CEBEDEAU • AquaBioSens sampling campaign launched

Background: biodiversity in our waterways

The Water Framework Directive (2000/60/EC) requires member states to ensure the quality of their water bodies, focusing on physico-chemical parameters on the one hand, and ecological quality on the other. It is in this context that CEBEDEAU is working on the AquaBioSens project.

This project focuses on natural waters such as rivers, swimming pools and ponds, and aims to develop a bioindicator of aquatic quality based on genomic analysis of periphyton. Periphyton is an aquatic biofilm adhering to substrates immersed in water. When under the influence of certain environmental factors, changes in the microbiological community are observed. This characteristic makes periphyton an interesting matrix for identifying pollution, or more generally, variations in water quality. The development of this bioindicator will complement current indices, such as the Global Biological Index, the Benthic Diatomic Index, the Macrophyte Index, the Biotic Index, the Integrated Piscicultural Index and so on. The latter are based on microscopic characterization of the organisms sought, but the analyses are relatively long, are carried out at low frequencies (every 2 to 6 years), and require - depending on the case - physical capture of the organisms, which may injure individuals or result in poor detection. Inventories of diatoms, for example, require skilled taxonomists, a profession in short supply in Europe.

Project objective: development of a biological index

The aim of the project is therefore to define a biological index calculation. To achieve this, a sampling campaign is planned for 2025 to collect as many samples as possible. The process is as follows: artificial supports (sampling based on passive accumulation of periphyton) will be immersed in the water for a period of one month, after which samples will be taken every 4 weeks. Samples will be taken by scraping periphyton off the sampler developed during the project, extracting DNA from the periphyton and, finally, sequencing the DNA using nanopore technology.

Project progress

2024: design of sampling plan and development of methods

The year 2024 was devoted to the design of (1) the sampler and (2) the sampling plan, and (3) the development of analysis methods.

The sampling system used for the sampling campaign will be a simple one, consisting of a PVC plate connected to a bridge via a stainless steel chain. A second sampling system is also being designed to compare different types of plastic and design for optimal periphyton recovery (this will be the subject of another article).

The sampling plan, meanwhile, has been designed to obtain samples that are diverse in terms of provenance and potential physico-chemical contamination.The first focus is on the influence of wastewater treatment plant discharges on biodiversity.Samplers will be placed upstream and downstream of wastewater treatment plants treating different types of effluent (domestic and hospital), as well as wastewater treatment plants undergoing rehabilitation.

A second axis concerns the biological quality of water, based on analyses carried out by SPW's Département de l'Etude du Milieu naturel et agricole (DEMNA). This organization carries out analyses of biological indices (diatoms, macrophytes, macro-invertebrates and fish) for the 356 water bodies in Wallonia. The sampling sites selected include an equal number of good and poor biological quality sites. Finally, a third axis concerns the type of water body studied: a mixture of streams and rivers was selected to make up the sampling plan.

The DNA extraction method was the subject of a fairly substantial study to determine which method is the most optimal for extracting DNA from periphyton communities of interest. A total of 3 cell lysis methods combined with 11 commercial DNA extraction kits were compared (see: https://www.cebedeau.be/fr/blog/article/projet-aquabiosens). DNA sequencing methodologies were also developed.

Genes coding for 16S and 18S ribosomal RNA will be sequenced during the sampling campaign to identify prokaryotic and eukaryotic organisms in the periphyton, respectively.

2025: sampling campaign

The aim for 2025 is to carry out the sampling campaign and apply the analysis methods developed. In February 2025, samplers will be installed at the various sampling sites. Various analyses will be carried out during this campaign:

• physico-chemical characterization of the water (temperature, pH, conductivity, oxygenation, water velocity, COD, BOD, nitrogen, phosphorus, etc.),
• taxonomic characterization of the periphyton (Nanopore sequencing of genes coding for 16S rRNA and 18S rRNA and calculation of the Diatom Biological Index),
• periphyton characterization (TSS, MVS, chlorophylls).

Thanks to the acquisition of these data, the aim is to create a metadata file that will eventually enable the bioindicator to be calculated.

Financing

AquaBioSens is an ERDF (European Regional Development Fund) project forming part of the SWaM@Sc (Smart Water Management Across Scales) project portfolio led by the University of Liège, in collaboration with CEBEDEAU. The project began in August 2023 and ends in December 2026.