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WP3WP3 State of the Art May 2004 CNR IAMC - Istituto per lAmbiente Marino Costiero Messina, Italy Università degli Studi di Messina CNR IAMC - Istituto.

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Presentation on theme: "WP3WP3 State of the Art May 2004 CNR IAMC - Istituto per lAmbiente Marino Costiero Messina, Italy Università degli Studi di Messina CNR IAMC - Istituto."— Presentation transcript:

1 WP3WP3 State of the Art May 2004 CNR IAMC - Istituto per lAmbiente Marino Costiero Messina, Italy Università degli Studi di Messina CNR IAMC - Istituto per lAmbiente Marino Costiero Messina, Italy Università degli Studi di Messina

2 16 rRNA Distribution at Family level In study: >830 sequences analysed 9 levels for Urania from 3.8% of Salinity to BB 7 Levels for Bannockfrom 3.9% to 28% of salinity 8 Levels for Discoveryfrom 6.2% of salinity to BB 7 Levels for LAtalantefrom 4% of salinity to BB 1st Statistical aproach:Family level (-aceae) 2nd Statistical aproach: Division Level

3 9,8 12,8 16,9 19 20,6 23,2 28 24.6 22.4 11 10,2 15,4 14,4 6 4,2 3,9 0 5 10 15 20 25 30 -300-250-200-150-100-50050 1,335 1,340 1,345 1,350 1,355 1,360 1,365 1,370 1,375 1,380 1,385 1,390 16S rRNA Bannock Sampling Profile Salinity % 4 4,8 4,4 5,2 7,8 12,8 19,4 21,2 23,8 24,2 25,6 25,4 26 27,6 0 5 10 15 20 25 30 -350-300-250-200-150-100-500 Eh 1,335 1,34 1,345 1,35 1,355 1,36 1,365 1,37 1,375 1,38 1,385 6,2 77 9 13 15,4 18 21,2 24 24,4 0 5 10 15 20 25 30 -300-250-200-150-100-50050 Eh 1,34 1,35 1,36 1,37 1,38 1,39 1,4 1,41 3,8 6,7 7,7 8,2 8,7 9 10,9 13,4 13,9 15 15,3 5,1 15,6 14,6 16 0 5 10 15 20 -350-250-150-5050 Eh 1,338 1,34 1,342 1,344 1,346 1,348 1,35 1,352 1,354 1,356 1,358 1,36 Refraction Index Eh LAtalanteDiscovery UraniaBannock

4 DHABs Diversity (families) Urania Bannock Discovery L'Atalante Stress: 0,18 nMDS analysis

5 Urania Bannock Discovery L'Atalante U3.8 U5.1 U8.2 U8.7 U10.9 U14 U14.6 U15.6 UBB B3.9 B12.8 B14.4 B16.9 B22.4 B24.6 B28 D6.2 D9 D15.4 D17* D21.2 D24.4 D26* DBB A4 A7,8 A12,8 A19,4 A23,8 A27.6 ABB Stress: 0,18 DHABs Diversity (families) nMDS analysis

6 Diversity indexes (families) High diversity – Low dominanceLow diversity, High dominance

7 7. Urania show a regular increasing of diversity along the gradient Family every basin seems to be a different world! 1. At level of bacterial Family every basin seems to be a different world! 3. Urania, Bannock, and LAtalante shows the highest diversity 4. Urania, Bannock, and LAtalante shows the lowest dominance 5. Discovery shows lower level of diversity 6. Discovery shows dominance of specific families at medium salinity concentration along the gradient 2. There is a grouping made by SW and BB layer………. Real o Bias?

8 Archeon DHABs [CD1] Thermotog. CFB Actinobacteria Candidate division sed 3 KB1 Verrucomicrobiales Clostridiales Flexibact. DHABs[CD2] DHABs[CD3] Spirochaet. Acidobact. Rhizob. Rhodobact. Magnetic Bacterium DHABs[CD4] Desulfuromon. Desulfobact. Desulfoarc. Geobact. Desulfohalob. Desulfobulb. Syntrophobact. SB1 Uncultured Helicobact. Campylobact. Moraxellac. Psudomonad. Oceanospirillales Enterobact. Pisciricketts. Alteromon. UnculturedG Dechlorimarinus Ectothiorhodospir. Chromat. Alcanivorax. UraniaBannockDiscoveryLAtalante

9 Urania Archeon DHABs [CD1] Thermotog. CFB Actinobacteria Candidate division sed 3 KB1 Verrucomicrobiales Clostridiales Flexibact. DHABs[CD2] DHABs[CD3] Spirochaet. Acidobact. Rhizob. Rhodobact. Magnetic Bacterium DHABs[CD4] Desulfuromon. Desulfobact. Desulfoarc. Geobact. Desulfohalob. Desulfobulb. Syntrophobact. SB1 Uncultured Helicobact. Campylobact. Moraxellac. Psudomonad. Oceanospirillales Enterobact. Pisciricketts. Alteromon. UnculturedG Dechlorimarinus Ectothiorhodospir. Chromat. Alcanivorax. ArcheonFlexibact. 0.793 ArcheonPisciricketts. 0.758 DHABs [CD1]Helicobact. 0.886 DHABs [CD1]Pisciricketts. 0.811 Thermotog.KB1 1 ActinobacteriaMoraxellac. 1 ActinobacteriaEnterobact. 0.834 Cand. Div. sed 3DHABs[CD3] 1 Cand. Div. sed 3Spirochaet. 1 Cand. Div. sed 3Magnetic Bacterium 1 Cand. Div. sed 3Desulfohalob. 0.837 KB1Acidobact. 0.796 ClostridialesRhizob. 1 ClostridialesGeobact. 0.735 ClostridialesEctothiorhodospir. 1 DHABs[CD3]Spirochaet. 1 DHABs[CD3]Magnetic Bacterium 1 DHABs[CD3]Desulfohalob. 0.837 Spirochaet.Magnetic Bacterium 1 Spirochaet.Desulfohalob. 0.837 Acidobact.Desulfobact. 0.766 Acidobact.Desulfobulb. 0.785 Rhizob.Geobact. 0.735 Rhizob.Ectothiorhodospir. 1 Magnetic BacteriumDesulfohalob. 0.837 Geobact.Ectothiorhodospir. 0.735 Helicobact.Campylobact. 0.736 Helicobact.Pisciricketts. 0.782 Moraxellac.Enterobact. 0.834 Psudomonad.Enterobact. 0.977 Direct and Inverse Correlations (Pearsons test, P<0.05 (uncorr))

10 UraniaBannock ArcheonDesulfobulb. 0,983 DHABs [CD1]Desulfobact. 0,794 CFBDesulfoarc. 1 CFBUnculturedG 1 ActinobacteriaPisciricketts. 0,735 Cand. Div. sed 3Clostridiales 0,936 Cand. Div. sed 3DHABs[CD2] 0,807 KB1Verrucomicrobiales 0,99 KB1DHABs[CD3] 0,938 KB1SB1 0,839 VerrucomicrobialesDHABs[CD3] 0,879 VerrucomicrobialesSB1 0,79 ClostridialesAcidobact. 0,729 ClostridialesGeobact. 0,729 Flexibact.Moraxellac. 1 Flexibact.Psudomonad. 0,821 DHABs[CD3]Desulfohalob. 0,704 DHABs[CD3]SB1 0,888 Acidobact.Geobact. 1 DHABs[CD3]Enterobact. -0,701 Desulfoarc.UnculturedG 1 Desulfohalob.Alteromon. 0,76 Moraxellac.Psudomonad. 0,821 SB1Psudomonad. -0,74 Direct and Inverse Correlations (Pearsons test, P<0.05 (uncorr)) Archeon DHABs [CD1] Thermotog. CFB Actinobacteria Candidate division sed 3 KB1 Verrucomicrobiales Clostridiales Flexibact. DHABs[CD2] DHABs[CD3] Spirochaet. Acidobact. Rhizob. Rhodobact. Magnetic Bacterium DHABs[CD4] Desulfuromon. Desulfobact. Desulfoarc. Geobact. Desulfohalob. Desulfobulb. Syntrophobact. SB1 Uncultured Helicobact. Campylobact. Moraxellac. Psudomonad. Oceanospirillales Enterobact. Pisciricketts. Alteromon. UnculturedG Dechlorimarinus Ectothiorhodospir. Chromat. Alcanivorax. 1

11 Discovery ArcheonClostridiales 1 ArcheonSpirochaet. 1 ArcheonDechlorimarinus 0,844 CFBAcidobact. 0,722 CFBEctothiorhodospir. 0,896 KB1Verrucomicrobiales 0,851 ClostridialesSpirochaet. 1 ClostridialesDechlorimarinus 0,844 Flexibact.Rhodobact. 1 Flexibact.Desulfobulb. 1 DHABs[CD2]UnculturedG 0,755 DHABs[CD2]Chromat. 1 Spirochaet.Dechlorimarinus 0,844 Rhodobact.Desulfobulb. 1 DHABs[CD4]Desulfuromon. 0,751 DHABs[CD4]Ectothiorhodospir. 0,722 Desulfobact.SB1 0,761 Desulfobact.Alcanivorax. 0,736 SB1Alcanivorax. 0,777 Psudomonad.Alteromon. 0,889 UnculturedGChromat. 0,755 DechlorimarinusAlcanivorax. 0,84 Direct and Inverse Correlations (Pearsons test, P<0.05 (uncorr)) Archeon DHABs [CD1] Thermotog. CFB Actinobacteria Candidate division sed 3 KB1 Verrucomicrobiales Clostridiales Flexibact. DHABs[CD2] DHABs[CD3] Spirochaet. Acidobact. Rhizob. Rhodobact. Magnetic Bacterium DHABs[CD4] Desulfuromon. Desulfobact. Desulfoarc. Geobact. Desulfohalob. Desulfobulb. Syntrophobact. SB1 Uncultured Helicobact. Campylobact. Moraxellac. Psudomonad. Oceanospirillales Enterobact. Pisciricketts. Alteromon. UnculturedG Dechlorimarinus Ectothiorhodospir. Chromat. Alcanivorax. 2

12 LAtalante DHABs [CD1]DHABs[CD2] 1 KB1Pisciricketts. -0,725 Rhodobact.Campylobact. 0,766 Campylobact.Pisciricketts. 0,828 OceanospirillalesAlcanivorax. 1 Direct and Inverse Correlations (Pearsons test, P<0.05 (uncorr)) Archeon DHABs [CD1] Thermotog. CFB Actinobacteria Candidate division sed 3 KB1 Verrucomicrobiales Clostridiales Flexibact. DHABs[CD2] DHABs[CD3] Spirochaet. Acidobact. Rhizob. Rhodobact. Magnetic Bacterium DHABs[CD4] Desulfuromon. Desulfobact. Desulfoarc. Geobact. Desulfohalob. Desulfobulb. Syntrophobact. SB1 Uncultured Helicobact. Campylobact. Moraxellac. Psudomonad. Oceanospirillales Enterobact. Pisciricketts. Alteromon. UnculturedG Dechlorimarinus Ectothiorhodospir. Chromat. Alcanivorax.

13 1. There are a specific DHABs Configuration 2. There are some unique and sporadic phylotypes (probably contamination?) 3. There are Specific direct correlations within tipical DHABs flora. 4. Gamma proteobacteria shows to be often inverse related with typical DHABs Flora 5. Discovery Basin show a peculiar configuration with two big groups(KB1 and Desulfobatcereaceae) domainig two different layer of the basin interface 6. In Discovery and Bannock Basin KB1 and Verrucomicrobiales go very well related along the basin interface gradient 8. In Atalante, there are very well defined bacterial distributions

14 Urania Bannock Discovery L'Atalante Stress: 0,16 2nd Statistical aproach: Divisions Level nMDS analysis DHABs Groups (Divisons)

15 Urania Bannock Discovery L'Atalante Stress: 0,16 U14 U14.6 UBB B3.9 B12.8 B14.4 B16.9 B22.4 B24.6 B28 D6.2 D9 DBB A4 A7,8 A12,8 A19,4 A23,8 ABB U3.8 nMDS analysis DHABs Groups (Divisons)

16 Urania Bannock Discovery L'Atalante Stress: 0,16 U5.1 U8.2 U8.7 U10.9 U14 U14.6 U15.6 UBB B3.9 B12.8 B14.4 B16.9 B22.4 B24.6 B28 D6.2 D9 DBB A4 A7,8 A12,8 A19,4 A23,8 ABB U3.8 nMDS analysis DHABs Groups (Divisons)

17 Urania Bannock Discovery L'Atalante Stress: 0,16 U5.1 U8.2 U8.7 U10.9 U14 U14.6 U15.6 UBB B3.9 B12.8 B14.4 B16.9 B22.4 B24.6 B28 D6.2 D9 D15.4 D17* D21.2 D24.4 D26* DBB A4 A7,8 A12,8 A19,4 A23,8 A27.6 ABB U3.8 nMDS analysis

18 functional group: At level of functional group: 1. There is a common significance that join the more saline Urania layers with the Bannock and Atalante basins 2. Discovery maintains its peculiar bizarre world 3. Some layers of Urania seems to drop out from the common distribution

19 DHABs [CD1] U3.8 U5.1 U8.2 U8.7 U10.9 U14 U14.6 U15.6 UBB B3.9 B12.8 B14.4 B16.9 B22.4 B24.6 B28 D6.2 D9 D15.4 D17* D21.2 D24.4 D26* DBB A4 A7,8 A12,8 A19,4 A23,8 A27.6 ABB Stress: 0,16 nMDS analysis

20 DHABs CFB U3.8 U5.1 U8.2 U8.7 U10.9 U14 U14.6 U15.6 UBB B3.9 B12.8 B14.4 B16.9 B22.4 B24.6 B28 D6.2 D9 D15.4 D17* D21.2 D24.4 D26* DBB A4 A7,8 A12,8 A19,4 A23,8 A27.6 ABB Stress: 0,16 nMDS analysis

21 DHABs Actinobactereaceae U3.8 U5.1 U8.2 U8.7 U10.9 U14 U14.6 U15.6 UBB B3.9 B12.8 B14.4 B16.9 B22.4 B24.6 B28 D6.2 D9 D15.4 D17* D21.2 D24.4 D26* DBB A4 A7,8 A12,8 A19,4 A23,8 A27.6 ABB Stress: 0,16 nMDS analysis

22 DHABs Candidate Division Sediment 3 U3.8 U5.1 U8.2 U8.7 U10.9 U14 U14.6 U15.6 UBB B3.9 B12.8 B14.4 B16.9 B22.4 B24.6 B28 D6.2 D9 D15.4 D17* D21.2 D24.4 D26* DBB A4 A7,8 A12,8 A19,4 A23,8 A27.6 ABB Stress: 0,16 nMDS analysis

23 DHABs KB1 U3.8 U5.1 U8.2 U8.7 U10.9 U14 U14.6 U15.6 UBB B3.9 B12.8 B14.4 B16.9 B22.4 B24.6 B28 D6.2 D9 D15.4 D17* D21.2 D24.4 D26* DBB A4 A7,8 A12,8 A19,4 A23,8 A27.6 ABB Stress: 0,16 nMDS analysis

24 DHABs Verrucomicrobiales U3.8 U5.1 U8.2 U8.7 U10.9 U14 U14.6 U15.6 UBB B3.9 B12.8 B14.4 B16.9 B22.4 B24.6 B28 D6.2 D9 D15.4 D17* D21.2 D24.4 D26* DBB A4 A7,8 A12,8 A19,4 A23,8 A27.6 ABB Stress: 0,16 nMDS analysis

25 DHABs [CD2] U3.8 U5.1 U8.2 U8.7 U10.9 U14 U14.6 U15.6 UBB B3.9 B12.8 B14.4 B16.9 B22.4 B24.6 B28 D6.2 D9 D15.4 D17* D21.2 D24.4 D26* DBB A4 A7,8 A12,8 A19,4 A23,8 A27.6 ABB Stress: 0,16 nMDS analysis

26 DHABs [CD3] U3.8 U5.1 U8.2 U8.7 U10.9 U14 U14.6 U15.6 UBB B3.9 B12.8 B14.4 B16.9 B22.4 B24.6 B28 D6.2 D9 D15.4 D17* D21.2 D24.4 D26* DBB A4 A7,8 A12,8 A19,4 A23,8 A27.6 ABB Stress: 0,16 nMDS analysis

27 DHABs Spiro U3.8 U5.1 U8.2 U8.7 U10.9 U14 U14.6 U15.6 UBB B3.9 B12.8 B14.4 B16.9 B22.4 B24.6 B28 D6.2 D9 D15.4 D17* D21.2 D24.4 D26* DBB A4 A7,8 A12,8 A19,4 A23,8 A27.6 ABB Stress: 0,16 nMDS analysis

28 DHABs Acidobactereaceae U3.8 U5.1 U8.2 U8.7 U10.9 U14 U14.6 U15.6 UBB B3.9 B12.8 B14.4 B16.9 B22.4 B24.6 B28 D6.2 D9 D15.4 D17* D21.2 D24.4 D26* DBB A4 A7,8 A12,8 A19,4 A23,8 A27.6 ABB Stress: 0,16 nMDS analysis

29 DHABs [CD4] U3.8 U5.1 U8.2 U8.7 U10.9 U14 U14.6 U15.6 UBB B3.9 B12.8 B14.4 B16.9 B22.4 B24.6 B28 D6.2 D9 D15.4 D17* D21.2 D24.4 D26* DBB A4 A7,8 A12,8 A19,4 A23,8 A27.6 ABB Stress: 0,16 nMDS analysis

30 DHABs Alpha Proteobacteria U3.8 U5.1 U8.2 U8.7 U10.9 U14 U14.6 U15.6 UBB B3.9 B12.8 B14.4 B16.9 B22.4 B24.6 B28 D6.2 D9 D15.4 D17* D21.2 D24.4 D26* DBB A4 A7,8 A12,8 A19,4 A23,8 A27.6 ABB Stress: 0,16 nMDS analysis

31 DHABs Delta Proteobacteria U3.8 U5.1 U8.2 U8.7 U10.9 U14 U14.6 U15.6 UBB B3.9 B12.8 B14.4 B16.9 B22.4 B24.6 B28 D6.2 D9 D15.4 D17* D21.2 D24.4 D26* DBB A4 A7,8 A12,8 A19,4 A23,8 A27.6 ABB Stress: 0,16 nMDS analysis

32 DHABs Epsilon Proteobacteria U3.8 U5.1 U8.2 U8.7 U10.9 U14 U14.6 U15.6 UBB B3.9 B12.8 B14.4 B16.9 B22.4 B24.6 B28 D6.2 D9 D15.4 D17* D21.2 D24.4 D26* DBB A4 A7,8 A12,8 A19,4 A23,8 A27.6 ABB Stress: 0,16 nMDS analysis

33 DHABs SB1 U3.8 U5.1 U8.2 U8.7 U10.9 U14 U14.6 U15.6 UBB B3.9 B12.8 B14.4 B16.9 B22.4 B24.6 B28 D6.2 D9 D15.4 D17* D21.2 D24.4 D26* DBB A4 A7,8 A12,8 A19,4 A23,8 A27.6 ABB Stress: 0,16 nMDS analysis

34 DHABs Gamma Proteobacteria U3.8 U5.1 U8.2 U8.7 U10.9 U14 U14.6 U15.6 UBB B3.9 B12.8 B14.4 B16.9 B22.4 B24.6 B28 D6.2 D9 D15.4 D17* D21.2 D24.4 D26* DBB A4 A7,8 A12,8 A19,4 A23,8 A27.6 ABB Stress: 0,16 nMDS analysis

35 > Gamma [] [] [] Epsilon Desulfobac>Desulfobulb Archaea–Desulfobact (D) Geobactereaceae HIGH DIVERSITY ZONE KB1+Verr>Desulfobact. (D) Desulfohalob> Desulfobact Desulfobulb>Desulfobact(B) KB1+Verr (D) > Gamma [] [] []

36 Proposed metabolic pathways in the DHABs 3380dbar e-e- e-e- e-e- e-e- e-e- e-e- e-e- e-e- e-e- Volatile fatty acids CO 2 Mn ++ Ba ++ SulphateSulphideSulphur Oxic Anoxic Acetate RedOx Potential NO 3 MnO 2 (FeOH) 3 SO 4 2- CO 2 O2O2O2O2 Eh:-232 -313 * Evidence of in situ active cells or functional genes involved in the pathways e-e- e-e- CO 2 CH 4 Methanogens * AOM and Methanohalophilus 16SrRNA SO 4 2- S-S- S0S0 SRB e-e- e-e- * dsrAB func.genes Fe(OH) 3 Fe 2+ FeRB e-e- * Geobacteriaceae 16SrRNA MnO 2 Mn 2+ Ba ++ Manganese oxidisers * Arcobacter 16SrRNA

37 Discovery Functional charachterisation In study: Distribution along the Discovery salinity graident of: Anaerobic Methane Oxidation related (ANME) bacteria (rRNA) Desulfobatcteraceae distribution (16rRNA) DSR Subunit A gene (RNA) DSR Subunit B gene (RNA)

38 0.1 Methanocaldococcus sp. Methanococcus voltae Methanococcus aeolicus Methanothermococcus okinawensis Halogeometricum borinquense Halophilic archaeon PalaeII AMO-11%-B24 Unidentified archaeon AMO11%-10 AMO11%-5 AMO11%-9 Deep Hidrotermal Vent Environment Group 3 Uncultured archaeon SAGMA-10 Unidentified Crenoarchaeota Uncultured Crenoarchaeota Uncultured archaeon 19a-14 A20-11%-B1 A20-6%-27 A20-11%-B2 A20-6%-12 A20-6%-8 A20-6%-29 Methanohalophilus euhalobius Methanohalophilus portucalensis AMO-19%-6 AMO-19%-7 AMO-11%-2 AMO-11%-3 A20-11%-19 A20-11%-11 AMO-19%-5 AMO-11%-1 ANME-2c Halophilic Metanogens and ANME Discovery Anaerobic Methane Oxidation related (ANME) - bacteria (16SrRNA)

39 0.1 6%-22 6%-3 Desulfobacterium phenolicum Uncultured clone DBBB-72 Desulfobacula toluolica 6%-7 6%-5 6%-36 Uncultured clone DLIB-25 11%-31 6%-35 Desulfobacterium vacuolatum 11%-20 Desulfotignum phosphitoxidans 6%-19 Desulfobacula sp. Desulfospira joergensenii Desulfobacula toluolica 11%-35 6%-21 6%-26 SRB clones at 6% salinity SRB clones at 11% salinity Discovery Desulfobactereaceae distribution clones - bacteria (16SrRNA)

40 Discovery basin SRB clones 6-11% Dissimilatory sulfite reductase Alpha subunit (dsrA) 11%-82 SRB clones at 6% salinity SRB clones at 11% salinity

41 Discovery basin SRB clones 6-11% SRB clones at 6% salinity SRB clones at 11% salinity Dissimilatory sulfite reductase Beta subunit (dsrB)

42 1. At 6% of salinity of Discovery basin we found only members related to crenoarcheota division 2. At 11% and 19% the new DHVE3 3. The anaerobic methanophiles was found at 11% and 19% of salinity - Bacteria 4. 16SrRNA from 6% and 11% of salinity shows mainly Desulfobacteraceae related bacteria 5. dsrA and dsrB distribution shows two levels of diversity (macro and micro) 6. the dsr clusters related to the Desulfobacteriaceae show an high ratio of microdiversity with two defined sub-clusters: i) 6% of salinity clones ii) 6% and 11% togeter - Archaea


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