164 DIFCO MANUAL 



BACTO 



KRUMWIEDE TRIPLE SUGAR AGAR (B85) 



DEHYDRATED 



Bacto-Beef Extract 1 g. 



Proteose Peptone No. 3, Difco . . 12 g. 



Bacto-Lactose 10 g. 



Saccharose, Difco 10 g. 



Bacto-Dextrose 1 g. 



Sodium Chloride 5 g. 



Bacto-Agar 15 g. 



Bacto-Phenol Red 0.025 g. 



Bacto-Krumwiede Triple Sugar Agar is a differential medium employed in the 

 primary identification of Gram-negative enteric pathogens. This medium permits 

 the determination of the formation of acid and gas from dextrose, lactose and 

 saccharose. It is employed to aid in the identity of pure cultures of colonies 

 picked from primary plating media such as MacConkey Agar, S S Agar, Bismuth 

 Sulfite Agar and others. 



Bacto-Krumwiede Triple Sugar Agar is prepared in accordance with the for- 

 mula of Krumwiede and Kohn,^ except in the choice of indicators. In place of 

 the Andrade indicator originally recommended, the more stable and sensitive 

 sulfonephthalein indicator, phenol red, is used. In addition to lactose and dex- 

 trose, as contained in Russell's Double Sugar Agar, this medium contains sac- 

 charose. This saccharose-containing medium has the advantage over Russell's 

 medium in giving an earlier and more accurate differentiation between the so- 

 called slow lactose fermenting strains of the coliform group and the Salmonella 

 group. The explanation for this increased rapidity of differential is that the slow 

 lactose fermenters attack the saccharose with greater avidity than they do lactose 

 under aerobic conditions. Salmonella are not influenced by the saccharose. There- 

 fore, with the Triple Sugar Agar many of the organisms giving a doubtful or 

 atypical Salmonella reaction on Russell's medium can promptly be excluded, 

 as a rule, as only slow fermenters of lactose. 



Like Russell's medium this Triple Sugar Agar is a differential tube medium 

 and inoculations are made from pure cultures by smearing the surface of the 

 slant and stabbing the butt. A properly inoculated tube showing, after incubation, 

 a red or cerise slope and a yellow butt, with or without gas formation, indicates 

 fermentation of the dextrose. Some strains of intestinal pathogens require 30-40 

 hours to produce an alkaline slope. A tube showing a yellow slant and butt, with 

 or without gas, indicates fermentation of the saccharose, or lactose, or both. A 

 tube showing no change indicates that none of the sugars has been fermented. 

 See the table on page 161 for the reactions of various intestinal bacteria on differ- 

 ential media. 



To rehydrate the medium, suspend 54 grams of Bacto-Krumwiede Triple 

 Sugar Agar in 1000 ml. of cold distilled water and heat to boiling to dissolve the 

 medium completely. The solution is distributed in tubes which are stoppered 

 with cotton plugs or loosely fitting caps. Sterilize in the autoclave for 15 minutes 

 at 15 pounds pressure (121°G.). Allow the tubes to solidify in a slanting position 

 so as to provide a deep butt. The final reaction of the medium will be pH 7.4. 



One pound of Bacto-Krumwiede Triple Sugar Agar will make 8.4 liters of 

 medium. 



ij. Med. Research, 37:225:1917. 



