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Motility Test Medium (Edwards and Ewing)

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M930
For testing motility of enteric bacteria.

Intended Use:

Recommended for testing motility of enteric bacteria.

Composition**

Ingredients g/L
Peptone 10.000
HM peptone B # 3.000
Sodium chloride 5.000
Agar 4.000
Final pH (at 25°C) 7.4±0.2

**Formula adjusted, standardized to suit performance parameters

# - Equivalent to Beef extract

Directions

Suspend 22.0 grams in 1000 ml purified/distilled water. Heat to boiling to dissolve the medium completely. Dispense 8 ml amounts in test tubes and sterilize by autoclaving at 15 lbs pressure (121°C) for 15 minutes. Cool the tubed medium in an upright position.

Principle And Interpretation

Bacterial motility can be observed directly on microscopic slide or it can be visualized on motility media having agar concentration of 0.4% or less (1). Use of such semisolid media to observe or detect motility was reported by Tittsler and Sandholzer (2). Motility Test Medium is the modification of the original formulation as per Edwards and Ewing and is used for testing motility of Enterobacteriaceae (3). Motility can be visualized as a diffused zone of growth flaring out from the line of inoculation (4,5,6).

Peptone and HM peptone B serve as sources of nitrogen, carbon, long chain amino acids and other essential growth nutrients required for bacterial metabolism. Sodium chloride maintains the osmotic equilibrium of the medium. Small amount of agar helps to create a semisolid medium.

Bacterial motility can be observed directly by examination of the tubes following incubation. Inoculation is done by stabbing through the centre of the medium. Incubate at appropriate temperature for 18 to 40 hours. Non-motile organisms grow only along the line of inoculation whereas motile organisms grow away from the line of inoculation or may show growth even throughout the medium. All weak or equivocal motility results should be confirmed by flagellum stain or by direct wet microscopy (hanging drop) (7,8). To enhance the visibility of bacterial growth 2,3,5 Triphenyl Tetrazolium Chloride (TTC)(FD057) may be added. Tetrazolium salts are colourless but are converted into insoluble formazan, a red coloured complex by the reducing properties of growing bacteria. In Motility Test Medium containing tetrazolium, the development of this red colour helps to trace the spread of bacteria from the inoculation line. The motility of Listeria monocytogenes is frequently best observed in medium without TTC.

Type of specimen

Pure isolates from clinical and non-clinical samples

Specimen Collection and Handling:

With inoculating needle, stab centre of medium to approximately one-half of depth (7).

For clinical samples follow appropriate techniques for handling specimens as per established guidelines (8,9).

After use, contaminated materials must be sterilized by autoclaving before discarding.

Warning and Precautions :

In Vitro diagnostic use. For professional use only. Read the label before opening the container. Wear protective gloves/ protective clothing/eye protection/face protection. Follow good microbiological lab practices while handling specimens and culture. Standard precautions as per established guidelines should be followed while handling specimens. Safety guidelines may be referred in individual safety data sheets.

Limitations :

  • Growth from an 18-24 hr pure culture should be used (7).

Performance and Evaluation

Performance of the medium is expected when used as per the direction on the label within the expiry period when stored at recommended temperature.

Quality Control

Appearance
Cream to yellow homogeneous free flowing powder

Gelling
Semisolid, comparable with 0.4% Agar gel.

Colour and Clarity of prepared medium
Light yellow coloured clear to slightly opalescent gel forms in tubes as butts

Reaction
Reaction of 2.2% w/v aqueous solution at 25°C. pH: 7.4±0.2

pH
7.20-7.60

Cultural Response
Cultural characteristics observed after an incubation at 35-37°C for 18-24 hours .

Organism Growth Motility
Escherichia coli ATCC 25922 (00013*) luxuriant positive, growth away from stabline causing turbidity
# Klebsiella aerogenes ATCC 13048 (00175*) luxuriant positive, growth away from stabline causing turbidity
Klebsiella pneumoniae ATCC 13883 (00097*) luxuriant negative, growth along the stabline, surrounding medium remains clear
Salmonella Enteritidis ATCC 13076 (00030*) luxuriant positive, growth away from stabline causing turbidity
Staphylococcus aureus subsp. aureus ATCC 25923 (00034*) luxuriant negative, growth along the stabline, surrounding medium remains clear
Vibrio cholerae ATCC 15748 luxuriant positive, growth away from stabline causing turbidity
Vibrio parahaemolyticus ATCC 17802 (00037*) luxuriant positive, growth away from stabline causing turbidity

Key: (*) Corresponding WDCM numbers

(#) Formerly known as Enterobacter aerogenes

Storage and Shelf Life

Store between 10-30°C in a tightly closed container and the prepared medium at 15-30°C. Use before expiry date on the label. On opening, product should be properly stored dry, after tightly capping the bottle in order to prevent lump formation due to the hygroscopic nature of the product. Improper storage of the product may lead to lump formation. Store in dry ventilated area protected from extremes of temperature and sources of ignition. Seal the container tightly after use. Product performance is best if used within stated expiry period.

Disposal

User must ensure safe disposal by autoclaving and/or incineration of used or unusable preparations of this product. Follow established laboratory procedures in disposing of infectious materials and material that comes into contact with sample must be decontaminated and disposed of in accordance with current laboratory techniques (8,9).

Reference

  1. Koneman E. W., Allen S. D., Janda W. M., Schreckenberger P. C., Winn W. C. Jr., (Eds.), 1992, Colour Atlas and Textbook of Diagnostic Microbiology, 4th Ed., J. B. Lippinccott Company.
  2. Tittsler R. P. and Sandholzer L. A., 1936, J. Bacteriol., 31:575.
  3. Edward P. R. and Ewing W. H. 1972, Identification of Enterobacteriaceae 3rd Ed., Minneapolis, Burgess.2
  4. Baron. E. J. and Finegold S. M. (Eds.), 1990, Bailey and Scott's 'Diagnostic Microbiology, 8th ed., The C. V. Mosby. Co, St., Louis, Missouri.
  5. Howard B. J. and Other (Eds.), 1994, Clinical and Pathogenic Microbiology, The C. V. Mosby. Year Book, Inc.
  6. DAmato R. F., and Tomfohrede K. M., 1981, J. Clin. Microbiol., 14 (3), 347-348.
  7. MacFaddin J. F., 1985, Media for Isolation--Cultivation-Identification-Maintenance of Medical Bacteria, Vol. 1, Williams and Wilkins, Baltimore.
  8. Isenberg, H.D. Clinical Microbiology Procedures Handbook 2nd Edition.
  9. Jorgensen, J.H., Pfaller, M.A., Carroll, K.C., Funke, G., Landry, M.L., Richter, S.S and Warnock., D.W. (2015) Manual of Clinical Microbiology, 11th Edition. Vol. 1.
More Information
Product Name Motility Test Medium (Edwards and Ewing)
SKU M930
Product Type Regular
Physical Form Powder
Origin Animal
Packaging type HDPE
References 1. Koneman E. W., Allen S. D., Janda W. M., Schreckenberger P. C., Winn W. C. Jr., (Eds.), 1992, Colour Atlas and Textbookof Diagnostic Microbiology, 4th Ed., J. B. Lippinccott Company.2.Tittsler R. P. and Sandholzer L. A., 1936, J. Bacteriol., 31:575.3.Edward P. R. and Ewing W. H. 1972, Identification of Enterobacteriaceae 3rd Ed., Minneapolis, Burgess.2.,,4.Howard B. J. and Other (Eds.), 1994, Clinical and Pathogenic Microbiology, The C. V. Mosby. Year Book, Inc.5.Baron. E. J. and Finegold S. M. (Eds.), 1990, Bailey and Scott's `Diagnostic Microbiology, 8th ed., The C. V. Mosby. Co,St., Louis, Missouri.6.DAmato R. F., and Tomfohrede K. M., 1981, J. Clin. Microbiol., 14 (3), 347-348.7.MacFaddin J. F., 1985, Media for Isolation-Cultivation-Identification-Maintenance of Medical Bacteria, Vol. 1, Williamsand Wilkins, Baltimore.
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