eDNA phytoplankton inventories in lakes distributed along an anthropogenic gradient
Based on two sampling campaigns conducted in 2022 and 2023, 186 lakes in France were sampled during one to four seasons each, for a total of 599 samples. These lakes are part of the Water Framework Directive (WFD) monitoring network under the Water Framework Directive (WFD). Water samplings for eDNA analysis of phytoplankton were synchronized with physico-chemical measurements (including ammonium, nitrates, orthophosphates, total phosphorus and transparency) to ensure temporal consistency for statistical analysis. Integrated water samples were collected from the euphotic zone in accordance with WFD guidelines for microscopic monitoring of phytoplankton (AFNOR 2017). Filter cartridges (SterivexTM with a porosity of 0.45 µm) were used in the field to filter 250 mL of water (or less if clogging). Lysis buffer (sucrose 0.75 M, Tris 50 mM pH 8, EDTA 40 mM) was added to the filter cartridges after filtration, to preserve DNA until laboratory storage at -20°C prior to DNA extraction. In only four lakes (Geneva, Bourget, Annecy, Aiguebelette), filtration methodwas differentdiffered. Water subsamples (250 mL) intended for molecular analysis were placed in cool box and filtered in the laboratory on using open filters (MF-Millipore®, cellulose nitrate membrane filter with a porosity of 0.45 µm) less than four hours after sampling in the field, and then immediately frozen at -20°C in a 2 mL Eppendorf tube until DNA extraction. DNA was extracted using a protocol combining the NucleoMag Water kit (Macherey-Nagel) and an automated platform (MagnetaPure 32 Nucleic Acid Purification System, Dutscher). The UPA region of the 23S rDNA gene (~420bp) was amplified by PCR with using the primers ECLA23S_F1 and ECLA23S_R1 (Canino et al. 2023). FastQ files were obtained after DNA sequencing based onusing the Illumina NextSeq2000 technology with the v3 reagent kint (2×250bp) at on the PGTB platform (Bordeaux, France).  AFNOR, XP T90-719 (2017). Qualité de l'eau – Échantillonnage du phytoplancton dans les eaux intérieures. Canino, A. et al. (2023). Which barcide to decipher freshwater microalgal assemblages? Tests on mock communities. Int. J. Lim., 59, 8. https://doi.org/10.1051/limn/2023008