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RECIPE meeting Scheyern 17-20 May 2005

Progress report AR-WSL, EPFL ECOS Andy Siegenthaler Emanuela Samaritani, Edward Mitchell, Alexandre Buttler. RECIPE meeting Scheyern 17-20 May 2005. Achievements since last meeting Andy (A), Edward (Ed), Emanuela (Em). A&Em: Methane emission in Chaux d’Abel, fall 2004

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RECIPE meeting Scheyern 17-20 May 2005

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  1. Progress report AR-WSL, EPFL ECOS Andy Siegenthaler Emanuela Samaritani, Edward Mitchell, Alexandre Buttler RECIPE meeting Scheyern 17-20 May 2005 RECIPE Meeting Scheyern 17-19.05.2005

  2. Achievements since last meetingAndy (A), Edward (Ed), Emanuela (Em) • A&Em: Methane emission in Chaux d’Abel, fall 2004 • Em: Photosynthesis and respiration measurements in Chaux d’Abel were started in May 2005 • A&Em: CH4-CO2 depth profiles continues (peepers) - Data set now complete for 2004. • A: Development of a new method for measuring soil respiration under realistic conditions • A&Em: Soil respiration and methane production measurements in WPIII to test the new method developed by Andy • A&Em: Soil respiration and methane production measurements in WPI: Selective activities measured using specific inhibitors RECIPE Meeting Scheyern 17-19.05.2005

  3. Achievements - continuedAndy (A), Edward (Ed), Emanuela (Em) • A&Em: Dendrochronological dating of the onset of regeneration • A&Em: Testing probes and protocols for analysing methanogenic Archea using FISH and CARD-FISH • Ed: WPI: Analyses of testate amoeba community structure in the Chaux d’Abel and Russey sites completed, ongoing for others sites • Ed: Establishment of the phylogenetic position of the Arcellinida (one of the two groups of testate amoebae) - Protist, in press • Ed, n.b. with Enrique Lara: Isolation of more Arcellinida species in view of continuing the phylogenetic work to increase the number of sequences in the data base. This will make it possible to analyse testate amoebae community profiles using molecular methods (c.f. also work by Antonis on the Euglyphid testate amoeba) RECIPE Meeting Scheyern 17-19.05.2005

  4. Dendrochronology Microscopic counting and measurement of tree rings, March 2005 Collection and preparation of birch cores, December 2004 RECIPE Meeting Scheyern 17-19.05.2005

  5. Birch recolonisation after abandonmentLa Chaux-d’Abel c b a Years (a) Regeneration stage Method: tree cores collected from the oldest birch trees found at the shortest distance from the centre of a selected stage Statistics: N=11, Kruskal-Wallis followed by Mann-Whitney RECIPE Meeting Scheyern 17-19.05.2005

  6. Gas profiles measurements Cylindrical peepers RECIPE Meeting Scheyern 17-19.05.2005

  7. Pipe to let air get down when peepers are retrieved Cylindrical peepers HT-Tuffryn membrane  50 mm (mesh: 0.2 µm) sealed to the PE cap with drinkable-water Coltgum Transparent PET 0.5 mm sheet to reduce possible leaching from the wood Removable handle      PE cap Cylinder (1m) Acryl 50 ml pot with water-tight PE cap Point 20 cm Insertion for the pot and the the cap Power-tape RECIPE Meeting Scheyern 17-19.05.2005

  8. Dissolved gases at different depths and stages (peepers)Chaux d’Abel, December 2004 RECIPE Meeting Scheyern 17-19.05.2005

  9. Dissolved gases at different depths and stages(peepers)Le Russey, autumn 2004 RECIPE Meeting Scheyern 17-19.05.2005

  10. Stable isotopes in peeper’s water: method summary 1) Samples were taken from the headspace of the degassed peat solutions found in the dialysis chambers (peepers). We used very fine needles (0.55 mm diam.) to sample through the 1 cm thick septae. 2) Separation between CH4 and CO2 was made using a infrared gas chromatograph equipped with a Varian poraplot Q coated column together with plot fused silica (25m x 0,32 mm). 3) The isotopic (12C-13C)/12C ratios were measured separately for CH4 and CO2 with help of a continuous-flow isotope ratio mass-spectrometer coupled to the gas chromatograph (GC-MS). RECIPE Meeting Scheyern 17-19.05.2005

  11. Delta 13C at various depths and stages (peepers) - Analyses by AndreasChaux-d’Abel, autumn 2004 Note: the CH4 concentrations were too low to measure d13C at H3 and, for A and C at H4. RECIPE Meeting Scheyern 17-19.05.2005

  12. 2D-clustering d13C-CH4 vs d13C-CO2 A C B RECIPE Meeting Scheyern 17-19.05.2005

  13. Repartition of delta 13C from dissolved gases(peepers), pooled depths Chaux d’Abel - December 2004 a b c D13C-CO2(0/00) Stats: N = 20, error bars = +/- 1 std error, Kruskal-Wallis followed by Mann-Whitney RECIPE Meeting Scheyern 17-19.05.2005

  14. Soil respiration profiles :A new method of laboratory incubation RECIPE Meeting Scheyern 17-19.05.2005

  15. Development of a new method to study soil respiration in mixed conditions Some problems with standard basal respirations: Because of the difficulties to separate anaerobiosis from aerobiosis in cores that are an assemblage of micro-sites and because it is also artificial to incubate peat in either N2 or air, we developed a method that allowed us to directly incubate the samples taken on-site. We measured all replicates of Jura sites at three different depths over a period of 15 days at 4-6 days interval. The advantages of this method are: reduced incubation times, get as close as possible to peat atmosphere conditions, no core disturbance to get reed of the roots, no need of extreme incubations in either air or N2, drastic reduction in the time of exposition to atmospheric conditions different than in-situ ones, increase of efficiency and cost effectiveness especially with further “selective” respirations using various microbial inhibitors. RECIPE Meeting Scheyern 17-19.05.2005

  16. The Monovette system RECIPE Meeting Scheyern 17-19.05.2005

  17. New setup for basal respiration Remove roots at one end of the Monovette 1.7 cm peat cores sampling at 3 depths, from 3 replicates and in 3 stages Transfer into a 9ml Monovette leave no air space Create artificial 1 ml headspace and 2 days incubation Flushing 1 ml of the triggered respiration Take 1 ml of gas for GC analysis, consider that loss Incubate the cores for 2, 6, 12, 18 days at 6°C Create an artificial 30% headspace seal properly Inject 2 ml of water or of inhibitor cocktail Short term, anaerobic and aerobic respirations RECIPE Meeting Scheyern 17-19.05.2005

  18. Selective inhibition cocktails RECIPE Meeting Scheyern 17-19.05.2005

  19. Preliminary tests for the WPIII basal respiration at depths in the control plots RECIPE Meeting Scheyern 17-19.05.2005

  20. WP1: Anaerobic respiration of methanogens Chaux-d’Abel a b b N.b. Interestingly, surface respiration measured in fall 2004 showed greater respiration in stage C (advanced) ! RECIPE Meeting Scheyern 17-19.05.2005

  21. Depth profiles: origin of the difference in the overall anaerobic respiration - Chaux-d’Abel Twice as much respiration in site A at 25 and 45 cm depths, very significant. RECIPE Meeting Scheyern 17-19.05.2005

  22. Trend for the autotrophic methanogensChaux-d’Abel (MA+) ° There seems to be less autotrophic methanogens activity in the intermediate stage. RECIPE Meeting Scheyern 17-19.05.2005

  23. Significantly lower basal respiration respiration in the advanced stage Trend towards lower eukaryotic (E*) (all but fungi) respiration in the advanced stage Approx. -14% RECIPE Meeting Scheyern 17-19.05.2005

  24. Steady increase of the methanogens’ respiration - Le Russey b a/b a • Respiration is higher in the advanced stage and mainly occurring near the surface! • Not many significant results are found at le Russey, probably because of the reduced number of replicates (3x). RECIPE Meeting Scheyern 17-19.05.2005

  25. WPIII. Basal respiration at various depths and for specific litters - preliminary results Depths CO2 respiration differs significantly among treatments only at one depth (H6)! Respiration is lower for Sphagnum as compared to either Eriophorum treatments RECIPE Meeting Scheyern 17-19.05.2005

  26. Litterbag experiment - species and water table effects on D13C-CO2 a b c a/b Position Method: surface gases collection with syringe after 8 min and 3h36 in the same dark chambers used for respiration. D13C analysed by A. Gattinger. Statistics: N = 9, error bars = +/- 1 std error(s), Kruskal-Wallis followed by Mann-Whitney RECIPE Meeting Scheyern 17-19.05.2005

  27. Description of microbial communities • Because of the variability throughout time, we must terminate the spring 2004 re-sampling for WPI community counts. • As soon as our data-set is complete, we will link microbial diversity/abundance with physico-chemical and gas production measurements and integrate them in multivariate statistical analyses. • CARD-FISH measurements should be finished by end of July. • According to the preliminary C-balance results, I would look for the important nominal variables (treatments) and reduce the number of samples for the experimental sites. • Greetings from India ! RECIPE Meeting Scheyern 17-19.05.2005

  28. Arcellinida phylogeny • Illustrations of the seven testate amoebae taxa analyzed : a & b : Heleopera sphagni (small green spheres inside the cyst are endosymbiotic algae), c : Nebela tincta var. major, d & e : Hyalosphenia papilio , f : Bullinularia indica, g : Trigonopyxis arcula, h : Arcella artocrea, i : Centropyxis laevigata. Scale bars indicate 50 µm. RECIPE Meeting Scheyern 17-19.05.2005

  29. Partial SSU rRNA data • MrBayes tree of partial SSU rRNA genes, showing the monophyly of Arcellinida. Numbers at nodes indicate posterior probabilities for MrBayes analysis (upper) and bootstrap values for ML analysis (lower). Arcella artocrea Centropyxis laevigata Trigonopyxis arcula Bullinularia indica Hyalosphenia papilio Nebela tincta major Heleopera sphagni RECIPE Meeting Scheyern 17-19.05.2005

  30. Tree of the Actin gene data Heleopera sphagni Arcellinida RECIPE Meeting Scheyern 17-19.05.2005

  31. Euglyphid testate amoebae Arcellinida RECIPE Meeting Scheyern 17-19.05.2005

  32. Testate amoeba species richness in the Jura sites RECIPE Meeting Scheyern 17-19.05.2005

  33. CA of testate amoebae samples RECIPE Meeting Scheyern 17-19.05.2005

  34. How will the community structure of different taxonomic group compare among the sites? Testate amoebae RECIPE Meeting Scheyern 17-19.05.2005

  35. Publications • In press: • Nikolaev SI, Mitchell EAD, Petrov NB, Berney C, Fahrni J, Pawlowski J, 2005. The testate lobose amoebae (order Arcellinida Kent, 1880) finally find their home within Amoebozoa. Protist, 156(2): (In press). • In prep, data available: • Emilie, Daniel, Hung, Andy, Edward: Microbial community methodological paper • Andy: Respiration methodological paper • Andy: WPI - Respiration, CH4 measurements with inhibitors • Andy: WPIII - Respirations and methanogenesis profiles • P. Steinmann, Andy: WPI - Modelling of gas profiles in peat • Fatima, Edward, Alexandre, Daniel G, etc.: WPI preliminary - peat OM, Bacteria, and testate amoebae in Jura sites RECIPE Meeting Scheyern 17-19.05.2005

  36. Publications (continued) • In prep, data partly available: • Andy, Emanuela: WPI Peepers gas profiles, delta 13C • Andy, Emanuela: WPI Archaea community profiles FISH/CARD-FISH • Emanuela, Andy, Mika: WPI Surface gas fluxes, photosynthesis, respiration, methanogenesis • Andy, Daniel Gilbert, etc.: Microbial community structure in WPI at all sites - Alone or with testate amoebae, vegetation and other data? • Andy, Daniel Gilbert etc.: WPI Microbial communities in the Jura sites - relationship with other data gases, FISH, vegetation… • Antonis, Edward: WPI Euglyphida diversity in Chaux d’Abel - comparison molecular and microscopic data RECIPE Meeting Scheyern 17-19.05.2005

  37. Publications (continued) • Planned • Emanuela: WPI Growth of Sphagnum and vascular plant biomass production at Chaux d’Abel • Edward, Enrique???: Further phylogenetic data on the Arcellinida • Possible / optional • Edward: WPI Paleoecology using testate amoebae RECIPE Meeting Scheyern 17-19.05.2005

  38. RECIPE Meeting Scheyern 17-19.05.2005

  39. RECIPE Meeting Scheyern 17-19.05.2005

  40. RECIPE Meeting Scheyern 17-19.05.2005

  41. Does under-pressure have a differential effect on microbial activity? “Mont-Blanc” 4808 m “Nanda Devi” 7828m N=9. Peat samples were incubated in under-pressured 9 ml air tight syringes for a period of 22 days. RECIPE Meeting Scheyern 17-19.05.2005

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