180 DIFCO MANUAL / 



Inoculate a Dextrose Broth tube. 



Acid fermentation — Suspect Shigella and proceed as in III B. 

 No reaction in dextrose — DISCARD. 



(Alkaligines and pseudomonas groups. An occasional pseudo- 

 monas strain produces a slight reaction in dextrose). 



III Non-motile types. 



A. Gram-stain all non-motile types. 



Gram-positive types — DISCARD. 



B. Gram-negative types — Proceed with tests for the Shigella group. (An 



occasional Sonne strain produces fermentation of lactose rather 

 rapidly. For this reason all non-motile, Gram-negative types are 

 put through suitable tests for this species). 



IV Motile cultures. 



A. Moderate motility, hydrogen sulfide production. 



Proceed with tests for S. typhosa. 



B. Active motility with or without hydrogen sulfide production. Inocu- 



late Bacto-Peptone or Bacto-Tryptone tube for indole test, and fer- 

 mentation tubes of Mannitol and Saccharose Broths. 



1. Mannitol fermenter, no fermentation of saccharose, indole nega- 



tive. 



a. Mannitol fermented (with acid and gas), no hydrogen 

 sulfide production — S. paratyphi and a few other Salmonella 

 types. 



b. Hydrogen sulfide production. 



(I) Mannitol fermented with acid and gas — Proceed with 



tests for Salmonella. 

 (II) Mannitol fermented with acid only — Proceed with tests 

 for S. typhosa. 



c. Indole positive and/or saccharose fermenting types (coli- 

 aerogenes, proteus groups) — DISCARD. 



2. Mannitol non-fermenting types (Morgan's bacillus and other 



proteus groups) — DISCARD. 



To prepare 200 ml. of the medium, suspend 7.2 grams of Bacto-Friewer 

 Shaughnessy Base No. 1 in 100 ml. of cold distilled water and heat to boiling to 

 dissolve the medium completely. To a second flask containing 100 ml. distilled 

 water, add 2.1 grams Bacto-Friewer Shaughnessy Base No. 2, and warm to 45°C. 

 When Base No. 2 is in complete solution add to the hot solution of Base No. 1. 

 Caution: To avoid a precipitate in the medium, it is necessary to have Base No. 

 1 in complete solution and hot when the warm solution of Base No. 2 is added. 

 Distribute in tubes to a depth of about 3 inches. Sterilize in the autoclave for 15 

 minutes at 15 pounds pressure (121°C.). Allow the sterile tubes to solidify in 

 an upright position. Final reaction of the medium will be pH 7.5. 



One pound of Bacto-Friewer Shaughnessy Base No. 1 will make 12.5 liters of 

 medium. 



One pound of Bacto-Friewer Shaughnessy Base No. 2 will make 43.2 liters 

 of medium. 



lAm. J. Clin. Path., Tech. Sect., 8:1:1944. 

 3 Personal Communication, 1947. 



