Background: Grasses release the most widespread aeroallergens with considerable sensitization rates, while different species produce several pollen concentration peaks throughout the season. This study analyzed the prevalence of grass species in three different European city areas and compared the flowering period of these species with daily pollen concentrations and the symptom loads of grass pollen allergy sufferers. Methods: The most prevalent grass species in Vienna (Austria), Berlin (Germany) and Turku (Finland) were studied and examined by use of three different approaches: phenology, pollen monitoring and symptom load evaluation. A mobile pollen exposure chamber was employed to observe reaction patterns of grass pollen allergy sufferers to three common grass species evaluated in this study versus placebo. Results: Common meadow grass (Poa pratensis) and the fescue grass species (Festuca spp.) are important contributors within the grass pollen season. The pollination period of orchard grass (Dactylis glomerata) and false-oat grass (Arrhenatherum elatius) indicated a greater importance in Berlin and Vienna, whereas a broader spectrum of grass species contributed in Turku to the main pollen season. The standardized provocation induced a nasal symptom load, reduction in nasal flow and increased secretion, in contrary to the placebo control group in grass pollen allergic subjects. Conclusion: The phenological observations, pollen measurements and symptom data evaluation provided unique insights into the contribution of multiple grass species in different European regions. All investigated grass species in the provocation induced rhinitis symptoms of comparable significance, with some degree of variation in symptom patterns.