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Ectomycorrhizal (ECM) fungi can influence the establishment and performance of host species by increasing nutrient and water absorption. Therefore, understanding the response of ECM fungi to expected changes in the global climate is crucial for predicting potential changes in the composition and productivity of forests. While anthropogenic activity has, and will continue to, cause global temperature increases, few studies have investigated how increases in temperature will affect the community composition of ectomycorrhizal fungi. The effects of global warming are expected to be particularly strong at biome boundaries and in the northern latitudes. In the present study, we analyzed the effects of experimental manipulations of temperature and canopy structure (open vs. closed) on ectomycorrhizal fungi identified from roots of host seedlings through 454 pyrosequencing. The ecotonal boundary site selected for the study was between the southern boreal and temperate forests in northern Minnesota, USA, which is the southern limit range for Picea glauca and Betula papyrifera and the northern one for Pinus strobus and Quercus rubra. Manipulations that increased air and soil temperature by 1.7 and 3.4 °C above ambient temperatures, respectively, did not change ECM richness but did alter the composition of the ECM community in a manner dependent on host and canopy structure. The prediction that colonization of boreal tree species with ECM symbionts characteristic of temperate species would occur was not substantiated. Overall, only a small proportion of the ECM community appears to be strongly sensitive to warming.