is indicated when
the pH of inoculated substrate is lowered after incubation. Detailed instructions for processing
anaerobic cultures can be found in the appropriate references.
pH Measurements: Once sufficient growth is achieved, the pH
can be read directly from the culture tube using a pH meter and a long thin
combination electrode. Thoroughly rinse
the electrode with distilled water into a container containing disinfectant
after reading each strain. Disinfect the
electrode before returning it to the storage solution.
Interpretation of pH: A pH value of 5.5 to 6.0 is considered a weak
acid production while a value of 5.5 or less is strong acid production. A pH above 6.0 is no acid production.
Esculin Hydrolysis
Detection: Add 2 to 3 drops of a 1%
solution of ferric ammonium citrate to an inoculated tube of PY-Esculin with
sufficient growth (7). Blackening of the
medium indicates that the esculin present has been hydrolyzed.
Starch Hydrolysis
Detection: Add 2 to 3 drops of diluted
1:5 of Gram’s iodine to an inoculated PY-Starch tube with sufficient
growth. No color change indicates hydrolysis
of starch, the formation of a blue / black color indicates the presence of
starch.
Gelatin Hydrolysis
Detection: Place a 24 – 72 hour culture
with an un-inoculated control tube in a 2 – 8oC environment. The un-inoculated tube will solidify, and, if the inoculated tube can no longer
solidify, then the gelatin present has been hydrolyzed.
PYG-Bile Growth
Determination: Turbidity in the medium
is compared to that of PYG to determine if the organism is inhibited or
stimulated by bile.
Materials Required But Not Provided
Standard microbiological supplies and equipment such as
loops, syringe needle, pipettes, saline blanks, disinfectant, slides, staining
supplies, microscope, incinerator or autoclave, incubators, anaerobic chamber
or anaerobic jars, disinfectant, other culture media and serological and
biochemical reagents.
Limitations
Peptone Yeast Extract Broth Based Media will not provide
complete information for the identification of bacterial isolates. Additional test procedures and culture media
are required for complete identification.
Consult reference materials for additional information.
Quality Control
Un-inoculated Peptone Yeast Extract Broth Based Media
should show only trace amounts, if any, of volatile and nonvolatile fatty acids
when tested with gas liquid chromatography as described (1,4). PYG cultures of appropriate control strains,
such as Bacteroides fragilis,
Fusobacterium necrophorum and Clostridium
perfringens, should have characteristic metabolic products when tested with
gas chromatography. If used properly, the
Peptone Yeast Extract Broth Based Media should support good growth of anaerobes
isolated from clinical specimens.
Growth performance:
The following organisms are routinely used at Anaerobe Systems for
quality assurance performance testing for growth and carbohydrate/biochemical
utilization of the Peptone Yeast Extract Broth Based Media.
|
Organism Tested
|
ATCC #
|
Results
|
Time (Hours)
|
|
Bacteroides vulgatus
|
8482
|
Growth
|
24 h
|
|
Bacteroides fragilis
|
25285
|
Growth
|
24 h
|
|
Prevotella melaninogenica
|
25845
|
Growth*
|
24 – 48 h
|
|
Fusobacterium necrophorum
|
25286
|
Growth*
|
24 – 48 h
|
|
Fusobacterium nucleatum
|
25586
|
Growth*
|
24 – 48 h
|
|
Clostridium perfringens
|
13124
|
Growth
|
24 h
|
|
Peptostreptococcus anaerobius
|
27337
|
Growth*
|
24 h
|
|
Clostridium novyi, Type A (PY and PYG only)
|
7659
|
Growth
|
48 – 72 h
|
|
Propionibacterium acnes (PY,
PY-Adonitol, PYG, and PY-Glycerol only)
|
6919
|
Growth
|
24 – 48 h
|
|
Staphylococcus aureus (PY and
PYG only)
|
25923
|
Growth
|
24 h
|
|
Peptoniphilus asaccharolyticus (PY-Fructose,
PYG-Gelatin, and PYG only)
|
29743
|
Growth*
|
24 – 48 h
|
*
No Growth for PYG-Bile
Biochemical utilization: The following additional organisms are
routinely used for each biochemical utilization at Anaerobe Systems and are
determined on the basis of pH with the production of acid. A strong acid will result in a pH of 5.5 or
below, weak acid results in a pH of 5.5 – 6.0 and no acid production results in
a pH of 6.0 or above.
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 capacity of this media is to be
tested for performance, it is recommended that the following ATCC organisms be evaluated for growth. Carbohydrate / biochemical utilization is
then determined by pH analysis.
|
Product
|
Organisms Tested
|
ATCC #
|
Pos
pH/hy
|
Organisms Tested
|
ATCC #
|
Neg
pH/hy
|
|
PY
|
Bacteroides fragilis
|
25285
|
G
|
|
|
|
|
Fusobacterium necrophorum
|
25286
|
G
|
|
|
|
|
PY-Adonitol
|
Klebsiella pneumoniae
|
49472
|
+
|
Bacteroides fragilis
|
25285
|
-
|
|
PY-Amygdalin
|
Bifidobacterium adolescentis
|
15703
|
+
|
Bacteroides vulgatus
|
8482
|
-
|
|
PY-Arabinose
|
Bacteroides vulgatus
|
8482
|
+
|
Bacteroides fragilis
|
25285
|
-
|
|
PY-Arginine
|
Eubacterium lentum
|
43055
|
G stimulated, compared to PY
|
|
|
|
PYG-Bile
|
Bacteroides vulgatus
|
8482
|
G
|
Peptoniphilus asaccharolyticus
|
29743
|
NG
|
|
PY-Cellobiose
|
Bifidobacterium breve
|
15700
|
+
|
Peptostreptococcus anaerobius
|
27337
|
-
|
|
PY-Dulcitol
|
Kluyvera intermedia
|
700722
|
+
|
Bacteroides fragilis
|
25285
|
-
|
|
PY-Erythritol
|
Eubacterium limosum
|
10825
|
+
|
Bacteroides vulgatus
|
8482
|
-
|
|
PY-Esculin
|
Bacteroides fragilis
|
25285
|
( /+)
|
Peptostreptococcus anaerobius
|
27337
|
( /-)
|
|
PY-Fructose
|
Bacteroides fragilis
|
25285
|
+
|
Peptoniphilus asaccharolyticus
|
29743
|
-
|
|
PY-Galactose
|
Bacteroides fragilis
|
25285
|
+
|
Fusobacterium necrophorum
|
25286
|
-
|
|
PYG-Gelatin
|
Clostridium perfringens
|
13124
|
+
|
Bifidobacterium longum
|
15707
|
-
|
|
PYG
|
Bacteroides fragilis
|
25285
|
+
|
Peptoniphilus asaccharolyticus
|
29743
|
-
|
|
PY-Glycerol
|
Propionibacterium acnes
|
6919
|
+
|
Bacteroides fragilis
|
25285
|
-
|
|
PY-Glycogen
|
Bacteroides vulgatus
|
8482
|
+
|
Fusobacterium necrophorum
|
25286
|
-
|
|
PY-Inositol
|
Clostridium perfringens
|
13124
|
+
|
Peptostreptococcus anaerobius
|
27337
|
-
|
|
PY-Inulin
|
Bifidobacterium adolescentis
|
15703
|
+
|
Peptostreptococcus anaerobius
|
27337
|
-
|
|
PY-Lactose
|
Clostridium perfringens
|
13124
|
+
|
Fusobacterium necrophorum
|
25286
|
-
|
|
PY-Maltose
|
Bacteroides vulgatus
|
8482
|
+
|
Fusobacterium necrophorum
|
25286
|
- |
|
|
|
|
|
|
|
|
Product
|
Organisms Tested
|
ATCC #
|
Pos
pH/hy
|
Organisms Tested
|
ATCC #
|
Neg
pH/hy
|
|
PY-Mannitol
|
Clostridium tertium
|
19405
|
+
|
Fusobacterium necrophorum
|
25286
|
-
|
|
PY-Mannose
|
Clostridium perfringens
|
13124
|
+
|
Fusobacterium nucleatum
|
25586
|
-
|
|
PY-Melezitose
|
Bifidobacterium longum
|
15707
|
+
|
Peptostreptococcus anaerobius
|
27337
|
-
|
|
PY-Melibiose
|
Bifidobacterium longum
|
15707
|
+
|
Peptostreptococcus anaerobius
|
27337
|
-
|
|
PY-Raffinose
|
Bacteroides vulgatus
|
8482
|
+
|
Fusobacterium necrophorum
|
25286
|
-
|
|
PY-Rhamnose
|
Bacteroides vulgatus
|
8482
|
+
|
Peptostreptococcus anaerobius
|
27337
|
-
|
|
PY-Ribose
|
Bifidobacterium longum
|
15707
|
+
|
Bacteroides fragilis
|
25285
|
-
|
|
PY-Salicin
|
Bifidobacterium adolescentis
|
15703
|
+
|
Peptostreptococcus anaerobius
|
27337
|
-
|
|
PY-Sorbitol
|
Bifidobacterium adolescentis
|
15703
|
+
|
Peptostreptococcus anaerobius
|
27337
|
-
|
|
PY-Starch
|
Clostridium perfringens
|
13124
|
(+/+)
|
Fusobacterium necrophorum
|
25286
|
(-/-)
|
|
PY-Sucrose
|
Clostridium perfringens
|
13124
|
+
|
Peptostreptococcus anaerobius
|
27337
|
-
|
|
PY-Trehalose
|
Bacteroides ovatus
|
8483
|
+
|
Peptostreptococcus anaerobius
|
27337
|
-
|
|
PY-Xylan
|
Bacteroides ovatus
|
8483
|
+
|
Bacteroides fragilis
|
25285
|
-
|
|
PY-Xylose
|
Bifidobacterium longum
|
15707
|
+
|
Fusobacterium nucleatum
|
15586
|
-
|
G
= Growth
Physical Appearance: Peptone Yeast Extract Broth Based Media should appear as clear golden-yellow to clear golden-brown
liquid within the 16mm x 100mm glass tube.
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.
Armfield. 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.
7.
Isenberg, H. D. 1992. Clinical
Microbiology Procedures Handbook.
American Society for Microbiology Publishing, Washington DC.
