Assessing the relevance of DNA metabarcoding compared to morphological identification for lake phytoplankton monitoring.

Phytoplankton is a key biological group used to assess the ecological status of lakes. The classical monitoring approach relies on microscopic identification and counting of phytoplankton species, which is time-consuming and requires high taxonomic expertise. High-throughput sequencing, combined with metabarcoding, has recently demonstrated its potential as an alternative approach for plankton surveys. Several studies have confirmed the relevance of the diatom metabarcoding approach to calculate biotic indices based on species ecology. However, phytoplankton communities have not yet benefited from such validation. Here, by comparing the results obtained with the two methods (molecular and microscopic counting), we evaluated the relevance of metabarcoding approach for phytoplankton monitoring by considering different metrics: alpha diversity, taxonomic composition, community structure and a phytoplankton biotic index used to assess the trophic level of lakes. For this purpose, 55 samples were collected in four large alpine lakes (Aiguebelette, Annecy, Bourget, Geneva) during the year 2021. For each sample, a metabarcoding analysis based on two genetic markers (16S and 23S rRNA) was performed, in addition to the microscopic count. Regarding the trophic level of lakes, significant differences were found between index values obtained with the two approaches. The main hypothesis to explain these differences comes from the incompleteness, particularly at the species level, of the barcode reference library for the two genetic markers. It is therefore necessary to complete reference libraries for using such species-based biotic indices with metabarcoding data. Besides this, species richness and diversity were higher in the molecular inventories than in the microscopic ones. Moreover, despite differences in taxonomic composition of the floristic lists obtained by the two approaches, their community structures were similar. These results support the possibility of using metabarcoding for phytoplankton monitoring but in a different way. We suggest exploring alternative approaches to index development, such as a taxonomy-free approach.

Phytoplankton morpho-functional trait dataset from French water-bodies.

In lake ecosystems, phytoplankton communities can be studied by adopting taxonomic-based approaches. However, these approaches suffer from identification issues and are sometimes of limited ecosystem ecological value. The recent development of functional approaches may allow an evaluation of other aspects of ecosystem quality, functions and interactions with abiotic parameters or other communities. Here, our aim was to create a phytoplankton trait database at the French scale. This database will be relevant for the analysis of phytoplankton communities that will lead to a better understanding of phytoplankton functional ecology in lakes of France and other European countries possessing similar biological communities. We used a French national database of phytoplankton occurrences sampled from 384 lakes over the entire French metropolitan territory. A final list of 636 taxa was used to compile 53 morpho-functional traits associated with taxonomic information. The traits encompassed variables such as shape, biovolume, motility, toxin production and Reynolds groups. With this new database, we aim to provide data for morpho-functional analyses of phytoplankton assemblages from local to European scale.

Defining chlorophyll-a reference conditions in European lakes.

The concept of "reference conditions" describes the benchmark against which current conditions are compared when assessing the status of water bodies. In this paper we focus on the establishment of reference conditions for European lakes according to a phytoplankton biomass indicator--the concentration of chlorophyll-a. A mostly spatial approach (selection of existing lakes with no or minor human impact) was used to set the reference conditions for chlorophyll-a values, supplemented by historical data, paleolimnological investigations and modelling. The work resulted in definition of reference conditions and the boundary between "high" and "good" status for 15 main lake types and five ecoregions of Europe: Alpine, Atlantic, Central/Baltic, Mediterranean, and Northern. Additionally, empirical models were developed for estimating site-specific reference chlorophyll-a concentrations from a set of potential predictor variables. The results were recently formulated into the EU legislation, marking the first attempt in international water policy to move from chemical quality standards to ecological quality targets.

Benefits and limitations of an intercalibration of phytoplankton assessment methods based on the Mediterranean GIG reservoir experience.

The status of European legislation regarding inland water quality after the enactment of the Water Framework Directive (WFD) originated scientific effort to develop reliable methods, primarily based on biological parameters. An important aspect of the process was to ensure that quality assessment was comparable between the different Member States. The Intercalibration process (IC), required in the WFD ensures the unbiased application of the norm. The presented results were developed in the context of the 2nd IC phase. An overview of the reservoir type definition of the Lake Mediterranean Geographical Intercalibration Group, where four types were considered divided by both alkalinity and climate, together with the results for selection of Maximum Ecological Potential sites (MEP) are presented. MEP reservoirs were selected based on pressure and biological variables. Three phytoplankton-based assessment methods were intercalibrated using data from Mediterranean countries. The Mediterranean Assessment System for Reservoirs Phytoplankton (Spain), the New Mediterranean Assessment System for Reservoirs Phytoplankton (Portugal and Cyprus) and the New Italian Method (Italy) were applied. These three methods were compared through option 3 of the Intercalibration Guide. The similarity of the assessments was quantified, and the Good/Moderate (GM) boundaries assessed. All three methods stood as comparable at the GM boundary except for the MASRP in siliceous wet reservoirs, which was slightly stricter. Finally, the main taxonomic groups represented in the phytoplankton community at MEP conditions were identified, as well as their main changes with an increasing trophic status. MEP sites are dominated by chrysophytes in siliceous wet reservoirs and by the diatoms Cyclotella and Achnanthes in calcareous ones. Cyanobacteria take over the community in both calcareous and siliceous wet reservoirs as eutrophication increases. In summary, the relevance and reliability of the quality assessment methods compared were confirmed both from an ecological perspective and a health risk management point of view.