Alpine ecosystems are special regions and are sensitive to global climate change. Revealing the succession of microbial communities along the altitude gradients, especially above the permanent snow line, is essential to understanding the influence of global climate change on the alpine ecosystem and the climate feedback of the alpine ecosystem. In this study, the abundance and community composition of bacteria and fungi in 12 soil and gravel samples collected from the northern slope of the Mount Everest (Tibetan Plateau) along an altitude gradient varying between 4000 and 6550m (above sea level, m.a.s.l.) were characterized by viable counting, real-time polymerase chain reaction (PCR), phospholipid fatty acid (PLFA), and denaturing gradient gel electrophoresis (DGGE), combined with clone sequencing. The results showed that both the viable count and rRNA gene abundance for bacteria and fungi decreased with increasing altitude. The PLFA analysis showed that the contents and diversity of bacterial, fungal, and total PLFA reduced with increasing altitude. Furthermore, none of the PLFA components representing gram-positive bacteria were detected in samples above snow line (5350m), but some PLFA components representing gram-negative bacteria and fungi were distributed in all samples for low and high-altitude areas, which suggested that gram-positive bacteria were more sensitive to the changes in altitude and altitude-dependent environmental variables. The unweighted pair-group method with arithmetic means (UPGMA) analyses of the DGGE profiles showed distinct clustering in the low-altitude (4000m) and high-altitude (≥ 5350m) samples for bacteria communities, but not for fungal communities. The DGGE band sequencing results suggested that Proteobacteria were the dominant bacteria in all samples, and that Gemmatimonadetes were dominant in higher altitude samples, whereas Actinobacteria were mainly found in lower altitude samples. Fungal DGGE band sequencing showed that Ascomycota predominated across the altitude gradient, whereas Cercozoa protists, identified by the fungal 18S rRNA gene primer, were only found in relatively higher altitude samples.