Supplementary Online Material for

Supplementary Online Material for Enhanced metabolic versatility of planktonic sulfur-oxidizing proteobacteria in an oxygen-deficient coastal upwelling ecosystem

Alejandro A. Murillo1, Salvador Ramírez-Flandes1, Edward F. DeLong2 and Osvaldo Ulloa1, * 1

Departmento de Oceanografía, Universidad de Concepción, Concepción, Chile

2

Department of Biological Engineering and Department of Civil and Environmental Engineering, Massachusetts Institute of Technology, Cambridge, USA

*To whom correspondence should be addressed. E-mail: [email protected]

1

Table of Contents Figure S1: Water-column nutrient and oxygen profiles during the upwelling period in January 2009………………………………………………………………………………3 Figure S2 to S7: Taxonomic classification of 16S rRNA gene sequences………….…4-9 Figure S8: lysR-cbbM-cbbQ-cbbO organization for the RuBisCO operon………..……10 Figure S9: Phylogenetic tree of SucB………………………………………….…..……11 Figure S10: Phylogenetic tree of cytochrome bc1 subunit and cytochrome oxidase coxA subunit………………………………………………………………………………..13-14 Figure S11: NADH:ubiquinone oxidoreductase………………………………………...15 Figure S12: Fumarate reductase………………………………………………………...16 Figure S13: F-type ATPase………………………………………………………..…….17 Figure S14: Sulfur oxidation (Sox) system…………...………………………..………..18 Figure S15: Sulfur oxidation (Sox) B subunit……..…………………………………....19 Figure S16: Dissimilatory sulfite reductase operon (dsr)…………...…………………..20 Figure S17: Organization of the aprMBA-sat genes……………………………..……...21 Figure S18: Dissimilatory nitrate reductase (nar) operon………………………………23 Table S1: Sequences classification information of the metagenomic data……………...24 Tables S2 to S15: Taxonomic binning for the analyzed contigs…………………….25-34 Table S16: Single sequences not assembled in to contigs…………………………...34-47 Tables S17: 16S rRNA gene sequences recovered from each metagenomic survey……48 Tables 18-19: Identity analysis for the studied contigs……………………………...…..48 Table S20: ABC and TRAP transport systems………………………………………….49 Tables S21 to S30: Identity analysis for the studied contigs…………………….…..50-53

2

Figure S1: Water-column nutrient and oxygen profiles during the upwelling period in January 2009. The percentages of sequences assigned to major taxonomic groups of organisms derived fron the protein-coding sequences are shown for the 85 m metagenome in the colour coded stacked bar.

3

A

B

Figure S3: Taxonomic classification of 16S rRNA gene sequences, 35 m March 2009. (A) Percentage of sequences, of total Bacterial sequences, associated to a given Bacterial Phyla. (B) Percentage o sequences, of total γ-Proteobacteria Class, associated to a given

4

order. ARCTIC96BD-19 and SUP05 were considered as “orders” artificially separated, to be compared. The color codes start counterclockwise.

A

B


5


A

B


6


A

B

7

Figure S6: Taxonomic classification of 16S rRNA gene sequences, 50 m March 2011. (A) Percentage of sequences, of total Bacterial sequences, associated to a given Bacterial Phyla. (B) Percentage o sequences, of total γ-Proteobacteria Class, associated to a given order. ARCTIC96BD-19 and SUP05 were considered as “orders” artificially separated, to be compared. The color codes start counterclockwise.

8

A

B

Figure S7: Taxonomic classification of 16S rRNA gene sequences, 85 m March 2011. (A) Percentage of sequences, of total Bacterial sequences, associated to a given Bacterial

9

Phyla. (B) Percentage o sequences, of total γ-Proteobacteria Class, associated to a given order. ARCTIC96BD-19 and SUP05 were considered as “orders” artificially separated, to be compared. The color codes start counterclockwise.

Figure S8: lysR-cbbM-cbbQ-cbbO organization for the RuBisCO operon retrieved from metagenomic data (year 2011 sampling). RuBisCO operon (yellow) described for the (A) uncultured GB SUP05 cluster bacterium gb2_c20, (B) the uncultured ESP-GSO cluster Contig14992, and (C) for uncultured SI SUP05 clone FGYC_13J7. The operons are in synteny, but present differences in the genetic context of the surrounding genes. The identity between each gene segment is shown in white numbers. Genes in black are hypothetical proteins. Genes in green are part of the nar operon.

10

11

Figure S9: Phylogenetic tree of SucB. Amino acid sequences, translated from the sucB gene, were compared against - and -proteobacteria representatives for the 2oxoglutarate dehydrogenase, E2 subunit, dihydrolipoamide succinyltransferase. The tree was constructed using the “Bosque” software (Ramírez-Flandes and Ulloa, 2008), with PhyML for maximum likelihood phylogenies, an optimized tree topology, and bootstrap of 100.

12

A

B

13

Figure S10: Phylogenetic tree of (A) cytochrome bc1 subunit and (B) cytochrome oxidase coxA subunit of the oxidative phosphorylation pathway. Amino acid sequences, translated from the cytb and coxA genes, were compared against -, -,  and proteobacteria representatives. The trees were constructed in the “Bosque” software, with PhyML for maximum likelihood phylogenies, an optimized tree topology, and bootstrap of 100.

14

Figure S11: NADH:ubiquinone oxidoreductase, complex

I of the oxidative

phosphorylation pathway. A) Comparison of the complex I (yellow) and the genomic context between the 2009 and 2011 contigs, and against the GB SUP05 subcluster gb1_c97 contig, SI SUP05 genomic scaffold (shown in green to denote de difference in size) and the symbionts Candidatus Ruthia magnifica and Candidatus Vesicomyosocius okutanii reference genomes. B) Specific annotation of the complex I and the flanking genes of contig generated with 2009 metagenomic data.

15

Figure S12: Fumarate reductase, complex II of the oxidative phosphorylation pathway. A) Comparison of the complex II (yellow) and the genomic context between the 2009 and 2011 contigs, and against the GB SUP05 subcluster gb1_c85 contig, SI SUP05 genomic scaffold and the symbionts Candidatus Ruthia magnifica and Candidatus Vesicomyosocius okutanii reference genomes. B) Specific annotation of the complex II and flanking genes of contig generated with 2009 metagenomic data.

16

Figure S13: F-type ATPase, complex V of the oxidative phosphorylation pathway. A) Comparison of the complex V (yellow) and the genomic context between the 2009 and 2011 contigs (99% of identity is represented by the black colour of synteny comparisons) and against the GB SUP05 subcluster gb1_c36 and gb2_65 contigs, SI SUP05 genomic scaffold

and

the

symbionts

Candidatus

Ruthia

magnifica

and

Candidatus

Vesicomyosocius okutanii reference genomes. B) Specific annotation of the complex V and flanking genes of contig generated with 2009 metagenomic data.

17

Figure S14: Sulfur oxidation (Sox) system. A) Incomplete soxAZYX cluster (yellow) associated to SUP05 lineage reconstructed in contigs from 2009 and 2011 metagenomic data, and compared with the GB SUP05 subcluster gb2_c35, SI SUP05 genomic scaffold and the symbionts Candidatus Ruthia magnifica and Candidatus Vesicomyosocius okutanii reference genomes. The soxB gene was recovered in a different contig suggesting a gen context different from the rest of the cluster, a classical organization described in other organisms. B) Specific annotation of sox genetic cluster coding in the contig retrieved from 2009 metagenomic data.

18

Figure S15: Sulfur oxidation (Sox) B subunit. A) Comparison of the soxB gene (yellow) and gene context of the contigs retrieved from 2009 and 2011 metagenomic data, against the GB SUP05 subclusters gb1_c23, gb1_c40 and gb2_c138 contigs, SI SUP05 genomic scaffold

and

the

symbionts

Candidatus

Ruthia

magnifica

and

Candidatus

Vesicomyosocius okutanii reference genomes. For 2009 data two contigs contain part of the soxB gene and present synteny with the described genomic context. B) Specific annotation of 2011 contig coding for soxB and the flanking genes.

19

Figure S16: Dissimilatory sulfite reductase operon (dsr). A) The dsr gene cluster generated with 2009 and 2011 metagenomic data (yellow, 99% of identity is represented by the black colour of synteny comparisons), compared against the reported operons for the GB SUP05 gb2_c106, SI SUP05 genomic scaffold and the symbionts Candidatus Ruthia magnifica and Candidatus Vesicomyosocius okutanii genomes. The dsr cluster retrieved from ESP SUP05 only lacks the dsrS gene compared to the canonical Allochromatium vinosum cluster and conserve synteny with the clusters described. B) Specific annotation of the dsr gene cluster and the flanking genes from the contig obtained with 2009 metagenomic data.

20

21

Figure S17: Organization of adenylyl-sulfate reductase aprMBA (yellow) and ATP sulfurylase sat (light cyan) genes, involved in the transformation of sulfite to sulfate + ATP. A) Comparison of de novo contigs generated from 2009 and 2011 metagenomic data, against the GB SUP05 subcluster gb1_c80, SI SUP05 genomic scaffolds and the symbionts Candidatus Ruthia magnifica and Candidatus Vesicomyosocius okutanii reference genomes. B) Specific annotation of the Sat-Apr complex and their flanking genes retrieved from 2011 data. C) Comparison of de novo contigs generated from 2009 and 2011 metagenomic data and the GB SUP05 subcluster gb1_c80, SI SUP05 genomic scaffolds, against the SCGC data of two ARCTIC96BD-19 representatives, AAA007O20 and AAA001-B15. The comparison shown low nuclotide identity of the sat-apr cluster between SUP05 and ARCTIC96BD-19 representatives and no synteny of the flanking genes, showing that our contig, for this gene cluster, are more closely related to SUP05 cluster.

22

Figure S18: Dissimilatory nitrate reductase (nar) operon. A) Organization of the membrane-bound nitrate reductase operon, the functional enzyme is encoding by narGHJI genes and is often linked to at least one narK gene encoding proteins belonging to the major facilitator superfamily of trans-membrane transporters. B) Histidine kinase nitrate/nitrite-specific regulators of nar operon, retrieved in a different contig and probable located in a different genomic context of the nar genes.

23

Table S1: Number of sequences and sequences classification information of the metagenomic data

Samples

Total seqs

March 2009, 5m

323,081

Putative proteincoding filtered sequences1 239,276

January 2009, 85m

273,690

187,726

203

38.8%

March 2009, 35m

517,392

485,388

241

65.0%

March 2009, 50m

522,769

423,413

240

51.3%

March 2009, 85m

489,998

433,417

247

59.0%

March 2011, 5m

1,207,582

1,189,834

386

60.4%

March 2011, 25m

1,279,798

1,264,855

375

61.2%

March 2011, 50m

1,327,086

1,312,034

378

67.4%

March 2011, 85m

1,328,955

1,313,720

387

72.1%

Average sequence length2

NCBInr3

231

48.7%

Contigs4

Largest contig

299

76,353 bp

1,497

76,610 bp

1

Total of sequences filtered at 98% of similarity after RNA sequences extracted

2

Average sequences length (in base pairs) of the putative protein-coding sequences de-replicated

3

Percentages relative to the putative protein-coding or unknown sequences, i.e., total unique sequences 4

Contigs assembled with metagenomic data enriched in sequences assigned to sulfur-oxidizing organisms. The oxygenated depths were excluded from the assembling.

24

Table S2: Taxonomic binning for contig00465, coding the soxB gene. Organisms Vesicomyosocius okutanii Vesicomyosocius okutanii Ruthia magnifica Ruthia magnifica Ruthia magnifica SUP05 SUP05 ARCTIC96BD-19 AAA007 ARCTIC96BD-19 AAA001 ARCTIC96BD-19 AAA001

Contig start 1 3050 1 1577 3053 1 1636 1865 501 3294

Contig end 2982 3924 1456 2983 4292 1471 4293 2924 602 3326

E-value 0.0 5e-84 0.0 2e-115 2e-95 0.0 0.0 6e-26 0.001 0.004

Bit score 1281 311 998 416 349 994 810 118 44.6 42.8

Table S3: Taxonomic binning for contig31297, coding the soxXAYZ cluster for the oxidation of thiosulfate to sulfur. Organisms SUP05 SUP05 SUP05 SUP05 SUP05 SUP05 SUP05 SUP05 SUP05 SUP05 SUP05 SUP05 SUP05 SUP05 SUP05 SUP05 SUP05 SUP05 SUP05 SUP05 SUP05 Ruthia magnifica Ruthia magnifica Ruthia magnifica Ruthia magnifica Ruthia magnifica

Contig start 44900 32580 2882 20195 8060 25370 38983 15399 27831 31309 43507 13552 24908 27069 13552 18301 14400 24128 42547 32757 22515 44900 24908 20187 38844 2840

Contig end 51536 38376 7942 23969 11719 27726 42074 18179 31151 32405 44695 14352 25275 28010 14286 18932 15378 24915 42966 32858 22811 51536 31151 23225 42073 5245

E-value 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 5e-172 2e-145 1e-128 8e-113 1e-97 3e-73 7e-57 3e-55 7e-25 3e-17 5e-08 5e-08 0.0 0.0 0.0 0.0 0.0

Bit score 4621 2910 2244 2089 1503 1422 1407 1404 982 607 518 462 410 360 279 224 219 118 93.3 62.6 62.6 4453 2778 1636 1317 1236

25

Ruthia magnifica Ruthia magnifica Ruthia magnifica Ruthia magnifica Ruthia magnifica Ruthia magnifica Ruthia magnifica Ruthia magnifica Ruthia magnifica Ruthia magnifica Ruthia magnifica Ruthia magnifica Ruthia magnifica Ruthia magnifica Vesicomyosocius okutanii Vesicomyosocius okutanii Vesicomyosocius okutanii Vesicomyosocius okutanii Vesicomyosocius okutanii Vesicomyosocius okutanii Vesicomyosocius okutanii Vesicomyosocius okutanii Vesicomyosocius okutanii Vesicomyosocius okutanii Vesicomyosocius okutanii Vesicomyosocius okutanii Vesicomyosocius okutanii Vesicomyosocius okutanii Vesicomyosocius okutanii Vesicomyosocius okutanii Vesicomyosocius okutanii ARCTIC96BD-19 AAA007 ARCTIC96BD-19 AAA007 ARCTIC96BD-19 AAA007 ARCTIC96BD-19 AAA007 ARCTIC96BD-19 AAA007 ARCTIC96BD-19 AAA007 ARCTIC96BD-19 AAA007 ARCTIC96BD-19 AAA007 ARCTIC96BD-19 AAA007 ARCTIC96BD-19 AAA007 ARCTIC96BD-19 AAA007 ARCTIC96BD-19 AAA007 ARCTIC96BD-19 AAA007

16815 35644 32631 5277 15481 13506 23344 43495 31883 24156 27280 22617 34953 21343 46564 24905 21037 2882 38829 16823 32572 35640 44899 31391 43495 15481 13425 24326 32591 42856 21876 2841 27 16835 7476 13477 46755 39314 15564 23338 20197 18468 24364 45944

18956 38252 34330 7864 16331 14347 23969 44695 32227 24878 27530 22718 35039 21924 51536 31168 23969 7900 41958 19154 34328 38406 46342 32373 44695 16330 14347 24806 32875 42971 21907 6104 1471 18158 7921 14385 47275 39922 16306 23992 20684 19167 24803 46075

0.0 0.0 0.0 0.0 3e-112 3e-111 1e-110 8e-107 3e-36 2e-32 1e-15 2e-07 2e-05 3e-04 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 7e-171 1e-135 9e-119 4e-110 8e-31 1e-09 2e-07 3e-04 0.0 0.0 0.0 8e-145 8e-107 2e-82 1e-73 1e-72 4e-66 3e-35 2e-31 7e-25 4e-22

1027 1005 962 737 408 405 403 390 156 143 87.8 60.8 53.6 50.0 3925 2742 1874 1671 1251 1050 1023 940 773 603 486 430 401 138 68.0 60.8 50.0 1195 843 664 517 390 309 280 277 255 152 140 118 109

26

ARCTIC96BD-19 AAA007 ARCTIC96BD-19 AAA007 ARCTIC96BD-19 AAA007 ARCTIC96BD-19 AAA007 ARCTIC96BD-19 AAA007 ARCTIC96BD-19 AAA007 ARCTIC96BD-19 AAA001 ARCTIC96BD-19 AAA001 ARCTIC96BD-19 AAA001 ARCTIC96BD-19 AAA001 ARCTIC96BD-19 AAA001 ARCTIC96BD-19 AAA001 ARCTIC96BD-19 AAA001 ARCTIC96BD-19 AAA001 ARCTIC96BD-19 AAA001 ARCTIC96BD-19 AAA001 ARCTIC96BD-19 AAA001 ARCTIC96BD-19 AAA001

46167 47284 759 38827 46581 765 49175 50797 50295 48203 31395 40877 39738 46576 47819 49809 13792 32121

46307 47536 904 39049 46669 844 49723 51091 50554 48487 31803 41117 39861 46751 47902 49921 13915 32334

6e-13 6e-13 2e-07 9e-05 0.001 0.004 1e-59 2e-32 9e-30 5e-27 5e-20 5e-20 3e-16 1e-15 2e-13 6e-13 8e-12 0.001

78.8 78.8 60.8 51.8 48.2 46.4 233 143 134 125 102 102 89.7 87.8 80.6 78.8 75.2 48.2

Table S4: Taxonomic binning for contig31001, coding the dissimilatory sulfite reductase (dsr) gene cluster. Organisms Ruthia magnifica Ruthia magnifica Ruthia magnifica Ruthia magnifica Ruthia magnifica Ruthia magnifica Ruthia magnifica Ruthia magnifica Ruthia magnifica Ruthia magnifica Ruthia magnifica Ruthia magnifica Ruthia magnifica Vesicomyosocius okutanii Vesicomyosocius okutanii Vesicomyosocius okutanii Vesicomyosocius okutanii Vesicomyosocius okutanii Vesicomyosocius okutanii Vesicomyosocius okutanii Vesicomyosocius okutanii

Contig start 2 19084 28044 33100 17696 24844 14287 22717 17210 24382 15757 18843 18833 610 19115 28047 22719 17210 33101 24844 14287

Contig end 13466 22593 31456 34034 18937 26527 15549 24024 17595 24628 16348 18924 18881 12998 22594 31403 24610 18937 33883 26527 15586

E-value 0.0 0.0 0.0 0.0 5e-139 6e-138 3e-116 2e-113 4e-32 6e-30 6e-24 8e-10 1e-06 0.0 0.0 0.0 2e-149 1e-139 7e-131 8e-130 2e-125

Bit score 7488 1921 1290 650 497 493 421 412 141 134 114 68.0 57.2 7068 1907 1272 531 499 470 466 452 27

Vesicomyosocius okutanii Vesicomyosocius okutanii Vesicomyosocius okutanii Vesicomyosocius okutanii Vesicomyosocius okutanii SUP05 SUP05 SUP05 SUP05 SUP05 SUP05 SUP05 SUP05 SUP05 SUP05 SUP05 ARCTIC96BD-19 AAA001 ARCTIC96BD-19 AAA001 ARCTIC96BD-19 AAA001 ARCTIC96BD-19 AAA001 ARCTIC96BD-19 AAA007 ARCTIC96BD-19 AAA007 ARCTIC96BD-19 AAA007

2 33913 15729 18830 18839 1 32786 28117 29960 25062 27142 33913 18846 26878 18854 18837 29231 18819 18840 18847 32786 22878 18854

426 34045 16350 18924 18881 6908 33884 29623 31455 26567 27428 34046 18924 27119 18881 18875 29527 18920 18875 18890 32932 22931 18881

4e-76 1e-39 1e-39 3e-08 6e-05 0.0 0.0 0.0 9e-155 4e-121 8e-48 3e-47 6e-11 1e-07 0.002 0.009 9e-22 5e-06 7e-04 7e-04 3e-08 3e-08 0.002

288 167 167 62.6 51.8 4643 901 682 549 437 194 192 71.6 60.8 46.4 44.6 107 55.4 48.2 48.2 62.6 62.6 46.4

Table S5: Taxonomic binning for contig00369, coding the sat-aprMBA gene cluster, involved in the sulfite oxidation to sulfate and the ATP production. Organisms Ruthia magnifica Ruthia magnifica Ruthia magnifica Vesicomyosocius okutanii Vesicomyosocius okutanii Vesicomyosocius okutanii SUP05 SUP05 SUP05 SUP05 ARCTIC96BD-19 AAA007 ARCTIC96BD-19 AAA007

Contig start 1311 46 4636 1324 46 4637 2543 39 1325 4634 2194 61

Contig end 4619 1270 5213 4619 1259 5209 4629 1270 2582 5218 4603 1061

E-value 0.0 0.0 2e-133 0.0 0.0 3e-106 0.0 0.0 0.0 1e-74 0.0 0.0

Bit score 3007 944 475 2996 897 385 1956 1209 1027 280 944 679

28

Table S6: Taxonomic binning for contig29238, coding the NADH:ubiquinone oxidoreductase gene cluster. Complex I of the oxidative phosphorylation pathway. Organisms Ruthia magnifica Ruthia magnifica Ruthia magnifica Vesicomyosocius okutanii Vesicomyosocius okutanii Vesicomyosocius okutanii SUP05 SUP05 SUP05 SUP05 SUP05 SUP05 ARCTIC96BD-19 AAA007 ARCTIC96BD-19 AAA007 ARCTIC96BD-19 AAA007 ARCTIC96BD-19 AAA007 ARCTIC96BD-19 AAA007 ARCTIC96BD-19 AAA007 ARCTIC96BD-19 AAA007 ARCTIC96BD-19 AAA007 ARCTIC96BD-19 AAA007 ARCTIC96BD-19 AAA007 ARCTIC96BD-19 AAA001 ARCTIC96BD-19 AAA001 ARCTIC96BD-19 AAA001

Contig start 2860 551 24402 2860 551 24402 2860 13099 578 26068 24404 22186 21769 19479 21348 24407 26409 27403 6236 27400 17408 6598 19817 13073 14515

Contig end 17065 2561 25460 17065 2534 25647 13092 17069 2561 26795 25278 22283 24204 20770 21681 24858 26680 27476 6363 27479 17481 6648 20184 13247 14659

E-value 0.0 0.0 8e-78 0.0 0.0 1e-89 0.0 0.0 0.0 3e-52 3e-46 0.002 0.0 0.0 1e-81 2e-54 6e-16 5e-11 2e-09 3e-07 1e-06 6e-04 3e-27 4e-12 2e-04

Bit score 7203 816 293 6937 765 333 5472 2426 890 208 188 46.4 971 865 306 215 87.8 71.6 66.2 59.0 57.2 48.2 125 75.2 50.0

Tables S7: Taxonomic binning for contig13071, coding the Succinate dehydrogenase/Fumarate reductase gene cluster. Complex II of the oxidative phosphorylation pathway. Organisms SUP05 SUP05 SUP05 SUP05 Vesicomyosocius okutanii Vesicomyosocius okutanii Vesicomyosocius okutanii Ruthia magnifica Ruthia magnifica

Contig start 2849 4 2 22 1 22 6455 1 22

Contig end 5448 1570 107 76 1587 76 6531 1405 76

E-value 0.0 0.0 3e-35 6e-06 0.0 5e-07 6e-06 0.0 5e-07

Bit score 1281 830 150 53.6 953 57.2 53.6 913 57.2

29

ARCTIC96BD-19 AAA001 ARCTIC96BD-19 AAA001 ARCTIC96BD-19 AAA001 ARCTIC96BD-19 AAA001 ARCTIC96BD-19 AAA001 ARCTIC96BD-19 AAA001

193 1098 26 192 7431 193

268 1412 76 222 7460 222

3e-16 5e-13 2e-11 2e-04 8e-04 8e-04

87.8 77.0 71.6 48.2 46.4 46.4

Table S8: Taxonomic binning for contig01121, coding the Cytochrome bc1 gene cluster. Complex III of the oxidative phosphorylation pathway. Organisms Vesicomyosocius okutanii Vesicomyosocius okutanii Vesicomyosocius okutanii Ruthia magnifica Ruthia magnifica Ruthia magnifica Ruthia magnifica ARCTIC96BD-19 AAA007 ARCTIC96BD-19 AAA007 ARCTIC96BD-19 AAA007 ARCTIC96BD-19 AAA007 ARCTIC96BD-19 AAA001 ARCTIC96BD-19 AAA001

Contig start 4702 208 2882 7274 33 2882 5119 7273 2883 38 6744 1009 661

Contig end 10707 2095 4598 10717 2095 4600 6192 10147 6313 2095 6872 1243 773

E-value 0.0 0.0 0.0 0.0 0.0 0.0 6e-119 0.0 0.0 0.0 6e-05 6e-24 1e-13

Bit score 1732 1012 958 1474 1162 1003 428 1312 1254 823 50.0 113 78.8

Table S9: Taxonomic binning for contig17720, coding the cytochrome c oxidase subunits I and II. Complex IV of the oxidative phosphorylation pathway. Organisms Ruthia magnifica Ruthia magnifica Vesicomyosocius okutanii Vesicomyosocius okutanii SUP05 ARCTIC96BD-19 AAA007 ARCTIC96BD-19 AAA007 ARCTIC96BD-19 AAA007

Contig start 1169 16 1179 13 4 148 475 4

Contig end 4325 703 4325 743 713 459 744 138

E-value 0.0 1e-21 0.0 2e-32 2e-57 4e-53 1e-47 1e-28

Bit score 1730 104 1579 140 223 208 190 127

30

Table S10: Taxonomic binning for contig13139, coding the ATP synthase (F-type ATPase). Complex V of the oxidative phosphorylation pathway. Organisms SUP05 SUP05 SUP05 SUP05 SUP05 SUP05 SUP05 SUP05 SUP05 Ruthia magnifica Ruthia magnifica Ruthia magnifica Ruthia magnifica Ruthia magnifica Ruthia magnifica Ruthia magnifica Ruthia magnifica Vesicomyosocius okutanii Vesicomyosocius okutanii Vesicomyosocius okutanii Vesicomyosocius okutanii Vesicomyosocius okutanii Vesicomyosocius okutanii Vesicomyosocius okutanii Vesicomyosocius okutanii Vesicomyosocius okutanii Vesicomyosocius okutanii Vesicomyosocius okutanii Vesicomyosocius okutanii Vesicomyosocius okutanii ARCTIC96BD-19 AAA007 ARCTIC96BD-19 AAA007 ARCTIC96BD-19 AAA007 ARCTIC96BD-19 AAA007 ARCTIC96BD-19 AAA007 ARCTIC96BD-19 AAA007 ARCTIC96BD-19 AAA007 ARCTIC96BD-19 AAA007 ARCTIC96BD-19 AAA007 ARCTIC96BD-19 AAA007

Contig start 22 22203 12624 16062 8010 11222 21539 10338 7031 40 18486 12601 16062 10793 8647 15405 10342 27 21539 4754 12601 16072 25105 10846 18486 8932 15405 20118 20919 10338 610 21540 4772 16098 13560 18537 25097 21 20100 12856

Contig end 6748 25804 15888 21360 10203 12452 22058 10679 7550 6796 25795 14448 17946 12310 10204 15890 10681 4453 24870 6759 14446 17945 25792 12293 19834 10201 15890 20663 21337 10686 4369 24875 6544 17919 14453 19762 25788 671 21337 13518

E-value 0.0 0.0 0.0 0.0 0.0 2e-160 1e-107 1e-50 4e-50 0.0 0.0 0.0 0.0 9e-166 1e-131 2e-73 4e-37 0.0 0.0 0.0 0.0 0.0 1e-158 2e-143 1e-132 2e-129 2e-84 1e-50 1e-38 2e-28 0.0 0.0 0.0 4e-177 6e-168 3e-166 2e-155 3e-128 7e-85 8e-78

Bit score 4376 1941 1878 1797 697 567 392 203 201 3990 2718 1002 839 585 471 279 158 3131 1452 1056 1003 884 562 511 475 464 315 203 163 129 2636 1261 719 623 592 587 551 461 316 293

31

ARCTIC96BD-19 AAA007 ARCTIC96BD-19 AAA007 ARCTIC96BD-19 AAA007 ARCTIC96BD-19 AAA001 ARCTIC96BD-19 AAA001 ARCTIC96BD-19 AAA001 ARCTIC96BD-19 AAA001 ARCTIC96BD-19 AAA001

15602 14991 7204 21554 15414 24322 13970 13967

15886 15484 7434 21883 15484 24375 14011 14028

3e-52 7e-34 3e-13 2e-27 8e-08 1e-05 1e-05 0.006

208 147 78.8 125 60.8 53.6 53.6 44.6

Table S11: Taxonomic binning for contig18758, coding the dissimilatory nitrate reductase pathway (nar) gene cluster, involved in the nitrate reduction to nitrite. Organisms SUP05 SUP05 Vesicomyosocius okutanii Vesicomyosocius okutanii Vesicomyosocius okutanii Vesicomyosocius okutanii Ruthia magnifica Ruthia magnifica ARCTIC96BD-19 AAA007

Contig start 164 7323 2105 6297 10660 5466 10660 9576 10665

Contig end 7316 10352 4936 7269 11212 5711 11191 9829 11146

E-value 0.0 0.0 0.0 1e-77 8e-74 4e-45 5e-82 1e-26 1e-70

Bit score 5663 2055 659 291 279 183 306 122 268

Table S12: Taxonomic binning for contig13237, coding the Nitrate reductase (nar) regulators narX/L and narQ. Organisms ARCTIC96BD-19 AAA007 ARCTIC96BD-19 AAA007 ARCTIC96BD-19 AAA007 ARCTIC96BD-19 AAA007 SUP05 SUP05 SUP05 SUP05 SUP05 Ruthia magnifica Ruthia magnifica Vesicomyosocius okutanii Vesicomyosocius okutanii ARCTIC96BD-19 AAA001 ARCTIC96BD-19 AAA001

Contig start 10226 6996 4501 9709 1 471 4495 1 3014 419 1 431 1 6431 5778

Contig end 14083 9519 6904 10008 4339 2813 5718 362 4130 4339 300 4339 257 6727 5853

E-value 0.0 0.0 0.0 2e-31 0.0 0.0 8e-100 8e-81 3e-61 0.0 3e-73 0.0 1e-72 3e-16 8e-05

Bit score 2327 1956 874 138 1687 836 365 302 237 1350 277 1330 275 87.8 50.0

32

Table S13: Taxonomic binning for contig11209, coding the Nitrite reductase nirBD genes, involved in the reduction of nitrite in to ammonium. Organisms SUP05 SUP05 Ruthia magnifica Vesicomyosocius okutanii ARCTIC96BD-19 AAA007 ARCTIC96BD-19 AAA007 ARCTIC96BD-19 AAA007

Contig start 362 5238 388 360 1 1 1

Contig end 3544 5641 3544 3544 63 63 55

E-value 0.0 1e-17 0.0 0.0 2e-16 5e-16 2e-09

Bit score 1487 91.5 1487 1402 87.8 86.0 64.4

Table S14: Taxonomic binning for contig14992, coding the RuBisCO operon genes. Organisms Vesicomyosocius okutanii Vesicomyosocius okutanii Vesicomyosocius okutanii Vesicomyosocius okutanii Vesicomyosocius okutanii Vesicomyosocius okutanii Ruthia magnifica Ruthia magnifica Ruthia magnifica Ruthia magnifica Ruthia magnifica Ruthia magnifica Ruthia magnifica Ruthia magnifica SUP05 SUP05 SUP05 SUP05 SUP05 SUP05 SUP05 ARCTIC96BD-19 AAA007 ARCTIC96BD-19 AAA007 ARCTIC96BD-19 AAA007 ARCTIC96BD-19 AAA007 ARCTIC96BD-19 AAA007 ARCTIC96BD-19 AAA007

Contig start 11479 8901 827 6181 4856 3799 11479 8901 968 4270 6209 3778 8470 14694 800 3770 4270 6120 8292 7327 3765 1208 6110 4270 8285 11580 3831

Contig end 14844 10632 2568 7296 5858 4173 14544 10632 2568 5861 7290 4174 8530 14755 3681 5861 5766 7283 9102 8071 4178 3146 7283 5849 9203 12130 4122

E-value 0.0 0.0 3e-162 6e-96 2e-94 3e-62 0.0 0.0 6e-165 1e-136 5e-71 4e-60 1e-09 2e-07 0.0 0.0 0.0 6e-140 3e-67 6e-64 3e-55 6e-115 1e-111 2e-107 7e-101 2e-64 1e-28

Bit score 2145 1245 572 352 347 241 1894 1139 581 488 269 233 66.2 59.0 937 879 762 499 257 246 217 416 405 390 369 248 129

33

ARCTIC96BD-19 AAA001

11586

11983

2e-44

181

Table S15: Taxonomic binning for contig19949, coding the sucB gene for the E2 subunit of the -ketoglutarate dehydrogenase complex. Organisms SUP05 SUP05 ARCTIC96BD-19 AAA001

Contig start 2902 4291 498

Contig end 4045 4961 703

E-value 9e-158 3e-49 1e-16

Bit score 556 196 87.8

Table S16: Single sequences not assembled into contigs, but with identity to some key functions in the uncultured ESP-GSO cluster bacterium. Sequence ID

Protein product

Taxonomic classification according BLASTX/NR

11159836

dissimilatory nitrite reductase

Pseudomonas entomophila L48

11181664


uncultured bacterium

11199038



11219913


Nitrosopumilus maritimus SCM1

11221723



11248601


Rhodopseudomonas palustris BisA53

11259125


uncultured SUP05 cluster bacterium

11273147



11278075



11280150



11291573



11325924


Maritimibacter alkaliphilus HTCC2654

11350238



11356056


Shewanella loihica PV-4

34

11369742



8032425


Chelativorans sp. BNC1

8035716


Afipia sp. 1NLS2

8038572



8057838


Afipia sp. 1NLS2

8070229


8097543


Rhodopseudomonas palustris CGA009

8098690


Silicibacter sp. TrichCH4B

8116699


Brucella suis 1330

8120022


Flavobacterium johnsoniae UW101

8120750


Candidatus Nitrosoarchaeum koreensis MY1

8130329



8147623



8163434


uncultured organism

8165180


uncultured marine crenarchaeote

8168758


Parvibaculum lavamentivorans DS-1

8173222



8173740


Afipia sp. 1NLS2

8191672


uncultured organism

8196490


Mesorhizobium opportunistum WSM2075

8200688


Flavobacteriaceae bacterium 3519-10

8207551


Afipia sp. 1NLS2

8219546



8223320



8231233



35

8247645


8257923



8280058



8286472



8305217


Phaeobacter gallaeciensis 2.10

8324263



8328512



8337452



8339144


Flavobacterium johnsoniae UW101

8342357



8353306



8362252


Nitrosomonas sp. AL212

8363598



8368866



8373915



8381021



8382760


Afipia sp. 1NLS2

8402569



8420320



8422412



8423452


Bradyrhizobium sp. BTAi1

8425568



8436312


Cardiobacterium hominis ATCC 15826

8453500



8462632



8462948



36

8491604



8496697


Phaeobacter gallaeciensis 2.10

8521757


uncultured marine crenarchaeote HF4000_APKG9M20

2324645


Brucella melitensis bv. 1 str. 16M

2334949



2361524



2383125



2395791


Nitrosococcus oceani ATCC 19707

2436641



2499135


Thermaerobacter marianensis DSM 12885

2537340



2561334



2670781


Maribacter sp. HTCC2170

2751723



2754496



2764310


Hyphomicrobium denitrificans

8946891



8975650



8982583


Afipia sp. 1NLS2

9052992



9119354


Leptospira biflexa serovar Patoc strain 'Patoc 1 (Ames)'

9137682


Chryseobacterium gleum ATCC 35910

9177387


Shewanella amazonensis SB2B

37

9188608



9246590



9260394



9266233



9287480



9301358


Nitrosomonas sp. AL212

9322765


Achromobacter xylosoxidans A8

9355166


Bradyrhizobium japonicum USDA 110

9355806



9404577


Flavobacteriaceae bacterium 3519-10

11242745

flavocytochrome c subunit fccA gamma proteobacterium HTCC5015

11246329

flavocytochrome c subunit fccA Pseudomonas fluorescens Pf0-1

11257122

flavocytochrome c subunit fccA Sulfurimonas autotrophica DSM 16294

11258364

flavocytochrome c subunit fccA uncultured SUP05 cluster bacterium

11285779

flavocytochrome c subunit fccA

11308428

flavocytochrome c subunit fccA Candidatus Vesicomyosocius okutanii HA

11323521


8092687

flavocytochrome c subunit fccA gamma proteobacterium HTCC2207

8119081

flavocytochrome c subunit fccA Nitrosococcus halophilus Nc4

8198572

flavocytochrome c subunit fccA Pseudomonas stutzeri

8215589


8241398

flavocytochrome c subunit fccA Psychromonas ingrahamii 37

8271633


8294160


Aeromonas hydrophila subsp. hydrophila ATCC 7966

Candidatus Ruthia magnifica str. Cm (Calyptogena magnifica)


38

8326594

flavocytochrome c subunit fccA Salinisphaera shabanensis E1L3A

8369349

flavocytochrome c subunit fccA Nitrosococcus oceani ATCC 19707

8498011

flavocytochrome c subunit fccA Photobacterium profundum SS9

8523317

flavocytochrome c subunit fccA Pseudomonas fluorescens WH6

2322874

flavocytochrome c subunit fccA Reinekea blandensis MED297

2327341

flavocytochrome c subunit fccA Methylophaga thiooxydans DMS010

2330440

flavocytochrome c subunit fccA Shewanella loihica PV-4

2343981

flavocytochrome c subunit fccA Neptuniibacter caesariensis

2374831


2375466


2376379


2389208


2443994

flavocytochrome c subunit fccA Pseudomonas fulva 12-X

2463254

flavocytochrome c subunit fccA Pseudomonas mendocina ymp

2491251


2499088


2504309


2536778


2549125


2580010


2601737

flavocytochrome c subunit fccA Shewanella loihica PV-4

2606017


2631764


2653539


2664755




39


2682055


2691848


2693110


2725821


2736012


2738672


2752758


8944836


8945195


9001406

flavocytochrome c subunit fccA Pseudomonas mendocina NK-01

9012287


9013113


9046437


9051215


9067921


9072026


9076749

flavocytochrome c subunit fccA Colwellia psychrerythraea 34H

9081830


9099316


9116415


9214593


9228153


9244952


9280540

flavocytochrome c subunit fccA Chloroherpeton thalassium ATCC 35110




40

9309643


9310844


9312058

flavocytochrome c subunit fccA Alcanivorax borkumensis SK2

9349829

flavocytochrome c subunit fccA Reinekea blandensis MED297

9370464


9372114


9379209


11319789

flavoprotein subunit of flavocytochrome c sulphide


11344843



11354712


Rhodobacterales bacterium HTCC2255

11365964



8156129


Sulfitobacter sp. NAS-14.1

8162154



8358740



8399672


Chlorobium phaeobacteroides DSM 266

8468178



8469424



2372352


Chloroherpeton thalassium ATCC 35110

2483877




41

2584149


Alkalilimnicola ehrlichii MLHE-1

2587797


Roseovarius sp. 217

9231232


Prosthecochloris aestuarii DSM 271

11185307

Nitric oxide reductase subunit, NorB

Magnetococcus sp. MC-1

11286654



11289072


gamma proteobacterium HdN1

8091847



8120821


Methylomonas methanica MC09

8441887


Aromatoleum aromaticum EbN1

2331831


Methylomicrobium album BG8

2336332


Gallionella capsiferriformans ES-2

2378688



2500966


Colwellia psychrerythraea 34H

2510708



2721593


Gallionella capsiferriformans ES-2

2721994



2727924

Nitric oxide reductase subunit,


42

NorB 8985531



9041128


Leptospira biflexa serovar Patoc strain 'Patoc 1 (Paris)'

9103588


Aromatoleum aromaticum EbN1

9140247



9158876


Bdellovibrio bacteriovorus HD100

9164437


Methylomicrobium album BG8

9235513


Pedobacter sp. BAL39

11249465

Nitric oxide reductase subunit, NorC


8115371


Methylococcus capsulatus str. Bath

8391088


Methylococcus capsulatus str. Bath

2316996



2350887



11232163

Nitrous-oxide reductase

Dechloromonas aromatica RCB

11234792


Magnetospirillum magnetotacticum MS-1

11235414


Dechloromonas aromatica RCB

11285643



11296017


Rhodothermus marinus DSM 4252

11381176


Hahella chejuensis KCTC 2396

43

11402898


Magnetospirillum magneticum AMB-1

8046329



8055048



8076239


Nitratiruptor sp. SB155-2

8085108


Gramella forsetii KT0803

8096494


Psychroflexus torquis ATCC 700755

8111312



8156165



8182399


Photobacterium profundum SS9

8277709


Wolinella succinogenes DSM 1740

8298441


Flavobacteriales bacterium ALC-1

8303932



8306195



8307783



8308094



8330361


Myxococcus xanthus DK 1622

8336016


Gramella forsetii KT0803

8377024


Ferroglobus placidus DSM 10642

8399485



8400245



8403529



8451430



2358589



2366243


Cellulophaga algicola DSM 14237

2376210



2382188



44

2422586



2438440


Sulfurimonas autotrophica DSM 16294

2443143



2469536



2477207


Shewanella denitrificans OS217

2485837


Moritella sp. PE36

2495695


Zobellia galactanivorans

2535266



2611256



2631356



2690590


Sulfurovum sp. NBC37-1

2694698



2712371



2720000


Magnetospirillum gryphiswaldense MSR1

2733021


Pirellula staleyi DSM 6068

2738612


Magnetospirillum gryphiswaldense MSR1

2747604


Campylobacterales bacterium GD 1

2752776



8981882



8985285


Magnetospirillum magnetotacticum MS-1

9006200



9077861


Photobacterium profundum 3TCK

9098179


Cellulophaga algicola DSM 14237

9159337



9233865



45

9249483


Symbiobacterium thermophilum IAM 14863

9279554



9290434


Shewanella denitrificans OS217

9302729


Sphaerobacter thermophilus DSM 20745

9335967


Runella slithyformis DSM 19594

9404653



9405883


Pirellula staleyi DSM 6068

9412732



9421709


Flavobacteriales bacterium ALC-1

11217942

Sulfide-quinone reductase


8512804



2319233



2343833


delta proteobacterium MLMS-1

2356450



2463820



2519689



2558454



2595635



2723841



2745672


2753815



2763953



8942671



46

8951760



8966234



8976071


Thiomicrospira crunogena XCL-2

9014270



9077682


Candidatus Vesicomyosocius okutanii HA

9086110



9126841



9173928



9188922


9228446



9277609



9325555



9337096



9354728



9396792



9420399



47

Table S17: Number of 16S rRNA gene sequences recovered from each metagenomic survey.

Metagenome

Total 16S seq

Total 16S seq Bacteria (% of total)

Total 16S seq SUP05 (% of Bac)

Total 16S seq ARCTIC96BD19 (% of Bac)

Total 16S seq Archaea (% of total)

341

299 (87.68)

9 (3.01)

42 (14.05)

42 (12.32)

494

469 (94.94)

63 (13.43)

104 (22.17)

25 (5.06)

506

494 (97.63)

66 (13.36)

92 (18.62)

12 (2.37)

1611

1501 (93.17)

144 (9.59)

190 (12.66)

110 (6.83)

1674

1579 (94)

408 (25.84)

294 (18.62)

95 (5.67)

2009 March_35m 2009 March_50m 2009 March_85m 2011 March_50m 2011 March_85m

Table S18: Contig14992 RuBisCO cluster related genes. Identity at the amino acids level of ESP-GSO against the databases, first four hits. Contig length 14,886 bp, 1,981 sequences recruited, average coverage 30.96X and GC content of 34.8%. Gene lysR cbbM cbbQ cbbO

SI SUP05 66.5 89.8 89.3 76.2

GB SUP05 67.7 77.4 76.2 73.0

C. R. magnifica 51.4 87.2 82.8 74.1

C. V. okutanii 80.2 88.3 82.1 72.5

Table S19: TCA cycle related genes. Identity at the amino acids level of ESP-GSO against the databases, first four hits. Gene mdh sdhB sdhA fumC sucB sucC sucD 2-OG-Fe II iso/isop

SI SUP05

GB SUP05

C. R. magnifica

C. V. okutanii

Contig length (bp)

Average coverage

N° of sequences

Contig GC%

73.5 76.2 76.8 74.2 § § §

¶ 70.3 71.0 61.9 § § §

73.1 ¶ ¶ 65.3 § 70.6 70.1

72.9 ¶ ¶ 62.8 § 68.2 66.9

11,490 11,490 11,490 51,536 5,264 13,924 13,924

1,647 1,647 1,647 7.083 830 2,147 2,147

37.7 37.7 37.7 40.6 37.3 36.5 36.5

§

§

§

§

23,849

32.65X 32.65X 32.65X 32.26X 37.39X 36.21X 36.21X 33.04X

3,358

36.9

82.7

§

76.9

76.6

5,093

10.07X

230

36.0 48

dh iso dh ¶ ¶ 76.6 76.9 22,464 acnB 81.4 ¶ 76.6 73.9 18,443 § The gene sequence is not described for this organism.

32.92X 35.01X

3,149 2,770

39.9 37.6

¶ There was not hit against this organism.

Table S20: ABC and TRAP transport systems associated to the sulfur-oxidizing γProteobacteria uncultured ESP-GSO cluster bacterium. Transporter

Contig

Total contig length (bp)

Function related CDSs (total length bp)

09616

10,548

3 (3,094)

17570

5,149

3 (1,632)

10689

24,523

2 (1,912)

22062

4,807

3 (2,421)

13071

11,490

5 (4,846)

17723

26,967

4 (3,687)

22770

10,191

2 (985)

32011

8,975

5 (5,255)

04646

45,476

3 (3,493)

30874

12,223

4 (4,240)

23558

6,689

5 (3,528)

14418

68,244

4 (4613)

21516

4,341

1(120)

20767

34,001

3 (3,075)

25898

32,563

3 (3,003)

15847

37,142

1 (1,020)

00446

4,061

1 (1,368)

31001

34,585

1 (957)

Amino Acids, general

Amino Acids, branched chain

Carbohydrates

Polyamines Di-peptides Trap Compatible Solutes Antiporter

49

Table S21: Contig29238 NADH:ubiquinone oxidoreductase, complex I of the oxidative phosphorylation pathway. Identity at the amino acids level of ESP-GSO against the databases, first four hits. Contig length 27,560 bp, 3,872 sequences recruited, average coverage 33.43X and GC content of 36%. SI GB C. R. C. V. SUP05 SUP05 magnifica okutanii nuoA 80.0 80.1 80.0 74.1 nuoB 96.2 79.8 94.3 94.3 nuoC 74.7 70.4 71.7 71.2 nuoD 89.9 68.7 88.7 88.0 nuoE 75.2 73.0 68.8 70.1 nuoF 86.6 75.0 85.9 84.5 nuoG 64.7 68.0 59.5 58.6 nuoH 76.9 70.6 73.3 73.3 nuoI 78.0 70.7 81.0 81.6 nuoJ 58.9 67.9 54.5 54.0 nuoK 82.2 71.9 77.2 74.3 ¶ nuoL 74.1 75.5 73.6 § nuoM 80.9 81.9 80.4 § nuoN 76.1 70.1 66.4 § The gene sequence is not described for this organism. Gene

¶ There was not hit against this organism. Table S22: Contig13071 Fumarate reductase, complex II of the oxidative phosphorylation pathway. Identity at the amino acids level of ESP-GSO against the databases, first four hits. Contig length 11,490 bp, 1,647 sequences recruited, average coverage 32.65X and GC content of 37.7%. SI GB SUP05 SUP05 sdhA 86.2 71.0 sdhB 72.8 70.3 sdhC 48.7 61.9 ¶ There was not hit against this organism. Gene

C. R. magnifica ¶ ¶ ¶

C. V. okutanii ¶ ¶ ¶

Table S23: Contig01121 Cytochrome bc1, complex III of oxidative phosphorylation pathway. Identity at the amino acids level of ESP-GSO against the databases, first four hits. Contig length 10,717 bp, 1069 sequences recruited, average coverage 37.21X and GC content of 40.9%. Gene SUP05 isp cytb cyt1

§ § §

GB SUP05 67.3 72.1 69.6

C. R. magnifica 72.0 74.4 46.4

C. V. ARCTIC96BDokutanii 19 73.0 71.0 73.0 72.3 54.8 58.9 50

§ The gene sequence is not described for this organism.

Table S24: Contig17720 Cytochrome c oxidase, complex IV of the oxidative phosphorylation pathway. Identity at the amino acids level of ESP-GSO against the databases, first four hits. Contig length 4325 bp, 183 sequences recruited, average coverage 9.90X and GC content of 41%. SI GB C. R. C. V. SUP05 SUP05 magnifica okutanii coxB § 71.3 59.7 60.5 coxA § 76.3 87.9 87.6 coxC § 71.2 63.8 61.9 ctaG/cox11 § 73.8 73.1 70.9 § The gene sequence is not described for this organism. Gene

ARCTIC96BD19 § § 78.6 §

Table S25: Contig13139 F-type ATPase, complex V of the oxidative phosphorylation pathway. Identity at the amino acids level of ESP-GSO against the databases, first five hits. Contig length 25835 bp, 4,151 sequences recruited, average coverage 38.18X and GC content of 39.4%. SI GB GB C. R. C. V. SUP05 SUP05 gb1 SUP05 gb2 magnifica okutanii a 77.3 74.0 75.4 78.0 77.3 c 97.9 80.5 82.8 71.4 70.1 b 71.5 75.0 74.0 66.3 67.6 δ 56.9 67.6 67.4 50.3 46.4 α ¶ 77.6 78.8 85.1 83.7 γ 84.0 76.6 76.4 77.0 76.5 β 89.5 78.3 77.1 87.5 87.9 ε 77.0 76.8 76.7 72.6 75.5 ¶ There was not hit against this organism. Gene

Table S26: Contig31297 Sulfur oxidation (Sox) system and contig26956 soxB. Identity at the amino acids level of ESP-GSO against the databases, first four hits. Contig length 51,536 bp, 7,083 sequences recruited, average coverage 32.26X and GC content of 40.6%. SI GB SUP05 SUP05 soxA 43.6 58.3 soxZ 74.7 69.8 soxY 45.9 70.6 soxX 71.1 71.8 ¥ soxB 87.3 77.0 76.4€ 75.8£

Gene

C. R. magnifica 40.0 70.9 43.9 63.2 84.4

C. V. okutanii 38.6 67.1 43.2 61.4 80.9 51

¥ € £

Compared to GB SUP05 gb1_c23 Compared to GB SUP05 gb1_c40 Compared to GB SUP05 gb2_c138

Table S27: Contig31001 Dissimilatory sulfite reductase operon (dsr). Identity at the amino acids level of ESP-GSO against the databases, first four hits. Contig length 34,585 bp, 5,068 sequences recruited, average coverage 34.33X and GC content of 36.1%. SI GB C. R. C. V. SUP05 magnifica okutanii SUP05 dsrA ¶ 76.3 80.6 80.0 dsrB 87.4 79.3 84.6 83.5 dsrE 83.6 76.2 82.8 85.7 dsrF 85.8 80.5 83.6 82.1 80.5 dsrH 84.7 82.7 81.6 dsrC 89.7 79.3 85.0 86.0 dsrM 67.9 72.6 63.6 62.1 dsrK 84.0 75.8 81.3 82.0 dsrL § 75.2 80.8 79.1 dsrJ § 66.0 ¶ 51.7 dsrO § 71.6 74.1¥ 71.2 ¥ dsrP § 74.1 65.7 64.8 dsrN § 67.0 63.3¥ 61.2 ¥ dsrR § § 66.4 66.4 § The gene sequence is not described for this organism. Gene

¥ The sequence is described as hypothetical protein in this organism. ¶ There was not hit against this organism. Table S28: Contig21217 adenylyl-sulfate reductase and ATP sulfurylase genes. Identity at the amino acids level of ESP-GSO against the databases, first six hits. Contig length 51,536 bp, 650 sequences recruited, average coverage 24.20X and GC content of 42.2%. SI GB SUP05 SUP05 sat 92.1 81.3 aprM 66.7 62.9 aprB 95.7 80.1 aprA 82.4 82.9 ¥ Compared to SCGC AAA001-B15 Gene

C. R. magnifica 82.4 70.0 97.8 84.5

C. V. okutanii 79.7 67.2 98.9 84.9

ARCTIC96BD19 72.7¥ 72.4€ 38.4¥ 37.4€ ¥ 69.8 70.4€ 73.0¥ 72.7€

€ Compared to SCGC AAA007-O20

52

Table S29: Contigs18758 and 13237 dissimilatory nitrate reductase (nar) pathway. Identity at the amino acids level of ESP-GSO against the databases, first three hits. Contig length (bp)

Average coverage

N° of sequences

Contig GC%

dsrC11,251 78.1 37.4 36.48 like narI 72.7 § § 38.7 11,251 narJ 71.2 § § 33.9 11,251 narH 85.0 § § 66.9 11,251 narG 89.1 § § 58.8 11,251 narK 85.2 § § § 11,251 narK2 72.8 § § § 11,251 narX/L 78.5 § 78.5 77.2 14,083 narQ 70.6 § 66.3 64.4 14,083 § The gene sequence is not described for this organism.

32.05X

1,604

38.3

32.05X 32.05X 32.05X 32.05X 32.05X 32.05X 36.64X 36.64X

1,604 1,604 1,604 1,604 1,604 1,604 2,184 2,184

38.3 38.3 38.3 38.3 38.3 38.3 37.7 37.7

Gene

SI SUP05

GB SUP05 §

C. R. magnifica

C. V. okutanii

Table S30: Contig11209 nitrite reductase (NAD(P)H) nirBD genes. Identity at the amino acids level of ESP-GSO against the databases, first four hits. Contig length 6,599 bp, 344 sequences recruited, average coverage 11.79X and GC content of 39.4%. SI GB C. R. C. V. SUP05 SUP05 magnifica okutanii NO3-/NO274.1 89.1 70.0 69.4 Ser/Thr PK 66.4 § 63.4 62.8 nirB 57.8 58.0 57.2 57.1 nirD § § § § § The gene sequence is not described for this organism. Gene

53