Enhanced contrast of bacteriophage plaques in Salmonella with ferric ammonium citrate and sodium thiosulfate (FACST) and tetrazolium red (TZR).
Visualization of bacteriophage plaques may be enhanced by addition of ferric ammonium citrate and sodium thiosulfate (FACST) or 2,3,5-triphenyltetrazolium chloride (tetrazolium red, TZR) to the soft agar layer of a traditional bacteriophage plaque assay. Background color from these reagents improved contrast between clear plaques and turbid host lawns in trypticase soy agar (TSA) plates. Enhancement by FACST is based on reaction with hydrogen sulfide gas (H2S) produced by some strains of bacteria and was tested here using H2S+ and H2S- strains of Salmonella enterica subsp. enterica with a bacteriophage (Podoviridae) isolated from swine lagoon effluent. Only the H2S+ strain produced dark brown-black color in FACST-amended agar. Both strains showed bright pinkish-red color in TZR-amended agar. Color intensity for both reagents decreased with decreasing concentrations of the reagents. Contrast in FACST-amended plates appeared greater than that with TZR, but diminished after 12 h, while contrast in TZR-amended plates remained constant. At the concentrations tested, neither reagent affected plaque counts in the H2S+ strain. The FACST should be useful in bacteriophage plaque assays with H2S+ strains of Salmonella and other H2S+ bacteria.
McLaughlin
M R
MR
United States Department of Agriculture, Agricultural Research Service, Waste Management and Forage Research Unit, Mississippi State, Mississippi, 39762 USA. mmclaughlin@ars.usda.gov
Balaa
M F
MF
D001431:Bacteriological Techniques; D003470:Culture Media; D005290:Ferric Compounds / Q000378:metabolism*; D006862:Hydrogen Sulfide / Q000378:metabolism; D007202:Indicators and Reagents; D017902:Podoviridae / Q000502:physiology*; D000644:Quaternary Ammonium Compounds / Q000378:metabolism*; D012483:Salmonella Phages / Q000502:physiology*; D019779:Salmonella enterica / Q000254:growth & development / Q000821:virology*; D013778:Tetrazolium Salts / Q000378:metabolism; D013885:Thiosulfates / Q000378:metabolism*; D010948:Viral Plaque Assay / Q000379:methods*
D003470:Culture Media; D005290:Ferric Compounds; D007202:Indicators and Reagents; D000644:Quaternary Ammonium Compounds; D013778:Tetrazolium Salts; D013885:Thiosulfates; C017717:sodium thiosulfate; C013531:ferric ammonium citrate; D006862:Hydrogen Sulfide