Nitrogen Fixation by New Species of Nocardia

SUMMARY: Two new species of the genus Nocardia ("rev.) have been isolated from the soil of chalk grassland. One of these, Nocurdiu cakureu n.sp., fixes atmospheric nitrogen in culture in amounts of 24.5 mg. N/g. glucose, sucrose or mannitol present in the medium. The second species, N. cetluluns n.sp., decomposes cellulose in culture and Axes atmospheric nitrogen in culture using glucose, sucrose, mannitol and cellulose as carbon source ; amounts of 12 mg. N fixed/g. cellulose decomposed were recorded. The properties of nitrogen fixation and cellulose decomposition have not previously been recorded for this genus. The two organisms are described. The two species of Nocardia to be described were isolated in the course of an examination of the nitrogen-fixing and cellulose-decomposing bacteria in the soil beneath a chalk grassland plant community. As the ability to fix atmo- spheric nitrogen shown by both these organisms and the ability to decompose cellulose shown by one are attributes not previously recorded for the genus Nocardia, a determinative description of the two organisms (which are regarded as new species) is given. It is proposed to use the name Nocardia celldam for the cellulose-decomposer and the name N. calcarea for the second organism, which as yet has been isolated only from chalk soil. METHODS Isolation of Nocardia calcarea. This organism appeared repeatedly in the course of isolations of species of Axotobacter in liquid enrichment media using chalk soil as inoculum. The ' nitrogen-free ' medium had the composition : 3.0 g. K,HPO,; 1.0 g. NaCl; 0.01 g. MgS0,.7H20; 0.01 g. FeCl,; 1-0 g. CaCO,; glass-distilled water, 1 1.; 1 p.p.m. Na,MoO,. 2H,O; 0.3 yo (w/v) sodium benzoate; pH 7.2. After three successive transfers at 20' in this medium the enriched cultures were plated on the same medium solidified with washed agar. The colonies of Nocardia were easily differentiated from those of Azoto- bacter by their pinkish colour, Final isolation was obtained as readily by using mannitol or glucose (1 yo, wlv) as the carbon supply. Isolation of the organism was also readily achieved by the ' spontaneous culture ' method of Winogradsky (1949, p. 609) following the addition of sodium acetate to soil samples, also by a modification of his plate insemination method (Winogradsky, 1949, p. 626) in which washed agar was used to solidify the basal medium and butanol added as carbon source. Isolation of Nocardia cellulans. A silica gel was used impregnated with basal medium (2-5 g. KNO,; 1.0 g. K,HPO,; 0.1 g. CaC1,; 0-3 g. MgSO, .7H20; 0.1 g. NaCl; 0.01 g. FeCl,; glass-distilled water, 1 1,; pH 7.0-7-2). Washed * Present address : Rothamsted Experimental Station, Harpenden, Herts.