Ectomycorrhizal fungi (EMF) form symbioses with a few plant species that comprise a large fraction of the arctic vegetation. Despite their importance, the identity, abundance and distribution of EMF in the Arctic, as well as the key drivers controlling their community composition are poorly understood. In this study, we investigated the diversity and structure of EMF communities across a bioclimatic gradient spanning much of the North American Arctic. We collected roots from two principal arctic ectomycorrhizal host plants, Salix arctica and Dryas integrifolia, typically growing intermingled, at 23 locations stratified across the five bioclimatic subzones of the Arctic. DNA was extracted from ectomycorrhizal root tips and the ITS region was sequenced and phylogenetically analyzed. A total of 242 fungal Operational Taxonomic Units (OTUs) were documented, with 203 OTUs belonging to the Basidiomycota and 39 to the Ascomycota, exceeding the number of previously morphologically described EMF in the Arctic. EMF communities were dominated by a few common and species-rich families such as Thelephoraceae, Inocybaceae, Sebacinaceae, Cortinariaceae, and Pyronemataceae. Both host plants showed similar species richness, with 176 OTUs on Salix arctica and 154 OTUs on Dryas integrifolia. Host plant identity did not affect EMF community composition. The ten most abundant OTUs had a wide geographic distribution throughout the Arctic, and were also found in boreal, temperate and Mediterranean regions, where they were associated with a variety of hosts. Species richness did not decline with increasing latitude. However, EMF community structure changed gradually across the bioclimatic gradient with the greatest similarity between neighboring bioclimatic subzones and locations. EMF community structure was correlated with environmental factors at a regional scale, corresponding to a complex of glaciation history, geology, soil properties, plant productivity and climate. This is the first large-scale study of EMF communities across all five bioclimatic subzones of the North American Arctic, accompanied by an extensive set of environmental factors analyzed to date. While our study provides baseline data to assess shifts of plant and fungi distribution in response to climate change, it also suggests that with ongoing climate warming, EMF community composition may be affected by northward shifts of some taxa.