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Alternaria spores are a common component of the bioaerosol. Many Alternaria species are plant pathogens, and their conidia are catalogued as important aeroallergens. Several aerobiological studies showing a strong relationship between concentrations of airborne spore and meteorological parameters have consequently been developed. However, the Alternaria airborne load variation has not been thoroughly investigated because it is difficult to assess their sources, as they are a very common and widely established phytopathogen. The objective of this study is to estimate the impact of vegetation and land uses as potential sources on airborne spore load and to know their influence, particularly, in cases of long-medium distance transport. The daily airborne spore concentration was studied over a 5-year period in León and Valladolid, two localities of Castilla y León (Spain), with differences in their bioclimatic and land use aspects. Moreover, the land use analysis carried out within a 30 km radius of each monitoring station was combined with air mass data in order to search for potential emission sources. The results showed a great spatial variation between the two areas, which are relatively close to each other. The fact that the spore concentrations recorded in Valladolid were higher than those in León was owing to prevailing winds originating from large areas covered by cereal crops, especially during the harvest period. However, the prevailing winds in León came from areas dominated by forest and shrubland, which explains the low airborne spore load, since the main Alternaria sources were the grasslands located next to the trap. Furthermore, the risk days in this location presented an unusual wind direction. This study reveals the importance of land cover and wind speed and direction data for establishing potential airborne routes of spore transport in order to improve the Alternaria forecasting models. The importance of conducting Alternaria aerobiological studies at a local level is also highlighted. [Display omitted] • There is a great spatial variation in Alternaria airborne load between nearby areas. • The analysis of land use around each trap is crucial to explain spore variation. • High frequencies of low wind speeds from Alternaria sources cause high spore levels. • Cereal crops and grasslands are the most important Alternaria emission sources. • Harvest season with favorable spore transport conditions is the period of greatest risk. [ABSTRACT FROM AUTHOR]