Cat. no. C6330 CRITERION™ MIO Medium 62gm
Cat. no. C6331 CRITERION™ MIO Medium 500gm
Cat. no. C6332 CRITERION™ MIO Medium 2kg
Cat. no. C6333 CRITERION™ MIO Medium 10kg
Cat. no. C6334 CRITERION™ MIO Medium 50kg


Hardy Diagnostics CRITERION™ MIO (Motility, Indole, Ornithine) Medium is recommended for use in testing motility, indole production, and ornithine decarboxylase activity of enteric bacilli.

This dehydrated culture medium is a raw material intended to be used in the making of prepared media products, which will require further processing, additional ingredients, or supplements.


Ederer and Clark, et al., developed Motility Indole Ornithine (MIO) Medium to be used as an aid in the identification of members of the Enterobacteriaceae family.(7) Motility, indole production, and ornithine decarboxylation are the three differentiating tests that are provided in the one culture tube.

The addition of agar to the medium allows for the detection of motility along the stab line of inoculation. Motile organisms extend from the stab line and produce turbidity or cloudiness throughout the medium. Non-motile organisms grow only along the stab line and leave the surrounding medium clear.

Tryptophan, present in the basal medium, is degraded by organisms that possess the enzyme tryptophanase. Degradation of tryptophan produces indole which is detected upon addition of Kovacs Reagent to the surface of the medium. Indole combines with p-dimethylaminobenzaldehyde and produces a red band at the top of the medium. A negative indole test results in no color change upon addition of Kovacs Reagent.

Bromcresol purple serves as the pH indicator which allows for detection of decarboxylase activity. Organisms that ferment dextrose will produce acids, thereby lowering the pH. A decreased pH results in the indicator changing from purple to yellow. The presence of acid also results in stimulation of enzyme activity. Once the enzyme has decarboxylated ornithine, the by-product diamine putrescine is produced. Production of putrescine causes an alkaline shift which turns the medium a dark purple. Organisms which do not produce decarboxylase remain acidic due to dextrose-fermentation, and the medium retains a yellow (acidic) color throughout or yellow with a purple band near the top of the tube.


Gram weight per liter: 31.0gm/L
Pancreatic Digest of Gelatin 10.0gm
Pancreatic Digest of Casein 10.0gm
L-Ornithine 5.0gm
Yeast Extract 3.0gm
Dextrose 1.0gm
Bromcresol Purple 0.02gm
Agar 2.0gm

Final pH 6.5 +/- 0.2 at 25ºC.

* Adjusted and/or supplemented as required to meet performance criteria.


Store the sealed bottle(s) containing dehydrated culture medium at 2-30ºC. Dehydrated culture medium is very hygroscopic. Keep lid tightly sealed. Protect dehydrated culture media from moisture and light. The dehydrated culture media should be discarded if it is not free-flowing or if the color has changed from its original beige.

Store the prepared culture media at 15-30ºC.



1. Suspend 31.0gm of the dehydrated culture media in 1 liter of distilled or deionized water.

2. Heat to boiling and mix to dissolve completely.

3. Dispense into tubes.

4. Sterilize in the autoclave at 121ºC. for 15 minutes.


For information on procedures and interpretation of results, consult listed references or refer to the prepared media Instructions for Use (IFU) for Cat. No. Q20.


Weak motile organisms or organisms that possess damaged flagella (due to heating, shaking, or other trauma) often result in false-negative motility tests. Motility results may be confirmed by performing a hanging drop motility test. Consult listed references for procedure.(2-4,6)

Motility and Ornithine results must be interpreted prior to addition of Kovacs Reagent.

A purple color near the top of the tube may result due to oxidation.

Erroneous results may occur if caps are not loose during incubation.


Standard microbiological supplies and equipment such as autoclaves, incinerators, and incubators, etc., are not provided.


Test Organisms Inoculation Method* Incubation Results
Time Temperature Atmosphere
Escherichia coli
ATCC ® 25922
D 18-48hr 35°C Aerobic Growth; motility positive, indole positive (turns pink after adding Kovacs Reagent), ornithine positive (purple throughout tube)
Klebsiella pneumoniae
ATCC ® 13883
D 18-48hr 35°C Aerobic Growth; motility negative, indole negative, ornithine negative (purple top layer, rest of tube yellow)

User Quality Control


CRITERION™ MIO Medium powder should appear homogeneous, free-flowing, and beige in color. The prepared media should appear clear, semi-solid, and purple in color.


1. Anderson, N.L., et al. Cumitech 3B; Quality Systems in the Clinical Microbiology Laboratory, Coordinating ed., A.S. Weissfeld. American Society for Microbiology, Washington, D.C.

2. Jorgensen., et al. Manual of Clinical Microbiology, American Society for Microbiology, Washington, D.C.

3. Tille, P., et al. Bailey and Scott's Diagnostic Microbiology, C.V. Mosby Company, St. Louis, MO.

4. Koneman, E.W., et al. Color Atlas and Textbook of Diagnostic Microbiology, J.B. Lippincott Company, Philadelphia, PA.

5. Isenberg, H.D. Clinical Microbiology Procedures Handbook, Vol. I, II & III. American Society for Microbiology, Washington, D.C.

6. MacFaddin, J.F. Biochemical Tests for Identification of Medical Bacteria,, Lipincott Williams & Wilkins, Philadelphia, PA.

7. Ederer and Clark. 1970. Appl. Microbiol.; 2:849.

ATCC is a registered trademark of the American Type Culture Collection.