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LYSINE IRON AGAR

Code: CM0381

A diagnostic medium for salmonellae including Salmonella arizonae.

Directions
Suspend 34g in 1 litre of distilled water. Bring to the boil to dissolve completely. Dispense into tubes and sterilise by autoclaving at 121° C for 15 minutes. Cool the tubes in an inclined position to form slants with deep butts.

Description
Lysine Iron Agar CM0381 is a differential medium which detects salmonellae (including lactose fermenting Salmonella arizonae) by lysine decarboxylase activity and H₂S production. Edwards & Fife¹ developed the medium to detect lactose-fermenting salmonellae which will produce pink colonies on lactose-containing media e.g. DCA and BGA. In the usual examination for enteric pathogens these organisms would be overlooked. Further, many of these cultures, when transferred to Triple Sugar Iron (TSI) Agar slants, produced acid conditions in the medium so quickly that the expected positive reaction for hydrogen sulphide was suppressed. Since Salmonella arizonae strains which ferment lactose rapidly are found occasionally in outbreaks of food infection, it is important to determine their occurrence.

The only recognised groups of Enterobacteriaceae which regularly decarboxylate lysine rapidly and which produce large amounts of hydrogen sulphide, are the salmonellae^2,3.

Lysine Iron Agar is therefore a sensitive medium for the detection of lactose-fermenting and non lactose-fermenting salmonellae.

Technique
The medium is tubed, sterilised and slanted so that a short slant and deep butt are formed. It is inoculated with a straight needle by stabbing to the base of the butt and streaking the slant. The caps of the tubes must be replaced loosely so that aerobic conditions prevail on the slant. Incubate at 35°C overnight.

Cultures which rapidly produce lysine decarboxylase cause an alkaline reaction (purple colour) throughout the medium. Those organisms that do not decarboxylate lysine produce an alkaline slant and an acid butt (yellow colour).

Cultures which produce hydrogen sulphide cause an intense blackening in the medium.

Due to deamination of the lysine, Proteus and Providencia cultures produce a red slant over an acid butt.

Reactions

Thatcher & Clark⁴ described a procedure for the isolation of salmonellae from foods in which suspect colonies from selective agar plates were purified and then inoculated into Lysine Iron Agar and Triple Sugar Iron Agar. Using this combination of media a greater discrimination can be made between the coliform organisms, e.g. Escherichia and Shigella.
Timms⁵ described the techniques of isolation and identification of salmonellae infection in turkeys, using Lysine Iron Agar.

Storage conditions and Shelf life
Store the dehydrated medium at 10-30°C and use before the expiry date on the label.
Store the prepared medium at 2-8° C.

Appearance
Dehydrated medium: Straw coloured, free-flowing powder
Prepared medium: Purple coloured gel

Quality control

* This organism is available as a Culti-Loop®

Precautions
Salmonella paratyphi A does not produce lysine decarboxylase and therefore will give an alkaline slant and an acid butt.
H₂S-producing Proteus species do not blacken this medium⁶.

References
1. Edwards P. R. and Fife Mary A. (1961) Appl. Microbiol. 9. 478-480.
2.Moeller V. (1954) Acta. Pathol. Microbiol. Scand. 355. 259-277.
3. Ewing W. H., Davis B. R. and Edwards P. R. (1960) Pub. Hlth Labs. 18. 77-83.
4. Thatcher F. S. and Clark D. S. (1968) University of Toronto Press, p.100.
5. Timms L. (1971) Med. Lab. Tachn. 28. 150-156.
6. Finegold S. M. & Martin W. J. (1982) Bailey & Scott’s Diagnostic Microbiology. 6th Edn. C. V. Mosby. St. Louis. p.63l.