photosynthetic picoeukaryotes in lakes Raw sequence reads

As the key contributor to plankton biomass and nutrient cycling in aquatic ecosystems, photosynthetic picoeukaryotes (PPEs) have been recently investigated in freshwater ecosystems. However, the limited access to remote areas creates challenges for PPE sample preservation before sorting and counting by flow cytometry in the laboratory. Here, we explored the effects of different preservation methods on the PPE community by combining flow cytometric (FCM) sorting and high-throughput sequencing. Our results showed that dimethyl sulfoxide (DMSO) cryoprotection could destroy the fluorescence and cell structure of the PPEs, making the subsequent FCM analysis and sorting difficult. Aldehyde fixation maintained the PPE fluorescence, and the fixed samples were of sufficient quality for abundance analysis and sorting by FCM. However, the sequencing results showed that after preservation by aldehydes, the proportion of PPEs dramatically decreased to approximately 10% in comparison to 90% in the fresh samples, and the sequences of Ascomycota significantly increased. In contrast, the fixation of F68 could not only maintain the PPE abundance close to the initial value but also keep the PPE community similar to that in the freshwater sample over a storage time of six months. Thus, F68 cryopreservation is the only proper preservation method for PPE community research.