Niche divergence suggests ecophysiological differences between cryptic phytoplankton species

Species-based ecology is fundamental to understanding and predicting the role of phytoplankton diversity in the production and biogeochemical cycles of marine ecosystems. However, microscopy fails to distinguish genetically distinct but morphologically identical (cryptic) or near-identical (pseudocryptic) species, which are lumped into species complexes under the assumption that they have similar ecological characteristics. To test this hypothesis, we analysed a 12-year V4-18S rDNA metabarcoding dataset (168 dates) from the Long-Term Ecological Research Site MareChiara (Gulf of Naples, Mediterranean Sea). Using Kernel Density Estimations and Outlying Mean Index niche analysis, we investigated the seasonality and ecology of 33 cryptic and pseudocryptic species or genetic variants, identified as amplicon sequence variants (ASVs). These belonged to eight species complexes, selected based on their abundance and frequency, within the diatoms Leptocylindrus, Pseudo-nitzschia, and Chaetoceros, the dinoflagellates Azadinium and Heterocapsa, and the chlorophyte Micromonas. Within each complex, ASVs exhibited distinct, annually recurrent seasonal dynamics, with peaks of the most abundant variants matching morphotype peaks for >60% of the timeline. Crucially, 75% of the sister ASVs rarely co-occurred (<30% of the samples), while their seasonality and ecological niches overlapped <60% in 72% and 56% of the couples, respectively. Divergence between species pairs suggests underrated ecophysiological differences and high adaptive potential within species complexes, highlighting the importance of identifying cryptic and pseudo-cryptic species. These findings demonstrate the power of long-term metabarcoding time-series in refining species-based ecological theories, while pointing to the need to uncover the molecular mechanisms driving phytoplankton phenology.