MIL (MOTILITY, INDOLE, LYSINE) MEDIUM

Cat. no. Q09 MIL Medium, 16x100mm Tube, 8ml Deep 20 or 100 tubes/Box

INTENDED USE

Hardy Diagnostics MIL Media is used to differentiate Enterobacteriaceae on the basis of motility, indole production, lysine-decarboxylation and lysine-deamination.

SUMMARY

MIL Medium is used to test for motility, lysine-decarboxylation, lysine-deamination and indole production. MIL Media gives equivalent reactions to those obtained by traditional single tube tests (i.e. Moeller's Lysine, Motility Test Medium). These tests are useful in differentiating within the Enterobacteriaceae.

MIL Media, when used in conjunction with TSI Agar (and, when necessary, Urea Agar) provide the necessary information for the presumptive identification of the enteric pathogens within the family Enterobacteriaceae. (2) For more information on differentiation within the family Enterobacteriaceae, consult listed references. (4,5)

FORMULA

Ingredients per liter of deionized water:*

Peptone 10.0gm
Tryptone 10.0gm
Lysine 10.0gm
Yeast Extract 3.0gm
Dextrose 1.0gm
Ferric Ammonium Citrate 0.5gm
Bromcresol Purple 20.0mg
Agar 2.0gm

Final pH 6.6 +/- 0.2 at 25ºC.

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

STORAGE AND SHELF LIFE

Storage: Upon receipt store at 15-30ºC. away from direct light. Media should not be used if there is any signs of deterioration (shrinking, cracking, or discoloration), contamination, or if the expiration date has passed. Product is light and temperature sensitive; protect from light, excessive heat, moisture, and freezing.

PRECAUTIONS

PROCEDURE

1. Prepare inoculum from an 18-24 hour isolate in pure culture.

2. Using an inoculating needle stab the medium to within one half inch from the bottom of the tube.

3. Incubate aerobically at 35ºC. for 18-24 hours.

4. Read and record results of lysine and motility.

5. Add 3-4 drops of Kovacs Reagent (Cat. no. Z67), read for indole reaction, and record result.

Note: Read lysine and motility prior to the addition of Kovacs Reagent

INTERPRETATION OF RESULTS

Motility
Growth away from the stab line is a positive test for motility. Growth only along the stab line is a negative test.

Lysine-Decarboxylation
A positive test for lysine-decarboxylase is a purple band and a purple butt. A negative test is a narrow purple band and a yellow butt.

Lysine-Deamination
A positive test for lysine-deamination is a deep red band with a yellow butt. A negative test is a purple band with a yellow butt.

Indole
A color change in the reagent layer from yellow to red or pink after addition of Kovacs reagent is a positive test. A negative test is a yellow to bright yellow color after the addition of the reagent.

LIMITATIONS

The indole test may produce false-negative or weak positive reactions. (1)

MATERIALS REQUIRED BUT NOT PROVIDED

Standard microbiological supplies and equipment such as loops, other culture media, swabs, applicator sticks, Kovacs Reagent (Cat. no. Z67), incinerators, and incubators, etc., as well as serological and biochemical reagents, are not provided.

QUALITY CONTROL

Test Organisms Inoculation Method* Incubation Results
Time Temperature Atmosphere
Escherichia coli
ATCC ® 25922
D 18-24hr 35°C Aerobic Motility: positive
Indole: positive
Lysine: positive

Proteus mirabilis
ATCC ® 12453
D 18-24hr 35°C Aerobic Motility: positive
Indole: negative
Lysine: negative

Klebsiella pneumoniae
ATCC ® 13883
D 18-24hr 35°C Aerobic Motility: negative
Indole: negative
Lysine: positive

User Quality Control

PHYSICAL APPEARANCE

MIL Medium should appear clear, and reddish-purple in color.

E. coli growing in MIL Medium

Escherichia coli (ATCC ® 25922) growing in MIL Medium (Cat. no. Q09). Incubated aerobically for 24 hours at 35ºC.

Positive Indole Test

Escherichia coli (ATCC ® 25922) growing in MIL Medium (Cat. no. Q09). Incubated aerobically for 24 hours at 35ºC. Five drops of Indole Kovacs Reagent (Cat. no. Z67) were added subsequent to incubation. The red color development is indicative of a positive reaction.

P. mirabilis growing in MIL Medium

Proteus mirabilis (ATCC ® 12453) growing in MIL Medium (Cat. no. Q09). Incubated aerobically for 24 hours at 35ºC.

Negative Indole Test

Klebsiella pneumoniae (ATCC ® 13883) growing in MIL Medium (Cat. no. Q09). Incubated aerobically for 24 hours at 35ºC. Five drops of Indole Kovacs Reagent (Cat. no. Z67) were added subsequent to incubation. No red color development is indicative of a negative reaction.



K. pneumoniae growing in MIL Medium

Klebsiella pneumoniae (ATCC ® 13883) growing in MIL Medium (Cat. no. Q09). Incubated aerobically for 24 hours at 35ºC.

Negative Indole Test

Klebsiella pneumoniae (ATCC ® 13883) growing in MIL Medium (Cat. no. Q09). Incubated aerobically for 24 hours at 35ºC. Five drops of Indole Kovacs Reagent (Cat. no. Z67) were added subsequent to incubation. No red color development is indicative of a negative reaction.



MIL Medium

Uninoculated tube of MIL Medium (Cat. no. Q09).





REFERENCES

1. MacFaddin, J.F. 1985. Media for Isolation, Cultivation, Identification, Maintenance of Bacteria, Vol. I. Williams & Wilkins, Baltimore, MD.

2. Reller, L.B. and Mirrett, S. 1975. Motility-Indole-Lysine medium for presumptive identification of enteric pathogens of Enterobacteriaceae. J. Clin. Microbiol.; 2 (3), 247-252.

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

4. Baron, E.J., L.R. Peterson, and S.M. Finegold. 1994. Bailey & Scott's Diagnostic Microbiology, 9th ed. Mosby, St. Louis, MO.

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


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

031016gr