DEHYDRATED CULTURE MEDIA 167 



In 1911 Russell^ described the use of two sugars in an agar medium to differ- 

 entiate Gram-negative organisms of intestinal origin. The ability of members of 

 this group to produce hydrogen sulfide was recognized as a valuable character- 

 istic and to detect its presence lead or iron salts have been added to the Russell 

 Medium by many investigators. Kligler--^ reported that by adding lead acetate 

 to Russell Double Sugar Agar, a medium capable of differentiating typhoid, para- 

 typhoid and dysentery could be obtained. A modification of this medium, Bacto- 

 Kligler Iron Agar, using phenol red as an indicator of acidity, and iron salts to 

 detect hydrogen sulfide production, has been used extensively for many years in 

 the differentiation of enteric organisms. Krumwiede and Kohn'* modified Russell 

 Double Sugar Agar by the addition of saccharose to the medium. This permitted 

 an earlier detection of those coliform organisms which ferment lactose slowly, 

 since many of these organisms attack saccharose more readily than lactose. The 

 added saccharose also permits the exclusion of certain coliform and proteus 

 organisms which have the ability to attack saccharose, but not lactose, in a 24-48 

 hour incubation period. 



In 1940 Sulkin and Willett^ described a triple sugar ferrous sulfate medium 

 for use in the identification of enteric organisms. This medium consisted of a 

 Beef Extract Tryptone Agar Base to which was added 1 per cent lactose, 1 per 

 cent saccharose, 0.1 per cent dextrose, 0.02 per cent ferrous sulfate, 0.015 per cent 

 sodium thiosulfate and brom thymol blue as an indicator of change in reaction. 

 In our laboratory, independently and concurrently with the work of Sulkin and 

 Willett, we prepared a similar medium by adding 1 per cent saccharose to Bacto- 

 Kligler Iron Agar. This latter medium contained phenol red as an indicator. 

 Bacto-Triple Sugar Iron Agar, so prepared, was distributed at that time to a 

 number of laboratories for comparative trials. Hajna^ described a similar medium 

 for the identification of bacteria of the intestinal group. The importance of sac- 

 charose, as already pointed out, is to eliminate certain saccharose fermenting 

 bacteria of the paracolon group which ferment lactose slowly as well as certain 

 proteus organisms capable of fermenting saccharose. Those laboratories especially 

 interested in recovery of paracolon bacilli or members of the proteus and para- 

 colon groups which ferment saccharose should use Bacto-Kligler Iron Agar, or 

 bear in mind when using Bacto-Triple Sugar Iron Agar that these organisms pro- 

 duce an acid and gas butt with an acid slant. 



The pathogenic significance of these saccharose fermenting organisms, mem- 

 bers of the paracolon and proteus groups, is not clearly defined, as has been 

 shown by many investigators. Parr'' showed the close relationship of the Morgan 

 bacillus, paracolons, anaerogenic Escherichia coli and slow lactose fermenting 

 coliform bacilli to the Shigella and Salmonella groups. Several outbreaks of gas- 

 troenteritis have been attributed to these normal or aberrant types. Stuart, 

 Wheeler, Rustigian and Zimmerman^ reported that paracolons are often associ- 

 ated with, and can cause, mild or acute gastroenteritis of short duration. Neter* 

 pointed out that the Triple Sugar Iron Agar does not eliminate all saccharose 

 fermenting strains of proteus and paracolons, and further that the pathogenic 

 significance of some of these organisms as incitants of diarrheal diseases is not 

 clearly known. He pointed out that these facts should be taken into consideration 

 when using the Triple Sugar Iron Agar as a diagnostic medium. 



Ewing and Bruner^^ used Triple Sugar Iron Agar as a differential tube medium 

 in isolation of Salmonella and Shigella for serological classification. Typical 

 suspicious colonies were picked from the primary media and were inoculated 

 into Triple Sugar Iron Agar. After overnight incubation at 37°G. the cultures 

 showing an alkaline slant and acid or acid and gas in the butt were transferred 

 to the Urea Medium of Christensen.^^ (A complete discussion of this medium, 

 Bacto-Urea Agar Base, is given on page 171). After incubation for 2-4 hours at 



