Biochemical Characterization of Citrobacter diversus (Burkey) Werkman and Gillen and Designation of the Neotype Strain

Abstract

The biochemical characteristics of 1 37 strains of Citrobacter diversus (Burkey) Werkman and Gillen were determined, and the resulting data are summarized. Members of this species produced indole and ornithine decarboxylase and fermented adonitol but failed to grow in KCN medium. Detectable amounts of hydrogen sulfide were not produced in triple sugar-iron-agar medium but were formed by some strains in peptone-iron-agar. The nomenclature and taxonomic position of these bacteria are discussed, and information that should be helpful for their differentiation from Citrobacter freundii and Enterobacter cloacae is included. Strain 3613-63 (ATCC 27 156) is designated as the neotype strain and is described herein. Werkman and Gillen (32) proposed the generic name Citrobacter for the "citrate-positive, coli-aerogenes intermediates", from a collection of fifteen strains, these investigators described seven species. Citro bacter Freundii (sic) Braak was designated as the type species of the new genus. A review of the literature [e.g., see Vaughn and Levine (31)l indicated that some, but by no means all, of the so-called intermediate bacteria belonged t o the genus Citrobacter. Tittsler and Sandholzer (30) and Carpenter and Fulton (7) suggested that "Escherichia-Aerobacter intermediate" strains, which did not produce acetylmethylcarbinol but utilized sodium citrate as a sole source of carbon, should be placed in the genus Escherichia. This suggestion, rather than that of Werkman and Gillen (32), was followed in the 1939, 1948, and 1957 editions of Bergey's Manual (2-4). Vaughn and Levine (31) deline-ated two divisions among the "intermediate" bacteria studied. Members of the first of these produced hydrogen sulfide, whereas constituents of the second did not. These investigators stated that the bacteria that produced hydrogen sulfide might be recognized as Escherichia freundii (Braak) Yale and that the hydrogen sulfide-negative microorganisms might be recognized as Escherichia inter-medium (Werkman and Gillen) comb. nov. The latter species was recognized in the 1948 edition of Bergey ' s Manual. Ewing and Edwards (15, 16) and Ewing (11, 12) considered members of the genus Citrobacter to be more closely related to the genera Salmonella and Arizona than to either Escherichia or Aero-bacter (now En terobacter). As presently constituted, the genus Citro-bacter is composed of similar bacteria that have been described under a variety of designations in addition t o those already mentioned. For example, the microorganism called Salmonella ballerup (22), "S. coli" 1 to 4 (21), and Salmonella hormaechei [ Monteverde (1944) cited by Bruner et al., (5)] originally were classified as salmonellae because of antigenic relationships and were included in the antigenic schema for those bacteria. Later, they were removed from the antigenic schema for Salmonella on the basis of their biochemical reactions [e.g., see Harhoff (20); Bruner et al. (5) ]. These microorganisms now are considered t o be C. freundii. Strains that closely resembled the 1401 1 biotype of Stuart et al. (29) were found t o be members of the Bethesda group characterized by Barnes and Cherry (1) and so named by Edwards et al. (1 0) and Moran and Bruner (28). West and Edwards (33) investigated 585 strains of Ballerup and Bethesda bacteria, combined these two into the so-called Bethesda-Ballerup group, and directed attention to the close biochemical relationships of members of the combined group and those of cultures of E. freundii. 12