Fungi play a crucial role in terrestrial Arctic ecosystems as symbionts of vascular plants and nutrient recyclers in soil, with many species persistently or temporarily inhabiting the phyllosphere of the vegetation. In this study we apply high-throughput sequencing to investigate the mycobiome of 172 samples of fresh (current year) and aged (3 year old) needles of Picea glauca from three sites over a distance of 500 km in Alaska (USA). We analysed Illumina-generated ITS2 sequences to relate mycobiome data with phenotypic tree traits, measures of genetic variation and climate variables obtained from long-term monitoring of the sites. Alpha-diversity declined with increasing environmental stress/climate harshness. Fungal communities differed in richness and taxonomic composition between sites, with a pronounced difference in the relative abundance of OTUs assigned to species of the rust genus Chrysomyxa, plant pathogens which seem to have been in an outbreak at two sites at the time of sampling. Beside climate parameters, needle age was the second strongest explanatory variable of the mycobiome composition, whereas we found no effect of tree genetic variation, indicating that environmental and tree trait specific variables mainly determined individual white spruce mycobiomes at Alaska's treelines.