The fungus-cultivating termites (Macrotermitinae) form part of diverse termite fauna in Africa, but information on their fungal symbionts is inadequate and poorly understood. In this study, the fungal communities and structure between termite gut, mound and surrounding soil were determined using the 454 pyrosequencing-based analysis of the internal transcribed spacer (ITS) gene sequences. Genomic DNA was extracted and purified from the guts of three termites (Odontotermes sp., Macrotermes michaelseni and Microtermes sp.), mound and surrounding soil samples for highthroughput sequencing. A total of 15,256 sequences were obtained and individual samples contained between 4 and 133 operational taxonomic units (OTUs). Termite gut had the least fungal diversity, dominated by members of the Basidiomycota (> 98%). More than 98% of the gut sequences were of the genus Termitomyces, while < 2% were related to the genera Chaetomium, Fusarium, Eupenicillium, Cladosporium, Curreya and Phaeosphaeria with between 95 and 98% pair-wise sequence identities. Members of Ascomycota (> 94%) were the most abundant in the mound and soil, but significantly differed (P value of 0.04; R value = 0.909) between the mound and soil environments. The results confirm that the genus Termitomyces exist in a tight association with their hosts and that Termitomyces species are scarcely present in the mound and soil. In addition, by altering soil properties; the fungus-cultivating termites modify the fungal community composition and structure in the mound and surrounding soil environments.