Abundances and community structure of archaea, bacteria and fungi in a red soil profile located at Xiangyin County, Hunan province, was investigated with the quantitative real-time PCR (qPCR) and 454 pyrosequencing approaches. Results showed that with increasing soil depth, soil clay increased in content, while organic matter, total nitrogen, and carbon to nitrogen ratio declined. Correspondingly, the numbers of gene copies per gram of dry soil also decreased within the range of10^7.09 ～10^9.30 for archaeal 16S rDNA, 10^8.10 ～10^9.70 for bacterial 16S rDNA, 10^6.54～10^7.95 for fungal 18S rDNA, 10^7.24～10^8.61for archaeal amoAgene, 10^4.76 ～10^6.25 for bacterial amoAgene, 10^5.94～10^7.88for nirK gene, 10^6.81～10^9.21for nirS gene, and 10^7.03～10^9.46 for nosZ gene. Pyrosequencing generated 6 459 archaeal 16S rDNA sequences with an average length of 496 bp, 28 626 bacterial 16S rDNA sequences with an average length of 448 bp, and 4 683 fungal 18S rDNA sequences with an average size of 534 bp. OTU (97% similarity) analysis revealed that the α-diversity was in the order bacteria>fungi>archaea, but there was no significant correlation between microbial α-diversity and soil properties determined. Analysis of the Jaccard dissimilarity indicated that microbial communities in different layers of the same soil profile were closer, compared with the dissimilarity between three surface replicates. Mantel test showed that clay content was the main soil factor explaining the variation of microbial communities. In all the soil samples, archaea was dominated (89%) with Thermoprotei (belonging to Crenarchaeota), of which the distribution was significantly related to soil clay content. The bacterial community consisted mainly of Acidobacteria (33%), Proteobacteria (17%), Chloroflexi (12%), Firmicutes (10%) ， Actinobacteria (7%) and Bacteria incertae sedis (11%) . Acidobacteria and Proteobacteria were found to be more abundant in the surface soils than in other soil layers, while the situations of Chloroflexi and Firmicutes were opposite. All fungal sequences belonged to three phyla, i.e., Ascomycota (87%), Basidiomycota (9%) and Glomeromycota (4%). This work demonstrated the great potential of pyrosequecing technique in revealing microbial diversity and presented background information of microbial communities in the red soil.