1. Dissolved inorganic carbon fixation of Thaumarchaeota vs. Bacteria in the meso- and upper bathypelagic waters of the world´s oceans differenciated with the use of metabolic inhibitors Marta M. Varela, Gerhard Herndl, Eva Sintes, Eva Teira, & Josep Gasol Instituto Español de Oceanografía-IEO. CO-Coruña Universidade de Viena Universidade de Vigo. Institut de Ciències del Mar. CMIMA, CSIC

2. Recent studies suggest that unidentified prokaryotes fix inorganic carbon at significant rates in the dark ocean (Reinthaler et al. 2010; Varela et al. 2011) Genomic and physiological studies have shown evidences that Thaumarchaeota should play an important role in the carbon cycle (Herndl et al. 2005; Walker et al. 2010) eastern-Atlanticmid-Atlantic WHY? Some Bacteria showed high chemoautotrophic activity Alonso-Sáez et al. 2010, Varela et al. 2011)

3.  Global ocean prokaryotic community composition:  Major groups of prokaryotes: Bacteria, Eury- and Thaumarchaeota  Bacteria: SAR 11, SAR202, Alteromonas, SAR 406, SAR324  Global ocean and deep water bulk uptake of bicarbonate, differentiating Bacteria and Thaumarchaeota contribution (Yokokawa et al.  Single-cell identification of active dark bicarbonate fixers To examine the contribution of Thaumarchaeota vs. Bacteria to total prokaryotic fixation of dissolved inorganic carbon (DIC) in the meso- and upper batypelagic waters of the world oceans. GOAL OF THE STUDY:

4. Radiolabeled Substrate ( 14 C) Incubation Fixation & Filtration Radioactive cell Non-Radioactive cell Prokaryotic community composition: by Fluorescence In Situ Hybridization (CARD-FISH): HOW ? Group Specific Activity: by microautoradiography combined with FISH (MICRO-CARD-FISH) Dissolved inorganic carbon (DIC) fixation: incorporation of 14 C-bicarbonate, by using Erythromycin to discriminate archaeal from bacterial activity (Yokokawa et al. 2012) Bicarbonate assimilation MICRO-CARD-FISH

5. Malaspina 2010 Circumnavigation Expedition: Global Change and Biodiversity Exploration of the Global Ocean" (December 2010-July 2011) STUDY SITE Pacific Ocean Atlantic Ocean Indian Ocean Indian Ocean: 46; 52; 57; 60; 66 Pacific Ocean: 86; 89; 93; 98; 105; 110; 116; 124 Atlantic Ocean: 130; 132; 135; 138; 143

6. Bacteria vs. Eury- and Thaumarchaeota + cells (% of DAPI counts): WHAT ? Indian Ocean Pacific OceanAtlantic Ocean n=9n= n= 4 Mesopelagic Bathypelagic

7. SAR11, SAR 202, Alteromonas, SAR 406, SAR 324 + cells (% of Eub + cell): Indian Ocean Pacific OceanAtlantic Ocean n= 11 Mesopelagic n= 4 Bathypelagic n=9 n= 13

8. DIC fixation by the prokaryotic community (bulk uptake of 14C-bicarbonate): Depth (m) DI 14 C fixation (µmol C m -3 d -1 ) Indian OceanPacific Ocean Atlantic Ocean

9. Contribution of Bacteria vs. Thaumarchaeota to DIC fixation :

10. % of Bacteria or Thaumarchaeota incorporating bicarbonate:

12. SUMMARY  Bacteria dominated throughout the water column in the three world oceans (54% of the total DAPI counts) and did not change with depth. By contrast, the abundance of Cren was generally higher in the bathypelagic layers than in the mesopelagic waters (reaching values up to 29% of the total DAPI counts in the Pacific Ocean).  SAR11 contributed 8, 33 and 18% to total prokaryotic inorganic carbon incorporation in the Indian, Pacific and Atlantic world oceans, respectively, over the entire water column.  Archaea contributed 8, 33 and 18% to total prokaryotic inorganic carbon incorporation in the Indian, Pacific and Atlantic world oceans, respectively, over the entire water column.  The Archaea contribution to total prokaryotic carbon incorporation tends to increases with depth, particularly at the Atlantic waters below 1000m and in the lower mesopelagic zone of the Pacific Ocean.  Single-cell analysis

14.  We thank all the people involved in the MALASPINA 2010 project who helped with the cruise preparation and sampling. ACKNOWLEDGMENTS  We also thank the officers and crew of the R/V Hespérides, as well as the staff of the Technical Support Unit (UTM), for their support during the work at sea.  This research was supported contracts MALASPINA (CSD 2008-00077), BIO-PROF and MODUPLAN.