Acanthostigma is a genus in the Tubeufiaceae family of fungi. Three new species were reported in 2010, growing from decomposing wood in Great Smoky Mountains National Park in the United States. The three new bitunicate ascomycetes belonging to the genus Acanthostigma are described from terrestrial decomposing wood collected from Great Smoky Mountains National Park, USA. Phylogenetic analyses of the nuclear ribosomal 28S large subunit and internal transcribed spacer region placed all three species in the Tubeufiaceae and confirmed morphological analyses that these are distinct species. Expanded phylogenetic analyses of 28S large subunit including taxa throughout the Dothideomycetes confirmed the placement of Acanthostigma in the Tubeufiaceae.