Hydrogen Sulfide (H2S) Production Test

Hydrogen Sulfide (H2S) Production Test

Objective of Hydrogen Sulfide (H2S) Production Test

To determine the ability of the organism to produce hydrogen sulphide.

Principle of Hydrogen Sulfide (H2S) Production Test

Hydrogen sulphide (H2S) production test is used for the detection of hydrogen sulphide (H2S) gas produced by an organism. It is used mainly to assist in the identification of members of family Enterobacteriaceae. H2S is produced by certain bacteria through reduction of sulphur containing amino acids like cystine, methionine or through the reduction of inorganic sulphur compounds such as thiosulfates, sulfates or sulfites. The hydrogen sulphide production can be detected by incorporating a heavy metal salt containing iron or lead as H2S indicator to a nutrient culture medium containing cystine and sodium thiosulfates as the sulfur substrates. Hydrogen sulphide, a colorless gas, if produced reacts with the metal salt forming visible insoluble black precipitate of ferrous sulphide.

Media Used in Hydrogen Sulfide (H2S) Production Test

SIM Medium

Composition per liter:

Pancreatic digest of casein………………………….. 20.0 gm

Peptic digest of animal tissue……………………….. 6.1 gm

Agar………………………………………………………….. 3.5 gm

Fe(NH4)2(SO4)2·6H2O…………………………………. 0.2 gm

Na2S2O3·5H2O……………………………………………. 0.2 gm

pH 7.3 ± 0.2 at 25°C

Procedure of Hydrogen Sulfide (H2S) Production Test

  1. Inoculate the organism into labeled tube by means of stab inoculation in SIM medium.
  2. Incubate the inoculated tubes at 37°C for 24-48 hours.
  3. Observe for the formation of black precipitate on the medium.

Result Interpretation of Hydrogen Sulfide (H2S) Production Test

Result Interpretation of Hydrogen Sulfide (H2S) Production Test

Positive result: blackening on the medium

Negative result: no blackening on the medium

Limitations of Hydrogen Sulfide (H2S) Production Test

  1. H2S production may be inhibited on TSI for organisms that utilize sucrose and
    suppress the enzyme mechanism that results in production of H2S.
  2. SIM is more sensitive in the detection of H2S than either TSI or KIA, because
    of its semisolid nature, its lack of interfering carbohydrates, and the use of
    peptonized iron as an indicator.
  3. Lead acetate paper is 10 times more sensitive than other media.
  4. Lead acetate is toxic to bacteria and may inhibit the growth of some bacteria. Do not allow the media to touch the strip.

Quality Control of Hydrogen Sulfide (H2S) Production Test

Positive control: Proteus vulgaris

Negative control: Escherichia coli

Hydrogen Sulfide (H2S) Production Test- Objective, Principle, Procedure, Media Used, Result Interpretation, Limitation and Quality Control.

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