The experiment was set up to investigate the effects of fertilization on soil microbial properties and its mechanism in the Loess Hilly Region. Soil microbial community structure and respiration under long-term fertilizations, including CK (control), N (nitrogen fertilizer), P (phosphorus fertilizer), M (manure), NP (nitrogen fertilizer + phosphorus fertilizer), MN (manure + nitrogen fertilizer), MP (manure + phosphorus fertilizer) and MNP (manure + nitrogen fertilizer + phosphorus fertilizer), were studied at Ansai Field Research Station. The results showed that microbial activities and PLFA contents in surface soil (0-20cm) were higher than those in the sub soil (20-40cm); Basal respiration, substrate-induced respiration and total PLFA contents in surface soil were 63.61%-116.78%, 53.45%-53.45% and 16.16%-43.67% higher than those in sub soil. Compared to the CK treatment, N fertilizer and P fertilizer increased surface soil basal respiration by 34.11% and 48.89%, respectively, and substrate-induced respiration intensities increased by 40.83% and 63.59%; In sub soil, the basal respiration intensities increased by 40.83% and 63.59%, and the substrate-induced respiration intensities increased by 14.70% and 20.49%, respectively. The effect of N fertilizer on G^-microbial community was significant and increased PLFA contents in 0-20cm and 20-40cm soil profile significantly by 63.19% and 53.07%. And P fertilizer also altered the microbial community structure significantly, however, the effects of the mixed fertilizer of NP on microbial community structure was not significant. The mixture of organic and inorganic fertilizers had a significant influence on soil respiration and microbial community structure. Variance analysis of three factors indicated that the effects of single N fertilizer factor on soil microbial properties were not significant; single N fertilizer had significant effect on the microbial respiration and PLFA contents. In the 0-20cm soil layer, the ratio of P fertilizer factor influencing these microbial characteristics was 11.4%-54.0%. The RDA analysis indicated that available P played a major role in driving the change in microbial characteristics. Long-term mixed manure, N and P fertilizers could improve soil microbial properties, thus contribute to the stability and sustainability of farmland ecosystem.