hemA Bpet4544 arsH2 Bpet4551 arsR Bpet4557 Bpet4559 Bpet4564 Bpet4565 traF2 virD2 sufI Bpet4570 Bpet4571 Bpet4572 Bpet4573 Bpet4579 Bpet4584 copB copA copS copR Bpet4596 Bpet4601 Bpet4603 Bpet4604 czcA czcB czcC Bpet4610 Bpet4612 Bpet4614 traG2 trbB2 trbE2 trbJ2 trbL2 trbF2 trbG2 trbI2

guaA 1262 1264 1266 1267 1268 1269 1270 1271 1272 1273 1275 1276 1277 1282 1283 1286 1288 1289 1291 1292 1293 1294 1295 1297 1298 1299 1301 1302 1303 1304 1305 1307 1308 1310 1311 1312 1313 1314 1316 1319 1320 1322 1323 1324 1325 1327

A0937 A0940 A0941 A0942 A0943 A0944 A0945 A0947 A0949 A0953 A0955 A0956 A0960 A0961 A0963 A0964 A0965 A0967 A0968 trbB A0973 trbJ trbL A0976 trbG A0978 A0982 A0983 A0984 A0985 A0988 A0989 A0990 A0991 A0993 A0997

4137 4140 4141 4142 4143 4144 4145 4146 4147 4148 4151 4152 4153 4154 4155 4156 4157 4159 4164 4166 4167 4169 4170 4171 4172 4173 4174 4175 4176 4177 4178 4179 4180 4181 4182 4184 4185 4186 4187 4188 4189 4191 4193 4194 4195 4198 4199 4200 4201 4202 4203

guaA RSMK00650 RSMK00654 parA3 traF RSMK00668 traG trbB RSMK00681 trbJ trbL trbF trbG trbI oprM

3675trbG36773678trbJ3681trbB368536863688368936903691369236933694369536963697369837003701370337043706370737123713nhaA37173718371937203721372237233725arsBarsCarsH3732

3663trbG36653666trbJ3669trbB3673367436763677367836793680368136823683368436853686368736893690369236933695369637003701nhaA37053706370737083709371037113713arsBarsCarsH3720

1244 1247 1248 1249 1250 1251 1252 1253 1254 1255 1256 1257 1261 1263 1264 1266 1267 1268 1269 1270 1275 1277 1278 1280 1281 1282 1285 1286 1288 1289 1290 1291

2601 2602 2603 2604 2605 2606 2607 2611 2612 2614 2618 2620 2621 2623 2624 2625 2627 2628 2629 2630 2631 2632 2633 2635 2637 2638 2639 2640 2641 2643 2644 2645 2648 2649 2650 2651 2652 2653 2655

int repA parA traF bphS bphE bphG bphF bphA1 bphB bphC bphD bphA4 traR traG trbB trbE trbJ trbL trbF trbG trbI

trbItrbGtrbFtrbLtrbJtrbEtrbBtraGSmlt1295Smlt1300Smlt1302Smlt1306Smlt1308Smlt1323Smlt1326Smlt1329Smlt1331Smlt1333Smlt1336

3497349834993500350235033506350735083510351135123514351535163517351835203521352235243526352835343535353835393540

2669267026712672267326742675267626772678267926802681268226832684268526862688269026912692269326952696269927002701270327052706270727092710271127122713271427152716271727182720272127242725272627272731273327342739274027412742274327442745

PD2939PD2942PD2943PD2944PD2945PD2946PD2947PD2948PD2950PD2953PD2954PD2959PD2960PD2962PD2964PD2965PD2968PD2970PD2973PD2974PD2975PD2977PD2978PD2981PD2983PD2984PD2990PD2992PD2998PD2999PD3003PD3004PD3005PD3006PD3007PD3008PD3009

guaA int 1467 1469 1470 1471 1472 1473 yupB parB 1479 1483 repA parA 1487 traF virD2 1490 tnpA cbbA1 cbbO cbbQ cbbL cbbR1 tnpA tnpA cbbF1 cbbR2 cbbE1 cbbF2 cbbP cbbT1 cbbZ1 cbbG1 pgk1 cbbY cbbA2 pykA2 cbbJ1 cbbI1 hoxF hoxU hoxY hoxH hoxI tnpA pntB hoxN hypB2 hypF2 hypD2 hypE2 hoxX hoxA tnpA traR traG 1549 trbB trbE trbJ trbL trbF trbG trbI 1561 1562

int 1237 ritA2 ritB2 ritC2 yupB parB 1246 1250 tnp 1252 repA parA 1257 traF virD2 1260 1261 tnpA tnpB tnpA 1265 1266 oprN ybhR ybhS ritA1 ritB1 ritC1 tnpA hypE1 hypD1 hypF1 hypB1 hoxV hoxT hoxQ hoxM hoxZ hupE hoxG hoxK 1300 tnpA patR 1306 1307 1308 1311 tmoE tmoA dmpH bphI mhpF xylJ dmpC 1323 dmpB tomA5 tomA3 tomA1 phaB2 chnA traR traG 1340 trbB trbE trbJ trbL trbF trbG trbI

04270429043004310432043404350438043904400442044404460448044904500451045204530454045504570458045904620464046704680473047404750476047704790480048104820483048804890490049204950497

int2 Bpet2169 Bpet2171 Bpet2174 Bpet2176 traF1 virD2 fadA2 Bpet2192 fadA3 Bpet2197 Bpet2198 Bpet2199 Bpet2201 Bpet2202 traG1 trbB1 trbE1 trbJ1 trbL1 trbF1 trbG1 trbI1

3695369736983699trbLtrbJ3703trbB3707370837103711kdpA1kdpB1kdpC13716371737193721372337243725372637273728373037313733373637403741374237433747

synonym="trbE"synonym="traG"30990310003101031030310403105031060310703108031090311003111031130311803119031200312403127031280

1865 1866 1867 1868 1869 1870 1876 1878 1879 1881 1882 1883 1884 1885 1886 1887 1888 1889 1890 1891 1893 1894 1895 1898 1899 1901 1902 1903 1904

290229032904290529072908291129122913291629182919292129232924292529262927292929302932293329352938294129432944294729492950295229532954295529562959

2483 2486 2488 2489 2491 2492 2496 2498 2499 2501 2502 2503 2504 2505 2506 2507 2509 2510 2511 2512 2514 2515 2516 2519 2520 2521 2522 2523 2524

2 3 4 5 6 7 8 9 10 radC 12 13 gene32 parB 16 17 arsC 19 20 21 22 23 24 25 26 27 xis rep parA 31 32 traF traI bphS tnp bphE bphG bphF bphA1 bphB bphC bphD bphW bphA4 traR traG 51 trbB trbE trbJ 57 trbL trbF trbG trbI 63 64 tnp

trbItrbGtrbFtrbLtrbJtrbEtrbBtraGRSc2588RSc2591RSc2593RSc2598RSc2601traFparA3RSc2606RSc2612RSc2617RSc2618RSc2619RSc2621RSc2622

trbGtrbFtrbLtrbJtrbEtrbB35740357603577035780357903580035820358303585035860358703588035890359003592035930traF35970359803600036050360703608036120

trbItrbGtrbFtrbLtrbJtrbEtrbBtraGtraRatp700898pcoDcopBcopAcusRcusStraItraF00908parArep009130091900920radC00925009270092800932htpX00935trxC009370093800939009400094200944ftsHcls0094800949009500095100953vsr009550095600957009580095900961

12350 12365 12370 12375 12380 12385 12390 12395 12405 12410 12415 12430 12440 12445 12465 12475 12480 12485 12500 12505 12510 12515 12520 12530 12540 12555 12560 12570 12575 12580 12585

0231802319023200232102323023240232702329023310233202333023340233502336023370233902340023430234402346023470234902350023520235502356023570236002361023620236302364

010200007819010200007834010200007859010200007889010200007909010200007969010200007994010200008004010200008009

0671 0672 0673 0675 0676 0679 0680 0683 0684 0686 0687 0689 0690 0691 0693 0694 0695 0696 0697 0698 0699 0700 0702 0704 0706 0708 0710 0713 0714 0715 0716 0717 0719 0720 0722 0724 0725 0726 0729 0730 0732 0733 0734 0735

0235cagXtrbFtrbL0240synonym="trbE"02440245traG0248025002510252traF025402550256025802590266026702710272027402750277027802790280

30 40 50 60 70 80 90 100

identity at amino-acid level (%)

75

76

77

78

79

81

82

83

84

87

88

89

90

91

92

93

94

95

96

98

99

100

101

102

104

105

107

108

109

110

111

112

4,803,647 - 4,889,901

1,324,211 - 1,387,456

896,365 - 966,129

12,616 - 82,006

331,248 - 372,343

30,615 - 80,249 (complementary)

136,375 - 186,009 (complementary)

1,458,447 - 1,503,679

2,715,199 - 2,769,318

1 - 55,374

1,324,222 - 1,367,730 (complementary)

3,746,937 - 3,791,764 (complementary)

2,938,269 - 3,016,679 (complementary)

3,207,911 - 3,280,947 (complementary)

1,584,594 - 1,683,445

1,359,262 - 1,460,899

459,522 - 526,276 (complementary)

2,250,540 - 2,297,729

3,829,571 - 3,884,857 (complementary)

2,677,690 - 2,721,399 (complementary)

1,988,398 - 2,031,631

3,075,815 - 3,129,205 (complementary)

9,143 - 56,453

1 - 61,807

2,781,376 - 2,825,985 (complementary)

3,634,262 - 3,681,100 (complementary)

954,457 - 1,035,107 (complementary)

133,371 - 184,030

2,603,984 - 2,652,521 (complementary)

40,334 - 84,863 (complementary)

730,667 - 790,510

1,853 - 44,008 (complementary)

Bordetella petrii DSM 12804

Acidovorax ebreus TPSY

Burkholderia pseudomallei MSHR346

Polaromonas naphthalenivorans CJ2

Ralstonia solanacearum MolK2

Burkholderia multivorans CGD2

Burkholderia multivorans CGD2M

Shewanella sp. ANA 3

Ralstonia pickettii 12J

Cupriavidus oxalaticus (Tn4371)

Stenotrophomonas maltophilia K279a

Acidovorax ebreus TPSY

Delftia acidovorans SPH 1

Comamonas testosteroni KF 1

Ralstonia metallidurans CH34

Ralstonia metallidurans CH34

Delftia acidovorans SPH 1

Bordetella petrii DSM 12804

Pseudomonas aeruginosa PA7

Pseudomonas aeruginosa UCBPP PA14

Thioalkalivibrio sp. HL-EbGR7

Acidovorax sp. JS42

Alicycliphilus denitrificans BC

Ralstonia solanacearum GMI1000

Azotobacter vinelandii DJ

Burkholderia multivorans ATCC 17616

Congregibacter litoralis KT71

Pseudomonas aeruginosa 2192

Pectobacterium carotovorumsubsp. brasiliensis PBR1692

Acidovorax avenae subsp. citrulli AAC00 1

Pseudomonas aeruginosa PACS171b

KKS102

Fig. S1 Comparative analysis of ICEs with traG that belonged to βand γ clusters. Coding sequences (CDSs) present in a range of genomicsequences from one organism were blastp-compared to those in another.Detected similarities with query coverage greater than 50% and identityscore greater than 30% are depicted with colored lines. The relationshipsbetween colors and identity scores are shown in the top-right area. CDSsin the int, parB, parI, and mpf blocks are shown in orange, cyan, green,and purple, respectively. radC is shown in pale cyan and traR is shown inpurple. At the left of each panel, the sequence number and exact sequencerange used for the analysis are indicated. See Supplementary Table S4 forsequence numbers, source organisms, and accession numbers. This figurewas drawn by using a computational tool, CompareSequences, which isavailable at: http://www.ige.tohoku.ac.jp/joho/gmProject/gmhome.html.

0 - 0

095409550958096109630968097509790989099020 10

7794

2 - 1

1129

42co

mpl

emen

tary

0 - 0

dnaE slt 0117 0120 0121 0122 0124 0125 0126 traG trbB trbE trbL trbG trbI mdtB

22 1119

95 -

1538

95

0 - 0

1470 1471 1474 1475 1492 1493 1496 trbE 1505 1508 1514

24 1557

517

- 160

1517

0 - 0

2241 2242 2243 2252 2255 2256 2258 2261 2264 2266 2267 2271 2273 2276

25 2434

824

- 247

8824

0 - 0

26012602260526082611261826192635263726392641264226472648265226592668

26 2742

596

- 281

2546

com

plem

enta

ry

0 - 0

2770 2771 2772 2776 2777 2779 2780 2781 2784 2798 2806 2809 2814 2818 2827 2830 2833 2837 2838

28 3034

579

- 310

2479

0 - 0

3147 3151 3160 3162 3163 3164 3174 3184 3192 3193 3196 3208 3215 3218 3221 3227

31 4779

24 -

5477

24

0 - 0

14721475147814811486149214931494149514961498149915031506150715121524152515261527152815331534154015411548

32 1429

721

- 152

3771

com

plem

enta

ry

0 - 0

06860690069306960701070207100720073007360737

34 7750

46 -

8258

96co

mpl

emen

tary

0 - 0

7706 7707 7708 7711 7717 7738 7739 trbE 7747 7750 7754

35 8096

791

- 814

5741

0 - 0

3795 3799 3801 3802 3803 3804 3806 3807 3808 3811 3824 3825 3830 3833 3836

36 4108

120

- 415

8920

879049 - 879049

07370740074307460751075207710772077307760780078107820785078607870789079207960797

37 8233

49 -

8891

49co

mpl

emen

tary

0 - 0

2851 2857 2858 2875 2876 2881 2884 2887

38 3180

920

- 321

7620

0 - 0

3987399139943997400140074008401540254026402940324034

39 4415

078

- 446

3278

com

plem

enta

ry

0 - 0

0568 0570 0571 0581 0588 0589 0591 0592 0593 0598 0601 0604 0607 0614 0617 0620 0622

40 6390

27 -

6968

27

0 - 0

3055 3056 3060 3065 3067 3076 3083 3084 3089 3092 3095 3098 3100 3102 3104 3105 3109 3112 guaA

41 3278

913

- 334

6313

0 - 0

3795 3799 3801 3802 3803 3804 3806 3807 3808 3811 3824 3825 3830 3833 3836 3846

42 4108

620

- 416

4420

0 - 0

41784183trbL4189419541964203421542204226422842344237

43 1861

06 -

2396

06co

mpl

emen

tary

0 - 0

3670 3672 3688 3689 3694 3697 3700 3706

44 4033

592

- 406

8292

0 - 0

6422trbL26427cadA26437643964416443645664636465guaA6468

46 2360

552

- 240

6452

com

plem

enta

ry

0 - 0

3122 3123 3128 3131 3148 3151 3179 3182 3187 3188 3197 3200 3203 3207

47 3043

19 -

3721

19

0 - 0

0134 0139 0142 0147 0150 0153 0160

48 1059

07 -

1425

07

0 - 0

3805 3806 3808 3809 3810 3819 3828 3842 3848 3849 3854 3857 3860

49 4108

085

- 416

7685

0 - 0

guaA0496050005030506051105120513051505160535053805400541

50 5501

51 -

5973

51co

mpl

emen

tary

0 - 0

087290877408799088040886908889

51 7166

- 39

766

com

plem

enta

ry

0 - 0

c1482 c1486 xerD c1493 c1494 c1498 c1508 c1509 trbE trbL trbI

52 1684

090

- 172

7690

0 - 0

guaARPA2211RPA2214RPA2215RPA2216RPA2218RPA2219RPA2220RPA2221trbI1trbLtrbE1traG2RPA2249

53 2480

697

- 255

6697

com

plem

enta

ry

0 - 0

10011004100710101016101710181019102310261047104810541055105610591061

55 1128

192

- 119

0392

com

plem

enta

ry

0 - 0

7386 7388 7391 7392 7399 7400 7401 7404 7405 7406 7409 7413 7421 7437 traG 7443 7446 7452

57 7763

075

- 782

7775

0 - 0

39943997399839994000400340054007400840124018402140264027404240434047

58 4462

219

- 453

3619

com

plem

enta

ry

0 - 0

3867387238743877388338863892389538983903390439103919392039263927

59 4296

505

- 436

3105

com

plem

enta

ry

0 - 0

3328 3329 3330 3333 3334 3351 3355 3356 3359 3360 3362 trbE 3370 3373 3378

60 3474

646

- 353

3446

0 - 0

3529353235353538354235433546354835533558356635673574

62 3919

200

- 397

6200

com

plem

enta

ry

0 - 0

29922995299830013006300730153026303130353036

63 3329

021

- 337

2821

com

plem

enta

ry

0 - 0

23152318232123272330234623482349

65 2465

574

- 250

0374

com

plem

enta

ry

0 - 0

12489 12509 12539 12554 12574 12594 12639

66 2537

765

- 257

3915

0 - 0

0359203617036270366703717

67 6429

15 -

6710

65co

mpl

emen

tary

1113036 - 1116436

05602 05632 05637 05642 05647 05652 05672 05682 05712 05757

68 1059

086

- 110

6936

0 - 0

16603 16688 16728 16733 16758

70 9086

8 - 1

2826

8

0 - 0

1807 1816 1821 1822 1831 1833 1838 1840 1843

71 1962

149

- 199

8599

0 - 0

1999 2006 2011 2017 2024 2025 2030 2032 2035

72 2139

520

- 218

1770

0 - 0

intSmsi003msi008fixIfixGfixNtrbItrbGtrbLtrbEtraGnodMvirAvirB4msi059virD4msi066

73 1 - 8

7584

com

plem

enta

ry

0 - 0

mlr6361 mlr6395 mlr6400 mlr6417

74 5184

649

- 525

7099

0 - 0

parB rep traI tnp traR traG trbB trbE trbL trbI tnp

1 - 6

1807

Fig. S2 Comparative analysis of putative ICEs carrying traGbelonging to α clusters I and II. See the legend to Fig. S1. TraGsequences not surrounded by typical ICE genes were omitted from thefigure.

75 Bordetella petrii DSM 12804 N. F.

76 Acidovorax ebreus TPSY N. F.

79 Ralstonia solanacearum MolK2 N. F.

92 Ralstonia metallidurans CH34 N. F.

107 Burkholderia multivorans ATCC 17616 N. F.

77 Burkholderia pseudomallei MSHR346 ATTTTGAG/TTTTTGAG

100 Acidovorax sp. JS42 ATTTTGAG/TTTTTGAG

101 Alicycliphilus denitrificans BC TTTTTGAG

98 Pseudomonas aeruginosa UCBPP PA14 GATTTTAAG

99 Thioalkalivibrio sp. HL-EbGR7 GATTTTAAG

102 Acidovorax sp. KKS102 GATTTTAAG

78 Polaromonas naphthalenivorans CJ2 TTTTTTAT

83 Shewanella sp. ANA 3 TTTTTTAT

108 Congregibacter litoralis KT71 TTTTTTAT

81 Burkholderia multivorans CGD2 TTTTTTGT/TTTTTTGA

82 Burkholderia multivorans CGD2M TTTTTTGT/TTTTTTGA

93 Ralstonia metallidurans CH34 TTTTTTGT

95 Bordetella petrii DSM 12804 TTTTTTGT/TTTTTTAC

88 Stenotrophomonas maltophilia K279a TTTTTTGT

96 Pseudomonas aeruginosa PA7 TTTTTTGT

105 Azotobacter vinelandii DJ TTTTTTGT/TTTTTTGA

112 Pseudomonas aeruginosa PACS171b TTTTTTAT

104 Ralstonia solanacearum GMI1000 TTTTTTAT/TTTTTTAG

109 Pseudomonas aeruginosa 2192 TTTTTTAT

110 Pectobacterium carotovorum subsp. brasiliensis PBR1692 N. F./TTTTTTAT

87 Cupriavidus oxalaticus TTTTTCAT

111 Acidovorax avenae subsp. citrulli AAC00 1 TTTTTCAT

94 Delftia acidovorans SPH 1 TTTTTCAT

84 Ralstonia pickettii 12J TTTTTCAT

91 Comamonas testosteroni KF 1 TTTTTCAT

89 Acidovorax ebreus TPSY TTCTTCAT/TTTTTCAT

90 Delftia acidovorans SPH 1 TTTTTCAT0.2

Fig. S3 Phylogenetic tree of Int proteins of Tn4371-like elements fromβ- and γ-proteobacteria Int protein sequences were aligned and aneighbor-joining tree was depicted by MAFFT (7). The strain names arefollowed by nucleotide sequences of att sites. In cases where the two sitesare not identical, both sites are shown. N. F.: not found.