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Three helicosporous hyphomycete collections representing two species were obtained from rotting wood found in freshwater and terrestrial habitats in the Guizhou and Guangxi Provinces, China. A new genus <i>Pseudotubeufia</i> (Tubeufiaceae, Tubeufiales), comprising <i>Ps</i>. <i>hyalospora</i> sp. nov. and <i>Ps</i>. <i>laxispora</i> sp. nov., was introduced with morphological characteristic and molecular data. In addition, the molecular evidence showed that <i>Helicomyces</i> sp. (G.M. 2020-09-19.1), <i>H</i>. <i>roseus</i> (CBS: 102.76), and the new genus <i>Pseudotubeufia</i> clustered together with high support based on a multi-gene (LSU, ITS, <i>tef1α</i>, and <i>rpb2</i>) phylogenetic analysis. Detailed descriptions, illustrations, and notes of the three new collections are provided.