River biofilms sampling for both downstream DNA analysis and microscopic counts v1
The objective of this protocol is to provide a reliable and replicable method for the sampling of river microphytobenthos and associated microbes in biofilms, to be used in both downstream DNA analysis and algal microscopic counts. The filed protocol is optimized for routine sampling and is in agreement with CEN guidance (NF EN 13946) and CEN technical report (CEN/TR 17245) for the analysis of benthic diatoms from rivers and lakes. The application proposed here in the context of EcoAlpsWater aims in comparing DNA inventories to traditional inventories (microscopy) This protocol is part of the deliverables provided by the WP1 of the Eco AlpsWater project. All members of the EcoAlpsWater consortium (involving 12 partners http:/www.alpine space.eu/projects/ecoalpswater/en/home) have contributed to the optimization of this protocol. Phytobenthos based ecological assessment of running waters is part of the EU Water Framework Directive 2000/60/EC (European Commission) allowing to evaluate the level of nutrients and organic matter pollution in rivers. According to the countries, phytobenthos assessment includes all groups of algae or is based only on diatoms communities. Other periphytic organisms such as fungi, bacteria or microbial eukaryotes as cilates or planktonic (metaphytic) algae are not considered in this assessment method. Further the group of charophytes is covered by the macrophyte ecology method assessment. The recent development of DNA metabarcoding has the potential to complement the traditional biological monitoring based on the direct observation of the organisms, both by reducing sample processing cost and time and by offering the opportunity to consider a larger taxonomic diversity (e.g. bacteria, heterotrophic micro eukaryotes The protocol described here is dedicated to the sampling of biofilms in rivers both for microscopic enumeration of the phytobenthic community and for downstream DNA analyses of microbial assemblages. This field protocol is based on routine methods used for biofilms sampling and is in agreement with: CEN, 2014. Water quality NF EN 13946 Guidance for the routine sampling and preparation of benthic diatoms from rivers and lakes. Afnor, 1 23. CEN 2018. Water quality CEN/TR 17245 Technical report for the routine sampling of benthic diatoms from rivers and lakes adapted for metabarcoding analyses. CEN, 1 8. THIS PROTOCOL ACCOMPANIES THE FOLLOWING PUBLICATION GUIDANCE ON THE MONITORING OF THE BIOLOGICAL QUALITY ELEMENTS PART A3 PHYTOBENTHOS (Jan 2015) Federal Ministry of Agriculture, Forestry, Environment and Water Management DOI dx.doi.org/10.17504/protocols.io.ben6jdhe PROTOCOL CITATION Frederic Rimet, Marine Vautier, Rainer Kurmayer, Nico Salmaso, Camilla Capelli, Agnes Bouchez, Peter Hufnagl, Isabelle Domaizon 2020. River biofilms sampling for both downstream DNA analysis and microscopic counts. protocols.io https:/dx.doi.org/10.17504/protocols.io.ben6jdhe 2 06/11/2020 Cita tion Frederic Rimet, Marine Vautier, Rainer Kurmayer, Nico Salmaso, Camilla Capelli, AgnA A s Bouchez, Peter Hufnagl, Isabelle Domaizon (06/11/2020) River biofilms sampling for both downstream DNA analysis and microscopic counts. https:/dx.doi.org/10.17504/protocols.io.ben6jdhe This is an open access protocol distributed under the terms of the Crea tive Com m ons Attribution License (https:/creativecommons.org/licenses/by/4.0/ which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited MANUSCRIPT CITATION please remember to cite the following publication along with this protocol GUIDANCE ON THE MONITORING OF THE BIOLOGICAL QUALITY ELEMENTS PART A3 PHYTOBENTHOS (Jan 2015) Federal Ministry of Agriculture, Forestry, Environment and Water Management