Pollen Morphometric Variation Among Seven Poaceae Species Highlights the Limitations of Routine Aerobiological Light Microscopy for Species-Level Identification

A comparative morphometric analysis was conducted on seven Poaceae species (Brachypodium pinnatum, Dactylis glomerata, Elymus repens, Festuca pratensis, Festuca rubra aggr., Lolium perenne, and Poa pratensis) to evaluate the potential for species-level discrimination under routine aerobiological light-microscopic conditions. The analysis included 1000 pollen grains per species from five independent individuals representing two geographically separated populations. Hydrated pollen grain size at inflorescence phenological stage 3 ranged from 21 to 40 µm among the examined species. To standardize pre-microscopic conditions, all inflorescences were stored for two weeks at 22 °C and 36% relative humidity, resulting in partial pollen dehydration. Pollen variation was characterized using the longest visible diameter (L), the diameter perpendicular to it (W), and their ratio (L/W). L ranged from 9 to 22 µm and W from 7 to 20 µm, with extensive overlap among species. Comparison with available literature reference values indicated an estimated reduction in characteristic pollen size of approximately 48–61%. Although statistically significant interspecific differences were detected for most species pairs, the extensive overlap in L, W, and L/W distributions prevented reliable species-level identification based on these morphometric parameters alone. Under basic-fuchsin-stained 400× light-microscopic conditions, additional potentially informative characters, such as fine aperture, annulus, and exine features, are not consistently accessible or quantifiable in randomly oriented grains. Thus, the morphometric parameters examined here provide insufficient diagnostic resolution for reliable species-level identification under routine aerobiological conditions.