SUPPORTING ONLINE MATERIAL. Material and methods.
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SUPPORTING ONLINE MATERIAL. Material and methods. Suppressive subtractive hybridization (SSH). The size of the library was estimated following Bogush et al. (BOGUSH et al. 1999) and Dr. Lev G. Nikolaev personal communication, using the formula: N = n/p, where N is the estimated library size (in clones), n is the number of clones sequenced, and p is the probability of finding x number of independent clones among the n sequenced clones. P was calculated as p = [(c1-1)+(c2-1)+(c3-1)+ ∑..+(cm-1)]/m, where c(i) is the number of observations of clone i, and m is the number of clones sequenced. For each strain we estimated the library size for the two different restriction enzymes independently. For TPRKU1-RsaI (N=267) the p value was 0.25 and a total library size of about 1080 clones, and for TP-RKU1-AluI (N=151) the p value was 0.13 and giving estimated library size of 1140 clones. The number of differential clones was estimated as 27% and 33% for TP-RKU1-RsaI and TP-RKU1-AluI, respectively. The number of differential genes was estimated by multiplying the number of differential clones with the average sequence length (TP-RKU1-RsaI: 423 bp, TP-RKU1-AluI: 422 bp) of the clones and dividing by the average gene length of T. maritima MSB8 genes (947 bp) (NELSON et al. 1999). Supplemental Table 1. Summary of fosmid-end sequences. Strain Na bp seq. b %G+C average bpc DNA identity to no DNA match in T. maritima MSB8 T. maritima MSB8 Thermotoga sp. RQ2 84 30418 46.6 358 95% 4 ( 5% )d Thermotoga petrophila RKU1 109 19636 45.9 180 93% 3 ( 3% ) Thermotoga naphthophila RKU10 22 2817 48.7 128 93% 0 (?%) Thermotoga neapolitana LA4 23 9112 47.0 396 80% 2 (8.7%) Thermotoga neapolitana LA10 36 13710 46.0 380 78% 2 (5.6%) Thermotoga sp. SG1 90 18495 46.8 205 77 % 12 (12%)e Thermotoga sp. RQ7 122 47839 47.8 402 76% 8 (6.5%)f Thermotoga sp. KOL6 26 6546 43.9 216 75% 10 (38%) 4 (15%)g a Number of high quality end sequences obtained. b Total number of bp sequenced. c Average sequence length obtained. d One of the sequences with no DNA match in T. maritima MSB8 had a protein-level match in the MSB8 genome e Four of the sequences with no DNA match in T. maritima MSB8 had protein-level matches. f Two of the sequences with no DNA match in T. maritima MSB8 had protein-level matches. g Sequences that did have a protein match in the T. maritima MSB8 genome not included in the estimate. Supplemental Table 2. Differential genes found among end-sequences of fosmid clones. clone bp first BLASTX hit organism exp. % indent note Thermotoga sp strain RQ2, Library: rq2f2 1.a01.for 404 VV10473 Cell wall-associated hydrolase Vibrio vulnificus 9.5E-42 80 The sequence from the reverse primer shows 95% and 99% DNA identity to TM0042 and TM0043. 1.b09.for 360 AAD32594.1 family 10 xylanase XynC Thermotoga sp. strain FjSS3-B.1 1E-153 94DNA No other hit in Genbank. The sequence from the reverse primer shows 99% and 100% DNA identity to TM1232 and TM1231. 1.g10.rev 325 TM1265 conserved hypothetical protein Thermotoga maritima MSB8 0.10 65 The sequence from the forward primer shows 100% and 98% DNA identity to TM1820 and TM1819. 1.h09 314 NP_782363.1 phosphoenolpyruvate- Clostridium tetani protein phosphotransferase 4E-17 42 Thermotoga petrophila RKU1 Library: rku1f1 2b08 154 no significant hit 2e03.for 41 no significant hit The sequence from the reverse primer shows 93% DNA identity to TM0908. 2f06.for 457 no significant hit The sequence from the reverse primer shows 93% DNA identity to TM0576. Thermotoga neapolitana LA4 1f04.for 438 no significant hit 1g03.for 482 a: BAB04420.1 C4-dicarboxylate transport system b: NP_772697.1| dicarboxylate transporter membrane protein Thermotoga neapolitana LA10 Library: tneLA4f1 The sequence from the reverse primer shows 97% DNA identity to TM0642. Bacillus halodurans 4E-04 31 Bradyrhizobium japonicum 1E-06 36 Library: LA10f1 2g01.for 460 CAB07809.1 permease Thermus sp. T2 4E-08 40 2d02.for 424 NP_435360.1 putative ABC transporter, ATP-binding protein Sinorhizobium meliloti 0.002 40 Thermotoga sp. strain SG1 Library: tspsg1f2 The sequence from the reverse primer shows 87% and 79% DNA identity to TM1172 and TM1173. 2a10.rev 419 a: no significant hit b: TM0281 alpha-Larabinofuranosidase Thermotoga maritima MSB8 2E-57 87 1c01.for 117 TM1541 flagellar protein FlgA, putative Thermotoga maritima MSB8 1E-05 57 1c01.rev 323 NP_996684.1 unknown Bacteriophage phi LC3 0.016 32 1c02.for 102 TM1060 conserved hypothetical protein Thermotoga maritima MSB8 0.24 69 1c07.for 533 PF0911 iron (III) ABC transporter, ATP-binding protein Pyrococcus furiosus 3E-30 67 1d09.for 181 TM0088 hypothetical protein Thermotoga maritima MSB8 1E-13 61 1d11.rev 500 NP_979594.1 DnaD domain protein Bacillus cereus 6E-18 52 Thermotoga sp. SG1 1E-72 98 1e05.for CAD89233.1 putative homing endonuclease Thermotoga sp. SG1 The first 188 bp have no significant hit in Genbank. The sequence from the forward primer shows 84% DNA identity to TM0322. Second hit: hypothetical protein TM0189. This fosmid was fully sequenced. From group I intron described in (NESBØ and DOOLITTLE 2003a). 1e07.rev 149 TM0162 hypothetical protein Thermotoga maritima MSB8 2E-05 65 1f08.rev 292 L42784 sugar hydrolase (yeeB) Lactococcus lactis 0.047 57 The sequence from the forward primer show 77% DNA identity to TM1042. 1g03.rev 437 ZP_00119049.1 hypothetical protein Cytophaga hutchinsonii 3E-07 29 The sequence from the forward primer show 72% DNA identity to TM1379. 1g07.rev 430 NP_173645.2| expressed protein 0.001 25 The sequence from the forward primer show 78% DNA identity to TM0643. Thermotoga sp. strain RQ7 1h02.rev, Arabidopsis thaliana Library: RQ7F1 169 no significant hit The sequence from the forward primer show 65% DNA identity to TM1831. 1h05.rev 1c06.rev, 749 ZP_00186152.1Ribose / xylose / arabinose / galactoside ABC-type transport systems, permease components Rubrobacter xylanophilus 4E-14 58 The sequence from the forward primer show 81% DNA identity to TM0310. 1d07.for 720 TM1786 hypothetical protein Thermotoga maritima MSB8 3E-05 57 The sequence from the reverse primer show 61% DNA identity to TM1573. 1g07.for 763 NP_070580.1 carbohydrate kinase, FGGY family Archaeoglobus fulgidus 2E-09 26 1g11.rev 117 TM1430 glycerol kinase Thermotoga maritima MSB8 1E-13 89 2e04.for 181 BH1487 sulfate adenylyltransferase Bacillus halodurans 5E-10 67 2e06.for 198 MA0407 predicted protein Methanosarcina acetivorans 5E-04 34 2f04.for 230 aq_372 putative protein Aquifex aeolicus 0.064 34 1c08.rev The sequence from the reverse primer show 82% DNA identity to TM1062. Thermotoga sp. strain KOL6 Library TsKOL6f1 1a03.for 247 TM1359 sensor histidine kinase Thermotoga maritima MSB8 1E-24 64 1a05.for 156 TM0806 hypothetical protein Thermotoga maritima MSB8 6E-05 45 1c02.for 86 no significant hit 1e05.for 201 no significant hit 1e05.rev 146 TM0754 oxidoreductase - Thermotoga maritima MSB8 0.2 70 1f04.for 143 NP_535223.1 ABC transporter, nucleotide binding/ATPase protein [sugar] Agrobacterium tumefaciens 0.007 41 1f05.for 210 NP_938431.1 Putative ABC transportCorynebacterium protein, membrane component diphtheriae 2E-06 40 The sequence from the reverse primer shows 75% DNA identity to TM0448. 1f07.rev 235 TM1393 conserved hypothetical protein Thermotoga maritima MSB8 1E-36 67 The sequence from the reverse primer shows 79% DNA identity to TM1363. 1g04.for 163 TM0366 endonuclease III Thermotoga maritima MSB8 4E-21 87 1g06.for 424 TM1799 conserved hypothetical protein Thermotoga maritima MSB8 2E-30 47 The sequence from the reverse primer shows 76% DNA identity to TM0030. a If more than one gene was covered by the clone, they are given as follows: a: gene X, b: gene Y, c: gene Z. b Percent identity refers to protein identity if not otherwise stated. Supplemental Table 3. Thermotoga petrophila RKU1 specific sequences with significant protein match in TIGR and/or NCBI protein databases. clonea bpb first BLASTX hitc organism exp. % identd note Transporters 227 a: TM1404 antibiotic ABC Thermotoga maritima transporter, transmembrane protein MSB8 249 b: no hit 1.2E-18 94 DNA 55.2f02 217 TM1067 oligopeptide ABC transporter, periplasmic oligopeptide-binding protein Thermotoga maritima MSB8 7.8E-09 43 55.2h02.2 185 TM0432 sugar ABC transporter, Thermotoga maritima periplasmic sugar-binding protein, MSB8 putative 2.0E-10 55 55.2.22g04 737 TM0432 sugar ABC transporter, Thermotoga maritima periplasmic sugar-binding protein, MSB8 putative 55.2.22g12 5.2E-71 62 63.3d08 290 TM0057, oligopeptide ABC Thermotoga maritima transporter, ATP-binding protein MSB8 2.0E-06 66 55.3d04 520 lin0218, similar to sugar ABC transporters, permease proteins 236 7.0E-09 43 Shows 92% DNA identity to RQ2.TAD15. Encoded on maga plasmid pSymB in Sinorhizobium meliloti. 55.2e04 55.2a04 AluI.2g11 Listeria innocua AluI.2b08 295 SMb20894,probable sugar uptake Sinorhizobium meliloti ABC transporter ATP-binding protein gguA 4.7E-26 58 AluI.2g08 291 TTE0205, Ribose /xylose Thermoanaerobacter /arabinose /galactoside ABC-type tengcongensis transport systems, permease components 5.2E-17 55 Two different overlapping sequences, probably duplication in RKU-1. Show patchy similarity to RQ2.TAC57, which shows 70% DNA identity to TM0432. Degradation of polysaccharides 55.2f04 533 BH1878, unknown conserved protein Bacillus halodurans 6.7E-10 49 Shows 90% DNA identity to bp 3771 – 4311 of the Thermotoga sp. strain RQ2 ‘arabinosidase island’. 206 BH1878~unknown conserved protein 596 Bacillus halodurans 1.0E-E Assembles into one 1071 bp contig, “ORF c” is rearranged compared to BH1878 as seen Thermotoga sp. strain RQ2, and the contig shows 94% DNA identity to bp 4303 – 5373 of the Thermotoga strain RQ2 ‘arabinosidase island’. 63.3a09 55.2a02 AluI.1e10 AluI.1c05 55.2g11 263 63.2g02 525 63.2g11 63.2h11 63.2a01a 39 AluI.1f08 506 Mdeg3957, hypothetical protein Microbulbifer degradans 3.0E-16 36 Second Blast hit: BH1878~unknown conserved protein. Shows 86 % DNA identity to bp 1434 – 1944 of the Thermotoga strain RQ2 ‘arabinosidase island’. 151 BS03926, similar to arabinan endo-1,5-alpha-L-arabinosidase Bacillus subtilis 9.1E-12 67 Second blastX hit: BH1878~unknown conserved protein, and shows 90% DNA identity to bp 2151 – 2299 of the Thermotoga strain RQ2 ‘arabinosidase island’. 63.2e08 400 BS03926, similar to arabinan endo-1,5-alpha-L-arabinosidase Bacillus subtilis 1.7E-19 65 DNA Second blastX hit: BH1878~unknown conserved protein. The 5’ end shows 86% DNA identity to bp 334-400 of the Thermotoga strain RQ2 ‘arabinosidase island’ 63.3g09 337 BH1878~unknown conserved protein Bacillus halodurans 6.0E-26 46 Shows 87% DNA identity to bp 585 – 921 of the Thermotoga strain RQ2 ‘arabinosidase island’. AluI.2c04 507 CAA52276, bglA gene for betaglucosidase Thermotoga maritima 0 55.2g02 648 SCO0488, putative hydrolase Streptomyces coelicolor 2.0E-33 34 AluI.1b11 63.3a07 55.22b12 55.22e04 55.22e12 55.5f11 96 DNA 63.2f02 63.2c08 Shows 98% DNA identity to RQ2.TAF85 and E6. Second blastX hit: rhamnosidase A Bacillus sp. GL1, 34% protein identity, exp. 9.0E-31. Probably part of the same gene as clone AluI.1d01. 55.3b08 55.3h08 795 AluI.1d01 458 BAB62314, rhamnosidase A Bacillus sp. GL1 1.0E-09 29 AluI.1c04 551 a: CAC3436, probable alphaarabinofuranosidase Clostridium acetobutylicum 2.0E-11 50 b: YPO2475, sugar ABC transporter, permease protein 0.006 Shows 99% DNA identity to RQ2.TAD18 and 98% DNA identity to RQ2.TAE82 53 Yersinia pestis 55.4f12 557 a: Chte1665 hypothetical protein Clostridium thermocellum 2.0E-21 31 AluI.1b07 720 b: TM0281 alpha-Larabinofuranosidase Thermotoga maritima MSB8 3.3 AluI.1d08 454 asdI, arabinosidase Bacteroides ovatus 4.0E-40 50 AluI.1f09 269 xynY, endo-1,4-beta-xylanase Clostridium thermocellum 2.0E-17 51 AluI.2f08 498 AF113969_1, pullulanase Thermococcus hydrothermalis 3.1E-04 32 Escherichia coli 1.3E-14 41 68 Surface polysaccharide biosynthesis AluI.2g09 329 AF172324_8, WbnE putative Assembles into one 1070bp contig. Shows 97% DNA identity to RQ2.KJB3 and 100% DNA identity to RQ.TXX2. glycosyl transferase Other AluI.1b05 85 63.3c10 55.2a08 TM0037 conserved hypothetical protein Thermotoga maritima MSB8 0.002 68 153 TM0638, hypothetical protein Thermotoga maritima MSB8 4.0E-13 76 240 TM0643, clostripain-related protein Thermotoga maritima MSB8 0.00042 48 55.2e08 55.2e12 55.2a11 774 a: TM1322 conserved hypothetical Thermotoga maritima protein MSB8 495 b: TM1323 hypothetical protein Thermotoga maritima MSB8 c: NP_391617; similar to hypothetical proteins Bacillus subtilis 3.0E-41 97 DNA 658 a: ald, alanine dehydrogenase Chlorobium tepidum 4.2 Thermotoga maritima MSB8 5.7E-58 90 DNA 270 AJ458646, Thermotoga sp. RQ2 subtraction clone TAD55 Thermotoga sp. RQ2 1.0E-17 86 DNA No significant hit in Genbank 63.2e06 259 AJ458663 and AJ458664, Thermotoga sp. RQ2 subtraction clone 2C1 and TAA14 Thermotoga sp. RQ2 9.0E-83 92 DNA No significant hit in Genbank 55.3d12.2 369 AJ458593 Thermotoga sp. RQ2 Thermotoga sp. RQ2 subtraction clone A5 and TAD45 1.0E-27 91 DNA No significant protein hit in Genbank, but RQ2.A5 and TAD45 shows similarity to TM0037 (NESBØ et al. 2002) 63.2h04 295 lmo1036, hypothetical protein (imported) Listeria monocytogenes 9.0E-07 41 55.2d12 276 YesW, conserved hypothetical protein Bacillus subtilis 4.0E-27 63 55.2g04 867 Q56216, 2-isopropylmalate synthase Thermus thermophilus 0.096 31 55.2h04 524 all4298, hypothetical protein Nostoc sp. PCC 7120 0.007 25 63.2d01 895 Chte0090, hypothetical protein Clostridium thermocellum 4.0E-04 29 63.2g09 AluI.2b06 AluI.1f11 b: TM1010, hypothetical protein 63.3h07 3.9E-09 83 DNA 1.0E-07 28 The clones assembles into a 904 bp contig. A sequence with similarity to TM1322 was also found in the Thermotoga sp. strain RQ2 library. Bp 1-97 95% identity to RQ2.TAF49. 27 55.2c05 63.3b05 55.3b10 55.3b12 573 a: Ddes2252, hypothetical protein Desulfovibrio 2.0E-06 37 desulfuricans b: FN0123, ATPase 0.002 44 Fusobacterium nucleatum 55.4c02 460 XCC1959, conserved hypothetical Xanthomonas campestris 0.054 45 Bp 38 – 417 shows 88% DNA identity to bp 48 – 522 of clone 55.2h04. Probably a duplication. protein 63.3e12 356 VCA0920, hypothetical protein Vibrio cholerae 0.002 34 63.3f02 526 a: lmo2712, highly similar to gluconate kinase Listeria monocytogenes 8.0E-4 29 Thermotoga maritima MSB8 6.0E-82 91 DNA b: TM0298, alcohol dehydrogenase, zinc-containing 63.3h02 222 NMB0507, hypothetical protein Neisseria meningitidis MC58 0.0029 64 DNA AluI.1c09 384 NTL02MP0372, lipoprotein Mycoplasma pulmonis UAB CTIP 2.2e-05 60 DNA Shows 89% DNA identity to RQ2. TAD30 No significant hit with blastX AluI.1g06 AluI.1h06 643 NTL01RP00104, unknown protein Rickettsia prowazekii 2.0E-06 57 DNA AluI.2h08 377 S77610, probable intercellular adhesion protein C Staphylococcus epidermidis 4.0E-02 40 AluI.1a05 683 a: ebiP1382 Anopheles gambiae str.PEST 0.029 25 0.029 45 Aquifex aeolicus 0.002 31 AluI.1c12 AluI.1d03 b: Desu1271, hypothetical protein First blastX hit: granule lattice protein 1 precursor , Tetrahymenathermophila, exp. 0.045, 32% ident. Desulfitobacterium hafniense 55.4b11 345 aq_373, hypothetical protein AluI.2d02 420 AluI.1e11 469 aq_035 , conserved hypothetical protein Aquifex aeolicus 8.0E-19 38 Second blastX hit: TM0972 Thermotoga maritima MSB8. Shows 99% DNA identity to RQ2.TAD44 AluI.2e12 258 aq_371, hypothetical protein Aquifex aeolicus 2.0E-09 51 Also in Thermotoga neapolitana NS-E (clone AluI.2d06). The clones have a 116 bp overlap and show 81% DNA identity. AluI.1e12 387 BB0701, hypothetical protein Borrelia burgdorferi 5.7E-05 57 DNA No significant hit with blastX AluI.1f04 186 long-chain primary alcohol dehydrogenase Thermoanaerobacter ethanolicus 6e-18 Shows 99% DNA identity to bp 3934-4118 of the Thermotoga strain RQ2 locus containing this adh gene (NESBØ and DOOLITTLE 2003b). 55.4b03 388 Chlo0556, hypothetical protein Chloroflexus aurantiacus 8.0E-28 49 Shows 99% DNA identity to bp 2144 – 2530 of the Thermotoga strain RQ2 locus containing MutS-3 homologs (NESBØ and DOOLITTLE 2003b). AluI.1c11 569 MutS-like ATPase involved in mismatch repair Thermoanaerobacter tengcongensis 9.0E-19 42 Shows 98% DNA identity to bp 1589 – 2153 of the Thermotoga strain RQ2 locus containing MutS-3 homologs. 467 MutS-like ATPase involved in mismatch repair Thermoanaerobacter tengcongensis 1.0E-41 53 Shows 99% DNA identity to bp 2665 – 3130 of the Thermotoga strain RQ2 locus containing 93 AluI.1f12 Assembles into one 557 bp contig. AluI.2e10 AluI.1f07 MutS-3 homologs. AluI.1c06 591 a: TM1001, conserved hypothetical protein Thermotoga maritima MSB8 7.9E-12 76 b: MA2365, predicted protein 0.002 24 Methanosarcina acetivorans 55.2b07a 432 a: TM0562, hypothetical protein 63.2a01b b: hypothetical protein (multidomain) Thermotoga maritima MSB8 4.5E-21 92 DNA 1.0E-36 61 AluI.1d10 261 Methanosarcina acetivorans 55.2g08 283 PH0175, hypothetical protein Pyrococcus horikoshii 2.0E-16 46 55.2b12 595 a: TM0136, conserved hypothetical protein Thermotoga maritima MSB8 1.7E-47 94 DNA 55.2g09 b: SSO3132, conserved hypothetical protein 1.4E-13 45 Sulfolobus solfataricus 63.2d05 354 MA4352, hypothetical protein (multi-domain) Methanosarcina acetivorans 2.0E-42 68 55.5c12 AluI.1h10 AluI.2a04 489 MK1259, MK0317, predicted Methanopyrus kandleri DNA methylase containing a Zn513 ribbon module 0.003 21 AluI.2e09 347 NP_558471.1, hypothetical protein Pyrobaculum aerophilum 5.7E-18 46 AluI.2g03 Methanosarcina mazei AluI.2h09 300 MM1048, D-alanine-D-alanine ligase related protein 172 2f10 457 a: 23S rRNA Thermotoga maritima MSB8 2.4E-10 28 1.8E-25 66 3.0E-49 98 DNA 3.0E-11 49 Pterosperma cristatum 309 a: NP_473343.1, hypothetical protein The clones do not overlap. 1.9E-07 50 b: AAL34315, putative protein AluI.2c07 The clones do not overlap. Plasmodium falciparum Thermotoga maritima b: TM1017 conserved hypothetical MSB8 protein 0.021 29 1.5E-08 75 DNA AluI.2e01 325 NP_504679.1, Putative protein Caenorhabditis elegans family member, with 2 coiled coil4 domains, of ancient origin 6.6E-03 36 55.3f09 598 a: TM0428 oxidoreductase, putative Thermotoga maritima MSB8 1.3E-74 94 DNA Thermotoga maritima MSB8 1.6E-19 90 DNA A putative site-specific DNA endonuclease from a group I intron (NESBØ and DOOLITTLE 2003a). The similarity to the Plasmodium falciparum covers the whole clone. b: no significant hit AluI.1a11 475 a: TM1005 transcriptional regulator, putative b: no significant hit a The clones were obtained using RsaI restricted Thermotoga petrophila RKU1 DNA except for clones labelled AluI.xxxx where AluI restricted DNA was used. For the RsaI two different hybridization temperatures were used - 63 C and 55 C – as indicated by the clone names 63.xxxx or 55. xxxx. For the AluI hybridizations we used 55C as hybridization temperature. b For identical clones, the length is given for the first clone listed. c If more than one gene was covered by the clone, they are given as follows: a: gene X, b: gene Y, c: gene Z. d Percent identity refers to protein identity if not otherwise stated. Supplemental Table 4. Recombination analyes of individual ORFs: Informative sites index (ISI), ratio of synonymous and non synonymous mutations and summary of phylogenetic analyses. ORF a ISI, pb Ds/Dnc Phylogenetic analysisd AU / SH -test if applicablee Comments TM0162, 0.54, p = 0.14 3.47 – 26.18 TM in clade with RKU1 and RKU10, but with no support. Split decomposition shows reticulate evolution between RQ2, RKU10 and RKU1. p (rRNA tree) = 0.01 / 0.04 As rRNA tree up to position 861 in alignment. Recombination TM-RKU10: 943 - 952, and RKU1-RQ2: 993 and extending into TM0163, RKU1 donor. TM0163, 837 bp 0.40, p = 0.19 3.0 – 6.4 RQ2 clusters with RKU1 and RKU10 with 86% bootstrap support. Split decomposition shows reticulate evolution between RQ2, TM, RKU10 and RKU1. p (rRNA tree) = 0.03 / 0.18 RQ2 - RKU1 recombination from TM0162 up to position 500, RKU1 donor. TM0164, 1.48, p = 0.08 5.1 – 10.7 RQ2 clusters with RKU1 and RKU10 with 66% bootstrap support. Split decomposition shows reticulate evolution between RQ2, TM, RKU10 and RKU1. p (rRNA tree) = 0.36 / 0.35 In the third position tree used in the ISI test had same topology as the rRNA tree. Recombination RQ2- RKU1/RKU10: 1 – 450, RKU1/RKU10 lineage donor. 0.64, p = 0.35 4.6 - 17.4 RQ2 clusters with RKU1 and RKU10 with 72% bootstrap support. Split decomposition shows reticulate evolution between RQ2, TM, RKU10 and RKU1. p (rRNA tree) < 0.001 / 0.01 Recombination RQ2 - RKU1: 67 – 202 and 380 – into TM0166, RQ2 likely donor in first and up to position 469 in second recombination event. -1.56 p = 0.97 6.3 – 17.6 RQ2 and RKU1 have identical sequences and cluster together with RKU10 with 81% bootstrap support. TM, RKU10 and RQ2-RKU1 form a star in split decomposition graph. p (rRNA tree) = 0.002 / 0.01 RQ2 and RKU1 identical. RKU1 lineage donor 1 - 421 and 748 – into TM0167, RQ2 lineage donor 463 – 720. Recombination RKU10 – TM: 385 – 421 and 1198- 1293. 0.85, p = 0.19 8.5 – 76.6 RQ2 clusters with RKU1 and RKU10. In bootstrap tree RKU1, RKU10 and TM together with 58% support. Split decomposition shows reticulate evolution between RQ2, TM, RKU10 and RKU1. p (rRNA tree) = 0.18 / 0.18 In the third position tree used in the ISI test TM, RQ2 and RKU10 cluster together. RQ2 and RKU1 identical 1-205 (recombination extending from TM0166). Recombination RQ2/TM – RKU1: 310 – 673, RQ/TM donor, RQ2- RKU1/RKU10: 862 and into TM0168 recombination, RKU1/RKU10 donor lineage. -2.7, p = 1.00 13.8 – 65.2 TM clusters with RKU1 and RKU10, but no bootstrap support. Split decomposition shows reticulate evolution between RQ2, TM, RKU10 and RKU1. p (rRNA tree) = 0.38 / 0.38 In the third position tree used in the ISI test TM clusters with RKU1 and RKU10 Recombination RQ2- RKU1/RKU10: 105 – 399, RKU1/RKU10 donor, TM RKU1/RKU10: 1293 – 1419, RKU1/RKU10 donor, RQ2 – RKU1: 1433 into TM0169 recombination, RQ2 donor. 0.0, p= 1.00 14.9 – 53.9 RKU10 clusters with RQ2 and TM with 67% support. Split decomposition shows reticulate evolution between RQ2, TM, RKU10 and RKU1. p (rRNA tree) = 0.17 / 0.19 Recombination RQ2 - RKU1: from TM0168 – position 201, RQ2 donor, RQ2 - NEAP 374 – 388, NEAP donor, RKU10 – TM/RQ2: 453 – 520, TM/RQ2 donor. TM0170, 218 bp 2.74, p = 0.99 38.8 – 80.0 As rRNA tree. Split decomposition shows reticulate evolution between RQ2, TM, RKU10 and RKU1. TM0171, 0.17, p = 0.38 4.2 - 16.7 RKU10 and RKU1 paraphyletic; (RKU10 (RKU1 (TM,RQ2))). Only support (52%) for RQ2-TM clade. Split decomposition shows reticulate evolution between RQ2, TM, RKU10 and RKU1. p (rRNA tree) = 0.33 / 0.32 Recombination RQ2 – RKU1: 107 – 183, RQ2 likely donor. Recombination TM – RKU10: 303 into TM0172, TM donor. -0.01, p = 0.51 5.7 - 30.0 TM and RKU10 are sisters and cluster together with RQ2 with 88% support. SG1 sister to LA4 and LA10 to the exclusion of RQ7 with 63% support. Split decomposition shows reticulate evolution between RQ2, TM, RKU10 and RKU1. p (rRNA tree) = 0.031 / 0.036 Recombination TM/RQ2 – RKU10 extending the whole ORF. -0.03, p = 0.63 7.1 – 41.8 RQ2 and RKU1 are sister taxa in the tree (100% support) and cluster together with TM (90% support). Split decomposition shows reticulate evolution between RQ2, TM, and RKU1. p (rRNA tree) = 0.015 / 0.025 Recombination RQ2 – RKU10: from TM0172 – 444, RQ2 donor. Recombination RQ2/TM – RKU1: 836 into TM0174, RQ2/TM donor. TM0174, 2169 bp 0.20, p = 0.09 31.6 – 122.1 TM, RQ2 and RKU1 cluster together to the exclusion of RKU10 with 89% support. Split decomposition shows reticulate evolution between RQ2, TM, RKU1 and RKU10. p (rRNA tree) = 0.062 / 0.070 Recombination TM/RQ2 - RKU1: from TM0173 – 1803, TM/RQ2 donor. TM0175, 0.24, p = 0.30 6.8 – 29.3 As rRNA tree. Split decomposition shows reticulate evolution between RQ2, TM, RKU1 and RKU10 as well as in the center of the tree. Recombination TM – RKU1/RKU10: 172 – 220, RKU1/RKU10 donor. 0.03, p = 0.43 23.0 - 27.5 As rRNA tree. Recombination TM - ?, or RQ2 – RKU1/RKU10 297 – 327. -0.23, p = 0.79 15.7 – 113.2 As rRNA tree. Split decomposition shows reticulate evolution between RQ2, TM, RKU1 and RKU10. RKU1 and RQ2 identical from position 576 in alignment – recombination extends into TM0178, RQ2 donor. -0.15, p = 0.81 7.5 – 17.6 RQ2 and RKU1 have identical sequences and cluster together with TM in phylogenetic trees (see Fig.1C). 2134 795 bp TM0165*, 573 bp TM0166*, 1293 TM0167*, 1170 TM0168, 2472 bp TM0169, 624 bp 465 bp TM0172, 1212 bp TM0173, 3315 bp 270 bp TM0176, Recombination TM/RQ2 – RKU10: 57 – 117, TM/RQ2 donor. 879 bp TM0177, 849 bp TM0178, 2202 16S p (rRNA tree) = 0.001 / 0.011 Recombination RQ2 – RKU1 extends the whole gene and into 16S, RQ2 donor. Recombination RQ2 – RKU1 from TM0178 – 217 (identical to RQ2) or 1043 (starts being most similar to RKU10), RQ2 donor and possible unknown donor. 23S Recombination RKU10 - ? or RKU1 – TM/RQ2: 24441 - end of the sequence. 5S TM0179, 0.30, p = 0.19 26.1 – 88.2 RKU1 and TM cluster together to the exclusion of RQ2, but with no bootstrap support. Split decomposition shows reticulate evolution between RQ2, TM, RKU1 and RKU10 as well as in the centre of the tree. -0.98, p = 0.94 0.49 – 9.5 As rRNA tree. The splits graph shows reticulate evolution. An alignment of variable in TM, RQ2 and RKU1 is shown in Fig. 4. Recombination RQ2 –RKU1: 1 – 180 and 298 – 470. Ds/dn values < 0 were observed in comparisons involving RQ7 and the two other neapolitana strains and was due to 4 mutations resulting in 2 aa replacements, probably due to recombination. -0.04, p = 0.58 12.3 – 53.3 As rRNA tree. RQ2, TM and RKU1 very similar in sequence: 1.5 – 2.9% divergence. Recombination RQ2 –RKU1: 243 – probably into TM0182, RKU1 donor. 0.08, p = 0.31 4.4 – 24.0 TM and RKU1 cluster together to the exclusion of RQ2 with 56% bootstrap support. Split decomposition indicates recombination between RQ2, TM and RKU1. -0.06, p = 0.58 9.9 –21.9 As rRNA tree. -0.60, p = 0.94 9.1 –19.1 As rRNA tree. 0.15, p = 0.37 7.7 – 27.0 As rRNA tree. Split decomposition show reticulate phylogeny, indicating recombination between RQ7 and SG1 and between RKU1 and TM. 0.24, p = 0.18 9.9 – 16.7 As rRNA tree. Split decomposition show reticulate phylogeny, indicating recombination between RKU1 and TM. -0.13, p =0.65 6.6 –16.3 As rRNA tree. -0.08, p = 0.56 16.1 – 99.2 As rRNA tree. Split decomposition show reticulate phylogeny. 0.06, p = 0.45 6.7 – 22.0 As rRNA tree. In SG1 this is a pseudo gene, however the full sequences corresponding to the complete ORF was included in the analysis TM0190, 1032 bp -0.17, p = 0.66 8.3 – 9.2 As rRNA tree. . TM0191, -1.21, p = 0.94 7.6 – 12.9 As rRNA tree. 0.13, p = 0.43 146 – 421 As rRNA tree. -0.15, p = 0.69 10.2 – 30.3 As rRNA tree. -0.23, p = 0.65 9.7 – 10.7 As rRNA tree 276 bp TM0180*, 570 bp TM0181, p (rRNA tree) = 0.17 / 0.18 972 bp TM0182, 827 bp TM0183, p (rRNA tree) = 0.05 / 0.08 Recombination TM – RKU1: 189 – 438, RQ2 –RKU1: 486 – end of ORF, RKU1 donor in both. 801 bp TM0184, 1281 bp TM0185, 339 bp TM0186, 1104 bp TM0187, 1183 bp TM0188 429 bp TM0189, 1038 bp 786 bp TM0192, 261 bp TM0193, 786 bp TM0194, 435 bp a Number of sites in the alignment is given for each ORF. A * indicates that the positions outlined in the comments refer to the reverse alignment. b The ISI test was done on the phylogenetic tree estimated from the ORF. For TM0161, TM0162, TM0164, TM0165, TM0166, TM0169, TM0171, TM0172, TM0173, TM0174, TM0178, TM0179, TM0182 the phylogeny used to do the ISI test was the recombinant tree. ISI is the informative sites index and is a measurement of the degree in which recombination has shaped the data and p is the significance of the recombination. c In several comparisons the ds/dn value could not be calculated, as either the ds or the dn value was 0 or because of mutational saturation. This is not shown. d Results from Splits decomposition analyses are described if they indicated reticulate evolution. TM: T. maritima MSB8, RQ2: Thermotoga sp strain RQ2, RKU1: T. petrophila RKU1, RKU10: T. naphthophila RKU10, SG1: Thermotoga strain SG1, RQ7: Thermotoga sp strain RQ7, NEAP: T. neapolitana lineage. e The approximately unbiased test. Supplemental Table 5. Recombination detected from concatenated alignments: Results from GENECONV, MAXCHI2 and RPD. Only events detected by 2 or more methods are listed. Gene fragment/ strains in alignment/ ISIa Strains involved b GENECONV (g=0) RPD 10 bp window, fragment, p-value fragment, p-value Maxchi 30 variable sites window, bpd ORFs, position Comment 132 -228 TM0148: 1126 - 1352 RKU10 donor 196 -1892 TM0148: 642 -1 RQ2 and TM show 99% identity. fragment, p-valuesc TM0146 – TM0162 TM, RQ2, RKU10 RQ2 – RKU10 1131 – 1263, 0.007 RQ2 - TM 1945 – 2141, 0.01 1131 – 1359, 0, 0.05 16478 bp 1842 – 3734, < 0.001 TM0149, TM0150: 183 -1 RKU10 – RQ2/? 7259 – 8182, < 0.001 6824 - 8331, < 0.001 8184 - 8489, < 0.001 RQ2 – RKU10 8719 – 8787, 0.001 6824 – 8331, 0, 0.04 305 - 1507 TM0153: 452 –1 7049 – 8295, 0, 0.008 TM0154, 7049 – 8331, 0, 0.008 TM0155: 1-946 8725 – 9065, 0, 0.03 68 - 340 TM0155: 649 - 994 9065 – 11454, 0, 0.01 228 - 4183 TM0155: 656 –1062, TM/RQ2 likely donor (RKU10 99% identity to TM). 8725 – 8952, 0.007 RQ2 – RKU10 8953 - 11415, < 0.001 8724 – 8952, < 0.001 9182 – 11377, 0, 0.007 RQ2 – RKU10 13802 – 15284, < 0.001 13857 - 15203< 0.001 13028 – 15513, 0, 0.03 13822 – 15513, < 0.001 13857 – 15203, 0, 0.03 13857 – 15504, 0.007 15294 – 16430, 0,0.04 502 - 2485 TM0156 – TM0160 RKU10 is almost identical to RQ2 and TM (98% identity to TM sequence), RQ donor. TM0161: 452 – TM/RQ2 likely donor. TM0162 13889 – 15203, < 0.001 15613 – 16478, 0.007 15882 – 16384, 0.002 TM0162 – 23S TM, RQ2, RKU1, RKU10, LA10, 26460 bp LA10 -? 45 – 450, < 0.001 1 – 527, 0, 0.1 TM0162: 179 - 1118 1 – 940, < 0.001 RKU10 – RQ2/? 1170 – 1446, < 0.001 1289 - 3460, < 0.001 Best ML tree: 2424 – 2553, 0.02 2032 – 4067, < 0.002 (LA10 (TM (RQ2 (RKU1, RKU10)))) 2188 - 3369, < 0.001 2733 – 2994, 0.002 AU – test: 2835 – 3115, < 0.001 p (rRNA tree) = 0.08 941 – 3358, 0, 0.03 2035 -3985 TM0162: 262 – TM0163, TM0164 TM0165: 576 - 205 Several recombinations between RKU1/ RKU10 and TM/RQ2 see Table 3. 3269 – 3369, < 0.001 ISI = 0.69, p = 0.001 RKU1 – TM/RQ2 506 – 858, 0.001 535 – 4537, < 0.001 1061 – 2031, < 0.001 941 – 2588, 0.03 276 - 4002 TM0162: 758 – TM0163 – TM0165, 1170 – 1446, < 0.001 Several recombinations between RKU1/ RKU10 and TM/RQ2, see Table 3. TM0166: - 1015 1529 – 2181, < 0.001 RKU1 – TM/RQ2 3380 – 3808, < 0.001 3422 – 4061, 0, 0.01 428 - 639 3499 – 4067, 0, 0.02 RQ2 – RKU10/LA10? 4068 - 5781, < 0.001 4121 – 5808, < 0.001 4088 – 5740, 0, 0.04 TM0164: 457 – RQ2 likely donor. TM0165: - 576 28 - 1713 TM0165: 202 – 4955 – 5319, < 0.001 4181 – 5764, 0, 0.04 TM0166 5354 – 5422, < 0.001 5059 – 5415, 0, 0.05 TM0167: - 948 Several recombinations between RKU1/ RKU10 and TM/RQ2, see Table 3. 5375 – 5403, < 0.001 RKU1 – TM/RQ2 6392 – 6432, 0.006 6189 – 8206, < 0.001 6184 – 8239, 0, 0.02 6416 – 6612, 0.01 6424 – 8524, < 0.001 6613 – 7930, 0, 0.02 6416 – 8112, < 0.001 40 - 2100 non-coding, RKU1/RKU10 likely donor. TM0168: 1- 1420 6421 – 7917, 0, 0.04 6421 – 6583, 0, 0.09 RQ2 – RKU10 6424 – 7420, 0.009 6421 – 7279, 0, 0.03 858 - 996 TM0167: 308 – RKU1/RKU10 likely donor TM0168: - 599 RKU1 – TM/? RKU1 – TM/RQ2/? RKU1 – RQ2 RKU1 – TM/RQ2 8240 – 8799, 0.03 7351 – 8206, 0.01 7666 – 9556, 0, 0.01 855 - 1890 TM0168: 531 -1945 8253 – 9556, 0.007 8239 – 9267, 0, 0.04 22 - 1303 TM0168: 1420 -2475 RQ2 likely donor. 85 - 1479 TM0169: 203 – Several recombinations between RKU1/ RKU10 (mostly RKU10) and TM/RQ2, see Table 3. 8801 – 9507, < 0.001 8253 – 9493, 0, 0.02 9187 – 9327, 0.01 8482 – 9505, 0, 0.03 9254 – 9276, 0.05 8320 – 9505, 0, 0.04 9506 – 9756, < 0.001 9508 – 10230, < 0.001 9557 – 9645, < 0.001 9508 – 9781, < 0.001 TM0170, TM0171, 9734 – 10074, < 0.001 9577 – 11056, < 0.001 TM0172: - 455 9980 – 10065, 0.03 9797 – 10172, 0.007 10445 – 10597, < 0.001 10538 – 11500, < 0.001 10449 – 11677, 0, 0.06 10599 – 10668, 0.02 10538 – 11011, < 0.001 10525 – 10687, 0, 0.04 10777 – 10904, 0.03 10985 – 11151, < 0.001 11003 – 11033, 0.03 11153 – 11274, < 0.001 9508 – 9739, 0, 0.12 30 - 1228 TM0172: 1- 1215 More likely recombination TM/RQ2 – RKU10, TM/RQ2 donor, see Table 3. 11222 – 11271, 0.02 11468 – 11538, 0.005 RQ2 – TM/RKU1 11294 – 12159, < 0.001 11113 – 12229, < 0.001 10420 – 12544, 0, 0.05 11362 – 12315, < 0.001 10538 – 12246, 0, 0.06 11906 – 12426, < 0.001 11314 – 12144, 0, 0.04 RKU1 – RKU10 12247 – 12548, < 0.001 RKU1 – TM/? 12736 – 13118, < 0.001 12699 – 14465, < 0.001 12992 – 13164, < 0.001 12857 – 14018, 0.02 14640 – 14675, 0.03 14243 – 16904, < 0.001 RKU1 – RQ2/? 14523 – 14745, 0.02 520 - 2124 TM0172: 1 – TM0173: - 623 More likely recombination TM/RQ2 – RKU10, TM/RQ2 donor, see Table 3. RKU1 and RKU10 identical. 12291 – 12544, 0, 0.06 253 -301 TM0173: 445 - 746 12540 – 13430, 0, 0.05 172 - 1766 TM0173: 690 - 2672 14621 – 1468, 0.016 35 - 2661 TM0173: 2799 – 14632 – 14691, 0, 0.18 TM0174: - 1753 14949 – 15220, 0.01 15729 – 15892, 0.004 RKU1 – RQ2/TM 16638 – 16903, 0.002 16940 – 19011, < 0.001 16904 – 18966, 0, 0.06 16638 – 16849, 0.002 16940 – 18966, < 0.001 16910 – 19011, 0, 0.06 16905 – 19067, < 0.001 16913 – 19011, < 0.001 16916 – 19141, 0, 0.22 211 - 2225 TM0174: 1538 – TM0177: - 649 Only one short recombination in TM0175 detected in phylogenetic analyses, Table 3. TM0177: 621 – RQ2 donor. 17943 – 18804, 0.002 RKU1 – TM/RQ2/? 19012 – 21990, < 0.001 19302 – 21693, < 0.001 19837 – 19997, 0.03 19721 – 21590, < 0.001 20046 – 22813, 0, 0.04 160 - 2978 TM0178 20723 – 20931, < 0.001 16S: - 215 20339 – 20679, < 0.001 20933 – 21237, 0.02 21068 – 21237, < 0.001 RKU1 – RQ2/LA10/? 21992 – 24562, < 0.001 22587 – 26459, < 0.001 21991 – 26418, 0, 0.03 2570 - 4427 16S 23S 22617 – 25205, < 0.001 22813 – 26459, 0.01 642 – 808, 0.01 838 – 921, 0.008 678 –830, 0, 0.12 14 - 546 TM0180: 570 -1 853 – 920, < 0.001 791 – 830, < 0.001 777 – 812, 0, 0.37 23S – TM0182 TM, RQ2, RKU10, KOL6, RQ7, LA10, LA4, SG1, TM - ? 2550 bp 872 – 886, 0.008 777 – 830, 0.30 777 - 894, 0, 0.24 Ml-tree used in PIST analysis: 777 – 908, 0, 0.24 (((LA10, LA4, RQ7) SG1) (KOL6 (TM (RQ2 RKU1)))) 777 – 1234, 0, 0.31 ISI = 0.07, p = 0.22 777 – 1323, 0, 0.35 May be artefact of higher conservation of rRNA genes. 854 – 932, 0.027 Tree including all sites and 5S has same topology as rRNA tree. TM0172 - TM0182 RKU1 – ? 872 – 886, 0.001 838 – 920, < 0.001 870 – 933, 0.030 14 - 90 TM0180: 362 - 279 877 – 894, 0.02 842 – 932, 0, 0.28 TM - ? 1081 – 1100, 0.02 1083 – 1134, 0, 0.13 19 -51 TM0180: 131 -78 RQ2 – TM/RKU1 2010 – 2501, < 0.001 1925 – 2978, < 0.001 1925 – 2813, 0, 0.05 145 - 1803 TM0173: 738 - 1753 8497 – 11074, < 0.001 8392 – 11320, 0, 0.02 208 - 2975 TM0177: 519 – TM, RQ2, RKU1, LA10, RQ7, SG1, KOL6 2121 2501, 0.001 2688 – 2833, 0.03 Best ML tree: (TM,((RQ2,RKU1),((SG1,(LA10,R Q7),KOL6))) RKU1 – RQ2/? 8393 – 11368, < 0.001 9721 – 10061, < 0.001 8458 – 11369, 0, 0.02 TM0178, 10105 – 10313, 0.03 16S: - 216 10315 – 10619, 0.001 AU – test: RQ2 - ? p (rRNA tree) = 0.41 18883 bp TM – RQ2/SG1/RKU1? ISI = -0.02, p = 0.66 11370 – 13940, 0.002 11369 – 16478, 0, 0.01 11631 – 12606, 0.008 11966 – 16478, 0, 0.02 16479 – 16645, 0.01 16628 – 16667, < 0.001 16331 – 16602, 0.017 16519 – 16523, 0.02 16615 – 16649, < 0.001 16483 – 16664, 0, 0.11 16636 – 16647, 0.02 975 - 5109 16S: 216 – 23S: - 3023 4 - 271 TM0179: 150 – TM0180: - 388 16614 – 16649, 0, 0.38 16614 – 16667, 0, 0.32 RKU1 – RQ2/RQ7/LA10? 16519 – 16523, 0.03 16515 – 16667, 0, 0.12 16690 – 16744, 0.003 16614 – 16731, 0, 0.02 16709 – 16723, 0.005 16614 – 16883, 0, 0.02 16714 – 16731, 0.05 16614 – 16885, 0, 0.31 4 - 271 TM0180: 540 - 166 16679 – 16769, 0, 0.30 16707 – 16770, 0, 0.02 SG1 – TM/? 16918 – 16937, 0.01 16756 – 17069, 0, 0.02 19 - 313 TM0180: 288 -1 TM – RKU1 18029 – 18286, 0.04 17458 – 18727, 0, 0.02 257 - 1269 TM0181: 336 – TM0182: - 662 Likely an artefact of high identity of sequences. TM0172 –TM0188 TM,RKU1, RQ7, SG1, KOL6 SG1 – TM/RKU1/? 10291 - 10595, < 0.001 10249 – 10559, 0.01 - 304 -310 TM0178: 1555 -1901 ISI = -0.05, p = 0.80 RQ7 – TM/RKU1/? 10291 – 10595, < 0.001 10273 – 10559, < 0.001 - 286 -304 TM0178: 1555 -1901 The same recombination as SG1 –TM/RKU1/ TM – RKU1/? 11610 – 12585, 0.02 11600 – 12629, < 0.001 - 975 -1029 16S: 480 - 1499 May be an artefact due to the high similarity of the rRNA genes 16569 – 16847, 0, 0 17 - 278 TM0180: 441 - 102 21870 - 21978, 0, 0.2 25 - 108 TM0186: 743-851 12 - 33 TM0184: 345 -379 11766 – 12629, 0.003 RKU1 – TM/? 16664 – 16678, 0.003 16583 – 16619, < 0.001 16669 – 16686, 0.03 16659 – 16712, < 0.001 16679 – 16712, 0.01 16795 – 16902, 0.01 RQ7 – SG1/? 21868 – 21893, 0.03 21873 – 21942, 0.006 TM – RKU1/? 2311 – 2323, 0.05 2313 – 2331, 0.006 TM0182 – TM0194 TM, RKU1, RQ7, SG1 2298 – 2331, < 0.001 ISI = -0.66, p = 1.0 RQ7 – SG1/ ? 3849 – 3874, 0.006 3854 – 3923, 0.001 3848 – 3905, 0, 0.21 25 - 69 TM0186: 854 - 779 RKU1 – ? 5744 – 5751, 0.03 5740 – 5753, 0.03 5730 – 5751, 0, 0.40 7 - 23 TM0187: 371 - 348 a Genes included in alignment. ISI test were done when more than 3 sequences were available by concatenating 3 position alignments. The approximately unbiased (AU) test was done where the fosmid tree disagreed with the rRNA tree. The alignments can be obtained from C. L. N. b TM = Thermotoga maritima MSB8, RQ2 = Thermotoga sp. strain RQ2, TPE = Thermotoga petrophila, TNAPH = Thermotoga naphtophila. e The Maxchi test gives two p-values, one for each predicted break point. d The range of the length of the fragments suggested to have been exchanged. BOGUSH, M. L., T. V. VELIKODVORSKAYA, Y. B. LEBEDEV, L. G. NIKOLAEV, S. A. LUKYANOV et al., 1999 Identification and localization of differences between Escherichia coli and Salmonella typhimurium genomes by suppressive subtractive hybridization. Mol Gen Genet 262: 721-729. GEVERS, D., F. M. COHAN, J. G. LAWRENCE, B. G. SPRATT, T. COENYE et al., 2005 Opinion: Re-evaluating prokaryotic species. Nat Rev Microbiol 3: 733-739. NELSON, K. E., R. A. CLAYTON, S. R. GILL, M. L. GWINN, R. J. DODSON et al., 1999 Evidence for lateral gene transfer between Archaea and bacteria from genome sequence of Thermotoga maritima. Nature 399: 323-329. NESBØ, C. L., and W. F. DOOLITTLE, 2003a Active self-splicing group I introns in the 23S rRNA genes of hyperthermophilic bacteria, derived from introns in eukaryotic organelles. PNAS 100: 10806-10811. NESBØ, C. L., and W. F. DOOLITTLE, 2003b Targeting clusters of transferred genes in Thermotoga maritima. Environmental Microbiology 5: 1144-1154. NESBØ, C. L., K. E. NELSON and W. F. DOOLITTLE, 2002 Suppressive subtractive hybridization detects extensive genomic diversity in Thermotoga maritima. J. Bacteriol. 184: 4475-4488.
