Formula

Columbia Agar Base, 42.5 g

Lead Acetate, 0.2 g

Hemin (0.1% soln), 5.0 ml

Menadione, 0.01 g

Glutathione, 1.0 g

Distilled Water, 1000.0 ml

 

Final pH 7.1 +/- 0.3 at 25 degrees C.

Final weight 16.0 g +/- 1.6 for AS-6430.

 

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 due to oxidation of media) 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:  OHO-C 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, possibly 3 – 5 days for hydrogen sulfide detection.  Detailed instructions for processing anaerobic cultures can be found in the appropriate references.  Black to grey halos surrounding colonies or black color in the center of colonies usually signifies positive hydrogen sulfide production.

 

Materials Required But Not Provided

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

 

Limitations

OHO-C will not provide complete information for 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.  Consult reference materials for additional information.

 

Quality Control

OHO-C should support good growth and hydrogen sulfide detection of oral anaerobic bacteria found clinical specimens.

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

Organism Tested	ATCC #	Results	Time (Hours)	Special Reaction
Bacteroides fragilis	25285	Growth	24 h
Prevotella melaninogenica	25845	Growth	24 h
Fusobacterium necrophorum	25286	Growth	24 h	+ H2S *
Fusobacterium nucleatum	25586	Growth	24 h	+ H2S *
Clostridium perfringens	13124	Growth	24 h	+ H2S *
Peptostreptococcus anaerobius	27337	Growth	24 h
Peptostreptococcus magnus	29328	Growth	24 h
Propionibacterium acnes   or  Clostridium difficile	6919      9686	Growth	24 – 48 h 24 h	+ H2S *
Enterococcus faecalis	29212	Growth	24 h
Campylobacter rectus	33238	Growth	24 – 48 h
Veillonella parvula	10790	Growth	24 h	+ H2S *

                                   * H₂S production may require 3 – 5 days of incubation.

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/reactivity capacity of this medium is to be tested for performance, it is recommended that the following ATCC organisms be evaluated for growth/reactivity.

 

Organism	Growth in Hours	Expected Results
F. necrophorum	24 hrs	+ H2S
F. nucleatum	24 hrs	+ H2S
C. perfringens	24 hrs	+ H2S
V. parvula	24 hrs	+ H2S
B. fragilis	24 hrs
P. anaerobius	24 hrs

 

Physical Appearance:  OHO-C should appear slightly opaque and yellowish green in color in a standard sized Petri dish (AS-6430) 

 

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

 

3.        Turng, B-F, Guthmiller, JM, Minhah, GE, Falkler, WA.  1996.  Development and Evaluation of a Selective and Differential Medium for the Primary Isolation of Peptostreptococcus micros.  Oral Microbiol Immunol 5: 356 – 361.

 

4.        Turng B-F, Minah GE, Falkler WA. 1997.  Development of an Agar Medium for the Detection of Oral H₂S Producing Organisms.  J Dent Res 76 (Spec Issue): 266, abstract # 1702.

 

5.        El-Halabi M, Minah G, Turng B, Zhang M.  1999.  Correlation between Volatile Sulfur Levels and Odorigenic Bacteria in Saliva.  J Dent Res 78 (Spec Issue): abstract # 1478.

 

6.        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.

 

7.        Engelkirk, P. G., Duben-Engelkirk, J. and Dowell, V. R.  1992.  Principles and Practices of Clinical Anaerobic Bacteriology.  Star Publishing Co., Belmont, CA 94002.

 

8.        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.