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Charophyte oospores and their features (size, shape, colour, outer wall ornamentation, etc.) are able to persist in seed banks of aquatic ecosystems. Thus, these characteristics can be used for taxonomic species identification, thereby providing knowledge about the specific composition of the charophyte community in a habitat that currently lacks, or has greatly altered, submerged vegetation. Several years ago, l’Albufera de València lost the charophyte meadows that covered the lake bottom in the past, which had never been studied in detail. Several sediment cores were extracted from the lagoon at three sites and the oospores and gyrogonites were isolated and characterized. The oospore/gyrogonite densities in the sediment layers were estimated. Ten different taxa have been identified: six belonging to genus Chara [Ch. vulgaris, Ch. Hispida var. major, Ch. fragilis (Ch. globularis), Ch. Hispida var. baltica, Ch. aspera, Ch. tomentosa]; one Nitella species (N. hyalina), two Tolypella species (T. glomerata and T. hispanica) and one Lamprothamnium species (L. papulosum). The latter, which indicates the period when water salinity was high in the lagoon, was found in the deepest sediment layers. An increase in the oospore abundance and diversity was observed in line with sediment depth. This indicates, on the one hand, that the lagoon went through different ecological stages (with important changes in salinity, mainly) and, on the other hand, the eutrophication process that the lagoon has been undergoing for at least the last 45 years, has progressively affected the abundance and diversity of the charophyte community.