1. Assessment of the morphological, genetic, and ecological diversity of coccolithophores along the BIOSOPE transect. Overarching Goal: Colomban de Vargas, Rutgers University, USA The Prymnesiophyceae are the most abundant organisms of the nannoplankton (2-20 μm) in the oligotrophic ocean, where they play a key role in directing flows of biogeochemically and climatically major elements and gasses. Coccolithophores, (Prymnesiophyceae) are one of the rare phytoplanktonic group to produce calcite (40 to 50% of global CaCO 3 production) We propose 1 - to examine, using innovative methods in molecular biology and microscopy, the genetic and morphological diversity of coccolithophores along the vertical and horizontal ecological gradients covered during the BIOSOPE cruise. 2- to quantify calcite production by coccolithophores along the BIOSOPE transect in order to establish relation between organic production and calcite production at specific level. Luc Beaufort, CNRS, CEREGE

2. Scientific Objectives: To estimate the coccolithophore diversity (taxonomy, abundance) at the species level in both the depth and biogeographic ecological gradients covered by BIOSOPE. To identify the genetic provinces of the key coccolithophore taxa in the stratified oligotrophic ocean, and compare the ecological partitioning between the morphological and genetic diversity. To estimate the coccolithophore CaCO3 production in the biogeographic areas covered by BIOSOPE, and particularly in extreme oligotrophic environments. To estimate the ecological importance of naked coccolithophores and other haptophytes in extreme oligotrophy. To analyze the global diversity of nannoplanktonic eukaryotes along BIOSOPE

3. Methods All proposed methods are based on materiel collected on different kind of filters : The morphological diversity of coccolithophores will be analyzed using: Scanning Electron Microscopy: SEM will allow the recognition of ultrastructural characters of the skeleton that allow species distinction in the biological, genetically defined species of coccolithophore. SYRACO: the “SYsteme de Reconnaissance Automatique de COccolithes”, which will allow a rapid –1000 individuals per minute- screening of (1) the morphological taxonomy, (2) the abundance of morphological species at each station and depths, (3) the weight of individual coccoliths and thus the production of CaCO3. The genetic diversity of coccolithophores will be analyzed using: DNA PCR, sequencing, RFLP: The use of specific DNA primers targetting different taxonomic levels within the coccolithophores will allow to (1) reconstruct a first, global phylogeny of the tropical coccolithophores, (2) detect the cryptic genetic diversity within the classical, morphological “species”, (3) analyse the biogeographic and depth partitioning of genetic diversity within the most important taxa. Fluorescence In Situ Hybridization : FISH will permit the identification of naked or non-calcifying coccolithophores and other haptophytes. SSU rDNA clones librairies : rDNA librairies will allow a rapid survey of the nannoplanktonic diversity present along BIOSOPE.

4. E. huxleyi U. tenuis Dollfus & Beauffort, Neur. Net., 1999 Beaufort & Dollfus, Mar. Mic. In Press

5. CaCO 3, growth rate, PP Beaufort, unpublished data Stoll et al., 2002

6. Sampling Strategy We propose to examine 6 different depths at each station: 2 samples below the Deep Chlorophyll Maximum (DCM) 1 in the DCM 1 just above the DCM 1 in the midle-photic zone 1 in the upper-photic zone For each depth, we will need 12 liters of sea water to accommodate the following protocol: Cellulose acetate membrane 12 liters 2-4 l Polycarbonate Membrane 0.8 μm 8 l 1 l SEM Genetics FISH SYRACO