Organic farming for sustainable agriculture depends on the effects of microbial activities in the soil. The present study evaluated seasonal changes in the soil microbial communities by fatty acid methyl ester (FAME) in an organic farming system (OFS) with rye rotation cropping management compared to those in a conventional farming system (CFS) with chemical fertilizers, pesticide and herbicide from May 2009 to October 2010 in an upland field. Concentrations of total bacteria, Gram-negative bacteria, Gram-positive bacteria, fungi, and arbuscular mycorrhizal fungi in the OFS were significantly higher than those in the CFS during soybean-growing stages (p<0.001). The communities of total bacteria, Gram-negative bacteria, Gram-positive bacteria, actinomycetes and arbuscular mycorrhizal fungi in the CFS soils were significantly higher than those in the OFS soils (p<0.001), whereas the community of fungi was significantly lower in the CFS soils than in the OFS soils (p<0.05) due to decreasing Gram-positive bacteria. OFS had a significantly lower ratio of $cy19:0/18:1{omega}7c$ compared with CFS, indicating microbial stress decreased (p<0.001). In principal component analyses of soil microbial communities, our findings suggested that Gram-positive bacteria should be considered as potential factor responsible for the clear microbial community differentiation observed between OFS and CFS in upland fields.