The API 20B microtube system to aid in the identification of N2-fixing Bacillaceae

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Abstract
N2-fixing bacterial isolates from soil have been identified successfully using the API 20E and 50E microtubes in conjunction with a computer-assisted biochemical profile search. The 20E and 50E result in few positive tests with N2-fixing Bacillaceae. Recently, a new battery of 20 microtube tests, the 20B, has been introduced for identifying aerobic, heterotrophic bacteria isolated from natural environments. This system was compared with the 20E and evaluated for N2-fixing bacillus species from soil. It was then used in conjunction with guanine-plus-cytosine analysis to identify the C-11-25 (5DN+) bacillus strain that fixes N2 in association with Canadian wheat cultivars. The API 20E and 20B gave similar results for many biochemical tests, but the 20E resulted in negative tests for carbohydrate utilization by bacillus strains. The use of different growth media or 0.85% NaCl to suspend these bacteria prior to inoculating the cupules did not alter the inability of the 20E in this manner. Carbohydrate utilization in the 20B system agreed well with that in the traditional utilization tests. Hence, I concluded that the API 20E is not suitable to evaluate carbohydrate utilization in N2-fixing Bacillaceae. The spore-forming, acetylene-reducing C-11-25 strain of bacillus was biochemically similar to the ATCC type culture of Bacillus polymyxa (strain 842). Its guanine-plus-cytosine ratio of 48.0 mol% was similar to that of B. polymyxa (43–46 mol%). A general procedure for the isolation and identification of N2-fixing Bacillus from soil is proposed.