manganese cycles in Shewanella algae C6G3. Axel Aigle1, Patricia Bonin1, Chantal .... 641632455 S. woodyi ATCC 51908. 2546892945 S. waksmanii ATCC ...
Physiological and transcriptional approaches reveal connection between nitrogen and manganese cycles in Shewanella algae C6G3 Axel Aigle1, Patricia Bonin1, Chantal Iobbi-Nivol2, Vincent Méjean2, Valérie Michotey1* Table S1 Presentation of target genes: primers sequences, fragment sizes, annealing temperatures (Tm) and references (ref.). Primers DGGE 300F1 Univ516R1 rpoD-F1 rpoD-R1
Target gene 16S rDNA rpoD
Primer sequences (5’ 3’)
Tm
Ref.
GCC TAC GGG AGG CAG CAG GTDTTACCGCGGCKGCTGRCA
Fragment size
55 °C
172 pb
(1)
TAC GCC GAA GGC CTG AAA AT GAT AGA CAG GCC GGT TTC GG
62 °C
90 pb
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50 °C
505 pb
(2)
58 °C
68 pb
(3)
60 °C
547 pb
62 °C
97 pb
50 °C
414 pb
(4)
58 °C
130 pb
(3)
60 °C
856 pb
62 °C
97 pb
60 °C
540 pb
62 °C
107 pb
60 °C
581 pb
62 °C
110 pb
60 °C
506 pb
62 °C
89 pb
60 °C
515 pb
62 °C
86 pb
nrfA-F1M2 GC WTG YTG GAG YTG TAA nrfA-R1M2 TAG GGC ATG TGA CAG TC nrfA 1 nrfA-2FM CAC GAG AAA GGT TCG CCT A 1 nrfA-2RM CC CAG GGT CTC CAT GGC 2 nrfA-2-F TGC CAA GGA GTA CCA TAG CC nrfA-2-R2 AC AAC TGA TAC CAG CCT TGG nrfA-2 1 nrfA-2-q-F ACT GAC ACT GCC AAG ACC TC 1 nrfA-2-q-R GCA AAC CTG TGA TTG CTG CAT 2 napA V66F TAYTTYYTNHSNAARATHATGTAYGG napA V67R2 DATNGGRTGCATYTCNGCCATRTT napA 1 napA-3FM CCC AAC GCC CGT CAC TG 1 napA-3RM CAT GTT GGA GCC CCA AAG 2 mtrC-F GGT AAT CCA GGT AAC CCA GG mtrC-R2 TCT TCT GTC CAC TTG CTG TT mtrC 1 mtrC-q-F GCT ACA CCT TCA GCA AGA CCT 1 mtrC-q-R GTA TTG TAA GCG CGC AGC AG mtrF-F2 CAT TGC CGG TAA CCA ATG AC 2 mtrF-R TAC TCA GCT TGA GCG AGA AG mtrF 1 mtrF-q-F CCG GAT TCC CCA AAC CCT TG 1 mtrF-q-R GCC ACA GGA CTC CAT GGT TG mtrF-2-F2 CTC CTG GAA CGC CAA TAT CT 2 mtrF-2-R AAC TTA ACG CTT GAA GTG GC mtrF-2 1 mtrF-2-q-F CCA ATA TCT ACC GCG AAG CCT mtrF-2-q-R1 CTG ACA TCG ACT GTC GTC GG 2 omcA-F GGT GCT GGA TAA AGA CGG TA 2 omcA- R GCA GTG TCT TAT CCG GTG TA omcA 1 omcA -q-F AGC CGT ATG ATA GTG GGC TG omcA -q-R1 TCA CTG AGA CGA ATA CGG CG 2 mtrH-F CTA CAG CAA CAA CGA CAG TG 2 mtrH-R TCA TTT CGG CAT GTT GAA CC mtrH 1 mtrH-q-F TCT GCC AAG ACC AGC GTT AC mtrH-q-R1 CCG TGC TTG ACA GTC ATG TTG 1Primers used for Q-PCR analysis, 2primers used to make clones.
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94 2507445795 S. baltica OS625 76
640831968 S. baltica OS185 648728399 S. baltica BA175 640118013 S. baltica OS155 95 100 643462532 S. baltica OS223 651656848 S. sp. HN-41 44 12 71 639725193 S. sp. ANA-3 99 100 2607294001 S. sp. MR-7 100 2608251605 S. sp. MR-4 100 639817576 S. sp. W3-18-1 2587190387 S. putrefaciens JCM 20190 2541006653 S. putrefaciens HRCR-6 100 2606816261 S. oneidensis MR-1 88 86 99 2526387065 S. sp. ZOR0012 59 2551867669 S. sp. POL2 99 2587645948 S. xiamenensis BC01 83 2529061620 S. decolorationis S12 2587183329 S. marina JCM 15074 82 641235376 S. pealeana ATCC 700345 88 641632455 S. woodyi ATCC 51908 95 90 2546892945 S. waksmanii ATCC BAA-643 84 2501720147 S. benthica KT99 640931779 S. sediminis HAW-EB3 66 643427295 S. piezotolerans WP3 92 100 2545749437 S. colwelliana ATCC 39565 71 99 2521614125 S. loihica PV-4 2546916276 S. sp. 38A GOM-205m 100 2568639512 S. algae C6G3 100 46 2587182142 S. algae JCM 21037 2518739822 S. amazonensis SB2B 100 2523119199 F. futtsuensis DSM 18154 2524107488 F. kyonanensis DSM 18153 100 648214825 F. balearica DSM 9799 76 2523949153 F. senticii DSM 18821 100 2523951175 F. senticii DSM 18821 93 2523950954 F. senticii DSM 18821 648214571 F. balearica DSM 9799 98 2524106756 F. kyonanensis DSM 18153 53 KU140670 S. algae C6G3 72 2587179128 S. algae JCM 21037 94 2546914905 S. sp. 38A GOM-205m 100 2566287942 S. haliotis JCM 14758 2510556861 S. alga ACDC 100 639817837 S. sp. W3-18-1 100 2587189830 S. putrefaciens JCM 20190 100 2526387478 S. sp. ZOR0012 41 2551867180 S. sp. POL2 100 80 2541006218 S. putrefaciens HRCR-6 41 639726958 S. sp. ANA-3 2524202538 S. frigidimarina NCIMB 400 2546891976 S. waksmanii ATCC BAA-643 77 100 641552037 S. halifaxensis HAW-EB4 86 641235790 S. pealeana ATCC 700345 54 97 2545621178 S. fidelis ATCC BAA-318 643426763 S. piezotolerans WP3 50 2545749784 S. colwelliana ATCC 39565 2521613776 S. loihica PV-4 52 43 2501721377 S. benthica KT99 83 640931532 S. sediminis HAW-EB3 100
Shewanella nrfA
Ferrimonas
Ferrimonas
nrfA-2 Shewanella
0.1
Figure S1 Phylogenetic distribution of NrfA and NrfA-2 through Shewanella and Ferrimonas genus. This Neighbor-joining tree is based on 747 amino acid characters aligned. The bootstrap percentages higher than 50% are indicated at the node after 1000 resampled data sets. The branch length corresponds to the sequence differences as indicated on the scale bar (substitutions per position).
2526387348 S. sp. ZOR0012 2551868100 S. sp. POL2 2606814039 S. oneidensis MR-1 2587644353 S. xiamenensis BC01 51 2529060131 S. decolorationis S12 639727393 S. sp. ANA-3 45 56 2607293303 S. sp. MR-7 96 100 2608253671 S. sp. MR-4 648727749 S. baltica BA175 100 643461759 S. baltica OS223 100 90 640829672 S. baltica OS185 43640118698 S. baltica OS155 252507443468 S. baltica OS625 100 2587187870 S. putrefaciens JCM 20190 55 639816810 S. sp. W3-18-1 83 651657967 S. sp. HN-41 44 17 54 2541005151 S. putrefaciens HRCR-6 2518741477 S. amazonensis SB2B 2510558050 S. alga ACDC 86 47 100 2587181122 S. algae JCM 21037 99 2566288371 S. haliotis JCM 14758 60 2546917183 S. sp. 38A GOM-205m 55 2568635873 S. algae C6G3 2545749367 S. colwelliana ATCC 39565 42 2521614198 S. loihica PV-4 100 640931846 S. sediminis HAW-EB3 67 643424297 S. piezotolerans WP3 68 61 2524200340 S. frigidimarina NCIMB 400 2612418659 S. sp. YQH10 648211772 F. balearica DSM 9799 2523118444 F. futtsuensis DSM 18154 41 100 2524105868 F. kyonanensis DSM 18153 648212717 F. balearica DSM 9799 641553610 S. halifaxensis HAW-EB4 53 91 641237336 S. pealeana ATCC 700345 100 2545619867 S. fidelis ATCC BAA-318 57 643426638 S. piezotolerans WP3 73 640932394 S. sediminis HAW-EB3 70 100 2546892719 S. waksmanii ATCC BAA-643 2545751073 S. colwelliana ATCC 39565 84 2521615886 S. loihica PV-4 65 2568635154 S. algae C6G3 99 2587180803 S. algae JCM 21037 100 2546916503 S. sp. 38A GOM-205m 49 92 2510557217 S. alga ACDC 2518740402 S. amazonensis SB2B 26 2507443580 S. baltica OS625 46 643461650 S. baltica OS223 49 648727855 S. baltica BA175 18 99 640118807 S. baltica OS155 100 640829776 S. baltica OS185 2541005044 S. putrefaciens HRCR-6 2606814139 S. oneidensis MR-1 99 651658096 S. sp. HN-41 44 18 100 45 2551866064 S. sp. POL2 100 2587644449 S. xiamenensis BC01 70 2526385095 S. sp. ZOR0012 2529060241 S. decolorationis S12 100 98 639727280 S. sp. ANA-3 100 2607293199 S. sp. MR-7 100 2608253565 S. sp. MR-4 38 90
0.1
100
mtrF-2
mtrF
Figure S2 Phylogenetic distribution of MtrF and MtrF-2 through Shewanella and Ferrimonas genus. This Neighbor-joining tree is based on 2,178 amino acid characters aligned. The bootstrap percentages higher than 50% are indicated at the node after 1000 resampled data sets. The branch length corresponds to the sequence differences as indicated on the scale bar (substitutions per position).
Figure S3 Follow-ups of S. algae ATCC 51192 growth (□) on nitrate (3A) or manganese oxide (3B) and concentrations of dissolved nitrite (●), nitrate (○) and manganese (Δ).
Figure S4. Follow-ups of the bacterial growth (□) and concentrations of dissolved nitrite (●) and manganese (Δ).
References 1.
Casamayor EO, Massana R, Benlloch S, Øvreås L, Díez B, Goddard VJ, Gasol JM, Joint I, RodríguezValera F, Pedrós-Alió C. 2002. Changes in archaeal, bacterial and eukaryal assemblages along a
salinity gradient by comparison of genetic fingerprinting methods in a multipond solar saltern. Environmental Microbiology 4:338-348. 2.
Mohan SB, Schmid M, Jetten M, Cole J. 2004. Detection and widespread distribution of the nrfA gene encoding nitrite reduction to ammonia, a short circuit in the biological nitrogen cycle that competes with denitrification. Fems Microbiology Ecology 49:433-443.
3.
Smith CJ, Nedwell DB, Dong LF, Osborn AM. 2007. Diversity and abundance of nitrate reductase genes (narG and napA), nitrite reductase genes (nirS and nrfA), and their transcripts in estuarine sediments. Applied and Environmental Microbiology 73:3612-3622.
4.
Nogales B, Timmis KN, Nedwell DB, Osborn AM. 2002. Detection and diversity of expressed denitrification genes in estuarine sediments after reverse transcription-PCR amplification from mRNA. Applied and Environmental Microbiology 68:5017-5025.
