Microorganisms are the driver of petroleum hydrocarbon degradation in soil micro-ecological systems. However, the distribution characteristics of microbial communities and hydrocarbon degradation dynamics during the remediation of petroleum-contaminated soil by enhancing moisture content are not clear. In this study, polymerase chain reaction and high-throughput sequencing of soil microbial DNA were applied to investigate the compositions of microorganisms and alpha diversity in the oil-polluted soil, and the hydrocarbon removal also being analyzed using ultrasonic extraction and gravimetric method in a laboratory simulated ex-situ experiment. Results showed the distribution of petroleum hydrocarbon degrading microorganisms in the petroleum-contaminated loessal soil mainly was Proteobacteria phylum (96.26%)-Gamma-proteobacteria class (90.03%)-Pseudomonadales order (89.98%)-Pseudomonadaceae family (89.96%)-Pseudomonas sp. (87.22%). After 15% moisture content treatment, Actinobacteria, Proteobacteria, and Firmicutes still were the predominant phyla, but their relative abundances changed greatly. Also Bacillus sp. and Promicromonospora sp. became the predominant genera. Maintaining 15% moisture content increased the relative abundance of Firmicutes phylum and Bacillus sp. As the moisture-treated time increases, the uniformity and the richness of the soil bacterial community were decreased and increased respectively; the relative abundance of Pseudomonas sp. increased. Petroleum hydrocarbon degradation by enhancing soil moisture accorded with the pseudo-first-order reaction kinetic model (correlation coefficient of 0.81; half-life of 56 weeks). The richness of Firmicutes phylum and Bacillus sp. may be a main reason for promoting the removal of 18% petroleum hydrocarbons responded to 15% moisture treatment. Our results provided some beneficial microbiological information of oil-contaminated soil and will promote the exploration of remediation by changing soil moisture content for increasing petroleum hydrocarbon degradation efficiency.