The phylogeny of Ptiliidae (Staphylinoidea, Coleoptera) – the smallest beetles and their evolutionary transformations

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The phylogeny of Ptiliidae (Staphylinoidea, Coleoptera) – the smallest beetles and their evolutionary transformations

A LEXEY A. P OLILOV *

^,^1,²

, I GNACIO R IBERA

³

, M ARGARITA I. Y AVORSKAYA

⁴

, A NABELA C ARDOSO

³

, V ASILY V. G REBENNIKOV

⁵

& R OLF G. B EUTEL

⁴

1

Department of Entomology, Biological faculty, Lomonosov Moscow State University, Moscow, Russia; Alexey A. Polilov * [polilov@gmail.com] —

²

Joint Russian-Vietnamese Tropical Research and Technological Center, Hanoi, Vietnam —

³

Institute of Evolutionary Biology (CSIC-Universitat Pompeu Fabra), Barcelona, Spain; Ignacio Ribera * [ignacio.ribera@ibe.upf-csic.es]; Anabela Cardoso * [anabela@um.es] —

⁴

Institut für Zoologie und Evolutionsforschung, FSU Jena, Jena, Germany; Margarita I. Yavorskaya * [margojavor@gmail.com]; Rolf G. Beutel * [rolf.beutel@uni- jena.de] —

⁵

Canadian Food Inspection Agency, Ottawa, Canada; Vasily v. Grebennikov * [v_grebennikov@mail.ru] — * Corresponding author

Supporting Information

Table S1. Studied specimens, localities and GenBank accession numbers.

Table S2. Primers used for DNA amplification.

Table S3. Data matrix of 68 morphological characters scored for 33 terminals and used for the phylogenetic analysis of Ptiliidae beetles.

Figures S1- S7. SEM habitus images of Ptiliidae.

Figure S 8 . Maximum Likelihood inference phylogram of Ptiliidae obtained with RAxML with

the molecular data only. Digits at internodes are bootstrap values when higher than 50%.

Figure S 9 . Majority rule consensus tree of the Bayesian analysis (obtained with BEAST) of the

molecular characters, with the simplest models (analysis C, see Methods). Numbers in nodes are

posterior probabilities when higher than 0.5.

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Table S1: Material used for the morphological (yellow) and molecular analyses (green – our sequences and grey – sequences obtained from Genbank), with current classification, voucher numbers, locality, collector and Genbank accession numbers (in bold, new sequences). "16S" refers to the

fragment16S+tRNA-Leu+NAD1.

(1) Sequences of both specimens were pooled in all analyses.

(2) Sequences were pooled in the combined analyses.

species voucher country locality collector morphology COI 12S 16S 18S 28S

F. Ptiliidae sF. Acrotrichinae T. Acrotrichini

Acrotrichis MNCN-AI413 Germany Thuringia, Jena, Pennickental, 19.7.2005 R.G. Beutel LT991584 - LT990832 LT991036 -

Acrotrichis atomaria GBK2 BMNH:676883 DQ156008 - - - -

Acrotrichis grandicollis Russia

Russia, Moscow region, ZBS MSU field station, 55°41′58″N 36°43′57″E, 18.10.2015, compost sifting

A.A. Polilov

Acrotrichis insularis GBK4 isolate 104 DQ313374 - - - -

Acrotrichis intermedia GBK5 BMNH:676119 DQ155930 - - - -

Acrotrichis intermedia Russia Russia, Moscow region, Morozki, 56°12′26″N

37°29′53″E, 2.10.2007, sifting A.A. Polilov

Acrotrichis sitkaensis GBK7 BMNH:676167 DQ155969 - - - -

Acrotrichis xanthocera GBK9 haplotype AA AY550881 - - - -

T. Nephanini

Baeocrara variolosa IBE-AN416 Russia Moscow region, ZBS MSU field station, 55°41′58″N

36°43′57″E, 18.10.2015, compost sifting A.A. Polilov LR742629 LR742613 - LR742664 LR742688 Nephanes titan IBE-AN415 Russia Moscow region, ZBS MSU field station, 55°41′58″N

36°43′57″E, 18.10.2015, compost sifting A.A. Polilov LR742639 - - LR742675 LR742697

Rioneta uluguruensis IBE-AN360 Tanzania East slope southern Uluguru Mts., S07º07'20"

E037º38'37", 18.xi.2010, 2038m, sifting17 V.V. Grebennikov LR742647 LR742626 - LR742683 LR742705 Rioneta uluguruensis MNCN-AI415 Tanzania Uluguru Mts. H. 2200 m W slope between

Tchenzema vil. and Lukvangule Plateau, 2002 V.V. Grebennikov LT991586 - - LR74264 -

Smicrus filicornis IBE-AN423 Russia Voronezh region, Venevitinovo, 51°48′48″N

39°23′5″E, 2.10.2006, light trap A.A. Polilov - LR742628 - - -

sF. Cephaloplectinae

Limulodes parki (1) IBE-AN819 Wisconsin (US) Co., Barcevel Prairie, Aphaenogaster / ant nest,

2006, 42.9926 / -89.9043, 343m J. Gruber LR742635 LR742617 LR742655 - -

Limulodes parki (1) GBK DDM0287 - - - - KJ844893

Limulodes parki USA USA, Illinois, Willow Springs, 1943 C. Seevers

sF. Ptiliinae

T. Discheramocephalini

Cissidium sp. IBE-AN362 Tanzania East slope southern Uluguru Mts., S07º07'20"

E037º38'37", 18.xi.2010, 2038m, sifting17 V.V. Grebennikov LR742630 LR742614 - LR742665 LR742689 Dacrysoma

usamabarensis IBE-AN359 Tanzania E USambara Mts., Amani NR, 5º10'34"S

38º36'01"E. 15.xii.2011, 1004m, sift. 05 V.V. Grebennikov LR742631 LR742615 LR742652 LR742666 LR742690 Discheramocephalus sp. IBE-AN361 Tanzania East slope southern Uluguru Mts., S07º07'20"

E037º38'37", 18.xi.2010, 2038m, sifting17 V.V. Grebennikov LR742632 LR742616 - LR742667 LR742691 T. Nanosellini

Cylindroselloides dybasi MNCN-AI564 Canada Eastern ON, vicinity of Westport, S.&J. Peck

cottage on Ridaeu Canal, from bracket fungus, 2005 V.V. Grebennikov LT991585 - LT990833 LT991037 LT991250

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Mikado sp. IBE-AN419 Vietnam

Cat Tien national park, 11º 26' 32"N 107º 25' 53"E, 27.09.2010, polypore №3 Abundisporus

fuscopurpureus

A.A. Polilov LR742637 LR742619 - LR742672 LR742695

Nanosellini gen. MNCN-AI414 Tanzania E. Usambara Mts. H 450 m. Amani Nat. Res., Zigo

Lodge, 2002 V.V. Grebennikov - - - LR742674 -

Porophila cedri IBE-AN417 Russia Far East, i. Russkiy, 43°3′3″N 131°51′39″E,

22.09.2015, polypore A.A. Polilov LR742642 LR742621 LR742659 LR742678 LR742700

Primorskiella anodonta IBE-AN418 Russia Far East, i. Russkiy, 43°3′3″N 131°51′39″E,

22.09.2015, polypore A.A. Polilov LR742643 LR742622 LR742660 LR742679 LR742701

Scydosella musawasensis GBK27 GBK27 KU302777 - KU302777 - -

Scydosella musawasensis IBE-AN424 Colombia Chicaque National Park, 4°37′8″N 74°18′43″W,

2200 m, 8.02.2015, polypore Steccherinum sp A.A. Polilov LR742650 LR742627 - LR742686 - T. Ptenidiini

Motschulskium

sinuatocolle IBE-AN357 California (US)

Del Norte Co., Coast Redwood St. Pk, False Klamath Cove, 2m., N41º36'13" W124º06'06", 17.vi.2006, sandy beach

A. Newton &

M. Thayer LR742638 LR742620 LR742656 LR742673 LR742696

Nossidium sp. GBK13 DDM0127 - - - KP419188 KJ845037

Nossidium pilosellum Krim Krim, Krimsky n.p., July 1956 M. Tihonravov

Sindosium sp. China

P.R. China, Yunnan, E slope Cangshan at Dali, N25º40'18.6" E100º08'00.7", 11.v.2010, 2690m, sifting 06

V.V. Grebennikov

Ptenidium sp. GBK22 BMNH:679236 DQ221987 - DQ202584 - DQ202645

Ptenidium formicetorum GBK14 BMNH:833149 HQ165580 - - - -

Ptenidium fuscicorne GBK15 BMNH:833160 HQ164997 - - - -

Ptenidium pusillum GBK21 BMNH:677424 HQ165269 - - - -

Ptenidium pusillum MNCN-AI515 Morocco 21.4.2001 /30º39’456”N 9º21’134”W /Immouzèr- des-Ida-Outanane/Assif Tanit, 550 m

I. Ribera &

A. Cieslak HE970847 LR742623 HE971079 HE970959 HE970997

Ptenidium pusillum Russia Russia, Moscow region, ZBS MSU field station,

55°41′58″N 36°43′57″E, 5.10.2015 compost sifting A.A. Polilov T. Ptiliini

Micridium vittatum IBE-AN427 Russia Moscow region, Snigeri, 55°54′27″N 37°2′21″E,

16.11.2015, Formica rufa nest A.A. Polilov LR742636 LR742618 - LR742671 LR742694

Ptilum myrmecophilum IBE-AN426 Russia Moscow region, Snigeri, 55°54′27″N 37°2′21″E,

16.11.2015, Formica rufa nest A.A. Polilov LR742644 LR742624 LR742661 LR742680 LR742702

Actidium boudieri Russia Russia, Moscow region, Zaharovo, 55°47′53″N

37°1′16″E, 26.06.2002, in flight A.A. Polilov Ptiliola kunzei Russia Russia, Far East, Lazovsky national park, 43°15′N

134°07E, 13.07.2005, Laetiporus A.A. Polilov Oligella foveolata Russia Russia, Voronezh region, Venevitinovo, 51°48′48″N

39°23′5″E, 2.10.2006, light trap A.A. Polilov

Ptiliolum sp. MNCN-AI649 Spain Córdoba, Cabra, La Nava, 22.1.2006 A. Castro

T. Ptinellini

Pteryx suturalis GBK23 724636 - - - EF362983 -

Pteryx suturalis Russia Russia, Moscow region, 94 km, 56°34′15″N

37°32′52″E, 30.06.2002, in rotting wood A.A. Polilov

Ptinella sp. (2) IBE-AN439 Australia QLD, Brisbane, St Lucia, Oct 2015, J. Maté LR742646 - - LR742682 LR742704

Ptinella aptera (2) GBK25 BMNH:833134 HQ165077 - - - -

Ptinella aptera Russia Russia, Moscow region, 94 km, 56°34′15″N

37°32′52″E, 30.06.2002, in rotting wood A.A. Polilov Ptinellini gen. IBE-AN356 Tanzania Uluguru Mts. at Bunduki vil., S07º03'50"

E037º36'55" 23.xi.2010 2263m, Sifting 22 V.V. Grebennikov LR742645 LR742625 - LR742681 LR742703

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F. Hydraenidae Adelphydraena

orchymonti MNCN-AI356 Venezuela 100m, Tobogan de la Selva, S Puerto Ayacucho,

2004 M. Balke HM588358 - HM588502 HM588573 HM588430

Hydraena monikae MNCN-AI455 South Africa W Cape 25.3.2001/Franschhoek, rd. R45/river 3 km SE Franschhoek

I. Ribera &

A. Cieslak HM588375 - HM588519 HM588580 HM588437

Hydraena paeminosa IBE-AN91 Suriname

SR10-0818-01A; Sipaliwini District; 2º19.280'N, 56º52.595'W, 224 m; Rapids on Kutari rRiver;

18.viii.2010

A. Short LR742634 - LR742654 LR742669 LR742693

Hydraena palawanensis MNCN-

AH133 Philippines

N Palawan, Taytay; Barangay, Manguao Stream, near sitio Danao, c.30m asl, 10°46'19''N 119°31'13'', 2007

H. Freitag HM588309 - HM588454 HM588546 HM588403

Hydraena palustris MNCN-AI309 Austria

02.07.2001 Voralberg / Pfänderstock, Bodensee.

Hörbranz vic. / Leiblach-Mündung / 400 m N47º32'01" E 9º43'46"

I. Ribera HM588342 - HM588486 HM588566 HM588423

Hydraena pygmaea MNCN-AI346 Austria

Nieder Österreich 6.8.2005 /Schwarzenbach, lat. rd.

18 W Sankt Veit an der Gölsen /448m N48º04’42.4”

E15º40’42.9”

I. Ribera &

Hydraena quadricollis IBE-RA323 Tanzania Nguru Mts. at Turiani, S06°06'24" E037°32'31",

30.x.2010, 933m, hand pick, V.V. Grebennikov HE970786 - HE971021 LR742670 HE970977

Hydraena scintillabella MNCN-AI688 Peru Lima. Huarochiri. Surco / Quebrada catarata Huanano / 11.09.2005

P. Aguilera &

Huamantinco HE970859 - HE971089 HE970969 HE971006

Hydraena testacea MNCN-AI566 Morocco 25.4.2000 / Oued Massa, Assif-Oumarhouz /

29º47'53"N 9º05'59"W / 1500m I. Ribera HM588386 - HM588530 HM588587 HM588444

Hydraena truncata MNCN-AI357 France Haute Provence 30.7.2005/rd. N202 2km N Moriez,

stream/834m N43º57’43.4” E6º26’14.4” I. Ribera &

Laeliaena sahlbergi MNCN-HI19 India

14.XI.2006 UTTARANCHAL 20: River Falda at Falda, ca. 15 km north of Bageshwar, Bageshwar District 1010 m a.s.l.; 29°55'11''N/79°52'08''E

M.A. Jäch LN995232 - LN995260 LN995286 LT991039

Limnebius cordobanus IBE-PA275 Spain Teruel, Beceite/r. Matarranya, El Parrisal/680m

N40º47’12.5” E0º12’13.5”/3.6.2006 I. Ribera HF931292 - HF931527 LN995319 LN995368

Limnebius myrmidon MNCN-AI703 Corsica (France) Corte, river Restonica, 1999 I. Ribera &

A. Cieslak LN995240 - LN995267 LN995293 LN995359

Limnebius nitidus MNCN-AI377 Spain

Barcelona 25.7.2005/La Pobla de Lillet. Te. de Solls/Rd. to S. Jaume. de Frontanyà/1055m N

42º12’26.2” E2º00”23.5” I. Ribera HE610238 - HF931415 LN995335 LN995392

Limnebius truncatellus MNCN-AI451 Spain

Avila, Puerto Pico de Gredos/N. side, rd. N502 pk 53, stream on left/1280m N40º20’14” W5º00’48”.

15.5.2005

I. Ribera HF931201 - HF931426 LN995338 LN995397

Limnebius wewalkai IBE-RA108 Oman 6.4.2010 J. Al-Akhdar, wadi Bani Awf residual pools, N23 13 42.9 E57 25 25.8 660m

I. Ribera, C. Hernando & A.

Cieslak

LN995241 - LN995269 LN995295 LN995365

Limnebius zaerensis MNCN-AC14 Morocco

10.4.2007/gorges Rd. Rommani-Ben Slimane/str. in Nerium, afl. Oued Korifla/265m N33º32'59.4"

W6º44'54.9"

I. Ribera, P. Aguilera &

C. Hernando

EU365868 -

EU365865+

EU365866+

EU365867

HE970937 HE970984

Asiobates aeneus MNCN-AI914 Morocco 24.4.2000 / Ou-Maghous, Amaghouz / 29º43'62.6"N 9º10'10.9"W / 600m

I. Ribera, P. Aguilera, C. Hernando & A.

Millán

HF931240 - HF931472 LT990865 LT991070

Asiobates dilatatus MNCN-AI792 Turkey

BARTIN 25.4.2006, Rd. to Topallar from Kakraz, calcareous stream in mixed forest, 25m

N41º45’50.5” E32º29’11”

I. Ribera HF931227 - HF931456 LT990849 LT991054

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Aulacochthebius exaratus MNCN-AI453 Tunisia 24.10.2001 /Rd. Jendouba-Makthar/O. 1 km NW cross to Jb. Kebbouch

I. Ribera &

A. Cieslak HF931202 - HF931427 LT990878 LT991085

Enicocerus aguilerai MNCN-AI387 Spain Avila, Arenas de S. Pedro/r. Pelayo, rd. AV924

/500m N40º12’06” W5º06’42”. 15.5.2005 I. Ribera GU143761 - GU143728 GU143770 GU143778

Ochthebius anxifer MNCN-AI945 Morocco 21.4.2001 /30º39’456”N 9º21’134”W /Immouzèr- des-Ida-Outanane/Assif Tanit, 550 m

I. Ribera &

Ochthebius gereckei MNCN-PA141 Sicily (Italy) Vallone Torto, Caltanissetta, 2007 P. Abellán &

F. Picazo FJ944176 - LT990803 LT990991 LT991204

Ochthebius

quadrifoveolatus MNCN-AI636 Gran Canaria (Spain)

15.4.2001 /Moya: Bco. de Azuaje/stream and ponds w. veget.

I. Ribera &

Ochthebius semisericeus MNCN-

AI1064 Spain Teruel, Beceite 7.8.2006/r. Matarranya, El

Parrisal/680m N40º47’12.5” E0º12’13.5” I. Ribera &

Ochthebius subpictus MNCN-AI452 France Bouches du Rhone/Salin du Girau 29.7.2005/0m N43º24’15.9” E4º45’07.5”

I. Ribera &

A. Cieslak HE970844 - HE971076 HE970957 HE970996

F. Leiodidae Anillochlamys

subtruncata NHM-IRC1 Spain Castellón, Montan, Cova Cirat, 1998 J. Fresneda GU356850 - GU356745 GU356904 GU356941

Catops subfuscus MNCN-AI550 France F-09, Illartein, Ravin de la Tire, MSS, 2004 Ph. Déliot &

A. Faille GU356861 - GU356757 GU356912 GU356951

Quaestus noltei NHM-IRC3 Spain Guipúzcoa, Olatz, Sima Kobeta, 1998 J. Fresneda HE572816 - HE576716 GU356928 HE572869

Speonemadus

angusticollis MNCN-AI551 Spain Jaén, Siles, Cueva Nacimiento de San Blas, 2002 M. Baena GU356889 - GU356787 GU356933 GU356980

Stygiophyes akarsticus NHM-IRC7 Spain Lleida, Altron, Forat del Tincatinc, 1999 J. Fresneda GU356899 - GU356798 GU356940 GU356990

F. Staphylinidae

Euaesthetus ruficapillus MNCN-AI831 England (UK) Moccas Park, Her. D. Lott LR742633 - LR742653 LR742668 LR742692

Micropeplus MNCN-AI521 Bhutan

Mo Chhu river, ca. 16 km NW of Punakha; Punakha Prov./Timphu Prov.; ca. 1450 m a.s.l.;

27°41'54''N/89°46'01''E; 28.XI.2005

M.A. Jäch HE572804 - HE576707 HE572837 HE572859

Lesteva MNCN-AI450 Spain

Avila, Puerto Pico de Gredos/N. side, rd. N502 pk 53, stream on left/1280m N40º20’14” W5º00’48”.

15.5.2005

I. Ribera HE572803 - HE576706 HE572836 HE572858

Osoriinae MNCN-AI647 Spain Córdoba, Sierras Sub-béticas, Carcabuey, suelo en

base de lentisco, 2006 A. Castro LR742641 - LR742658 LR742677 LR742699

Thinodromus MNCN-AI554 Switzerland Pfyn 2.8.2005 /stream in village/408m N47º35’43.8”

E8º57’17.4”

I. Ribera &

Paederus MNCN-AI552 Italy Liguria 31.7.2005 lat. rd SS28/2Km E Cosio di

Arriscia, stream/819m N44º04’41.6” E7º50’41.5” I. Ribera &

Phloeocharis diecki MNCN-AI880 Spain Guipuzkoa, Pagoeta, pista a Legarrola, 2006 C. Hernando HE572813 - HE576714 HE572842 HE572864

Scydmaenus insidiosus MNCN-AI646 Spain Córdoba, Cabra, 2006 A. Castro LR742649 - LR742662 LR742685 LR742706

Oiceoptoma thoracicum MNCN-AI837 England (UK) Norfolk, Ludham 6.5.2006 , How Hill Marsh I. Ribera LR742640 - LR742657 LR742676 LR742698

Thanatophilus MNCN-AI838 England (UK) Norfolk, Ludham 6.5.2006 , How Hill Marsh I. Ribera LR742651 - LR742663 LR742687 LR742707

Coprophilus striatulus MNCN-AI839 England (UK) Norfolk, Ludham 6.5.2006 , How Hill Marsh I. Ribera HE572812 - HE576713 HE572841 HE572863 Tachyporus hypnorum MNCN-AI555 Switzerland Pfyn 2.8.2005 /stream in village/408m N47º35’43.8”

E8º57’17.4” I. Ribera &

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Table S2. Primers used in the amplificantion and sequencing.

Gene Name Sense Sequence Ref.

COI Jerry (M202) F CAACATTTATTTTGATTTTTTGG 4 Pat (M70) R TCCA(A)TGCACTAATCTGCCATATTA 4 Chy F T(A/T)GTAGCCCA(T/C)TTTCATTA(T/C)GT 2 Tom R AC(A/G)TAATGAAA(A/G)TGGGCTAC(T/A)A 2 16S-tRNA-NAD1 16saR (M14) F CGCCTGTTTA(A/T)CAAAAACAT 4 ND1A (M223) R GGTCCCTTACGAATTTGAATATATCCT 4

16Sbi F ACATGATCTGAGTTCAAACCGG 4

FawND1 R TAGAATTAGAAGATCAACCAGC 4

12S 12Sai F AAACTAGGATTAGATACCCTATTAT 4

12Sbi R AAGAGCGACGGGCGATGTGT 4

18S 5’ F GACAACCTGGTTGATCCTGCCAGT 3

b5.0 R TAACCGCAACAACTTTAAT 3

28S Ka F ACACGGACCAAGGAGTCTAGCATG 1

Kb R CGTCCTGCTGTCTTAAGTTAC 1

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A B C

D E F

G H I J

Fig. S1. General views, SEM: A-C – Sindosium sp.; D, E – Nossidium pilosellum; F –

Motschulskium sinuatocolle; G,H – Limulodes parki; I, J – Cephaloplectus sp. Scale: A-H –

200 µm; I-J – 500 µm.

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A

B

C

D

E

F

Fig. S2. General views, SEM: A-C – Discheramocephalus sp.; D-F – Ptenidium pusillum.

Scale: A-C – 100 µm; D-E – 250 µm.

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A B C

D E

F

G H I

Fig. S3. General views, SEM: A-C – Mikado sp.; D, E – Porophila cedri; F – Scydosella

musawasensis; G-I – Primorskiella anodonta. Scale: A-C, G-I – 100; D-I – 150 µm; F – 50

µm.

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A B

C

D

E

Fig. S4. General views, SEM: A – Cissidium sp. B – Actidium boudieri; C- E – Oligella

foveolata. Scale: A,B – 200 µm; C-E – 150 µm.

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A B C

D E

F G H

Fig. S5. General views, SEM: A,B – Micridium vittatum; C-E – Ptilium myrmecophilum;

F-H – Ptiliola kunzei. Scale: 200 µm.

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A

B

C D

E F G

I J H

Fig. S6. General views, SEM: A, B – Smicrus filicornis; C,D – Acrotrichis grandicollis; E-G –

Baeocrara variolosa; H-J – Nephanes titan. Scale: A-G – 200 µm; H-J – 150 µm.

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A

B

C D E

F G

Fig. S7. General views, SEM: A,B – Pteryx suturalis; C-E – Ptinellini gen.; F-G – Rioneta

uluguruensis. Scale: 200 µm.

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TABLE S3. Data matrix of 68 morphological characters scored for 33 terminals and used for the phylogenetic analysis of Ptiliidae beetles

10 20 30 40 50 60 68

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0000000000 0000000000 0000000000 000000000? 0000000000 000000-0-0 0-00??10 0100000000 0000000000 0000000002 000000000? 0000000000 00000000-0 0-000?00 0001012011 1000100000 0200230102 000100000? 1000000000 00000000-0 10012000 0001012011 1000100000 0200200100 0001000000 1000000000 00000000-0 10010000 0110000121 1010101010 1000100000 0000001000 1000011000 1011100110 10011000 0110000121 1010101010 1101100001 0000001000 1000011000 1011100110 10011000 0110000121 1010101010 1000000001 0010001000 1000011000 1011100110 10011000 0110000121 1010101110 1100110000 0002400020 2010011000 1121211220 10012020 0110000121 1010201110 1201100002 0002100020 2010011000 1121211320 10011021 0110000121 1010101110 1001100002 0002100020 2010011000 1121211320 10113020 0110000121 1011201110 1201100002 0002100110 2010011000 1121211320 10014021 0110000121 1010101110 1101100002 0002100010 2010011000 1121211320 10113020 0110000121 1010101110 1201000002 0002100110 2010011000 1121211320 10114021 0110000121 1010201110 1101120002 0002100110 2010011000 1121211320 10114021 0110100121 1010101110 1101100002 1002100020 2010011001 1121211320 10013020 0110100121 1010101110 1001100002 1002100020 2010011001 1121211320 10013020 0110100121 1010101110 1101100002 1002100020 2010011001 1121211320 10013020 0110100121 1010101110 1001100002 1002100020 2010011002 ---0-0 10014020 0110000121 1010101110 1200000002 0012100020 2010011000 1121211320 10011020 0110000121 1010101110 1101120002 0012400000 2000011000 1121211320 10012020 0110000121 1010101110 1100010002 0012100020 2010011000 1121211320 10011020 0110000121 1110301111 1001100012 0002500031 2020011000 1121221220 10015020 0110000121 1110301111 1001100012 0002300231 2020011000 1121222220 10215020 0110001121 2110311211 1001100002 0102300241 2020011010 1121222220 10215020 0110001121 2110311211 1101100002 0102300241 2021011010 1121222220 10215020 0110001121 2110311211 1101100002 0102200241 2020011010 1121222220 10215020 0110000121 1010101110 1001100002 1002100020 2110011000 1121211320 10316122 0110000121 1010101110 1101100002 1002100020 2110011000 1121211320 10317122 0110000121 1010101110 1001100002 1002100020 2110011000 1121211320 10317122 Necrophilus

Catops Ochthebius Hydraena Nossidium Motschulskium Sindosium Ptenidium Actidium Ptiliola Oligella Ptilioloum Micridium Ptilium Ptinella Pteryx

Ptinellini sp.

Rioneta Dacrysoma

Discheramocephalus Cissidium

Mikado Porophila Primorskiella Scydosella

Cylindroselloides Nephanes

Smicrus Baeocrara

Acrotrichis intermedia 0110000121 1011101110 1001100002 1002100020 2110011000 1121211320 10317122

Acrotrichis grandicollis 0110000121 1011101110 1001100002 1002100020 2110011000 1121211320 10317122

Limulodes 1110221121 1110101010 1011100003 1002410020 1030111002 ---0-1 11014020

Cephaloplectus 1110221231 1110101010 1011101003 1000410020 1030111102 ---0-1 11014020

(15)

0.06 Ochthebius gereckei PA141 Ochthebius subpictus AI452

Osoriinae AI647

Cissidium AN362

Baeocrara variolosa AN416

Euaesthetus AI831 Ptenidium formicetorum GBK14

Coprophilus AI839 Thinodromus AI554 Ochthebius anxifer AI945

Thanatophilus AI838

Micridium vitalum AN427

Limnebius truncatellus AI451

Rioneta uluguroensis AI415

Asiobates dilatatus AI792

Scydmaenus AI646

Scydosella musawasensis AN424

Hydraena truncata AI357

Oiceoptoma AI837

Cylindroselloides dybasi AI564

Hydraena testacea AI566

Limnebius nitidus AI377

Ochthebius semisericeus AI1064 Laeliaena sahlbergi HI19

Limnebius wewalkai RA108

Discheramocephalus AN361

Limnebius myrmidon AI703

Ochthebius quadrifoveolatus AI636

Primorskiella anodonta AN418 Scydosella musawasensis GBK27

Pteryx suturalis GBK23

Stygiophies IRC7

Ptinella aptera GBK25

Adelphydraena orchymonti AI356

Acrotrichis atomaria GBK2

Hydraena paeminosa AN91

Acrotrichis xanthocera GBK9

Speonemadus angusticollis AI551

Rioneta uluguroensis AN360

Ptenidium fuscicorne GBK15

Aulacochthebius exaratus AI453 Motschulskium AN357

Smicrus filicornis AN423 Acrotrichis insularis GBK4

Hydraena scintillabella AI688

Asiobates aeneus AI914 Nossidium sp GBK13

Porophila cedri AN417 Mikado AN419

Quaestus IRC3 Anillochlamys IRC1

Catops tristis AI550

Ptenidium sp GBK22

Dacrysoma AN359

Lesteva AI450

Paederus AI552 Hydraena palawanensis AH133

Phloeocharis AI880 Micropeplus AI521

Limnebius zaerensis AC14 Hydraena monikae AI455

Tachyporus AI555

Hydraena pygmaea AI346

Hydraena quadricollis nr RA323

Limnebius cordobanus PA275

Enicocerus aguilerai AI387 Hydraena palustris AI309

Nephanes titan AN41 5 Ptinellini AN356

Ptinella AN439

Nanosellini AI414

Acrotrichis AI413

100 83

86

100

80

97

90

52

55

78 Limulodes parki AN819 GBK Ptilium myrmecophilum AN426 Ptenidium AI515

100 Ptenidium pusillum GBK21

100 98 100

100 100 99

70

77

100

67

72

89

99

58

69 100 97

100

84 77 73

59

100

55

98

100 92 35

97 92 100

96 89 Acrotrichis intermedia GBK5

Acrotrichis sitkaensis GBK7

81

99

47

100

73

88

69

32

85

57

100

93 Figure S8. Maximum Likelihood inference phylogram of

Ptiliidae obtained with RAxML with the molecular data only.

Digits at internodes

are bootstrap values

when higher than

50%.

(16)

0 5 0

100 150

200 Ma

Acrotrichis AI413

Micropeplus AI521

Hydraena pygmaea AI346 Motschulskium AN357

Paederus AI552

Limnebius nitidus AI377

Anillochlamys IRC1 Ptenidium sp GBK22

Quaestus IRC3

Porophilla cedri AN417 Mikado AN419

Stygiophies IRC7

Ochthebius quadrifoveolatus AI636 Cissidium AN362

Enicocerus aguilerai AI387 Hydraena quadricollis nr RA323 Acrotrichis xanthocera GBK9 Baeocrara variolosa AN416

Hydraena palawanensis AH133

Asiobates dilatatus AI792

Thinodromus AI554 Oiceoptoma AI837

Hydraena monikae AI455 Ptinella AN439

Catops tristis AI550

Limnebius cordobanus PA275 Ptenidium fuscicorne GBK15

Ochthebius gereckei PA141

Asiobates aeneus AI914 Limulodes parki AN819 GBK

Hydraena palustris AI309

Ochthebius anxifer AI945 Acrotrichis intermedia GBK5

Laeliaena sahlbergi HI19

Aulacochthebius exaratus AI453 Ptinellini AN356

Ptinella aptera GBK25

Ochthebius semisericeus AI1064

Ochthebius subpictus AI452 Scydosella musawasensis AN424

Ptenidium pusillum GBK21 Rioneta uluguroensis AN360

Nanosellini AI414

Coprophilus AI839

Ptenidium formicetorum GBK14

Hydraena truncata AI357 Ptilum myrmecophilum AN426

Euaesthetus AI831

Discheramocephalus AN361

Nossidium sp GBK13 Acrotrichis sitkaensis GBK7

Smicrus filicornis AN423

Pteryx suturalis GBK23

Lesteva AI450

Limnebius zaerensis AC14 Rioneta uluguroensis AI415

Micridium vitalum AN427

Hydraena testacea AI566

Adelphydraena orchymonti AI356 Acrotrichis insularis GBK4

Dacrysoma AN359

Scydosella musawasensis GBK27

Speonemadus angusticollis AI551 Limnebius wewalkai RA108

Tachyporus AI555 Ptenidium AI515

Cylindroselloides dybasi AI564

Thanatophilus AI838

Hydraena scintillabella AI688

Scydmaenus AI646

Hydraena paeminosa AN91

Limnebius myrmidon AI703

Phloeocharis AI880 Nepanes titan AN415

Acrotrichis atomaria GBK2

Limnebius truncatellus AI451 Primorskiella anodonta AN418

Osoriinae AI647

1 0.77

1

0.98

1

0.98 0.49 0.37

0.68 1 0.91

1

0.98

1

1 1 0.48

1 1

0.73

1

1 1 1

1

0.38

1

0.52

0.99

0.89

0.77

0.89 1 1 0.54

0.79

1

0.38

1 1 1

1

1 1 1

0.61

1

0.58 1 0.98

1 0.96

1

0.99

0.99 1 1

0.99

0.99 0.84 0.98

1

0.46 1 1

0.39

1 1 0.95

1 1

0.51

1 0.77 113.4

175.1

109.5

32.2

123.36 17.4 96.4

70.7 30.3 48.5

32.3 102.2

54.8 122.9 33.8

0

45.8

141.8

157.2 80.7 72.8

47.7 43.5

118.7

93.0

187.8

47.6 61.5 131.0

28.3

148.6

110.1 112.9 110.7

57.8 81.7

117.8

132.6

60.2

103.6

14.4

79.6

64.3

27.3 21.7 96.2

0.15 57.7 60.0

93.7 42.0 39.2

39.2 59.5

12.9

46.2 38.1 32.9

83.0 100.5

131.6 150.5

51.6

1.9 37.7 71.9

133.9 43.4 100.8

145.5

154.6 0.37 55.8

164.8 79.4

57.9 Figure S9. Majority rule consensus tree of the Bayesian analysis (obtained with BEAST) of the molecular characters, with the simplest models (analysis C, see Methods).