SIM test microbiology procedure

The SIM test (Sulfide, Indole, Motility test) is a microbiological test used to identify and differentiate bacteria based on sulfide production, indole production, and motility.

The SIM test is commonly used for identifying members of the family Enterobacteriaceae, which includes pathogens such as Escherichia coli, Proteus, and Salmonella species.

SIM test procedure steps

• Using isolated colonies from an 18-24 hour culture on solid media, inoculate the SIM Medium by stabbing the center of the medium to a depth of 1/2 inch.
• Incubate the inoculated medium aerobically at 35ºC. for 18-24 hour.
• Observe for H2S production and motility.
• Once H2S and motility reaction have been read and recorded, apply three drops of Kovacs Reagent (Cat. no. Z67) tothe surface of the medium.
• Observe for the development of a pink to red color.

Interpretation of result

• A positive H 2S test is denoted by a blackening of the medium along the line of inoculation.
• A negative H2S test isdenoted by the absence of blackening.
• A positive motility test is indicated by a diffuse zone of growth flaring from the line of inoculation.
• A negative motility test is indicated by growth confined to the stab line.
• A positive test for indole is denoted when a pink to red color band is formed at the top of the medium after addition ofKovacs Reagent. A yellow color denotes a negative indole test after addition of Kovacs Reagents.

LIMITATIONS of SIM test

The inoculum should be taken from a solid medium. Use of an inoculum from a liquid or broth suspension will delay
the initiation of growth and may result in erroneous results.

When inoculating semi-solid media, it is important that the inoculating needle be removed along the exact same lineused to inoculate the medium.

A fanning motion may result in growth along the stab line that may result in falsepositive interpretation.

Motility and H2S results must be interpreted prior to addition of Kovacs Reagent.

Weakly motile organisms or organisms that possess damaged flagella (due to heating, shaking, or other trauma) oftenresult in false-negative motility tests.

Motility results may be confirmed by performing a hanging drop motility test.