Ferric ammonium citrate decomposition--a taxonomic tool for gram-negative bacteria

The iron uptake test of Szabó and Vandra has been modified and used for the differentiation of Gram-negative bacteria. Nutrient agar containing 20 g per litre of ferric ammonium citrate was distributed into narrow tubes and solidified so as to form butts and slants. Considering the localization of the rusty-brown coloration produced after seeding and incubation, 2367 strains were classified into four groups. (1) Unchanged medium: Escherichia coli, Shigella spp., Yersinia spp., Hafnia alvei and Morganella morganii 100% each, Klebsiella spp., 50%, Enterobacter cloacae 37%, Proteus vulgaris 59%, Acinetobacter spp. 42%, Pseudomonas fluorescens 19%, some other bacteria 2-12%. (2) Rusty-brown slant, unchanged butt: Salmonella subgenera II, III and IV 98%, Citrobacter freundii 65%, E. cloacae 55%, P. vulgaris 41%, Proteus mirabilis 98%, Providencia rettgeri 100%, urease-negative Providencia 96%, Acinetobacter spp. 58%, Pseudomonas aeruginosa 100%, P. fluorescens 81%, UFP (unclassified fluorescent pseudomonads) 100%, other Pseudomonas spp. 55%. (3) Unchanged slant, brown butt: S. typhi 88%, Salmonella subgenus I 3%, Klebsiella spp. 31%, some other bacteria 2-3%. (4) Rusty-brown slant, brown butt: Salmonella subgenus I 75%, C. freundii 20%, Klebsiella spp. 12%, some other bacteria 1-5%. Colour reactions in ferric ammonium citrate agar are associated with the accumulation of ferric hydroxide: bacteria giving positive reactions on the slant took up as an average, 63 times more iron than those with negative test. The localization of colour reaction correlated partly with aerobic and anaerobic citrate utilization or decomposition in Simmons' minimal and in Kauffmann's peptone water medium.