Abstract: The rate of organic nitrogen decomposition could be not controlled after organic fertilizer application in tobacco-planted soils. This problem could affect the normal growth and quality of tobacco leaves. In this study, chemical fertilizer, cow manure, maize straw and rapeseed cake were applied to the root-zone soil under tobacco. The study attempted to isolate highly active local ammonifiers and nitrosobacteria, and to determine the effects of the two bacteria types on the decomposition of organic nitrogen and ammonia. The study was to make possible the preparation of a microbial agent for the efficient decomposition of organic nitrogen. The results showed that the numbers of bacteria, fungi, actinomycetes and ammonifiers treated with organic fertilizer were higher than those treated with chemical fertilizer during the vigorous growth period of flue-cured tobacco. The numbers of ammonifiers, nitrosobacteria, fungi and actinomycetes treated with organic fertilizer were higher than those treated with chemical fertilizer in flue-cured tobacco at maturity period. Through analysis of soil samples of various strains, the dominant strains with high ammoniation intensity were all derived from soil samples treated with organic fertilizer. It was shown that soils treated with organic fertilizer had higher active ammonifiers than soils treated only with chemical fertilizer. The results showed that decline in organic nitrogen content was highest in Bacillus pumilus, Geobacillus stearothermophilus and B. megaterium, which was 84.74%, 92.74% and 79.52% lower than that of initial organic nitrogen. Stenotrophomonas sp. had the highest nitrosation and nitrification activity. After 7 days of culturing, nitrate nitrogen content in cultured medium was 0.617 mg·L^-1. S. maltophilia had the strongest nitrosation activity. After 7 days of culturing, nitrite content in the cultured medium was 0.518 mg·L^-1. Organic nitrogen decomposition activity of ammoniated bacteria decreased after 48 h of culturing. Then the nitrification and nitrification activities were still high 7 days after culturing. Thus the separation of different functions of bacteria in the preparation of nitrogen-containing composite microbial agents should be done at different times by adding fermentation strain to achieve high activity of microbial agents. In this study, eight strains (Naxibacter sp., Stenotrophomonas sp., B. pumilus, G. stearothermophilus, B. stratosphericus, Cellulosimicrobium cellulans, B. altitudinis and B. megaterium) were identified as high-efficiency strains of microbial agents for the decomposition of soil organic nitrogen.