Plant Systematics and Evolution

, Volume 285, Issue 3, pp 209–232

Closing the gaps: phylogenetic relationships in the Brassicaceae based on DNA sequence data of nuclear ribosomal ITS region

Authors

    • Eastern Cereal and Oilseed Research CentreAgriculture and Agri-Food Canada
  • Klaus Mummenhoff
    • FB Biologie/Chemie, BotanikUniversität Osnabrück
  • Connie A. Sauder
    • Eastern Cereal and Oilseed Research CentreAgriculture and Agri-Food Canada
  • Marcus A. Koch
    • Heidelberg Institute of Plant Sciences, Biodiversity and Plant SystematicsUniversity of Heidelberg
  • Ihsan A. Al-Shehbaz
    • Missouri Botanical Garden
Original Article

DOI: 10.1007/s00606-010-0271-8

Cite this article as:
Warwick, S.I., Mummenhoff, K., Sauder, C.A. et al. Plant Syst Evol (2010) 285: 209. doi:10.1007/s00606-010-0271-8

Abstract

Sequence data from the nuclear encoded ribosomal internal transcribed spacer (ITS) region were used to determine monophyly of tribes, tribal limits, and tribal relationships of 96 so far unassigned or tentatively assigned genera (represented by 101 taxa/accessions) within the Brassicaceae. Maximum-parsimony and maximum-likelihood analyses of 185 ITS Brassicaceae sequences, which also included representatives of each of the 34 currently recognized tribes, supported the separate phylogenetic distinctness of these tribes and permitted the tribal assignment of all but 12 of the unassigned genera into tribal clades. The data support the recognition of eight new, well-resolved, uni- or oligogeneric tribes recognized herein as the Alyssopsideae [96% bootstrap support (BS); including the central and southwestern Asian Alyssopsis and Calymmatium], Asteae (100% BS; including the Mexican Asta), Eudemeae (97% BS; South American Brayopsis, Eudema, and Xerodraba), Kernereae (96% BS; European Kernera and Rhizobotrya), Notothlaspideae (100% BS; New Zealandic Notothlaspi), Oreophytoneae (100% BS; eastern African Oreophyton and southern European Murbeckiella), and Yinshanieae (100% BS; Chinese Yinshania), as well as the moderately supported Microlepidieae (75% BS; Australian Microlepidium and Carinavalva). Furthermore, the results fully support the recent findings that the tribes Schizopetaleae and Thelypodieae ought to be recognized as two distinct tribes instead of a single tribe, as well as provide some support for the re-establishment of the tribe Cremolobeae, bringing the total number to 44 tribes in the family. Nearly 92% (308) of the 336 genera in the family have been assigned to a tribe. The earlier-published Anastaticeae is taken here to replace the Malcolmieae.

Keywords

BrassicaceaeITS regionPhylogenySystematicsTribal assignments

Introduction

The Brassicaceae is a large family of ca. 338 genera and over 3,700 species distributed throughout the world, primarily in temperate and alpine regions (Al-Shehbaz 1984; Al-Shehbaz et al. 2006; Warwick et al. 2006b). It is a natural family, easily distinguished by floral and fruit morphology, i.e., cruciform corolla, tetradynamous stamens, and characteristic siliques. Tribal classification of the family, however, has long been problematic and not well understood phylogenetically. The characters traditionally used in the delimitation of tribes are few and include orientation of the radicle in relation to the cotyledons in the embryo, fruit length-to-width ratio, fruit compression and dehiscence, number of rows of seeds in each locule, trichome type, and features of the nectaries. Five tribal systems were proposed in the past century (Hayek 1911; Schulz 1936; Janchen 1942; Avetisian 1983; Al-Shehbaz et al. 2006), and with the exception of the Brassiceae, few if any traditional tribes have proven monophyletic (Hedge 1976; Al-Shehbaz 1984; Koch et al. 2003a; Al-Shehbaz et al. 2006). Thirty-four tribes are currently recognized in the family (Al-Shehbaz et al. 2006; Al-Shehbaz and Warwick 2007; German and Al-Shehbaz 2008; Koch and Al-Shehbaz 2009; Warwick et al. 2009) on the basis of molecular phylogeny and re-evaluation of morphological features. Twenty-five of these tribes were established primarily on the ndhF-based phylogeny of Beilstein et al. (2006), and most of them were supported in ITS-based phylogenetic studies (reviewed in Bailey et al. 2006), the mitochondrial DNA-based study of Franzke et al. (2009) and multigene supernetworks (Koch et al. 2007), while the additional nine tribes are based on subsequent ITS-based phylogenetic studies and larger taxon sampling that included many Asian and New World genera not previously studied (Al-Shehbaz and Warwick 2007; German and Al-Shehbaz 2008; Khosravi et al. 2009). The majority of the tribes are in the Old World, two tribes (Cardamineae and Lepidieae) are reported from all continents except Antarctica, whereas the Boechereae, Halimolobeae, Physarieae, Thelypodieae, and Schizopetaleae are exclusively or almost exclusively New World tribes.

The ITS region is currently the most widely used marker in phylogenetic analyses of the Brassicaceae (Heenan et al. 2002; Koch 2003; Koch and Mummenhoff 2001; Koch et al. 2003a; Lihová et al. 2006a; Mummenhoff et al. 2005, 2009; Warwick et al. 2002, 2004a, 2004b, 2006a, 2006c, 2007, 2008, 2009; Warwick and Sauder 2005; Warwick and Hall 2008; Bailey et al. 2006). Hence, this paper utilizes this general advantage of the marker system to determine the monophyly status and tribal affiliations of 96 genera that either had not been included in earlier molecular studies or were studied but whose tribal assignments remained unresolved within the family. This analysis closes most of the generic gaps in Brassicaceae phylogeny and systematics. Almost two-thirds of the 338 genera have been previously studied for ITS sequences (Supplementary Table 1). Herein, sequences of the internal transcribed spacers of nuclear ribosomal DNA and the 5.8S rRNA gene (collectively, the ITS region) of 96 previously not assigned genera (101 species/accessions) are included in the analysis, along with representatives of all 34 currently recognized tribes, for a total of 164 genera and 185 taxa/accessions. The sampling covers at least two core species/representatives of each actually accepted or proposed tribe in order to provide a comprehensive tribal framework.
Table 1

Taxa sequences, collector, and collection number for each voucher specimen, geographical origin of the species, and GenBank numbers for ITS

Taxon

Herbarium source (Collector No. or date; (herbarium code) or Citation)

Geographical origin

ITS GenBank accession no.

Not assigned to tribe

 Alyssopsis mollis (Jacq.) O.E. Schulz

Rechinger et al. 5419; (MO)

Iran

GQ497845

 Alyssopsis trinervis Botsch. & Sejfulin

Sejfulin; 08.10.1979; (MO)

Turkmenistan

GQ497846

 Ammosperma cinerea (Desf.) Hook.f.

Lambinon & Margot 99/152; (B)

Tunisia

GQ424606

 Anastatica hierochuntica L.

Neuffer & Hurka; 03.03.1989; (OSBU)

Egypt

GQ424524

 Aplanodes doidgeana Marais

Hilliard & Burrt 17903;13.12.1984; (E)

South Africa

GQ497847

 Arcyosperma primulifolium (Thomson)  O.E. Schulz

Polunin, Sykes & Williams; 16.06.1952; (A)

Nepal

GQ424525

 Asperuginoides axillaris (Boiss. & Hohen.) Rauschert

Warwick et al. (2008)

EF514626

 Asta schaffneri (S. Watson) O.E. Schulz (received as Asta_fendlerii_1)

Fuentes-Soriano 48; (MO)

Mexico

GQ497848

 Asta schaffneri var. pringlei (O.E. Schulz) Rollins

Bridges & Woodruff 13113; (TEX)

Mexico

GQ424526

 Baimashania pulvinata Al-Shehbaz

Al-Shehbaz & Yue 20026; (MO)

China

GQ497849

 Bivonaea lutea (Biv.) DC.

Plantae florae Siculae, leg. Citarda 1860 (JE 2007-0002)

Italy, Sicily

GQ497850

 Boreava aptera Boiss. & Heldr.

INIA 78-1811-70; (HEID)

Algeria

GQ497851

 Brossardia papyracea Boiss.

Grant 17710; (MO)

Iran

GQ497852

 Callothlaspi lilacinum (Boiss. & A. Huet) F.K. Mey.

Sinter: Iter orientale 1890, No. 2278; 15.05.1890; (JE)

Turkey

GQ497853

 Calymmatium draboides (Korsh.) O.E. Schulz

Anders 8130; 06.08.1971; (E)

Afghanistan

GQ497854

 Carinavalva glauca Ising

Symon s.n.; (CANB 257161)

Australia

GQ424527

 Catenulina hedysaroides (Botsch.) Soják

Kinzikaeva & Junussov; 18.04.1978; (B)

Tajikistan

GQ424607

 Chalcanthus renifolius (Boiss. & Hohen.) Boiss.

Zarre & Moazzeni; 27.05.2005; (TUH)

Iran

GQ424528

 Chartoloma platycarpum (Bunge) Bunge

Nikitin s.n.; 21.04.1954; (WAG)

Turkmenistan

GQ424529

 Chaunanthus acuminatus (Rollins) R.A. Price & Al-Shehbaz

Sanders, Daniel & Phillips; 21.05.1991; (GH)

USA, California

GQ497855

 Chlorocrambe hastata (S. Watson) Rydb.

Tuhy; 10.08.1985; (GH)

USA, Utah

GQ497856

 Chrysochamela velutina (DC.) Boiss.

Koch et al. (2007)

DQ249856

 Coluteocarpus vesicaria (L.) Holmboe

Gagnidze & Shetekauri 256; (MO)

Georgia

GQ497857

 Cremolobus chilensis (Lag. ex DC.) DC.

Weigend et al. 8426; (Herbarium Mummenhoff)

Peru

GQ424530

 Cymatocarpus pilosissimus (Trautv.) O.E. Schulz

Kurbanov 1649; (MO)

Turkmenistan

GQ497858

 Cyphocardamum aretioides Hedge

Kasy 28; 08.06.1965; (E)

Afghanistan

GQ497859

 Delpinophytum patagonicum (Speg.) Speg.

Anegliezo s.n.; 1899; (LP)

Argentina, Patagonia

GQ497887

 Desideria flabellata (Regel) Al-Shehbaz [=Solms-laubachia (Regel) Yue et al. (2008]

Neuffer, Hurka & Friesen; 01.08.2004; (OSBU)

Kyrgyzstan

GQ497886

 Didesmus aegyptius (L.) Desv.

Raus & Schiers 16349; (B)

Greece

GQ424531

 Didesmus bipinnatus (Desf.) DC.

INIA 1853-70; (DAO)

Algeria

GQ497860

 Dielsiocharis kotschyi (Boiss.) O.E. Schulz

Collector unknown; 25.4.1997; (TUH)

Iran

GQ424532

 Dipoma iberideum Franch

Forrest 28853; (E)

China

GQ497861

 Douepea tortuosa Cambess

Lamond 1507; 21.05.1965; (E)

Pakistan

GQ497862

 Eigia longistyla (Eig.) Soják (=Stigmatella longistyla Eig.)

Frey & Kürschner 80-576; 13.04.1980; (E)

Jordan

GQ497863

 Elburzia fenestrata (Boiss. & Hohen.) Hedge

Collector unknown; 1977; (TUH)

Iran

GQ424533

 Eremodraba intricatissima (Phil.) O.E. Schulz

Werdermann 731; (E)

Chile

GQ424534

 Eremophyton chevallieri (Barrate ex L. Chevall.) Bég.

Staudinger 6619; (OSBU)

Morocco

GQ424535

 Eunomia andersonii (Hook.f. & Thomson) Al-Shehbaz (in press)

Ludloff, Sherriff & Elliott 15191; 08.06.1947; (E)

China, Tibet

GQ497864

 Eunomia oppositifolia (Pers.) DC

Al-Shehbaz s.n.; (MO)

GQ497865

 Fezia pterocarpa Pit. ex Batt.

INIA 1459-68; (Herbarium Mummenhoff)

Morocco

GQ424536

 Foleyola billotii Maire

INIA 1481-68; (DAO)

Morocco

GQ497866

 Gagria lobata M. Král (=Pachyphragma macrophyllum (Hoffm.) N. Busch)

Mummenhoff et al. (2001)

AF283488 and AF283489

 Heldreichia bupleurifolia Boiss.

Mummenhoff et al. (2005)

AJ628291 and AJ628292

 Henophyton deserti (Coss. & Durieu) Coss. & Durieu

Podlech 34139; (WAG)

Algeria

GQ424537

 Hesperidanthus jaegeri (Rollins) Al-Shehbaz

Beilstein 01-74; (MO)

USA, California

GQ424569

 Hesperidanthus suffrutescens (Rollins) Al-Shehbaz

Beilstein 01-54; (MO)

USA, Utah

GQ424567

 Horwoodia dicksoniae Turrill

Rechinger 157; (WAG)

Iraq

GQ424538

 Idahoa scapigera (Hook.) A. Nelson & J.F. Macbr.

Baum 365; (A)

USA, Washington

GQ497867

 Iodanthus pinnatifidus (Michx.) Steud.

Beilstein 01-01; (MO)

USA

GQ424539

 Kernera saxatilis (L.) Sweet

Koch and Heenan, unpublished

AF401118 and AF401119

 Kotschyella stenocarpa (Boiss.) F.K. Mey.

Zarre, Moazzeni, & Fritsch; 04.06.2005; (TUH)

Iran

GQ497888

 Lachnocapsa spathulata Balf.f.

Kilian, Hein & Kürschner YP3719; (B)

Yemen

GQ424540

 Lachnoloma lehmannii Bunge

Dahaghin; 20.05.1987; (TUH)

Iran

GQ497889

 Leavenworthia crassa Rollins

Beck 512; (MO)

USA, Alabama

GQ424541

 Lepidostemon glaricola (H. Hara) Al-Shehbaz

Miehe & Miehe 00-185-07; (MO)

Bhutan

GQ424542

 Lithodraba mendocinensis (Hauman) Boelcke

Boelcke et al. 11121; (US)

Argentina

GQ497890

 Lunaria redeviva L.

Neuffer & Hurka; 17.05.2002; (OSBU)

Germany

GQ424543

 Megadenia pygmaea Maxim.

Makryi & Kazanovski; 16.08.2002; (OSBU)

Russia, Siberia

GQ424544

 Menonvillea cuneata (Gillies & Hook.) Rollins

Hellwig 10026; (OSBU)

Chile

GQ497868

 Microlepidium pilosulum F.Muell.

Murfet 3924; (Herbarium Mummenhoff)

Australia

GQ497869

 Moriera spinosa Boiss.

Kurbanov 403; (MO)

Turkmenistan

GQ424545

 Murbeckiella huetii (Boiss.) Rothm.

Neuffer; 24.07.1999; (OSBU)

Russia, Caucasus

GQ424546

 Myagrum perfoliatum L.

Iranian-American Exped.; 02.05.2004; (TUH)

Iran

GQ424547

 Nasturtiopsis coronopifolia (Desf.) Boiss.

Danin et al. 17.013; (B)

Israel

GQ424548

 Noccidium hastulatum (Steven) F.K. Mey.

Koch and Mummenhoff (2001)

AF336164 and AF336165

 Ochthodium aegyptiacum (L.) DC.

Alziar et al. 1134; (B)

Cyprus

GQ497870

 Octoceras lehmannianum Bunge

Collector unknown; 5.05.2002; (TUH)

Iran

GQ424609

 Oreophyton falcatum (Hochst ex A. Rich) O.E.Schulz

Harmsen 3; (WAG)

Kenya

GQ424549

 Ornithocarpa torulosa Rollins

Bye; 23.07.1977; (G)

Mexico

GQ424550

 Pachypterygium brevipes Bunge

Rechinger 56266; (B)

Iran

GQ424551

 Parlatoria rostrata Boiss. & Hohen.

Iranian-American Exped.; 26.05.2004; (TUH)

Iran

GQ424552

 Pegaeophyton scapiflorum (Hook.f. & Thomson) C. Marquand & Airy Shaw

Koch et al. (2007)

DQ518398

 Peltariopsis planisiliqua (Boiss.) N. Busch

Rauss 4231; (B)

Turkey

GQ424553

 Petrocallis pyrenaica (L.) W. T. Aiton

Koch 2001; (HEID)

Austria

GQ497871

 Phaeonychium jafrii Al-Shehbaz [=Solms-laubachia jafrii (Al-Shehbaz) J.P. Yue, Al-Shehbaz & H. Sun; Yue et al. (2008)]

Miehe & Miehe; 03-056-30; (MO)

China, Tibet

GQ497872

 Phlebolobium maclovianum (d’Urv.) O.E. Schulz

Nürnberger s.n.; (HEID)

Falkland Islands

GQ497873

 Phoenicaulis cheiranthoides Nutt.

Hurka; 27.05.1986; (OSBU)

USA, California

GQ497874

 Physocardamum davisii Hedge

A. Tali; (E 5806)

Turkey

GQ497875

 Planodes virginicum (L.) Greene

Slemmer 34; (MO)

USA, Missouri

GQ424554

 Pseuderucaria teretifolia (Desf.) O.E. Schulz

Neuffer, 02.03.1989; (OSBU)

Egypt

GQ497891

 Pseudofortuynia esfandiarii Hedge

INIA 3760-75 (Herbarium Mummenhoff)

Iran

GQ497876

 Pseudosempervivum aucheri (Boiss.) Pobed. (as Cochlearia aucheri Boiss.)

Koch and Mummenhoff (2001)

AF33620

 Pseudovesicaria digitata (C.A. Mey.) Rupr.

Gagnidze & Shetekauri 808; (MO)

Georgia

GQ497877

 Pycnoplinthopsis bhutanica Jafri

Miehe & Miehe 00-134-05; (MO)

Bhutan

GQ497878

 Pycnoplinthus uniflora (Hook.f. & Thomson) O.E. Schulz

Miehe & Miehe 9399/01; (MO)

China

GQ497879

 Rhizobotrya alpina Tausch

Kropf et al. (2003)

AJ440315

 Ricotia cretica Boiss. & Heldr.

Hurka & Neuffer; 16.05.2005; (OSBU)

Greece, Crete

GQ497880

 Robeschia schimperi (Boiss.) O.E. Schulz

Furse 1769; (E)

Iran

GQ497881

 Sameraria nummularia Bornm.

INIA 3776-75; (Herbarium Mummenhoff)

Iran

GQ424555

 Sarcodraba dusenii (O.E. Schulz) Al-Shehbaz

Arroyo et al.; 20.01.77; (BAA)

Argentina

GQ424568

 Schimpera arabica Hochst. & Steud.ex Steud.

Rawi et al. 239; (ETH)

Kuwait

GQ424556

 Selenia dissecta Torr. & A. Gray

Worthington 28283; (OSBU)

USA, New Mexico

GQ424557

 Shangrilaia nana Al-Shehbaz, J.P. Yue & H. Sun

Yue 0366; (MO)

China

GQ424558

 Sibaropsis hammittii S. Boyd & T.S. Ross

Boyd & Ross 6775; 01.04.1992; (GH)

USA, California

GQ424559

 Sisymbrella aspera (L.) Spach

Valdes et al. 2925/88; (B)

Spain

GQ424560

 Sobolewskia caucasica N. Busch

Merello, Schmidt & Chibooshvili 2114; (MO)

Georgia

GQ424561

 Spryginia winkleri (Regel) Popov

Kurbanov 1945; (MO)

Turkmenistan

GQ424563

 Subularia monticola A. Braun ex Schweinf.

Katende K3293; (MO)

Uganda

GQ424565

 Tchihatchewia isatidea Boiss. [=Neotchihatchewia isatidea (Boiss.) Rauschert]

Rix & Martyn 1589; 05.05.1970; (E)

Turkey

GQ497882

 Tropidocarpum gracile Hook.

Ross; 21.03.1997; (OSBU)

USA, California

GQ497883

 Xerodraba pectinata (Speg.) Skottsb.

Arroyo et al. 841013; (CONC)

Argentina

GQ497884

Tribes or major ITS clades

 Aethionemeae (AETH)

  Aethionema arabicum (L.) Andrz. ex O.E. Schulz

Hong et al. (2003)

AY254539

  Aethionema grandiflorum Boiss. & Hohen.

Koch et al. (2007)

DQ249867

 Alysseae (ALYS)

  Alyssum montanum L.

Mengoni et al. (2003)

AY237938

  Berteroa incana (L.) DC.

Warwick et al. (2008)

EF514632

 Anchonieae (ANCH)

  Anchonium billardieri DC.

Warwick et al. (2007)

DQ357512

  Matthiola incana (L.) W.T. Aiton

Mummenhoff et al., unpublished

AJ628339 and AJ628340

  Matthiola sp.a

Hindukusch expedition; 35 Herbar Baschant acc. 1959; (B)

Afghanistan

GQ424566

 Aphragmeae (APHR)

  Aphragmus eschscholtzianus Andrz.

Warwick et al. (2006a)

DQ165334

  Aphragmus hobsonii (H. Pearson) Al-Shehbaz & S.I. Warwick

Warwick et al. (2006a)

DQ165357

 Arabideae (ARAB)

  Arabis alpina L.

O’Kane and Al-Shehbaz (2003)

AF137559

  Aubrieta deltoidea (L.) DC.

Koch et al. (1999b)

AJ232909

  Draba aizoides L.

Koch and Al-Shehbaz (2002)

AF146512

 Biscutelleae (BISC)

  Biscutella didyma L.

Bailey et al. (2006)

DQ452058

  Biscutella laevigata L.

Bailey et al. (2006)

DQ452056

 Boechereae (BOEC)

  Boechera retrofracta (Graham) A. Löve & D. Löve

Roy (2001)

AF183105

  Sandbergia whitedii (Piper) Greene

Mummenhoff et al., unpublished

AJ628295 and AJ628296

 Brassiceae (BRAS)

  Brassica oleracea L.

Warwick and Sauder (2005)

AY722423

  Cakile maritima Scop.

Warwick and Sauder (2005)

AY722494

  Crambe maritima L.

Francisco-Ortega et al. (1999)

AF039970 and AF040013

  Hirschfeldia incana (L.) Lagr.-Foss.

Warwick and Sauder (2005)

AY722470

 Buniadeae (BUNI)

  Bunias erucago L.

Koch, unpublished

GQ497885

  Bunias orientalis L.

Koch et al. (2007)

DQ249863

 Calepineae (CALE)

  Calepina irregularis (Asso) Thell.

Koch et al. (2007)

DQ249822

  Goldbachia laevigata (M.Bieb.) DC.

Warwick et al. (2007)

DQ357545

 Camelineae (CAME)

  Camelina microcarpa Andrz. ex DC.

O’Kane and Al-Shehbaz (2003)

AF137574

  Pseudoarabidopsis toxophylla (M.Bieb.) Al-Shehbaz, O’Kane & R.A. Price

O’Kane and Al-Shehbaz (2003)

AF137558

 Cardamineae (CARD)

  Cardamine pratensis L.

Sheridan et al., unpublished

X98637

  Nasturtium officinale W.T. Aiton

Hong et al. (2003)

AY254531

 Chorisporeae (CHOR)

  Chorispora tenella (Pall.) DC.

Warwick et al. (2007)

DQ357526

  Diptychocarpus strictus (Fisch. ex M. Bieb.) Trautv.

Warwick et al. (2007)

DQ357534

 Cochlearieae (COCH)

  Cochlearia megalosperma (Maire) Vogt

Koch and Mummenhoff (2001)

AF336208 and AF336209

  Ionopsidium acaule (Desf.) DC. ex Rchb.

Koch and Mummenhoff (2001)

AF336210 and AF336211

 Conringieae (CONR)

  Conringia clavata Boiss. [as C. perfoliata (C.A. Mey.) N. Busch]

Warwick and Sauder (2005)

AY722505

  Zuvanda crenulata (DC.) Askerova

Warwick et al. (2007)

DQ357606

 Descurainieae (DESC)

  Descurainia sophia (L.) Webb ex Prantl

Goodson et al. (2006)

DQ418727

  Hornungia petraea (L.) Rchb.

Mummenhoff et al., unpublished

AJ628293 and AJ628294

 Dontostemoneae (DONT)

  Clausia aprica (Stephan) Korn.-Trotzky

Warwick et al. (2007)

DQ357529

  Dontostemon integrifolius (L.) Ledeb.

Warwick et al. (2007)

DQ357536

 Erysimeae (ERYS)

  Erysimum capitatum (Douglas ex Hook.) Greene

Warwick et al. (2007)

DQ357540

  Erysimum cheiranthoides L.

Kress et al. (2005)

DQ005989

 Euclidieae (EUCL)

  Braya alpina Sternb. & Hoppe

Warwick et al. (2004a)

AY353095

  Neotorularia torulosa (Desf.) Hedge & J. Léonard

Warwick et al. (2004a)

AY353164

 Eudemeae (EUDE-NEW)

  Brayopsis colombiana Al-Shehbaz

Warwick et al. (2009)

EU620283

  Eudema nubigena subsp. nubigena Humb. & Bonpl.

Warwick et al. (2009)

EU620297

 Eutremeae (EUTR)

  Eutrema altaicum (C.A. Mey.) Al-Shehbaz & S.I. Warwick

Warwick et al. (2006a)

DQ165364

  Eutrema edwardsii R.Br.

Warwick et al. (2006a)

DQ165350

 Halimolobeae (HALI)

  Halimolobos lasiolobus (Link) O.E. Schulz

Bailey et al. (2002)

AF307647

  Pennellia longifolia (Benth.) Rollins

Bailey et al. (2002)

AF307627

 Heliophileae (HELI)

  Chamira circaeoides (L.f) Zahlbr.

Mummenhoff et al. (2005)

AJ862719 and AJ862720

  Heliophila arenaria Sond.

Mummenhoff et al. (2005)

AJ863600 and AJ864811

  Heliophila subulata Burch. ex DC.

Mummenhoff et al. (2005)

AJ863580 and AJ864835

 Hesperideae (HESP)

  Hesperis matronalis L.

Warwick et al. (2007)

DQ357547

  Hesperis siberica L.

Warwick et al. (2007)

DQ357548

 Iberideae (IBER)

  Iberis amara L.

Kropf et al. (2003)

AJ440311

  Iberis spathulata Bergeret

Kropf et al. (2003)

AJ440312

  Teesdalia nudicaulis (L.) W.T. Aiton

Koch and Mummenhoff (2001)

AF336214 and AF336215

 Isatideae (ISAT)

  Boreava orientalis Jaub. & Spach

Koch et al. (2007)

DQ249859

  Isatis tinctoria L.

Koch et al. (2007)

DQ249851

  Tauscheria lasiocarpa Fisch. ex DC.

Koch et al. (2007)

DQ249843

 Lepidieae (LEPI)

  Lepidium latifolium L.

Mummenhoff et al. (2004)

AJ582447 and AJ582521

  Stubendorffia gracilis (Pavlov) Botsch. & Vved.

Bailey et al. (2006)

DQ780944 and DQ780945

 Malcolmieae (MALC)

  Cithareloma lehmannii Bunge

Warwick et al. (2007)

DQ357528

  Malcolmia triloba (L.) Spreng.

Warwick et al. (2007)

DQ357561

 Megacarpaeeae (MEGA)

  Megacarpaea delavayi Franch.

Mummenhoff et al., unpublished

AJ628325 and AJ628326

  Megacarpaea gracilis Lipsky

Mummenhoff et al., unpublished

AJ628327 and AJ628328

 Noccaeeae (NOCC)

  Noccaea cochleariformis (DC.) A. Löve & D. Löve

Koch et al. (2007)

DQ249838

  Noccaea fendleri (A. Gray) Holub

Koch and Al-Shehbaz (2004)

AY154824

  Vania campylophylla F.K. Mey.

Koch and Mummenhoff (2001)

AF336168 and AF336169

 Notothlaspi clade (NOTO)

  Notothlaspi australe Hook.f.

Mitchell and Heenan (2000)

AF100689

  Notothlaspi rosulatum Hook.f.

Mitchell and Heenan (2000)

AF100690

 Physarieae (PHYS)

  Physaria didymocarpa (Hook.) A. Gray

O’Kane and Al-Shehbaz (2003)

AF137583

  Synthlipsis greggii A. Gray

O’Kane and Al-Shehbaz (2003)

AF137590

 Schizopetaleae (SCHIZ)

  Mathewsia foliosa Hook. & Arn.

Warwick et al. (2007)

DQ357563

  Schizopetalon walkeri Sims

Warwick et al. (2009)

EU620315

 Sisymbrieae (SISY)

  Sisymbrium altissimum L.

Warwick et al. (2002)

AF531560

  Sisymbrium septulatum DC.

Warwick et al. (2002)

AF531600

 Smelowskieae (SMEL)

  Smelowskia alba (Pall.) Regel

Warwick et al. (2004b)

AY230562

  Smelowskia jacutica (Botsch. & Karav.) Al-  Shehbaz & S.I. Warwick

Warwick et al. (2004b)

AY230646

 Thelypodieae (THEL)

  Hesperidanthus linearifolius (A. Gray) Rydb.

Warwick et al. (2002)

AF531612

  Thelypodium laciniatum (Hook.) Endl.

Warwick et al. (2009)

EU620328

 Thlaspideae (THLA)

  Peltaria alliacea Jacq.

Koch et al. (2007)

DQ249855

  Thlaspi arvense L.

Koch and Mummenhoff (2001)

AF336152S1 and S2

 Yinshania clade (YINS)

  Yinshania acutangula (O.E. Schulz) Al-Shehbaz, G. Yang, L.L. Lu & T.Y. Cheo

Koch and Al-Shehbaz (2002)

AH007969

  Yinshania acutangula subsp. wilsonii (O.E. Schulz) Al-Shehbaz, G. Yang, L.L. Lu & T.Y. Cheo

Koch and Al-Shehbaz (2002)

AH007968

Outgroup (OUTG)

 Cleome lutea Hook.

O’Kane and Al-Shehbaz (2003)

AF137588

Herbarium codes from Holmgren et al. (1990)

INIA GCC, Gómez-Campo Collection, Instituto Nacional de Investigaciones Agrarias, Madrid, Spain

aThis is a misidentification of Veselskya griffithiana (Boiss.) Opiz. This specimen is Matthiola sp., det. by Ihsan Oct 08; the species could not be determined because the fruits were missing

With the present study we are able to unravel the systematic neighborhood of more than 95% of all currently accepted genera within the crucifer family and screened almost all available plant material collections and resources on a worldwide scale. Previous studies have indicated, however, that ITS-based phylogenies do not provide basal resolution of the family (e.g., Bailey et al. 2006), and this is not a goal of this paper.

Materials and methods

Plant material

Collection data, taxa, vouchers, and GenBank accession numbers are listed in Table 1. The present study included 47 new ITS Brassicaceae sequences, as well as 138 ITS Brassicaceae sequences and one ITS Cleomaceae sequence taken from GenBank. Generic types are indicated in Supplementary Table 1 for all genera in the family.

The ITS analysis included 186 taxa/accessions in total: 185 sequences in the ingroup and 1 outgroup sequence (Table 1). The total Brassicaceae ingroup included 96 genera (101 taxa/accessions) not assigned to tribes and 68 genera (84 taxa/accessions) representing the 34 currently recognized tribes in the family and distinct Notothlaspi Hook.f. and Yinshania Y.C. Ma & Y.Z. Zhao, ITS-based clades (Al-Shehbaz et al. 2006; Bailey et al. 2006; Al-Shehbaz and Warwick 2007; German and Al-Shehbaz 2008; Khosravi et al. 2009). Cleome lutea Hook. served as the outgroup (Table 1), as the Cleomaceae is considered sister to the Brassicaceae (Hall et al. 2002, 2004). Although only poor resolution and support for the backbone phylogeny of the Brassicaceae has been obtained in ITS analyses (Bailey et al. 2006), ndhF (Beilstein et al. 2006, 2008), and nad4 data sets (Franzke et al. 2009), these studies have in general supported the recognition of clades corresponding to tribal groupings. All sequences representing the other Brassicaceae tribes and the outgroup taxa were obtained from GenBank (Table 1).

DNA extraction, amplification, and sequencing

DNA was extracted from approximately 100 mg or less of dried leaf tissue using Bio 101 FastDNA kit (Qbiogene Inc., Carlsbad, CA, USA) following manufacturer instructions. All polymerase chain reaction (PCR) amplifications were performed using Ready-To-Go PCR beads (GE Healthcare, Little Chalfont, Buckinghamshire, UK) and approximately 50 ng genomic DNA in a total of 25 μL. The entire ITS region (including ITS-1 and ITS-2 of nuclear ribosomal DNA and the 5.8S rRNA gene) was amplified and sequenced as a single fragment using the primers ITS1-18S described by O’Kane et al. (1997) and ITS4 (White et al. 1990). PCR amplification, purification, and sequencing of ITS followed Warwick et al. (2004a, b). For various taxa (e.g., Bivonaea lutea) a cloning strategy described in Koch et al. (2003b) was used prior to sequencing to test for multiple ITS copies and improve sequence quality.

Data scoring and analysis

The sequences were assembled and edited using Sequencher 4.7 software (Gene Codes Corporation, Ann Arbor, MI, USA), aligned using the CLUSTAL method in MEGALIGN (DNA Star, Madison, WI, USA), and further adjusted extensively by eye. Almost all previous ITS phylogenetic studies of the Brassicaceae have been based on CLUSTAL alignments of the data set (e.g. Warwick et al. 2008, 2009). Other alignment programs, such as MUSCLE and MAFFT, have been applied to other ITS Brassicaceae data sets and compared with both CLUSTAL and alignment by hand, but the results did not differ greatly and the MUSCLE and MAFFT alignments did not generally result in better resolved trees (co-author Mummenhoff, personal observation). Sequences of the ITS region are deposited in GenBank (Table 2), and the full alignment of the data set is either available from the authors upon request or can be found at TreeBASE (Study submission number S2662, Matrix accession number M5130). Boundaries of the ITS-1 and ITS-2 regions were determined by comparisons with the published sequences of Sisymbrium irio L. (Warwick et al. 2002). Lacking 5.8S positions were treated as missing data. Some ITS sequences that were missing the 5.8S region were included in the analysis, as these were the only sequences available in GenBank. Insertions and deletions (indels) in the ITS region were also treated as missing data to retain phylogenetic information from taxa not having the deletion. No additional gap coding was performed in this study. Gaps have been coded before in previous Brassicaceae phylogenetic papers that focused on select genera (e.g., Warwick et al. 2004a) or a single tribe (Warwick and Sauder 2005). However, in all previous higher-level ITS Brassicaceae studies (all other Warwick et al. studies cited herein; Bailey et al. 2006; Khosravi et al. 2009; Couvreur et al. 2010; German et al. 2009), gaps were not coded. Coding for gaps in such a divergent data set would be difficult while ensuring homeologous character scoring.
Table 2

Tribal placement of previously unassigned genera

Genus

Tribe (Schulz 1936)

Al-Shehbaz earlier 1980, 1990s papers

Tribe (Al-Shehbaz et al. 2006)

Revised tribal assignment, present study (tribe, code, parsimony % BS)

Moriera Boiss.

Lepidieae–Iberidinae

Lepidieae

Aethionemeae

Aethionemeae (AETH, 100% BS)

Physocardamum Hedge

Lepidieae

Unassigned

Alysseae (ALYS, 69% BS)

Alyssopsis Boiss.

Arabideae

Arabideae

Unassigned

Alyssopideae (ALYP-NEW, 95% BS)

Calymmatium O.E. Schulz

Sisymbrieae–Brayinae

Sisymbrieae

Unassigned

Alyssopideae (ALYP-NEW, 95% BS)

Delpinophytum Speg.

Lepidieae–Lepidiinae

Lepidieae

Lepidieae

Alyssopideae (ALYP-NEW, 95% BS)

Anastatica L.

Euclidieae

Euclidieae

Euclidieae?

Anastaticeae (ANAS, 95% BS)

Eigia Soják

Anchonieae (Hes)

Unassigned

Anastaticeae (ANAS, 95% BS)

Lachnocapsa Balf.f.

Lepidieae–Iberidinae

Lepidieae

Unassigned

Anastaticeae (ANAS, 95% BS)

Arcyosperma O.E. Schulz

Sisymbrieae–Pachycladinae

Sisymbrieae

Unassigned

Arabideae (ARAB, 79% BS)

Baimashania Al-Shehbaz

Arabideae

Arabideae (ARAB, 79% BS)

Dielsiocharis O.E. Schulz

Sisymbrieae–Pachycladinae

Sisymbrieae

Unassigned

Arabideae (ARAB, 79% BS)

Asta Klotzch ex O.E. Schulz

Lepidieae–Cochleariinae

Unassigned

Asteae (ASTE-NEW, 100% BS)

Megadenia Maxim.

Lepidieae–Iberidinae

Lepidieae

Unassigned

Biscutelleae (BISC, 59% BS)

Phoenicaulis Nutt.

Arabideae

Arabideae

Boechereae

Boechereae (BOEC, 84% BS)

Ammosperma Hook.f.

Brassiceae

Brassiceae

Brassiceae

Brassiceae (BRAS, <50% BS)

Didesmus Desv.

Brassiceae

Brassiceae

Brassiceae

Brassiceae (BRAS, <50% BS)

Douepea Cambess. ex Jacquem.

Brassiceae

Brassiceae

Brassiceae

Brassiceae (BRAS, <50% BS)

Eremophyton Bég.

Brassiceae

Brassiceae

Brassiceae

Brassiceae (BRAS, <50% BS)

Fezia Pit. ex Batt.

Brassiceae

Brassiceae

Brassiceae

Brassiceae (BRAS, <50% BS)

Foleyola Maire

Brassiceae

Brassiceae

Brassiceae

Brassiceae (BRAS, <50% BS)

Henophyton Coss. & Durieu

Brassiceae

Brassiceae

Brassiceae

Brassiceae (BRAS, <50% BS)

Pseuderucaria (Boiss.) O.E. Schulz

Brassiceae

Brassiceae

Brassiceae

Brassiceae (BRAS, <50% BS)

Nasturtiopsis Boiss.

Sisymbrieae–Brayinae

Sisymbrieae

Sisymbrieae?

Brassiceae? (BRAS, <50% BS)

Chrysochamela (Fenzl) Boiss.

Sisymbrieae–Camelineae

Sisymbrieae

Unassigned

Camelineae (CAME, <50% BS)

Noccidium F.K. Mey.

Lepidieae

Noccaeeae

Camelineae (CAME, <50% BS)

Aplanodes Marais

Arabideae

Unassigned

Cardamineae (CARD, 94% BS)

Iodanthus (Torr. & A. Gray) Steud.

Matthioleae

Arabideae

Cardamineae

Cardamineae (CARD, 94% BS)

Leavenworthia Torr.

Arabideae

Arabideae

Cardamineae

Cardamineae (CARD, 94% BS)

Ornithocarpa Rose

Schizopetaleae

Anchonieae (Hes)

Cardamineae

Cardamineae (CARD, 94% BS)

Planodes Greene

Arabideae

Arabideae

Cardamineae

Cardamineae (CARD, 94% BS)

Selenia Nutt.

Lunarieae

Arabideae? Alysseae?

Cardamineae

Cardamineae (CARD, 94% BS)

Sisymbrella Spach

Arabideae

Arabideae

Unassigned

Cardamineae (CARD, 94% BS)

Cremolobus DC.

Cremolobeae–Cremolobinae

Lepidieae–Cremolobeae

Separate tribe with Menonvillea?

Cremolobeae (CREM-NEW, <50% BS)

Menonvillea DC.

Cremolobeae–Menonvilleinae

Lepidieae–Cremolobeae

Separate tribe with Cremolobus?

Cremolobeae (CREM-NEW, <50% BS)

Robeschia Hochst. ex O.E. Schulz

Sisymbrieae–Descurainieae

Sisymbrieae

Descurainieae

Descurainieae (DESC, 81% BS)

Tropidocarpum Hook.

Lepidieae–Tropidocarpinae

Lepidieae

Descurainieae

Descurainieae (DESC, 81% BS)

Catenulina Soják

Hesperideae?

Unassigned

Euclidieae (EUCL, 73% BS)

Cymatocarpus O.E. Schulz

Sisymbrieae–Arabidopsidinae

Sisymbrieae

Unassigned

Euclidieae (EUCL, 73% BS)

Desideria Pamp.

Arabideae

Arabideae? Sisymbrieae?

Euclidieae

Euclidieae (EUCL, 73% BS)

Lachnoloma Bunge

Euclidieae

Euclidieae

Unassigned

Euclidieae (EUCL, 73% BS)

Lepidostemon Hook.f. & Thomson

Arabideae

Arabideae

Unassigned

Euclidieae (EUCL, 73% BS)

Octoceras Bunge

Euclidieae

Euclidieae

Unassigned

Euclidieae (EUCL, 73% BS)

Phaeonychium O.E. Schulz

Arabideae

Arabideae

Unassigned

Euclidieae (EUCL, 73% BS)

Pycnoplinthopsis Jafri

Not Hesperideae

Unassigned

Euclidieae (EUCL, 73% BS)

Pycnoplinthus O.E. Schulz

Hesperideae

Not Hesperideae

Unassigned

Euclidieae (EUCL, 73% BS)

Shangrilaia Al-Shehbaz, J.P. Yue & H. Sun

Euclidieae

Euclidieae (EUCL, 73% BS)

Spryginia Popov

Brassiceae

Anchonieae

Unassigned

Euclidieae (EUCL, 73% BS)

Xerodraba Skottsb.

Sisymbrieae–Brayinae

Sisymbrieae

Schizopetaleae?

Eudemeae (EUDE-NEW, 97% BS)

Chalcanthus Boiss.

Brassiceae

Brassiceae

Eutremeae?

Eutremeae (EUTR, 93% BS)

Pegaeophyton Hayek & Hand.-Mazz.

Arabideae

Arabideae

Unassigned

Eutremeae (EUTR, 93% BS)

Tchihatchewia Boiss.

Alysseae–Lunarieae

Unassigned

Hesperideae (HESP, 99% BS)

Boreava Taub. & Spach

Euclidieae

Euclidieae

Isatideae?

Isatideae (ISAT, 97% BS)

Chartoloma Bunge

Lepidieae–Isatidinae

Lepidieae

Isatideae?

Isatideae (ISAT, 97% BS)

Myagrum L.

Euclidieae

Euclidieae

Isatideae

Isatideae (ISAT, 97% BS)

Pachypterygium Bunge

Lepidieae–Isatidinae

Lepidieae

Isatideae?

Isatideae (ISAT, 97% BS)

Sameraria Desv.

Lepidieae–Isatidinae

Lepidieae

Isatideae?

Isatideae (ISAT, 97% BS)

Schimpera Hochst. & Steud. ex Endl.

Euclidieae

Euclidieae

Isatideae?

Isatideae (ISAT, 97% BS)

Kernera Medik.

Drabeae

Alysseae–Drabeae

Unassigned

Kernereae (KERN-NEW, 96% BS)

Rhizobotrya Tausch

Drabeae

Alysseae–Drabeae

Unassigned

Kernereae (KERN-NEW, 96% BS)

Cyphocardamum Hedge

Lepidieae

Unassigned

Lepidieae (LEPI, 99% BS)

Lithodraba Boelcke

Sisymbrieae

Schizopetaleae?

Lepidieae (LEPI, 99% BS)

Carinavalva Ising

Lepidieae

Unassigned

Microlepidieae (MICR-NEW, 75% BS)

Microlepidium F. Muell.

Lepidieae–Capsellinae

Lepidieae

Unassigned

Microlepidieae (MICR-NEW, 75% BS)

Brossardia Boiss.

Lepidieae–Thlaspidinae

Lepidieae

Unassigned

Noccaeeae (NOCC, 78% BS)

Callothlaspi F.K. Mey.

Noccaeeae

Noccaeeae (NOCC, 78% BS)

Coluteocarpus Boiss.

Lepidieae–Physariinae

Alysseae

Unassigned

Noccaeeae (NOCC, 78% BS)

Eunomia DC.

Lepidieae–Thlaspidinae

Unassigned

Noccaeeae (NOCC, 78% BS)

Kotschyella F.K. Mey.

Lepidieae

Noccaeeae

Noccaeeae (NOCC, 78% BS)

Pseudosempervivum (Boiss.) Grossh.

Lepidieae–Cochleariinae

Lepidieae

Unassigned

Noccaeeae (NOCC, 78% BS)

Murbeckiella Rothm.

Sisymbrieae

Sisymbrieae?

Oreophytoneae (OREO-NEW, 100% BS)

Oreophyton O.E. Schulz

Sisymbrieae–Pachycladinae

Sisymbrieae

Unassigned

Oreophytoneae (OREO-NEW, 100% BS)

Chaunanthus O.E. Schulz

Sisymbrieae–Sisymbriinae

Arabideae

Schizopetaleae

Thelypodieae (THEL, 94% BS)

Chlorocrambe Rydb.

Stanleyeae

Thelypodieae

Schizopetaleae

Thelypodieae (THEL, 94% BS)

Eremodraba O.E. Schulz

Sisymbrieae–Pachycladinae

Sisymbrieae

Schizopetaleae?

Thelypodieae (THEL, 94% BS)

Hesperidanthus (B.L. Rob.) Rydb.

Matthioleae

Not Matthioleae

Schizopetaleae

Thelypodieae (THEL, 94% BS)

Phlebolobium O.E. Schulz

Sisymbrieae–Sisymbriinae

Sisymbrieae

Unassigned

Thelypodieae (THEL, 94% BS)

Sarcodraba Gilg & Muschl.

Sisymbrieae–Pachycladinae

Sisymbrieae

Schizopetaleae?

Thelypodieae (THEL, 94% BS)

Sibaropsis S. Boyd & T.S. Ross

Schizopetaleae?

Thelypodieae (THEL, 94% BS)

Elburzia Hedge

Alysseae–Drabeae

Unassigned

Thlaspideae (THLA, 76% BS)

Gagria M. Král

Lepidieae

Unassigned

Thlaspideae (THLA, 76% BS)

Parlatoria Boiss.

Sisymbrieae–Alliariinae

Sisymbrieae

Thlaspideae

Thlaspideae (THLA, 76% BS)

Peltariopsis (Boiss.) N. Busch

Lunarieae

Alysseae

Unassigned

Thlaspideae (THLA, 76% BS)

Pseudovesicaria (Boiss.) Rupr.

Drabeae

Alysseae–Drabeae

Unassigned

Thlaspideae (THLA, 76% BS)

Sobolewskia M.Bieb.

Sisymbrieae–Alliariinae

Sisymbrieae

Thlaspideae?

Thlaspideae (THLA, 76% BS)

Asperuginoides Rauschert

Alysseae?

Unresolved

Bivonaea DC.

Lepidieae–Thlaspidinae

Lepidieae

Unassigned

Unresolved

Dipoma Franch.

Lepidieae–Cochleariinae

Lepidieae

Unassigned

Unresolved

Heldreichia Boiss.

Lepidieae–Iberidinae

Lepidieae

Unassigned

Unresolved

Horwoodia Turrill

Lepidieae

Unassigned

Unresolved

Idaho A. Nelson & J.F. Macbr.

Drabeae

Alysseae–Drabeae

Unassigned

Unresolved

Lunaria L.

Lunarieae

Alysseae

Unassigned

Unresolved

Ochthodium DC.

Euclidieae

Euclidieae

Unassigned

Unresolved

Petrocallis W.T. Aiton

Drabeae

Alysseae–Drabeae

Unassigned

Unresolved

Pseudofortuynia Hedge

Brassiceae

Brassiceae

Brassiceae

Unresolved

Ricotia L.

Lunarieae

Alysseae

Unassigned

Unresolved

Subularia L.

Lepidieae–Subulariinae

Lepidieae

Cardamineae?

Unresolved

Two phylogenetic analyses were conducted on the ITS data set: (1) maximum parsimony analyses using PAUP*, version 4.0b10 (Swofford 2002) and (2) maximum-likelihood analyses using PHYML (Guindon and Gascuel 2003; Guindon et al. 2005). In the maximum-parsimony analysis, the most parsimonious trees were generated using the heuristic search algorithm, with tree bisection-reconnection (TBR) branch-swapping, equal-weighted characters, with gaps treated as missing data, 500 random additions of the sampled taxa, and 200 trees saved per replicate. A strict consensus tree was computed and a total of 100 bootstrap replicates, respectively, were generated in PAUP* using a full heuristic search, with options MULTREES and TBR branch-swapping, and each replicate generated with simple addition sequence of taxa, with 100 trees saved per replicate, to test the stability of particular nodes in the parsimony analysis. Fifty-six sequence evolution models for maximum-likelihood analyses were tested with the aid of Modeltest v3.6 (Posada and Crandall 1998) to establish which model of DNA substitution provided the optimal fit for the data. Maximum-likelihood analyses were run using PHYML, under the TrN + I + G model of substitution, which was selected by the tests above as the most appropriate model for both data sets. Maximum-likelihood bootstrap values from 100 replicates were also generated for both data sets using the TrN + I + G model of substitution using PHYML. Analyses were conducted on 186 aligned ITS sequences, which included 185 ingroup sequences and 1 outgroup accession.

Results and discussion

Alignment

A total of 186 accessions, including the outgroup, were used in the analysis (Table 1). The resulting multiple alignment of the ITS region, including the 5.8S rRNA gene, was 773 bp long, and although the region between 150 and 200 bp was highly variable, it was not eliminated from the analysis. Of the 773 bp, 246 bp were parsimony uninformative, 131 bp were variable but not parsimony informative, and 396 bp were potentially parsimony informative. The ITS-1 region was variable at 297 of 362 sites (82.0%), the ITS-2 region at 174 of 238 sites (73.1%), and the 5.8S rRNA gene at only 56 of the 173 sites (32.4%). The alignment required the introduction of several indels, most of which were 1–4 bp, but also indels of 5, 7, and 13 bp. Few of these served as tribal markers, for example, Schizopetaleae taxa, Eudema nubigena subsp. nubigena Humb. & Bonpl. and Brayopsis colombiana Al-Shehbaz, share a 31-bp deletion; while Asta spp. share an 8-bp deletion. Indels that were unique to all members of a tribe occurring within the highly variable region between 150 and 200 bp include Smelowskieae and Sisymbrieae (see TreeBASE alignment).

The equal-weight maximum-parsimony analysis, based on 186 ITS sequences (1 outgroup and 185 ingroup accessions, each representing a unique sequence) and 396 informative characters, yielded 200 most parsimonious trees of 4,984 steps [Consistency Index (CI) 0.18, Retention Index (RI) 0.54]. Figure 1 shows the strict consensus tree, with bootstrap values. Figure 2 shows the maximum-likelihood tree. Parsimony bootstrap values appear above branches; branches in bold indicate parsimony and ML bootstrap values >80%; a dashed line indicates where only ML bootstrap values were >80%.
https://static-content.springer.com/image/art%3A10.1007%2Fs00606-010-0271-8/MediaObjects/606_2010_271_Fig1_HTML.gif
Fig. 1

A broad ITS-based phylogenetic backbone of Brassicaceae that focuses on the placement of 96 unassigned genera as defined in Table 1. Strict consensus tree of the 200 most parsimonious trees generated from analysis of 186 ITS sequences. Tree length = 4,984 (based on 396 parsimony-informative characters); CI 0.18; RI 0.54. Parsimony bootstrap values appear above branches; branches in bold indicate parsimony and ML bootstrap values >80%; a dashed line indicates where only ML bootstrap values were >80%. Major clades are indicated to the right, and codes correspond to tribal codes listed in Table 2

https://static-content.springer.com/image/art%3A10.1007%2Fs00606-010-0271-8/MediaObjects/606_2010_271_Fig2_HTML.gif
Fig. 2

A broad ITS-based phylogenetic backbone of Brassicaceae that focuses on the placement of 96 unassigned genera as defined in Table 1. Maximum-likelihood tree showing ML bootstrap values >80%. Major clades are indicated to the right, and codes correspond to tribal codes listed in Tables 1 and 2

Phylogenetic trees

Several well-supported clades were obtained in the analyses (Figs. 1, 2). These correspond to tribal groups (Tables 2, 3); the composition (including the position of previously unassigned genera) and support for each clade containing these genera will be reviewed separately below. The position of 12 genera remains unresolved and these will be discussed individually. In order to simplify the naming of tribes, especially in Figs. 1 and 2, four-letter abbreviations are adopted below for the previously recognized and herein proposed new tribes. Tribal assignments of all remaining genera in the family are summarized in Supplementary Table 1. A total of 308 (92%) of the 336 genera in the family have been assigned to a tribe. The latter table also cites all previous ITS studies for each genus. ITS data is available for 318 of the 336 genera.
Table 3

Brassicaceae tribes and percentage bootstrap support (% BS) from parsimony and maximum-likelihood (ML) analyses of ITS data

Tribes (code)

Parsimony (% BS)

ML (% BS)

Aethionemeae (AETH)

Mono (100)

Mono (95)

Alysseae (ALYS)

Mono (69)

Mono (61)

Alyssopsideae (ALYP-NEW)

Mono (95)

Mono (100)

Anastaticeae (ANAS)

Mono (95)

Mono (96)

Anchonieae (ANCH)

Mono (50)

Poly

Aphragmeae (APHR)

Mono (99)

Mono (96)

Arabideae (ARAB)

Mono (79)

Mono (86)

Asteae (ASTE-NEW)

Mono (100)

Mono (100)

Biscutelleae (BISC)

Mono (59)

Mono (65)

Boechereae (BOEC)

Mono (84)

Mono (71)

Brassiceae (BRAS)

Mono (<50)

Mono (16)

Buniadeae (BUNI)

Mono (100)

Mono (99)

Calepineae (CALE)

Mono (72)

Mono (86)

Camelineae (CAME)

Mono (<50)

Mono (16)

Cardamineae (CARD)

Mono (94)

Mono (89)

Chorisporeae (CHOR)

Mono (100)

Mono (100)

Cochlearieae (COCH)

Mono (100)

Mono (100)

Conringieae (CONR)

Mono (60)

Mono (72)

Cremolobeae (CREM-NEW)

Mono (<50)

Mono (47)

Descurainieae (DESC)

Mono (81)

Mono (78)

Dontostemoneae (DONT)

Poly

Mono (69)

Erysimeae (ERYS)

Mono (100)

Mono (100)

Euclidieae (EUCL)

Mono (73)

Mono (60)

Eudemeae (EUDE-NEW)

Mono (97)

Mono (97)

Eutremeae (EUTR)

Mono (93)

Mono (90)

Halimolobeae (HALI)

Mono (99)

Mono (100)

Heliophileae (HELI)

Poly

Poly

Hesperideae (HESP)

Mono (99)

Mono (100)

Iberideae (IBER)

Mono (<50)

Mono (3)

Isatideae (ISAT)

Mono (97)

Mono (98)

Kernereae (KERN-NEW)

Mono (96)

Mono (96)

Lepidieae (LEPI)

Mono (99)

Mono (91)

Megacarpaeeae (MEGA)

Mono (100)

Mono (100)

Microlepidieae (MICR-NEW)

Mono (75)

Mono (14)

Noccaeeae (NOCC)

Mono (78)

Mono (73)

Notothlaspideae (NOTO-NEW)

Mono (100)

Mono (100)

Oreophytoneae (OREO-NEW)

Mono (100)

Mono (100)

Physarieae (PHYS)

Mono (100)

Mono (100)

Schizopetaleae (SCHZ)

Mono (92)

Mono (88)

Sisymbrieae (SISY)

Mono (99)

Mono (99)

Smelowskieae (SMEL)

Mono (100)

Mono (100)

Thelypodieae (THEL)

Mono (94)

Mono (97)

Thlaspideae (THLA)

Mono (76)

Mono (67)

Yinshanieae (YINS-NEW)

Mono (100)

Mono (100)

It should be noted here that deeper nodes or the backbone of the family is only poorly resolved, and its resolution is out of the scope of the present manuscript. However, maximum-parsimony and ML analyses are largely in congruence, recognizing groups on the tribal level.

Tribes

Aethionemeae (AETH)

The ITS results confirm the placement of Moriera Boiss. along with Aethionema in the Aethionemeae [100% and 95% parsimony and ML bootstrap support (BS), respectively], as suggested in Al-Shehbaz et al. (2006).

Alysseae (ALYS)

ITS data support the inclusion of the monospecific genus Physocardamum Hedge (Turkey) in the Alysseae (69%, 61% BS). The genus had been tentatively placed in the Lepidieae (Al-Shehbaz 1986), but was not assigned to a tribe in Al-Shehbaz et al. (2006).

Warwick et al. (2008) did not include the genus in their ITS study of 85 species previously assigned to the tribe.

Alyssopsideae (ALYP-NEW)

Two previously unassigned genera, Alyssopsis Boiss. (2 spp., Middle East to Central Asia) and Calymmatium O.E. Schulz (2 spp., Central Asia), and Delpinophytum Speg. (1 sp., Patagonia), tentatively assigned to the Lepidieae (Appel and Al-Shehbaz 2003; Al-Shehbaz et al. 2006), formed a well-supported clade (95%, 95% BS). The odd and unexpected tribal position for the South American genus Delpinophytum requires further ITS sampling and re-examination of its morphology to test the present findings. In the Khosravi et al. (2009) paper, Alyssopsis deflexa Boiss. (a synonym of our Alyssopsis mollis) was nested in the Camelineae, a position not supported in the current study, where the well-supported Alyssopsis is quite remote from the poorly supported heterogenous Camelineae (sensu Beilstein et al. (2006, 2008); such inconsistencies may reflect errors in identification or sampling.

Anastaticeae (ANAS, re-established)

ITS data strongly confirms (95%, 96% BS) the recent inclusion of Anastatica L., tentatively assigned to the Euclidieae (Al-Shehbaz et al. 2006) and to the Malcolmieae (Khosravi et al. 2009), and supported the assignment of two previously unassigned monospecific Middle Eastern genera, Eigia Soják and Lachnocapsa Balf.f., to this tribe. When the tribe Malcolmieae was recently proposed as new (Al-Shehbaz and Warwick 2007), Anastatica had not been included in any molecular study. Our present findings agree with Khosravi et al. (2009) that one tribe is involved here, but the earliest name for it should be Anastaticeae, a name proposed by de Candolle and ignored by subsequent authors. Therefore, the tribal name Malcolmieae should be reduced to synonymy of Anastaticeae.

Aphragmeae (APHR)

This monogeneric tribe was monophyletic in this study (Figs. 1, 2), and no additional taxa are added to it following its recognition as new by German and Al-Shehbaz (2008).

Arabideae (ARAB)

ITS data provided good support (79%, 86% BS) for the inclusion of Baimashania Al-Shehbaz (2 spp., China) assigned to the Arabideae in Al-Shehbaz et al. (2006), and two previously unassigned genera: Arcyosperma O.E. Schulz (1 sp., Himalayas) and Dielsiocharis O.E. Schulz (2 spp., Iran-Tajikistan). The latter two genera had been considered morphologically similar and placed by Schulz (1936) in the subtribe Pachycladinae of the Sisymbrieae. In Khosravi et al. (2009), however, Dielsiocharis was close to Alyssopsis in the Camelineae, a relationship not supported in this study; again such inconsistencies may reflect errors in identification or sampling.

Asteae (ASTE-NEW)

The Mexican Asta Klotzch ex O.E. Schulz (2 spp.) was highly resolved in this study (100%, 100% BS) as a clade related to the New World tribes Schizopetaleae, Eudemeae, and Cremolobeae. The genus had been assigned to Lepidieae–Cochleariinae (Schulz 1936) and Lepidieae (Al-Shehbaz 1986), but was unassigned in Al-Shehbaz et al. (2006). This new tribe very likely includes the Mexican Scoliaxon Payson (1 sp.), a genus that resembles Asta by having extremely well-developed “stipule-like” glands.

Biscutelleae (BISC)

The monospecific Asian genus Megadenia Maxim. (China and Siberia) fell in a weakly supported clade (59%, 65% BS) with Biscutella L. (Figs. 1, 2), providing only weak support for its inclusion in the newly described tribe, Biscutelleae (German and Al-Shehbaz 2008; German et al. 2009).

Boechereae (BOEC)

The ITS data is consistent (84%, 71% BS) with the inclusion of the monospecific New World genus Phoenicaulis Nutt. (W. USA) in the Boechereae, as proposed earlier by Al-Shehbaz et al. (2006) and shown by Kiefer et al. (2009).

Brassiceae (BRAS)

The Brassiceae, a large tribe of ca. 52 genera, was extensively studied for ITS sequences (Warwick and Sauder 2005, and references therein). It formed a weakly supported ITS clade in the present and above studies (monophyletic but with <50% parsimony BS and only 16% ML BS). The present ITS data confirm the tribal assignment of previously unstudied Brassiceae genera: Ammosperma Hook.f. (1 sp.), Didesmus Desv. (2 spp.), Douepea Cambess. ex Jacquem. (2 spp.), Eremophyton Bég. (1 sp.), Fezia Pit. ex Batt. (1 sp.), Foleyola Maire (1 sp.), Henophyton Coss. & Durieu (1 sp.), and Pseuderucaria (Boiss.) O.E. Schulz (3 spp.). Although Nasturtiopsis Boiss. formed a weakly supported clade (56% BS) with the genus Fezia (Figs. 1, 2), its position in the Brassiceae is rather odd because it does not have conduplicate cotyledons or segmented fruits, both of which are characteristic of the tribe. Nasturtiopsis was most recently assigned to the Sisymbrieae, sister tribe to the Brassiceae, in Al-Shehbaz et al. (2006). Further sampling and re-examination of its morphology are needed to determine its tribal assignment.

Camelineae (CAME)

Chrysochamela (Fenzl) Boiss. (3 spp., Middle East) and Noccidium F.K. Mey. fell in a weakly supported clade (<50% parsimony BS and only 16% ML BS) with the Camelineae. The inclusion of Noccidium in the Camelineae had also been recently suggested by Khosravi et al. (2009). The data surprisingly did not support inclusion of Noccidium in the Noccaeeae with other segregate Thlaspi L. genera (Appel and Al-Shehbaz 2003) that had been assigned (Al-Shehbaz et al. 2006). Recent studies on the Camelineae (Bailey et al. 2006; Koch and Al-Shehbaz 2009; German et al. 2009) demonstrated that the tribe is heterogeneous and should perhaps be split into at least two or three smaller tribes.

Cardamineae (CARD)

The ITS data confirmed (94%, 89% BS) the previous assignment by Al-Shehbaz et al. (2006) of Iodanthus (Torr. & A. Gray) Steud., Leavenworthia Torr., Ornithocarpa Rose, Planodes Greene, and Selenia Nutt. to the Cardamineae. The data also supported the inclusion in this tribe of two previously unassigned genera: Aplanodes Marais (2 spp., South Africa) and Sisymbrella Spach. (2 spp., western Mediterranean).

Cremolobeae (CREM, re-established)

The South American Cremolobus DC. (8 spp.) and Menonvillea DC. (24 spp.) formed a weakly supported (<50% parsimony BS and 47% ML BS) ITS clade separate from other tribal clades. Although the Cremolobeae was proposed by Robert Brown (see the appendix in Denham and Clapperton 1826) as a new tribe, it was ignored by subsequent authors until treated as a tribe by Schulz (1936) and a subtribe, Cremolobinae of the Lepidieae, by Al-Shehbaz (1986). Both genera were not assigned to a tribe in Al-Shehbaz et al. (2006). Despite the weak support (Figs. 1, 2), the present ITS results confirm their close relationship and monophyletic status.

Descurainieae (DESC)

ITS data confirmed the assignment by Al-Shehbaz et al. (2006) of the monospecific Middle Eastern Robeschia Hochst. ex O.E. Schulz and the North–South American Tropidocarpum Hook. (4 spp.) to the Descurainieae (81%, 78% BS). Al-Shehbaz et al. (2006) described this new tribe as consisting of about six genera and some 60 species distributed in the Americas, Eurasia, and Africa. The tribe includes Descurainia (48 spp., including Hugueninia Rchb.) and the smaller Eurasian Hornungia (3 spp.), central Asian Ianhedgea Al-Shehbaz & O’Kane (1 sp.), and Patagonian Trichotolinum O.E. Schulz (1 sp.). The tribe is monophyletic based on the studies by Beilstein et al. (2006, 2008) and Goodson (personal communication). Descurainia is centered in the Canary Islands (7 spp.) and North and South America.

Euclidieae (EUCL)

ITS data confirmed (73%, 60% BS) the previous assignment by Al-Shehbaz et al. (2006) of Desideria Pamp. (12 spp.) and Shangrilaia Al-Shehbaz, J.P. Yue & H. Sun (1 sp.) to the Euclidieae. The data also confirmed Schulz’s (1936) previous assignment of the monospecific Lachnoloma Bunge and Octoceras Bunge to the Euclidieae. In the analysis of Khosravi et al. (2009), Octoceras has also been classified in the Euclidieae whereas Lachnoloma was assigned to the Anchonieae, which appear as sister to the Euclidieae in the molecular tree. The data also supported the placement of seven additional previously unassigned genera to this tribe: Catenulina Soják (1 sp.), Cymatocarpus O.E. Schulz (3 spp.), Lepidostemon Hook.f. & Thomson (6 spp.), Phaeonychium O.E. Schulz (7 spp.), Pycnoplinthopsis Jafri (1 sp.), Pycnoplinthus O.E. Schulz (1 sp.), and Spryginia Popov (7 spp.). Desideria taxa and Phaeonychium jafrii Al-Shehbaz have been recently transferred to Solms-laubachia (Yue et al. 2008), a genus previously assigned to the Euclidieae (Al-Shehbaz et al. 2006).

Eudemeae (EUDE-NEW)

The ITS data strongly support (97%, 97% BS) the inclusion of Xerodraba Skottsb. (8 spp., Argentina) in the South American clade containing Eudema Humb. & Bonpl. (6 spp.) and Brayopsis Gilg & Muschl. (6 spp.) and recently designated as Schizopetaleae—SCHIZ I clade (Warwick et al. 2009). Xerodraba had been tentatively assigned to the Schizopetaleae in Al-Shehbaz et al. (2006). The three genera are placed herein in the new tribe Eudemeae Al-Shehbaz et al. (see below).

Eutremeae (EUTR)

The ITS data supported (93%, 90% BS) the inclusion of the monospecific genus Chalcanthus Boiss. (Afghanistan, Iran, and Central Asia) in the primarily Asian Eutremeae, as tentatively proposed by Al-Shehbaz et al. (2006), as well as supported the inclusion of the previously unassigned Asian Pegaeophyton Hayek & Hand.-Mazz. (6 spp.). The latter genus was placed in the Arabideae (Schulz 1936; Al-Shehbaz 1988a) but had not been assigned to a tribe in Al-Shehbaz et al. (2006). Warwick et al. (2006a) showed that Neomartinella (3 spp.), Platycraspedum O.E. Schulz (2 spp.), Taphrospermum C.A. Mey. (7 spp.), and Thellungiella O.E. Schulz (3 spp.) were nested within the earlier-published Eutrema R.Br. (25 spp.), in which they were subsequently united (Al-Shehbaz and Warwick 2005). Chalcanthus and Taphrospermum were also shown to be sister taxa in the ndhF- and phyA-based phylogenetic studies of Beilstein et al. (2006, 2008), respectively.

Hesperideae (HESP)

The ITS data strongly supported (99%, 100% BS) the inclusion of monospecific Tchihatchewia Boiss. (=Neotchihatchewia Rauschert) (Turkey) in the Hesperideae. The genus was tentatively placed in the Alysseae–Lunarieae (Al-Shehbaz 1987) and was not assigned in Al-Shehbaz et al. (2006). The Hesperideae was formerly recognized as unigeneric, and its type genus, Hesperis L. (46 spp.), is centered in the Middle East and Europe. The tribe is readily distinguished from the rest of the Brassicaceae by having unicellular glands on uniseriate stalks, as well as simple and forked trichomes. However, several species of Hesperis are eglandular, and the placement herein of Tchihatchewia, which has simple, two-forked, and dendritic trichomes and lacks the glands (Appel and Al-Shehbaz 2003), does not disrupt the tribal boundaries.

Isatideae (ISAT)

The ITS data in this study strongly supported (97%, 98% BS) the inclusion of Boreava Jaub. & Spach (1 sp.), Chartoloma Bunge (1 sp.), Myagrum L (1. sp.), Pachypterygium Bunge (3 spp.), Sameraria Desv. (9 spp.), and Schimpera Hochst. & Steud. ex Endl. (1. sp.) in the Isatideae, as tentatively proposed by Al-Shehbaz et al. (2006). The Isatideae (ca. 90 spp., 8 genera) also includes Isatis L. (79 spp.) and monospecific genera Glastaria Boiss. and Tauscheria Fisch. ex DC. ndhF (Beilstein et al. 2006) and phyA (Beilstein et al. 2008) sequence data showed that Isatis and Myagrum L. (1 sp.) formed a highly supported monophyletic group. In a comprehensive analysis of tribe Isatideae (Moazzeni et al. 2010), Pachypterygium together with Boreava, Sameraria, and Tauscheria, were all nested within Isatis. In agreement with the molecular studies, analysis of seed-coat characters does not support the separation of genera Isatis, Pachypterygium, Sameraria, and Tauscheria (Moazzeni et al. 2007).

Kernereae (KERN-NEW)

The present study strongly supports (96%, 96% BS) the inclusion of two European monospecific genera, Kernera Medik. and Rhizobotrya Tausch, in a new tribe, the Kernereae. Both genera were unassigned in Al-Shehbaz et al. (2006) and were placed in the Drabeae by Schulz (1936). This tribe formed a well-supported sister clade (85% BS) to the Buniadeae, a recently resurrected monogeneric tribe including two species in Bunias L. (Al-Shehbaz and Warwick 2007).

Lepidieae (LEPD)

The monospecific genera Cyphocardamum Hedge (Afghanistan) and Lithodraba Boelcke (Argentina) fell in the Lepidieae clade (99%, 91% BS). The Lepidieae, as designated in Al-Shehbaz et al. (2006), consists of three to five genera and over 240 species. It is represented by 231 spp. in the core genus Lepidium L. (including Cardaria Desv., Coronopus Zinn, and Stroganowia Kar. & Kir.), which is distributed on all continents except Antarctica, and the Middle Eastern and Central Asian Stubendorffia Schrenk ex Fisch. & Avé-Lall. (8 spp.) and Winklera Regel (3 spp.).

A phylogenetic analysis of nuclear ITS and noncoding cpDNA of 124 taxa representing the genus Lepidium and related genera was performed by Mummenhoff et al. (2009). Both phylogenies clearly demonstrate that the genera suggested to be closely related to Lepidium, i.e., Cardaria, Coronopus, Stroganowia, Stubendorffia, and Winklera, are all well nested within the three main lineages of a broader defined Lepidium, and only one of these related genera (Winklera) is monophyletic. Instead of being separate from Lepidium, species of these genera group with Lepidium species of the same continental distribution (Fig. 1).

This current study confirms the tentative assignment of the genus Cyphocardamum by Al-Shehbaz (1986) to the Lepidieae, and does not support the assignment of Lithodraba to the Schizopetaleae. The data also support the exclusion of Delpinophytum from this tribe, as indicated above. The monospecific Acanthocardamum Thell. (Iran), previously assigned to the Lepidieae (Schulz 1936, Al-Shehbaz et al. 2006), has recently been shown to be nested within Aethionema (Khosravi et al. 2008b).

Microlepidieae (MICR-NEW)

Two previously unassigned Australian genera, Carinavalva Ising (1 sp.) and Microlepidium F. Muell. (2 spp.), fell in a well-supported ITS clade separate from other tribal clades (75%, 14% BS). Both were previously placed in the Lepidieae (Schulz 1936; Al-Shehbaz 1986).

Noccaeeae (NOCC)

The ITS data confirm (78%, 73% BS) the inclusion of the former Thlaspi segregate genera Callothlaspi F.K. Mey. and Kotschyella F.K. Mey. in the Noccaeeae, as proposed in Al-Shehbaz et al. (2006). The data also supported the inclusion in the Noccaeeae of three previously unassigned Middle Eastern genera Brossardia Boiss. (1 sp.), Coluteocarpus Boiss. (1 sp.), and Pseudosempervivum (Boiss.) Grossh. (4 spp.), as well as two species originally described in Eunomia DC. and Iberidella Boiss. Iberidella andersonii Hook.f. & Anderson was transferred to Noccaea Moench (Al-Shehbaz 2002), but the transfer of Eunomia oppositifolia (Pers.) DC. has not yet been done. Brossardia has also recently been transferred to Noccaea (Khosravi et al. 2008a). The genus Noccaea, represented by N. andersonii (Hook.f. & Thomson) Al-Shehbaz, N. cochleariformis (DC.) A. Löve & D. Löve, and N. fendleri (A. Gray) Holub, was monophyletic in the present study, whereas E. oppositifolia formed a separate well-supported clade (100% BS) with Vania F.K. Mey. and Kotschyella taxa. Extensive molecular studies (for summaries, see Koch 2003; Koch and Al-Shehbaz 2004) support the clear distinction between the Noccaeeae and Thlaspideae. However, in earlier studies (Koch 2003), many of Meyer’s segregate genera of Thlaspi L. (Meyer 1973, 1979) grouped with Noccaea in a well-supported clade, and only a few of his ten segregates [e.g., Neurotropis (DC.) F.K. Mey. and part of Microthlaspi F.K. Mey.] merit separate recognition. The ITS data presented herein also support the separate recognition of Callothlaspi and Kotschyella.

Notothlaspideae (NOTO-NEW)

This monogeneric tribe was monophyletic in this study (Figs. 1, 2), as in the studies of Mitchell and Heenan (2000), Bailey et al. (2006), Warwick et al. (2007), and German et al. (2009), and is herein recognized as a new tribe.

Oreophytoneae (OREO-NEW)

Two previously unassigned genera, Murbeckiella Rothm. (5 spp., Mediterranean) and Oreophyton O.E. Schulz (1 sp., tropical East Africa), fell in a well-supported ITS clade separate from other tribal clades (100%, 100% BS). Both had been previously been placed in the Sisymbrieae (Schulz 1936).

Schizopetaleae (SCHIZ)

Although the tribe was broadly circumscribed to include all genera presently assigned to the Thelypodieae (Al-Shehbaz et al. 2006), more recent studies (e.g., Beilstein et al. 2008; Warwick et al. 2009) amply demonstrate that the two tribes are not closely related. As delimited herein, the Schizopetaleae includes only Schizopetalon Sims (Chile and Argentina; 10 spp.) and Mathewsia Hook. & Arn. (N Chile and Peru; 7 spp.).

Thelypodieae (THEL)

The ITS data strongly support (94%, 97% BS) the inclusion in the Thelypodieae of several genera that were either assigned or tentatively assigned to the Schizopetaleae in Al-Shehbaz et al. (2006). These include Chaunanthus O.E. Schulz (3 spp.), Chlorocrambe Rydb. (1 sp.), Eremodraba O.E. Schulz (2 spp.), Hesperidanthus (B.L. Rob.) Rydb. (5 spp.), Sarcodraba Gilg & Muschl. (4 spp.), and Sibaropsis S. Boyd & T.S. Ross (1 sp.). The data also supported the inclusion of the monospecific Phlebolobium O.E. Schulz (Falkland Islands) that had previously been assigned to the Sisymbrieae (Schulz 1936; Al-Shehbaz 1988c). Hesperidanthus appears polyphyletic in this study, but generic resolution is not strong in the tribe (Warwick et al. 2002, 2006c, 2009).

Thlaspideae (THLA)

The ITS data support (76%, 67% BS) the inclusion of Parlatoria Boiss. (2 spp.) and Sobolewskia M.Bieb. (4 spp.) in the Thlaspideae, as previously indicated in Al-Shehbaz et al. (2006). The data also supported the inclusion of previously unassigned genera Elburzia Hedge (Iran; 1 sp.), Gagria M. Král (Georgia, 1 sp.), Peltariopsis (Boiss.) N. Busch (Armenia, Iran, Turkey; 2 spp.), and Pseudovesicaria (Boiss.) Rupr. (Caucasus; 1 sp.) in the Thlaspideae. Gagria is a synonym of Pachyphragma (DC.) Rchb., a genus already assigned to Thlaspi s. str. (Mummenhoff et al. 2001b) and the Thlaspideae (Al-Shehbaz et al. 2006).

Yinshanieae (YINS-NEW)

This monogeneric tribe was monophyletic in this study (Figs. 1, 2), and in the studies of Koch et al. (1999a), Koch and Al-Shehbaz (2000), Bailey et al. (2006), and German et al. (2009), and is herein recognized as a new tribe.

Unresolved genera

The tribal relationships of 12 genera remain unresolved in this study.

Asperuginoides Rauschert

The position of this monospecific genus (central Asia, Middle East) was not resolved in this study. It was tentatively assigned to the Alysseae in Al-Shehbaz et al. (2006) and was excluded from that tribe by Warwick et al. (2008). It formed a weakly supported (<50% BS) sister clade to the Cochlearieae, but in the analysis of Iranian Brassicaceae (Khosravi et al. 2009) was found along with Cochlearia and Ionopsidium in the Cochlearieae. The genus is rather odd in the family because of the presence of glochids on the fruits, and more studies are needed to resolve its tribal affiliation.

Bivonaea DC. and Horwoodia Turrill

The monospecific Bivonaea (Mediterranean) and Horwoodia (Middle East) fell in a weakly supported clade (<50% parsimony BS) that was separate from other tribal clades and sister to the primarily Mediterranean Brassiceae clade. Both genera were previously placed in the Lepidieae (Schulz 1936) but were unassigned in Al-Shehbaz et al. (2006).

Dipoma Franch.

The position of this monospecific Chinese genus was unresolved, falling within a weakly supported polytomy (<50% BS) that included the well-supported tribes Alyssopsideae, Microlepidieae, and Physarieae (Figs. 1, 2). The genus was placed in the Lepidieae (Schulz 1936, Al-Shehbaz 1986) but was unassigned in Al-Shehbaz et al. (2006). Further studies are needed to elucidate its tribal affiliation.

Heldreichia Boiss.

This Middle Eastern genus (Iran, Lebanon, Turkey; 4 spp.) fell in a weakly supported clade (<50% BS) with the Aphragmeae. It was previously placed in the Lepidieae (Schulz 1936; Al-Shehbaz 1986) but was unassigned to a tribe in Al-Shehbaz et al. (2006).

Idahoa A. Nelson & J.F. Macbr., Petrocallis W.T. Aiton, and Subularia L.

The genera Idahoa (1 sp., W USA), Petrocallis (1 sp., Europe), and Subularia (2 spp., Canada, USA, Europe, Africa) fell into a weakly supported clade (<50% BS) separate from other tribal clades. Both Idahoa and Petrocallis had been placed in the Drabeae by Schulz (1936) and the Alysseae–Drabeae (Al-Shehbaz 1987) and were unassigned to a tribe in Al-Shehbaz et al. (2006). Subularia was placed in the Lepidieae (Schulz 1936; Al-Shehbaz 1986), and because of the aquatic habitats its species occupy, it was suspected to belong to the Cardamineae (Al-Shehbaz et al. (2006). None of these relationships were supported in this study, and all three genera require further studies.

Lunaria L.

The tribal position of Lunaria (3. spp., Europe) was unresolved in this study. It was sister to the Biscutelleae clade, but this relationship was only weakly supported (<50% BS). The genus had been previously assigned to the Lunarieae by Schulz (1936), placed in the Alysseae by Al-Shehbaz (1987), and was unassigned in Al-Shehbaz et al. (2006). In the analysis of Iranian Brassicaceae, Khosravi et al. (2009) found Lunaria as sister to the Cochlearieae. The genus is unique in the family for a combination of long gynophores, long funicles united with a broad septum, and opposite leaves. Its position was not resolved in ndhF (Beilstein et al. 2006) and phyA (Beilstein et al. 2008) phylogenetic studies in the Brassicaceae. Evidently, Lunaria needs further studies to show its tribal disposition.

Ochthodium DC. and Pseudofortuynia Hedge

The previously unassigned and monospecific Ochthodium (Mediterranean region) and Pseudofortuynia (Iran) formed a weakly supported clade (55%, 58% BS). Pseudofortuynia, an Iranian monotypic genus, was originally described as Sisymbrium hesperidiflorum (Boissier and Buhse 1860). Pseudofortuynia was placed in Flora Iranica by Hedge in the tribe Brassiceae, and its synonym (Sisymbrium hesperidiflorum) by Rechinger in the tribe Hesperideae (Hedge and Rechinger 1968). Our phylogenetic tree and the analysis of Khosravi et al. (2009) indicate indeed that Pseudofortuynia is closely related to Old World Sisymbrium L. species. There is also morphological evidence supporting the close relationship between Pseudofortuynia and Sisymbrium: plants of both genera are glabrous or covered with simple hairs, the siliques are linear or subcylindrical, the fruit valves are convex with three nerves with the middle one most conspicuous, the seeds are oblong, uniseriate, and wingless, the cotyledons are incumbent or oblique, and the chromosome base number is x = 7 (Khosravi et al. 2009). The position of Ochthodium requires further study, and its association with Pseudofortuynia and Sisymbrium raises serious questions, because they are drastically different morphologically in nearly all aspects of the plants; Ochtodium for example, has woody, indehiscent, silicles, versus nonwoody, dehiscent siliques in the other two genera (see Appel and Al-Shehbaz 2003).

Ricotia L.

The tribal position of the genus Ricotia (9 spp., SE Europe and adjacent Middle East) was unresolved in this study. It was sister to the Arabideae clade, but this relationship was only weakly supported (<50% BS). Similar to Lunaria above, Ricotia had been previously assigned to the Lunarieae by Schulz (1936), placed in the Alysseae (Al-Shehbaz 1987), and was unassigned in Al-Shehbaz et al. (2006).

New tribes

Tribe Alyssopsideae

Al-Shehbaz, Warwick, Mummenhoff and Koch, trib. nov. Type genus: Alyssopsis Boiss., Ann. Sci. Nat. Bot. II. 17: 57. 1842.

Herbae perennes vel annuae, pilis bi- vel multifurcatibus stipitatis; folia caulina dentata, sessilia, basi auriculata vel exauriculata; racemi ebracteati; sepala patula vel ascendens, petala flava vel rarissime alba, stigma integra; ovula 8–40; fructus siliquae vel rarissime siliculae, teretes; semina uniseriata, saepe nonmucilaginosa; cotyledones accumbentes vel incumbentes.

This Central Asian and Middle Eastern tribe also includes four species, Alyssopsis mollis (Jacq.) O.E. Schulz, A. trinervis Botsch. & Sejfulin, Calymmatium draboides (Korsch.) O.E. Schulz, and C. notorrhizum (Gilli) Botsch. (see above).

Tribe Asteae

Al-Shehbaz, Warwick, Mummenhoff and Koch, trib. nov. Type genus: Asta Klotzsch ex O.E. Schulz, Bot. Jahrb. Syst. 66: 91. 1933.

Herbae annuae vel biennae, glabrae; folia caulina dentata, brevi petiolata vel sessilia, basi exauriculata; racemi ebracteati; sepala erecta, petala alba, stigma integra; ovula 4–20; fructus siliculae, teretes; semina uniseriata, mucilaginosa; cotyledones incumbentes.

Although currently including only Asta schaffneri (S. Watson) O.E. Schulz and A. stricta Rollins, this Mexican tribe may well also include Scoliaxon mexicanus (S. Watson) Payson (see above). However, when this is shown to be the case, the tribal diagnosis above should be amended to have branched, two- or several-rayed trichomes, flexuous racemes, and accumbent cotyledons.

Tribe Eudemeae

Al-Shehbaz, Warwick, Mummenhoff and Koch, trib. nov. Type genus: Eudema Humb. & Bonpl., Pl. Aequinoct. 2: 133. 1809.

Herbae perennes pulvinatae scaposae, glabrae vel pilis simplicibus vel furcatis stipitatis; folia basalia sessilia, rosulata, exauriculata; flores solitares, sepala erecta vel patula, petala alba vel flava, stigma integra; ovula 2–60; fructus siliquae vel siliculae, teretes, latiseptates vel angustiseptates; semina uniseriata, nonmucilaginosa; cotyledones incumbentes.

As discussed above, this South American tribe of 20 spp. includes Eudema, Brayopsis, and Xerodraba.

Tribe Kernereae

Al-Shehbaz, Warwick, Mummenhoff and Koch, trib. nov. Type genus: Kernera Medik., Pfl.-Gatt. 1: 77. 1792, nom. cons.

Herbae perennes, pilis simplicibus; folia caulina, sessilia, basi exauriculata; racemi ebracteati vel bracteati; sepala patula, petala alba, stigma integra; ovula 6–16(–24); fructus siliculae, teretes vel latiseptates; semina biseriata, nonmucilaginosa; cotyledones accumbentes.

This European tribe includes two species, Kernera saxatilis (L.) Rchb. and Rhizobotrya alpina Tausch.

Tribe Microlepidieae

Al-Shehbaz, Warwick, Mummenhoff and Koch, trib. nov. Type genus: Microlepidium F. Muell., Linnaea 25: 371. 1852.

Herbae annuae, glabrae vel pilis simplicibus et 2–multifurcatis; folia caulina integra, dentata vel pinnatisecta, sessilia vel petiolata, basi exauriculata; racemi ebracteati; sepala erecta vel patula, petala eburnea, stigma integra vel biloba; ovula 4–160; fructus siliculae, angustisepates, carinates; semina biseriata, mucilaginosa; cotyledones incumbentes.

This Australian tribe of three species includes Microlepidium alatum (J.M. Black) E. Shaw, M. pilosulum F. Muell., and Carinavalva glauca Ising.

Tribe Notothlaspideae

Al-Shehbaz, Warwick, Mummenhoff and Koch, trib. nov. Type genus: Notothlaspi Hook.f. in Bentham & Hooker, Gen. Pl. 1: 90. 1862.

Herbae perennae, scaposae, glabrae vel pilis simplicibus; folia basalia dense rosulata, folia caulina carens vel petiolata, exauriculata, dentata; racemi multiflori ebracteati vel basi bracteati; sepala suberecta, petala alba, stigma biloba; ovula 30–70; fructus siliculae, angustisepates, carinates, alates; semina biseriata; cotyledones incumbentes.

The tribe incudes Notothlaspi australe Hook.f. and N. rosulatum Hook.f., both of which are endemic to New Zealand.

Tribe Oreophytoneae

Al-Shehbaz, Warwick, Mummenhoff and Koch, trib. nov. Type genus: Oreophyton O.E. Schulz in Engler, Pflanzenreich IV. 105(Heft 86): 183. 1924.

Herbae perennes glabrae vel pilis simplicibus et multifurcatis stipitatis; folia basalia pinnatisecta vel pinnatifida, folia caulina carens vel sessilia pinnatifida et basi auriculata; floris solitares et in racemi ebracteati; sepala ascendens, petala alba vel rubella, stigma integra; ovula 20–60; fructus siliquae teretes vel latiseptates lineares; semina uniseriata vel biseriata, saepe nonmucilaginosa; cotyledones vel incumbentes.

Except for the alpine East African (Ethiopia, Kenya, Tanzania, Uganda) Oreophyton falcatum (Hochst. ex A. Rich.) O.E. Schulz, the remaining eight species of Murbeckiella are Mediterranean and Caucasian.

Tribe Yinshanieae

Al-Shehbaz, Warwick, Mummenhoff and Koch, trib. nov. Type genus: Yinshania Y.C. Ma & Y.Z. Zhao, Acta Phytotax. Sin. 17: 113. 1979.

Herbae annuae vel rarissime perennes, glabrae vel pilis simplicibus vel rarissime 2-furcatis stipitatis; folia caulina pinnatisecta, trifoliolata, vel 5–9-foliolata, rarissime simplicia, petiolata, basi exauriculata; racemi ebracteati vel bracteati; sepala patula, petala alba patula, stamina patula, stigma integra; ovula 6–20; fructus siliculae, teretes, latiseptates vel angustiseptates; semina uniseriata, mucilaginosa; cotyledones incumbentes.

This unigeneric tribe includes 13 species (Al-Shehbaz et al. 1998; Koch and Al-Shehbaz 2000) and is primarily endemic to China, with the range of Yinshania rivulorum (Dunn) Al-Shehbaz, G. Yang, L.L. Lu & T.Y. Cheo extending into Taiwan and that of Y. paradoxa (Hance) Y.Z. Zhao barely reaching into northern Vietnam.

Conclusion and final remarks

Recently, four major tasks have been defined in Brassicaceae systematics, taxonomy, and evolution (Koch and Al-Shehbaz 2009): (1) achieving a new infrafamiliar classification system based on phylogenetically circumscribed new tribes, (2) recognition and assignment of monophyletic genera, (3) unraveling the principles in crucifer evolution and exploring detailed examples for species- or genus-specific evolutionary histories, and (4) phylogenetic circumscription of the order Capparales and the determination of Cleomaceae as the closest and sister family to the Brassicaceae. With this contribution we present a big step forward to achieving task 2 and provide substantial information to continue unraveling an infrafamiliar classification (task 1), providing also information not only on the existence and composition of tribes but also on their relative relationships to each other. Phylogenetic analysis of ITS sequence data using a select tribal framework allowed us to analyze tribal positions of several genera whose tribal assignment could not be clarified in previous studies. The new tribal framework of 44 tribes presented herein (Supplementary Table 1) should serve as a useful guide in studying the few remaining unassigned genera in the family. It will also be useful in further studies of familial and tribal age estimation (e.g., Mitchell-Olds et al. 2005; Koch and Mummenhoff 2006; Franzke et al. 2009; Couvreur et al. 2010), genome evolution (e.g., Lysak and Lexer 2006; Lysak et al. 2009), and trait evolution in the Brassicaceae. However, the ITS marker does not resolve basal resolution in the family, nor do other single- or even multimarker-based phylogenies (Bailey et al. 2006; Beilstein et al. 2006, 2008; Couvreur et al. 2010). In the latter analysis, Couvreur et al. concatenated all checked and reliable sequences available from GenBank as well as new sequences for a total of 207 currently recognized genera and eight molecular markers representing comprehensive coverage of all three genomes. The resulting tree, the largest in terms of number of genera and markers sampled to date and covering the whole family in a representative way, provided important insights into the evolution of the family on a broad scale. The lack of backbone resolution in the tree was interpreted as the consequence of early rapid radiation within the family after the split with the early diverging tribe Aethionemeae. The supermatrix, dated under an uncorrelated relaxed molecular clock using a direct fossil calibration approach, put the family origin at 37.6 (24.2–49.4) Ma, and thus under a warm and humid climate. The authors further speculate that the rapid radiation detected was caused by global cooling during the Oligocene coupled with a genome duplication event which could have allowed the family to adapt rapidly to the changing climate.

Acknowledgments

Fieldwork by Al-Shehbaz in South America was supported by National Geographic Society grant 8139-06. We wish to thank Dr. Mark Beilstein, The Arnold Arboretum Harvard University, for providing two unpublished ITS sequences, and Ulrike Coja for technical assistance.

Supplementary material

606_2010_271_MOESM1_ESM.xls (137 kb)
Supplementary material (XLS 137 kb)

Copyright information

© Springer-Verlag 2010