As a kind of biomarker, phospholipid fatty acid (PLFA) was used to detect the microbial community diversity in soil both qualitatively and quantitatively. The diversity of PLFA in the tobacco soils was tested. The PLFA data was analyzed, using Shannon\|wiener（H1）,Brillouin（H2） and McIntosh（H3） diversity indices，the abundance(S)，Pielou evenness（J）and Simpson indices（D）. The result of the ecological evaluation showed that: 43 PLFAs in total were detected in soils during the whole growing periods of tobacco. Total PLFAs reached the highest in soils that were collected after growing tobacco for 90 days. The 43 PLFAs biomarkers tested could be divided into 5 groups by means of the cluster analysis according to diversity indices. It was as follows: Group Ⅰ was characterized with high quantity, high frequency and high diversity for the biomarkers in the soil, the dominant biomarker was 18:1ω9c belonging to fungal biomarker; Group Ⅱ was rich with 16:0, characterized with high frequency and high diversity, belonging to pseudomonas; Group Ⅲ had 16:1ω5c with characteristics of high quantity, high frequency and moderate diversity, belonging to methanotrophs; Group Ⅳ had i15:0, a15:0, i16:0 and a17:0 which were related to aerobic bacterial biomarkers, 18:1ω7c which was anaerobia, 16:0 10Me which was sulfate reducing bacterial PLFA biomarkers, those of PLFA biomarkers were moderate quantity and diversity, high frequency; Group Ⅴ had i 15:0 3OH, 15:1 i G, a16:0, i16:1 G, i16:1 H, 17:0, i17:0, 15:0 2OH, 15:0 3OH, 17:0 and 17:0 2OH which were aerobic bacteria, 18:3ω6c (6,9,12) which was fungus, 17:0 10Me and 18:0 10Me which were actinomyces, 20:4ω6,9,12,15c which was protozoa. Group V PLFAs biomarkers were low quantity, frequency and diversity. Therefore, the order of microbes which played an important role to the tobacco soil properties was fungus, pseudomonas, methanotrophs, aerobic bacteria, anaerobe, and then sulfate reducing bacteria. It could instruct the equitable adjustment of soil miroecological system which would be a new method to assess the diversity of microbial community in soils.