340 



ORDER IV. EUBACTERIALES 



Edwards, West and Bruner (Jour. Bact., 

 55, 1948, 711), Bruner, Edwards and Hopson 

 (Jour. Inf. Dis., 85, 1949, 290) and West and 

 Edwards (U. S. Pub. Health Service Mono- 

 graph 22, 1954), 32 O groups and 87 H anti- 

 gens have been established. When the sera 

 used in the classification of Bethesda- 

 Bellerup cultures were used in the examina- 

 tion of normal strains of E. freundii, it was 

 found that either or both the O and H anti- 

 gens of the majority of E. freundii cultures 

 could be recognized. Thus, in view of the 

 similarity of their biochemical reactions 

 and their close serological relationships, it 

 seems better to regard the Bethesda-Bal- 

 lerup group as a variety of the species E. 

 freundii. Subsequent biochemical investiga- 

 tions have confirmed this view. 



Comments: In 1932, Werkman and Gillen 

 (Jour. Bact., 23, 1932, 177), following the 

 custom prevalent at that time, established 

 the citrate-positive coliform organisms in a 

 separate genus, Citrobacter, and subdivided 

 this genus into seven species on the basis of 

 action on gelatin and on differences in their 

 fermentation of sucrose, esculin, salicin, 

 dulcitol and similar compounds. Five of the 

 seven species described by Werkman and 

 Gillen produced H2S in proteose peptone- 

 ferric citrate agar and, following Vaughn 

 and Levine (Jour. Bact., U, 1942, 502), are 

 all regarded here as belonging to Esch- 

 erichia freundii. 



Source: Isolated from canal water in 

 Holland. 



Habitat: Normally found in soil and 

 water and, to a varying degree, in the intes- 

 tinal canals of man and other animals. 

 Widely distributed in nature. 



4. Escherichia intermedia (Werkman 

 and Gillen, 1932) Vaughn and Levine, 1942. 

 (Citrobacter intermedium Werkman and 

 Gillen, Jour. Bact., 23, 1932, 178; Vaughn 

 and Levine, Jour. Bact., U, 1942, 498.) 



in.ter.me'di.a. L. adj. iniermedius inter- 

 mediate. 



Short rods, with rounded ends, occur- 

 ring singly, in pairs and in short chains in 

 young nutrient agar or broth cultures. 

 Activel}^ motile by means of peritrichous 

 fiagella or non-motile. Gram-negative. 



Gelatin stab: No liquefaction after 60 

 days at 20° C. 



Agar slant: Smooth to wrinkled surface; 

 grayish white, abundant, raised, butyrous 

 growth. 



Levine's eosine-methylene blue agar: 

 Well isolated colonies vary from 1 to 4 mm 

 in diameter. No confluence of neighboring 

 colonies. Colonies are slightly to moderately 

 raised with surfaces varying from flat to 

 conve.x and usually smooth and glistening 

 but sometimes dull, rough and granular. 



By transmitted light two types of colonies 

 have been observed: (1) colonies having 

 almost the same appearance throughout 

 but with a distinctly lighter center, the 

 color being similar to the medium; (2) 

 colonies having a dark brownish central 

 area which diffuses out to a lighter margin. 



By reflected light three types of colonies 

 have been observed: (1) dark, button-like, 

 concentrically ringed colonies possessing a 

 strong, greenish metallic sheen so char- 

 acteristic for Escherichia coli; (2) colonies 

 with dark, purplish, wine-colored centers 

 surrounded by a light pink zone; some 

 colonies are concentrically ringed; (3) pink 

 colonies with no suggestion of sheen but 

 sometimes concentrically ringed. 



Nutrient broth: Turbid; slight ring at 

 surface. 



Litmus milk : Acid ; sometimes coagulated 

 and reduced; no proteolysis. 



Potato: Abundant, white to ivory- col- 

 ored growth. 



Indole maj' or may not be produced. 



Hj'drogen sulfide not detected in proteose 

 peptone ferric citrate agar. 



Acid or acid and gas produced from 

 xj'lose, arabinose, rhamnose, glucose, fruc- 

 tose, mannose, galactose, lactose, maltose, 

 trehalose and mannitol. No acid or gas from 

 melezitose, amygdalin or erj'thritol. Su- 

 crose, raffinose, cellobiose, a-methyl-gluco- 

 side, adonitol, dulcitol, glycerol, inositol, 

 sorbitol, starch, aesculin, salicin and sodium 

 malonate may or may not be fermented. 



Fermentation of glucose: The end prod- 

 ucts characteristic for the genus Escherichia 

 are produced. Carbon dioxide and hydrogen 

 gases are produced in approximately equi- 

 molar proportions (gas ratio, 1:1) besides 



