|Comparative H2S production by species on TSI medium. A
- Citrobacter youngae, B - Proteus penneri, C - Salmonella
Typhimurium, D - Salmonella Gallinarum
False-positive H2S production on TSI medium (left).
|Hydrogen Sulfide Production
Some bacteria have the enzymatic capability to degrade amino acids (cysteine,
cystine etc.) that contain sulfhydryl group (-SH) producing hydrogen sulfide. Hydrogen
sulfide reacts with heavy metals such as lead or iron forming a black precipitate. You
can use TSI medium (contains iron) or prepare a nutritive agar with lead acetate (1g
Pb acetate to 100 ml nutritive agar).
Peptic peptone 10g, Casein peptone 10g, Meat extract 3g, Yeast extract 3g, Lactose
10g, Sacharose 10g, Dextrose 1g, NaCl 5g, Fe2SO4 0.2g, Na-thyosulphate 0.3g,
Phenol red 0.024g, Agar 12g, H2O ad 1000 ml.
Harvest a well isolated colony and inoculate a TSI tube by stabbing the medium.
Incubate at 37 °C, 24 hours. Reaction is positive if a black color appears.
Bacteria growing in TSI degrade aminoacids forming ferrous sulfide which blackens
1. Helgomar Raducanescu, Valeria Bica-Popii,1986. Bacteriologie veterinara, Ed.
2. Margaret Barnett, 1992. Microbiology Laboratory Exercises. Wm. C. Brown
|TSI Medium blackening due to H2S production
(left tube) - Salmonella Choleraesuis
(c) Costin Stoica
Do not incubate more than 24 hours. False-positive or false-negative results may
The amount of hydrogen sulfide produced varies by species (see picture 3).
Klebsiella oxytoca strains can produce a dark brown pigment when growth on media
containing gluconate and ferric citrate. The TSI slant is darkened by pigment, not by
hydrogen sulfide production.