Fig. S1. Schematic Representation of the Phylogenetic Relation Between the Thermococcales, the Methanococcales and Their Closest Archaeal Relatives

Fig. S1. Schematic representation of the phylogenetic relation between the Thermococcales, the Methanococcales and their closest archaeal relatives. In this schema the different genus of the Thermococcales and the Methanococcales were coloured in blue and green respectively Original organisation before MGE integration tRNA Arg TK_RS02830 TK_RS03040

TK_RS01875 TK_RS02080 tRNA Glu

~ 150kb

Integrations

Original organisation after the multiple MGE integration tRNA TK_RS01880 Arg TK_RS02830 TK_RS02070 TK_RS03040

TK_RS01875 TKVA TK_RS02840 TK_RS02080tRNA TKVB TK_RS03035 Glu

attLA attRA attLB attRB

Inversion

Organisation in the type strain genome tRNA TK_RS01880 TK_RS02070 TK_RS02830 Arg TK_RS03040

TK_RS01875 TKV2 tRNATK_RS02080 TK_RS02840 TKV3 TK_RS03035 Glu

320,075-122 347,135-87 499,291-338 526,817-69

attL2 attR2 attL3 attR3

Fig. S2. TKV2 and TKV3 inactivation by large DNA inversion in T. kodakarensis genome. 1 1 1 1 1 P P P I I P I 1 1 P I _ _ I _ P P _ 3 4 _ I I 6 1 6 2 1 1 9 1 5 _ _ 1 P 1 1 2 1 1 1 9 P 9 2 1 3 I P 0 1 8 - I I 1 P 1 P 7 0 P 7 P _ 0 E E 1 I 6 I P _ I I _ 0 1 i 1 2 4 4 _ 1 _ _ 1 0 _ 7 _ 1 1 2 _ 1 _ R 5 7 5 5 2 M 4 B 1 M M 2 5 1 S V V M V 1 A 2 l E E C H S H S M S C S A 7 H 6 7 V − o C X K O 1 1 F 1 3 2 F S G G C D C 6 D J N D N C C P C C a a a K V V E V V V p h a u a 2 e p u h p b a D g t i V M M i s m m M M m c G b K c g K s b k H s y l T M a T P T T T T T T T TPV1 TKV1 M p p Afu M M M M M M M M M M P P P P P Mva_IV1 Mok_IV1 MfeAG86_IV1 P Ta Tb Tc Td Te Tf Tg Th Ti Tj Tk Tl Tm Tn To Tp A C Ma B F Mb Mc Md Me Mf Mg Mh Mi Mj Mk Ml D E Ta PabGE2_IP1 Tb TGV1 Tc PHV1 Td PspNA2_IP1 Te PyaCH1_IP16 Tf PchGC74_IP1 Tg PkuNCB100_IP1 Th TbaCH5_E2 Ti TKV3 Tj TceDSM17994_IP1 Tk TguDSM11113_IP1 Tl pT26−2 Tm TKV2 Tn TspJCM11816_IP1 To PliDSM5473_IP1 Tp TbaCH5_E1 Ma MVV1 Mb MthDSM2095_IP1 Mc MigKol5_IP1 Md pMEFER01 Me MspFS406-22_IP1 Mf MmaKA1_IP1 Mg MmaOS7_IP1 Mh MMC7V2 Mi MMPV1 Mj MMC7V1 Mk MmaX1_iP1 Methanococcales virus 50 Ml MMC6V1 A TPV1 Methanococcales pT26-2 40 B Mva_IV1 C Afu Thermococcales pT26-2 30 D TKV1 E Mok_IV1 Thermococcales virus 20 F MfeAG86_IV1 Unknown 10

0 Number of BBH Fig. S3. Conservation of plasmids of the pT26-2and their connections with viruses by heatmap. Results of Bidirectional Best-Hit are represented as a heatmap. In the heatmap the number of conserved proteins is indicated in the scale. The heatmap analysis revealed two distinct groups, one containing the elements in Thermococcales and the other those of Methanococcales. The heatmap also show that some plasmid of the pT26-2 family shared genes with viruses. Protein group Core

in 2 to 4 pT26-2-related plasmid

Singleton

Other

Fig. S4. Size protein encoded by plasmids of the pT26-2 family. The protein set were divided in 4 different groups. For each group the density of protein is indicated at the different protein size. Fig. S5. Organisation of the pT26-2 putative replication protein and its comparison with other replication proteins. In this schematic representation the Primpol and the P-loop NTPase domain are indicated in green and blue respectively. KXA90124_candidate_divison_MSBL1_archaeon_SCGC_AAA259B11 98.6/100KXB04138_candidate_divison_MSBL1_archaeon_SCGC_AAA261G05 100/100 KUO41305_Hadesarchaea_archaeon_DG_33_1 96.1/99 KUO42083_Hadesarchaea_archaeon_YNP_N21 100/100KYC45172_Arc_I_group_archaeon_B03fssc0709_Meth_Bin005 96.4/100 KYC54749_Arc_I_group_archaeon_ADurb1013_Bin02101 KYK36871_1_Theionarchaea_archaeon_DG_70_1_no_comment 272844_MCM1_NP_127115_Pyrococcus_abyssi_GE5 65.1/82 98.6/10070601_MCM1_NP_142570_Pyrococcus_horikoshii_OT3 96.2/10096/100342949_MCM_shell_YP_004424138_Pyrococcus_sp_NA2 96.8/99186497_MCM1_NP_578211_Pyrococcus_furiosus_DSM_3638 529709_MCM1_YP_004623548_Pyrococcus_yayanosii_CH1 98.6/1001042877_MCM1_YP_004762586_Thermococcus_sp_4557 99.7/10090.8/100523850_MCM1_YP_002306424_Thermococcus_onnurineus_NA1 78/82 99.4/10069014_MCM1_YP_184033_Thermococcus_kodakarensis_KOD1 86.5/100246969_MCM1_YP_002582218_Thermococcus_sp_AM4 95.5/100593117_MCM1_YP_002960390_Thermococcus_gammatolerans_EJ3 100/100523849_MCM1_ZP_09730841_Thermococcus_litoralis_DSM_5473 73.4/99 604354_MCM1_YP_002994932_Thermococcus_sibiricus_MM_739 90.8/96 391623_MCM1_YP_004070279_Thermococcus_barophilus_MP Tbaro_E2_TBCH5v1_2333_ALM76228_1 97.1/100 100/100WP_011250312_TKV4_Thermococcus_kodakarensis_no_comment 30.3/90 WP_048151234_STIV_pTN3fam_Palaeococcus_ferrophilus_no_comment AEY69051_TPV1_Thermococcus_prieurii_virus_1_no_comment 92.4/88 98.7/100 WP_099209188_Thermococcus_sp_EXT12c_no_comment100/100 98.4/100 AGX15318_pTN3_plasmid_Thermococcus_nautili_no_comment 342949_MCM1_YP_004424245_PspNA2_MGE_Pyrococcus_sp_NA2 100/100 TKV1_TK0096_BAD84285_1 644295_MCM1_YP_003726351_Methanohalobium_evestigatum_Z_7303 12.9/76 224325_MCM1_NP_069353_Archaeoglobus_fulgidus_DSM_4304 99.5/100589924_MCM1_YP_003435419_Ferroglobus_placidus_DSM_10642 82.5/100693661_MCM1_YP_004340760_Archaeoglobus_veneficus_SNP6 98/99 AGK61979_Archaeoglobus_sulfaticallidus_PM70_1 572546_MCM1_YP_003400528_Archaeoglobus_profundus_DSM_5631 100/100 572546_MCM_shell_YP_003401418_Archaeoglobus_profundus_DSM_5631 99.9/100 572546_MCM_shell_YP_003400568_Archaeoglobus_profundus_DSM_5631 78.3/99188937_MCM_shell_NP_615641_Methanosarcina_acetivorans_C2A 100/100192952_MCM1_NP_633860_Methanosarcina_mazei_Go1 269797_MCM1_YP_305120_Methanosarcina_barkeri_str_Fusaro 55.8/74 679901_MCM1_YP_004615679_Methanosalsum_zhilinae_DSM_4017 28/97 97.1/100259564_MCM1_YP_567033_Methanococcoides_burtonii_DSM_6242 99.8/9488.3/97 547558_MCM1_YP_003541798_Methanohalophilus_mahii_DSM_5219 644295_MCM_shell_YP_003726241_Methanohalobium_evestigatum_Z_7303 188937_MCM1_NP_618700_Methanosarcina_acetivorans_C2A 990316_MCM1_YP_004385287_Methanosaeta_concilii_GP6 87.1/65 77.6/100 100/100 349307_MCM1_YP_842696_Methanosaeta_thermophila_PT 1110509_MCM1_AET65277_Methanosaeta_harundinacea_6Ac 84.7/98 100/100OFV66637_Candidatus_Syntrophoarchaeum_butanivorans 89.1/9295/100 OFV68247_Candidatus_Syntrophoarchaeum_caldarius PXF52786_ANME_1_cluster_archaeon 94.6/92 PPA79667_ANME_2_cluster_archaeon_HR1 95.8/100304371_MCM1_YP_003356305_Methanocella_paludicola_SANAE 100/1001041930_MCM1_YP_005381018_Methanocella_conradii_HZ254 351160_MCM1_YP_686686_Methanocella_arvoryzae_MRE50 86.9/75 410358_MCM1_YP_001030268_Methanocorpusculum_labreanum_Z 368407_MCM1_YP_001047160_Methanoculleus_marisnigri_JR1 98.8/9256.9/83 100/10057.8/99 882090_MCM1_ZP_09043156_Methanolinea_tarda_NOBI_1 58.7/100 521011_MCM1_YP_002467125_Methanosphaerula_palustris_E1_9c 97.5/10043.4/100456442_MCM1_YP_001404833_Methanoregula_boonei_6A8 90.6/99 323259_MCM1_YP_502454_Methanospirillum_hungatei_JF_1 937775_MCM1_ZP_09700740_Methanoplanus_limicola_DSM_2279 99.9/100679926_MCM1_YP_003894184_Methanolacinia_petrolearia_DSM_11571 OKY78610_Candidatus_Methanohalarchaeum_thermophilum 797209_MCM1_ZP_08042510_Haladaptatus_paucihalophilus_DX253 99.5/93 309800_MCM1_YP_003534297_Haloferax_volcanii_DS2 57.3/6495.9/100 87.8/100469382_MCM1_YP_004035124_Halogeometricum_borinquense_DSM_11551 98.9/100 362976_MCM1_YP_659323_Haloquadratum_walsbyi_DSM_16790 49.8/97756883_MCM1_YP_004809424_halophilic_archaeon_DL31 88.1/89 84.7/48 416348_MCM_shell_YP_002567153_Halorubrum_lacusprofundi_ATCC_49239 797210_MCM1_YP_004596122_Halopiger_xanaduensis_SH_6 21.1/84547559_MCM_shell_YP_003480325_Natrialba_magadii_ATCC_43099 100/10075.3/100797304_MCM_shell_ZP_08967542_Natronobacterium_gregoryi_SP2 89.8/78543526_MCM1_YP_003401890_Haloterrigena_turkmenica_DSM_5511 84.3/66 795797_MCM1_YP_003735228_Halalkalicoccus_jeotgali_B3 64.7/77 348780_MCM_shell_YP_331276_Natronomonas_pharaonis_DSM_2160 634497_MCM1_YP_004794671_Haloarcula_hispanica_ATCC_33960 99.9/100272569_MCM_shell_YP_137231_Haloarcula_marismortui_ATCC_43049 97.4/78 99.6/100 98.6/78 485914_MCM1_YP_003176874_Halomicrobium_mukohataei_DSM_12286 519442_MCM1_YP_003131067_Halorhabdus_utahensis_DSM_12940 93.3/78 1033806_MCM1_ZP_08560916_Halorhabdus_tiamatea_SARL4B 89.2/90 478009_MCM1_YP_001689961_Halobacterium_salinarum_R1 12.6/32 99.5/98 100/100751944_MCM1_ZP_09027959_Halobacterium_sp_DL1 99.4/100797303_MCM1_ZP_08964560_Natrinema_pellirubrum_DSM_15624 100/100 348780_MCM1_YP_327587_Natronomonas_pharaonis_DSM_2160 99.5/100 272569_MCM_shell_YP_136848_Haloarcula_marismortui_ATCC_43049 416348_MCM1_YP_002565410_Halorubrum_lacusprofundi_ATCC_49239 100/100 797304_MCM_shell_ZP_08968377_Natronobacterium_gregoryi_SP2 KYK28479_Thermoplasmatales_archaeon_SG8_52_1 98.6/100 WP_020449728_Candidatus_Methanomassiliicoccus_intestinalis 100/100WP_049796361_Methanomassiliicoccus_luminyensis 439481_MCM1_YP_003483546_Aciduliprofundum_boonei_T469 99.3/100 333146_MCM1_ZP_05571039_Ferroplasma_acidarmanus_fer1 82.9/100 99.9/100 31/90 263820_MCM1_YP_023995_Picrophilus_torridus_DSM_9790 100/100 EQB71237_Thermoplasmatales_archaeon_A_plasma 95.6/100 273075_MCM1_NP_394261_Thermoplasma_acidophilum_DSM_1728 100/100273116_MCM1_NP_111551_Thermoplasma_volcanium_GSS1 274854_MCM1_EHR75878_uncultured_marine_group_II_euryarchaeote 1072681_MCM1_ZP_09209841_Candidatus_Haloredivivus_sp_G17 99.5/100 KYK25047_Euryarchaeota_archaeon_SM23_78 91.8/100 PKM92164_Euryarchaeota_archaeon_HGW_Euryarchaeota_1 ODS38582_Candidatus_Altiarchaeales_archaeon_WOR_SM1_86_2 96.6/100 OYT42372_Candidatus_Altiarchaeales_archaeon_ex4484_43 99.7/100 PIX48385_Candidatus_Altiarchaeum_sp__CG_4_8_14_3_um_filter_33_2054 100/100OIQ05030_Candidatus_Altiarchaeum_sp__CG2_30_32_3053 0/76PIV27464_Candidatus_Altiarchaeum_sp__CG03_land_8_20_14_0_80_32_618 76.2/59 0/43PIN68127_Candidatus_Altiarchaeum_sp__CG12_big_fil_rev_8_21_14_0_65_33_22 PIZ30941_Candidatus_Altiarchaeum_sp__CG_4_10_14_0_8_um_filter_32_851 60.4/96 243232_MCM_shell_NP_247337_Methanocaldococcus_jannaschii_DSM_2661 97.5/99 419665_MCM_shell_YP_001325414_Methanococcus_aeolicus_Nankai_3 PhMferAG86_MEFER_RS07965_WP_015792115_1 63.8/95243232_MCM2_NP_248495_Methanocaldococcus_jannaschii_DSM_2661 55.2/94644281_MCM2_YP_003457928_Methanocaldococcus_sp_FS406_22 96.5/98573064_MCM2_YP_003128462_Methanocaldococcus_fervens_AG86 87.3/76 48.5/95579137_MCM2_YP_003247103_Methanocaldococcus_vulcanius_M7 573063_MCM2_YP_003615830_Methanocaldococcus_infernus_ME 647171_MCM2_ZP_09707424_Methanotorris_formicicus_Mc_S_70 99.8/100880724_MCM2_YP_004484748_Methanotorris_igneus_Kol_5 402880_MCM2_YP_001098161_Methanococcus_maripaludis_C5 91.5/97 97.7/61 8.7/59444158_MCM2_YP_001548966_Methanococcus_maripaludis_C6 98.7/100426368_MCM2_YP_001330245_Methanococcus_maripaludis_C7 267377_MCM2_NP_987150_Methanococcus_maripaludis_S2 98.9/89 100/1001053692_MCM2_YP_004741666_Methanococcus_maripaludis_X199.9/100 406327_MCM2_YP_001323542_Methanococcus_vannielii_SB 1053692_MCM_shell_YP_004742497_Methanococcus_maripaludis_X1 100/100 97.7/59 15.7/51MMV7_V1_MMARC7_RS00120_WP_048060482_1 100/10074.7/100MmarS2_MMP_RS03900_WP_011170692_1 36.8/37 97.2/100 MmarC6_MMARC6_RS00155_WP_012192845_1 444158_MCM_shell_YP_001548164_Methanococcus_maripaludis_C6 60/97 267377_MCM_shell_NP_987590_Methanococcus_maripaludis_S2 444158_MCM_shell_YP_001548488_Methanococcus_maripaludis_C692.3/100 99.5/94 100/100402880_MCM_shell_YP_001097634_Methanococcus_maripaludis_C5 100/100 92.8/100444158_MCM_shell_YP_001548188_Methanococcus_maripaludis_C6 94.1/58 98.5/59 444158_MCM_shell_YP_001548471_Methanococcus_maripaludis_C6 402880_MCM_shell_YP_001097170_Methanococcus_maripaludis_C5 18.4/77 444158_MCM_shell_YP_001549879_Methanococcus_maripaludis_C6 100/100 426368_MCM_shell_YP_001329286_Methanococcus_maripaludis_C799.8/100 406327_MCM_shell_YP_001322788_Methanococcus_vannielii_SB 96.6/100 419665_MCM2_YP_001325333_Methanococcus_aeolicus_Nankai_3 88.6/95 647113_MCM2_YP_004576461_Methanothermococcus_okinawensis_IH1 79.7/93 456320_MCM2_YP_003708030_Methanococcus_voltae_A3 pMEFER01_Mefer_1610_ACV25413_1 69.8/72 ph_MokiH1_Metok_0291_AEH06281_1 573064_MCM1_YP_003128165_Methanocaldococcus_fervens_AG86 86.6/89579137_MCM1_YP_003247785_Methanocaldococcus_vulcanius_M7 82.2/89644281_MCM1_YP_003458623_Methanocaldococcus_sp_FS406_22 98.6/100100/100243232_MCM1_NP_247956_Methanocaldococcus_jannaschii_DSM_2661 573063_MCM1_YP_003616672_Methanocaldococcus_infernus_ME 419665_MCM1_YP_001324688_Methanococcus_aeolicus_Nankai_3 98.8/100647113_MCM1_YP_004577256_Methanothermococcus_okinawensis_IH1 100/100 402880_MCM1_YP_001097099_Methanococcus_maripaludis_C5 43.5/100444158_MCM1_YP_001549679_Methanococcus_maripaludis_C6 99.8/10046/99 99.3/100426368_MCM1_YP_001329487_Methanococcus_maripaludis_C7 267377_MCM1_NP_988144_Methanococcus_maripaludis_S2 90/10052.4/991053692_MCM1_YP_004742798_Methanococcus_maripaludis_X1 33.3/8097.5/100406327_MCM1_YP_001322869_Methanococcus_vannielii_SB 456320_MCM1_YP_003707474_Methanococcus_voltae_A3 647171_MCM1_ZP_09708126_Methanotorris_formicicus_Mc_S_70 99.6/100880724_MCM1_YP_004484620_Methanotorris_igneus_Kol_5 523846_MCM1_YP_004003611_Methanothermus_fervidus_DSM_2088 99.8/100877455_MCM1_YP_004289753_Methanobacterium_lacus 100/100 42.9/99868131_MCM1_YP_004520673_Methanobacterium_paludis 91.6/100 339860_MCM1_YP_447408_Methanosphaera_stadtmanae_DSM_3091 634498_MCM1_YP_003423334_Methanobrevibacter_ruminantium_M1 99.2/100 100/100420247_MCM1_YP_001273083_Methanobrevibacter_smithii_ATCC_35061 187420_MCM1_NP_276876_Methanothermobacter_thermautotrophicus_str_Delta_H 99.9/10079929_MCM1_YP_003849261_Methanothermobacter_marburgensis_str_Marburg 272557_MCM1_NP_147033_Aeropyrum_pernix_K1 100/100 666510_MCM1_YP_003816359_Acidilobus_saccharovorans_345_15 399550_MCM1_YP_001041229_Staphylothermus_marinus_F1 59.3/95 100/100 91.3/100 591019_MCM1_YP_003669227_Staphylothermus_hellenicus_DSM_12710 633148_MCM1_YP_003649354_Thermosphaera_aggregans_DSM_11486 100/100 99.7/100768672_MCM1_ZP_09026152_Desulfurococcus_fermentans_DSM_16532 100/100490899_MCM1_YP_002428380_Desulfurococcus_kamchatkensis_1221n 765177_MCM1_YP_004176369_Desulfurococcus_mucosus_DSM_2162 415426_MCM1_YP_001013097_Hyperthermus_butylicus_DSM_5456 98.1/100694429_MCM1_YP_004780157_Pyrolobus_fumarii_1A 933801_MCM1_YP_004457710_Acidianus_hospitalis_W1 100/10099.3/1001006006_MCM1_YP_004408837_Metallosphaera_cuprina_Ar_4 98.2/98 99.8/100399549_MCM1_YP_001192107_Metallosphaera_sedula_DSM_5348 74.5/97 49.9/99 671065_MCM1_EHP68655_Metallosphaera_yellowstonensis_MK1 330779_MCM1_YP_255556_Sulfolobus_acidocaldarius_DSM_639 98.2/100273063_MCM1_NP_376352_Sulfolobus_tokodaii_str_7 425944_MCM1_YP_003419527_Sulfolobus_islandicus_LD85 100/100 0/2083.4/99419942_MCM1_YP_002840492_Sulfolobus_islandicus_YN1551 15.9/66 0/16429572_MCM1_YP_002832115_Sulfolobus_islandicus_LS215 81.4/98439386_MCM1_YP_002837550_Sulfolobus_islandicus_YG5714 427317_MCM1_YP_002829419_Sulfolobus_islandicus_M1425 100/1000/84426118_MCM1_YP_002914633_Sulfolobus_islandicus_M164 94.7/79 427318_MCM1_YP_002843345_Sulfolobus_islandicus_M1627 273057_MCM1_NP_342281_Sulfolobus_solfataricus_P2 453591_MCM1_YP_001435880_Ignicoccus_hospitalis_KIN4_I 583356_MCM1_YP_003860177_Ignisphaera_aggregans_DSM_17230 92.6/84 178306_MCM1_NP_558926_Pyrobaculum_aerophilum_str_IM2 37.7/1001104324_MCM1_YP_005085994_Pyrobaculum_sp_1860 92.5/100340102_MCM1_YP_001152380_Pyrobaculum_arsenaticum_DSM_13514 80.7/9898.4/100698757_MCM1_AFA40403_Pyrobaculum_oguniense_TE7 100/100410359_MCM1_YP_001054990_Pyrobaculum_calidifontis_JCM_11548 384616_MCM1_YP_930666_Pyrobaculum_islandicum_DSM_4184 36.2/88 89/100444157_MCM1_YP_001793508_Pyrobaculum_neutrophilum_V24Sta 100/100999630_MCM1_YP_004336914_Thermoproteus_uzoniensis_768_20 99.9/100 768679_MCM1_YP_004892030_Thermoproteus_tenax_Kra_1 100/100572478_MCM1_YP_003900969_Vulcanisaeta_distributa_DSM_14429 90.6/100 80.5/99 985053_MCM1_YP_004244902_Vulcanisaeta_moutnovskia_768_28 397948_MCM1_YP_001540218_Caldivirga_maquilingensis_IC_167 368408_MCM1_YP_919907_Thermofilum_pendens_Hrk_5 64.5/78 PSN90812_Candidatus_Marsarchaeota_G2_archaeon_OSP_D 100/100 PSN87651_Candidatus_Marsarchaeota_G2_archaeon_OSP_D PSN82363_Candidatus_Marsarchaeota_G1_archaeon_BE_D99.7/100 100/100 72.2/100PSN96608_Candidatus_Marsarchaeota_G2_archaeon_ECH_B_2 99.5/93 99.8/100PSO03905_Candidatus_Marsarchaeota_G2_archaeon_BE_D 99.9/100PSO07514_Candidatus_Marsarchaeota_G2_archaeon_BE_D PSN83932_Candidatus_Marsarchaeota_G1_archaeon_OSP_D 100/100311458_MCM1_BAJ50918_Candidatus_Caldiarchaeum_subterraneum 100/100 BAJ48116_Candidatus_Caldiarchaeum_subterraneum PUA31189_Aigarchaeota_archaeon_NZ13_MG1 OGD53557_Candidatus_Bathyarchaeota_archaeon_RBG_13_38_9 90.2/98 100/100PVX26717_Candidatus_Bathyarchaeota_archaeon 70.7/96 79/99 PVX27672_Candidatus_Bathyarchaeota_archaeon 98.9/100KPV64382_Candidatus_Bathyarchaeota_archaeon_BA2 OGD44080_Candidatus_Bathyarchaeota_archaeon_RBG_13_46_16b 93.8/9694.1/100 99.9/100 KON33837_miscellaneous_Crenarchaeota_group_6_archaeon_AD8_1 KYH37295_Candidatus_Bathyarchaeota_archaeon_B24 KON30514_miscellaneous_Crenarchaeota_group_15_archaeon_DG_45 100/100KYH38443_Candidatus_Bathyarchaeota_archaeon_B23 66.1/100886738_MCM1_ZP_08257443_Candidatus_Nitrosoarchaeum_limnia_SFB1 78.6/93WP_007402344_Candidatus_Nitrosoarchaeum_limnia PHY09592_Candidatus_Nitrosoarchaeum_sp_ 94.2/97 99.8/100 1001994_MCM1_EGP92971_Candidatus_Nitrosoarchaeum_koreensis_MY1 88.6/100WP_048109323_Candidatus_Nitrosoarchaeum_koreensis 99.8/100EGP92971_Candidatus_Nitrosoarchaeum_koreensis_MY1 97/100436308_MCM1_YP_001581576_Nitrosopumilus_maritimus_SCM1 97.5/100ABX12138_Nitrosopumilus_maritimus_SCM1 69.5/7393.1/100WP_048071375_Marine_Group_I_thaumarchaeote_SCGC_AAA799_P11 68.7/72 859350_MCM1_ZP_09606757_Candidatus_Nitrosopumilus_salaria_BD31 414004_MCM1_YP_876639_Cenarchaeum_symbiosum_A 78.6/94 ABZ09285_uncultured_marine_crenarchaeote_HF4000_APKG7F11 93.3/96KRT61695_Thaumarchaeota_archaeon_CSP1_1 99.7/99WP_081844828_Thaumarchaeota_archaeon_N4 100/100 WP_101477893_Nitrosotalea_sp__SbT1 AFU57673_Candidatus_Nitrososphaera_gargensis_Ga9_2

0.3 Fig. S6. MCM history within the Archaea. Maximum Likelihood phylogeny of the MCM protein within the Archaea. The Thermococcales chromosomic MCM are indicated in darkblue, the MCM encoded by Thermococcales MGEs are indicated in lightblue. The two Methanococcales chromosomic MCM are indicated with two different green. The Methanococcales MGEs encoded MCM are indicated in brown. The scale-bar represent the average number of substitutions per site. Values at nodes represent support calculated by aLRT and ultrafast bootstrap approximation (1,000 replicates). Fig. S7. MCM history within Thermococcales. Maximum Likelihood phylogeny of the MCM protein within Thermococcales using the Theionarchaea and the Methanofastidiosa as an out group. The Thermococcales chromosomic MCM are indicated in dark blue, the MCM encoded by MGEs are indicated in and the purple dots correspond to plasmids of the pT26-2 family. The scale-bars represent the average number of substitutions per site. Values at nodes represent support calculated by aLRT and ultrafast bootstrap approximation (1,000 replicates). Fig. S8. MCM history within Methanococcales. Maximum Likelihood phylogeny of the MCM protein within Methanococcales using the Methanobacteriales as an out group. The two Methanococcales chromosomic MCM are indicated in dark green by the names MCM1 and MCM2. the MCM encoded by MGEs are indicated with several other colours and are all within three groups highlighted in grey. The MCM encoded by Methanococcales plasmids of the pT26-2 family are indicated in red. The scale-bars represent the average number of substitutions per site. Values at nodes represent support calculated by aLRT and ultrafast bootstrap approximation (1,000 replicates). Fig. S9. Putative Replication origin prediction by GC-skew (left) and dotplot (right) in plasmid of the pT26-2 family. For integrated elements, the zero coordinate correspond to the first nucleotide of attL. Dotplots were drawn with Gepard. pT26-2 plasmid

Thermococcus kodakarensis TKV1

Thermococcus kodakarensis TKV2 Thermococcus kodakarensis TKV3

Thermococcus guaymasensis

Thermococcus litoralis Thermococcus barophilus E1

Thermococcus barophilus E2

Thermococcus sp. JCM11816 Thermococcus gammatolerans TGV1

Thermococcus celericrescens

Pyrococcus chitonophagus Pyrococcus kukulkanii

Pyrococcus korikoshii

Pyrococcus kukulkanii Pyrococcus sp. NA2

Methanococcus maripaludis MMC6V1 Methanococcus maripaludis MMC7V2

Methanococcus maripaludis S2 MMPV1

Methanococcus maripaludis KA1 Methanococcus maripaludis OS7

Methanococcus voltae MVV1

Methanothermococcus thermolithotrophicus Methanotorris igneus

Methanocaldococcus sp. FS406-22

pMEFER01 a. b. Halo_plasmid Haemophilus_phage pNOB8-type SNJ2-type

Halo_plasmid pNOB8-type Methano-pT26-2 Halo_plasmid SSV-type XerC

SNJ2-type pTN3-type XerC Halo_plasmid Methano-pT26-2 Thermo-pT26-2 Thermo-pT26-2 SSV-type

pTN3-type

Fig. S10. Integrase similarity network view and corresponding family attribution. The results of all integrases versus all integrase protein by BlastP (expect >0.001) are represented as a network with two additional different criteria in the limits of similarity in a the similarity is >25% among 65% of the protein, and in b >25% among 40% of the protein. WP_011225135_1_pKEF9_pNOB8like_Sulfolobus_islandicus 91.3/100 74.3/98WP_011225175_1__pARN3_pNOB8like_Sulfolobus_islandicus 94/100WP_012548831_1_pHA1_pNOB8like_Acidianus_hospitalis WP_020936647_1_int_E_Sulfolobus_solfataricus_P2 98.3/100 ARM76103_1_integrase_Acidianus_manzaensis 99/100 CDF47326_1_pARN4_pNOB8like_Sulfolobus_islandicus 99.6/100 WP_012386964_1_pSOG1_pNHOB8like_Sulfolobus_islandicus 56.7/82 CAA09137_1_pNOB8_Sulfolobus_sp_NOB8H2 pNOB8-like 95.1/90 ABP94320_1_integrase_Metallosphaera_sedula_DSM_5348 99.9/100 100/100 WP_080512783_1_integrase_Metallosphaera_sedula WP_066798157_1_pNOB8like_Caldivirga_sp_MU80 75.2/96 KUO92513_1_pNOB8like_Int_Thermocladium_sp_ECH_B 96.7/93 WP_052885762_1_pNOB8like_Vulcanisaeta_distributa 93.7/74 WP_066797986_1_integrase_pNOB8like_Caldivirga_sp_MU80 3.5/32 WP_081228025_1_pNOB8like_Vulcanisaeta_sp_EB80 95.7/97 WP_081227140_1_pNOB8like_Vulcanisaeta_sp_EB80 XerC_Thermococcus_kodakarensis 96.5/99 15.2/54WP_012571973_1_XerC_Thermococcus_onnurineus 76.7/60 WP_010867504_1_XerC_Pyrococcus_abyssi 99.2/100 99.7/100WP_013467433_1_XerC_Thermococcus_barophilus XerC 93.6/99 WP_048164945_1_XerC_Palaeococcus_pacificus 89.5/98 KYC56616_1_XerC_like_Arc_I_group_archaeon_ADurb1113_Bin01801 XerD_Sulfolobus_islandicus pHSR2_01_AKH98641_1_Halanaeroarchaeum_sulfurireducens 78.3/89 AHG00321_1_SNJ2_like_Halostagnicola_larsenii_XH_48 99.2/100 ACV10974_1_SNJ2_like_Halorhabdus_utahensis_DSM_12940 AFO55992_1_SNJ2_like_Natrinema_sp__J7_2 99.7/100 75.4/90 ELZ11643_1_SNJ2_like_Haloterrigena_thermotolerans_DSM_11522 74.1/97 AAV47109_1_SNJ2_like_Haloarcula_marismortui_ATCC_43049 99.6/100 96.8/100 42.2/87 WP_004515348_1_SNJ2_like_Haloarcula_vallismortis SNJ2-like 100/100 95/96 CAI50775_1_SNJ2_like_Natronomonas_pharaonis_DSM_2160 ACV47094_1_SNJ2_like_Halomicrobium_mukohataei_DSM_12286 76.7/86 ADE02447_1_SNJ2_like_Haloferax_volcanii_DS2 AGB16629_1_SNJ2_like_Halovivax_ruber_XH_70 79.9/99 CAI49276_1_SNJ2_like_Natronomonas_pharaonis_DSM_2160 M_maripaludis_S2 75.6/98 97/98 M_maripaludis_C7_MMC7V12 85.2/95 98.3/100 M_maripaludis_C7_MMC6V12 M_maripaludis_X11 M_maripaludis_C7_MMC6V13 M_maripaludis_C7_MMC7V2 94.9/100 76.6/100M_maripaludis_KA11 99.3/100 M_maripaludis_OS7 99.8/100 93.9/100 M_vannielli_integrated_phage2 M_vannielli_integrated_phage1 54.4/73 97.2/100 M_maripaludis_C7_MMC7V11 100/100 M_maripaludis_C7_MMC6V11 88.4/74 M_vannielli_integrated_phage3 M_maripaludis_X12 M_voltae 33/55 M_thermolithotrophicus 69.9/70 97.8/100 80.1/58 M_aeolicus_integrated_phage 27.9/46 M_okinawensis_integrated_phage 34.2/63M_sp_FS406_22 M_vulcanicus_integrated_phage 94.8/91 M_igneus 83/95M_jannashii_integrated_phage 98.5/98 M_fervens TPV1_prieuri_virus 98.8/99 T_litoralis T_gammatolerans 52.3/96TKV2_Tkodakarensis 98.3/100 99.8/100T_sp_JCM11816 65.5/95 intpT26_2 PspNA2_IP 82.9/82 85/99 Tbaro_E1 87.3/88 P_kulkukani 97.7/100 P_horikoshii 65.4/97 93.9/93 84/88 P_abyssi_GE2 intpTN3like_T_barophilus 5.5/41 T_guaymasensis 52/79 100/100 T_EXT12C_other_element TKV1_Tkodakarensis 86.1/96 Tbaro_E2 90.7/89 100/100 T_thioreducens_other_element P_yayanosii 100/100 P_chitonophagus 85.3/96 TKV3 pTN3_YP_008619357_1_Thermococcus_nautili 23.8/66 86.3/75 intpTN3_T_eurythermalis 100/99 100/100 intpTN3_T_guaymasensis pTN3-like intTKV4_Tkodakarensis intpTN3_T_CL1 99.2/100 intSSV1 99.7/100 intASV1 61.4/57 intSSV2 87.1/91 intSSV7 98.8/100 intSSV4 97.5/95 76.9/62 intSSVK1 88.6/65 intSSV3 SSV-like intSSVRH 80.9/91 intSSV5 99/100 intSVV6

0.4

Fig. S11. Two integrases family encoded by plasmid of the pT26-2 family Maximum Likelihood phylogeny of thepT26-2 encoded integrases protein using several other known integrases family as an outgroup. a.

Fig. S12. Integrase phylogeny and targeted tRNA gene for Thermococcales (top) and Methanococcales (bottom). Maximum Likelihood tree of integrases proteins (see uncollapsed version in supplementary Fig. S11). The tRNA gene in which the att site is located is indicated by coloured symbols. tRNA corresponding to the same amino-acid but with different anti-codon are indicated by the same symbol but varying colour. Thermococcales integrases target 9 different tRNA genes while Methanococcales integrases target 4 different tRNA genes. Most parsimonious ancestral tRNA gene target is indicated when possible. Integrase homologs found in integrated element that differ from pT26-2 are included.

Table S1. The putative replication origins coordinates in the plasmids of the pT26-2 family. Two complementary methodologies were used to determine the origin of replication : GC- skew and dotplot element Host ori localisation methodology name Themococcus sp 26-2 pT26-2 between t26-20p and t26-21p GC-skew and dotplot

Thermococcus kodakarensis KOD1 TKV1 Not found

Thermococcus kodakarensis KOD1 TKV2 from TK_RS02005 to TK_RS02015 GC-skew and dotplot

Thermococcus kodakarensis KOD1 TKV3 inside TK_RS02910 GC-skew

Thermococcus guayamensis from X802_RS00960 to X802_RS00965 GC-skew DSM11113

Thermococcus litoralis DSM 5473 Not found

Thermococcus barophilus CH5 E1 Not found

Thermococcus barophilus CH5 E2 from TBCH5V1_RS11215 to TBCH5V1_RS11225 GC-skew

Thermococcus sp. JCM11816 between Ga0128353_102299 and Ga0128353_102300 GC-skew and dotplot CONTIG 00002

Thermococcus gammatolerans EJ3 TGV1 from TGAM_RS03400 to TGAM_RS03405 GC-skew

Thermococcus celericrescens from the end of APY94_04115 to APY94_04120 and GC-skew and dotplot DSM17994 CONTIG 013 downstream

Pyrococcus chitoniphagus GC74 between A3L04_06365 and A3L04_06370 GC-skew

Pyrorcoccus kukulkanii sp. NCB100 between TQ32_RS02635 and TQ32_RS02645 GC-skew

Pyrococcus horikoshii OT3 PHV1 between PH_RS05530 and PH_RS05540 GC-skew

Pyrococcus yayanosii CH1 between PYCH_RS07010 and PYCH_RS07015 GC-skew and dotplot

Pyrococcus sp. NA2 immediatly downstream of PNA2_RS06700 GC-skew and dotplot

Pyrococcus abyssi GE2 pGE2 In the intergenic region two ORFs upstream the putative GC-skew and dotplot replication protein Methanococcus maripaludis C6 MMC6V1 between MMARC6_RS00125 and MMARC6_RS00130 GC-skew and dotplot

Methanococcus maripaludis C7 MMC7V1 Not found

Methanococcus maripaludis C7 MMC7V2 from MMARC7_RS07665 to MMARC7_RS07670 dotplot

Methanococcus maripaludis S2 MMPV1 between MMP_RS03905 and MMP_RS08880 GC-skew

Methanococcus maripaludis KA1 Dotplot: between tRNAser and MMKA1_04820 (or GC- skew between MMKA1_04910 and MMKA1_04970) Methanococcus maripaludis OS7 between MMOS7_04780 and MMOS7_04790 dotplot

Methanococcus maripaludis C5 Not found

Methanococcus maripaludis X1 Not found

Methanococcus voltae A3 MVV1 between MVOL_RS08870 and downstream of GC-skew MVOL_RS07785 Methanothermococcus Dotplot: between F555_RS08905 and F555_RS0101670 thermolithotrophicus GC skew: from F555_RS0101630 to F555_RS0101635 Methanotorris igneus Kol5 between METIG_RS02530 and METIG_RS02535 GC-skew

Methanocaldococcus sp. FS406-22 between MFS40622_1105 and MFS40622_1106 GC-skew and dotplot

Methanocaldococcus fervens AG86 pMEFER01 between MEFER_RS08100 and MEFER_RS08105 GC-skew and dotplot Table S2. List of primers

Name Sequence (5’->3’) Ref SP-ISC913 This study ASP-ISC913 This study SP-pGE2-CDS7 ATGAATACCGGAGTGTTCCTGAAGC This study ASP-pGE2-CDS7 AACGATGGCGTAACTTACGGTAAGA This study SP-pGE2-CDS29 TTGCTGCGTTTAGAATTAGCTCGTT This study ASP-pGE2-CDS29 TGGGTTGGGAGTACACCATAAAGAA This study Arc344F ACGGGGYGCAGCAGGCGCGA Raskin et al., 1994 Uni516R GTDTTACCGCGGCKGCTGRCA Takai & Horikoshi 2000

Raskin L, Stromley JM, Rittmann BE, Stahl DA (1994) Group-specific 16S rRNA hybridization probes to describe natural communities of methanogens. Appl Environ Microbiol 60: 1232–1240. Takai & Horikoshi (2000) Rapid detection and quantification of members of the archaeal community by quantitative PCR using fluorogenic probes. Appl Environ Microbiol 66 : 5066-5072