Cladistics · 2018. 1. 10. · 2 Flores et al. / Cladistics 0 (2017) 1–25. diagnoses and testing previous ideas about character evolution. In addition, the contribution to and conﬂict

Combined phylogenetic analysis of the subclass Marchantiidae(Marchantiophyta): towards a robustly diagnosed classification

Jorge R. Floresa,b,*, Santiago A. Catalanoa,b, Jesus Mu~nozc and Guillermo M. Su�areza,b

aUnidad Ejecutora Lillo (UEL; FML-CONICET), Miguel Lillo 251, S.M. de Tucum�an 4000, Argentina; bFacultad de Ciencias Naturales e

Instituto Miguel Lillo, Universidad Nacional de Tucum�an, Miguel Lillo 205, S.M. de Tucum�an 4000, Argentina; cReal Jard�ın Bot�anico

(RJB – CSIC), Plaza de Murillo 2 Madrid, 28014, Spain

Accepted 22 August 2017

Abstract

The most extensive combined phylogenetic analyses of the subclass Marchantiidae yet undertaken was conducted on the basisof morphological and molecular data. The morphological data comprised 126 characters and 56 species. Taxonomic samplingincluded 35 ingroup species with all genera and orders of Marchantiidae sampled, and 21 outgroup species with two genera ofBlasiidae (Marchantiopsida), 15 species of Jungermanniopsida (the three subclasses represented) and the three genera ofHaplomitriopsida. Takakia ceratophylla (Bryophyta) was employed to root the trees. Character sampling involved 92 gameto-phytic and 34 sporophytic traits, supplemented with ten continuous characters. Molecular data included 11 molecular markers:one nuclear ribosomal (26S), three mitochondrial genes (nad1, nad5, rps3) and seven chloroplast regions (atpB, psbT-psbH, rbcL,ITS, rpoC1, rps4, psbA). Searches were performed under extended implied weighting, weighting the character blocks against theaverage homoplasy. Clade stability was assessed across three additional weighting schemes (implied weighting corrected for miss-ing entries, standard implied weighting and equal weighting) in three datasets (molecular, morphological and combined). Thecontribution from different biological phases regarding node recovery and diagnosis was evaluated. Our results agree with manyof the previous studies but cast doubt on some relationships, mainly at the family and interfamily level. The combined analysesunderlined the fact that, by combining data, taxonomic enhancements could be achieved regarding taxon delimitation and qual-ity of diagnosis. Support values for many clades of previous molecular studies were improved by the addition of morphologicaldata. The general idea that morphology may render spurious or low-quality results in this taxonomic group is challenged. Themorphological trends previously proposed are re-evaluated in light of the new phylogenetic scheme.© The Willi Hennig Society 2017.

Introduction

Among embryophytes (land plants), liverworts(Marchantiophyta) are recognized as the basal group(Qiu et al., 1998; Nickrent et al., 2000; Karol et al.,2001). The complex thalloid liverworts (Marchantiidae)encompass about 345 species and 36 genera (Villarrealet al., 2016) which are remarkably different from otherembryophytes. Complex thalloid liverworts are charac-terized by a combination of both plesiomorphic charac-ters and novelties in their structure. On the one hand,several of these morphological features, such as a

haplodiploid life cycle and the absence of vascular tis-sue, are widespread among bryophytes. On the other,highly specialized fertile branches (gametangiophores),internal schizogenous air chambers and pegged rhizoidsare traits exclusive to the complex thalloid liverworts(Schuster, 1984a,b; Bischler, 1998; Crandall-Stotler andStotler, 2000). Given their position within embryophytesand the morphological peculiarities, Marchantiidaestand as crucial taxa to disentangle the evolutionary his-tory of land plants.The first classifications within Marchantiidae, based

on phylogenetic systematic principles, were presentedbased on morphological data. However, the subsequentclassifications relied exclusively on molecular data. Themost comprehensive morphological phylogeny (Bischler,

*Corresponding author.E-mail address: [email protected]

CladisticsCladistics (2017) 1–25

10.1111/cla.12225

© The Willi Hennig Society 2017

1998) supported the distinction of a basal Ricciineae anda derived group comprising Corsiineae + Targioniineae+ Marchantiineae. One of the few combined analyseswas published by Boisselier-Dubayle et al. (2002) andincluded 27 out of 35 genera of Marchantiidae. Thecombined trees obtained in that study rendered a com-pletely different pattern of relationships from that pub-lished by Bischler (1998). Subsequent molecular analyseslargely agree with the relationships obtained by Boisse-lier-Dubayle et al. (2002), except in the relationshipsamong morphologically simple families (Wheeler, 2000;Crandall-Stotler et al., 2005; Forrest et al., 2006; He-Nygr�en et al., 2006).Boisselier-Dubayle et al. (2002) analysed the conflict

between morphological data and molecular data, show-ing that both sources of evidence produced very differenttrees, but also that support values could be improved byconcatenating different data types. Crandall-Stotleret al. (2005) reported a high degree of homoplasy withinmorphological data. Nevertheless, they were able to findsynapomorphies for most of the nodes. Subsequently,many diagnoses were modified and several more diag-nostic characters were found to define higher taxonomicgroups (e.g. lenticular apical cell for Metzgeriidae; Cran-dall-Stotler et al., 2009) than in previous classifications.Contrasting with Boisselier-Dubayle et al. (2002) andCrandall-Stotler et al. (2005), He-Nygr�en et al. (2006)did not carry out an explicit survey on character conflict.Even so, they performed the most extensive combinedanalysis in terms of morphological data inclusion. Inaddition, the results obtained by He-Nygr�en et al. (2006)displayed no significant differences when different opti-mality criteria were considered. Although He-Nygr�enet al.’s study was the first attempt to reach a reliablediagnosed classification, Marchantiidae taxonomic sam-pling included only 13 genera (37% of the accepted gen-era in the subclass).The current accepted classification (Crandall-Stotler

et al., 2009) was achieved as a result of several multi-locus phylogenies produced during the last decade.Compared to the previous morphology-based classifi-cation, several taxonomic categories were omitted. Inaddition, many genera of Marchantiidae were re-located in completely different categories (e.g. Mono-carpus D.J. Carr (Forrest et al., 2015) or Neohodgsonia(Perss.) Perss. (Crandall-Stotler et al., 2009)). As Cran-dall-Stotler et al. (2009) pointed out, diagnosing theseunexpected groups—from the morphological point ofview—represented a real challenge.Despite the advances in the morphological knowl-

edge of Marchantiidae accomplished in the last decade(e.g. Duckett et al., 2014), there was no attempt toinclude this new information into quantitative phylo-genetic analyses. Although recently performed phylo-genies sampled a considerable number of molecularmarkers (Forrest et al., 2006, 2015; Villarreal et al.,

2016), support values for several nodes were still low.On the one hand, some taxonomic changes have beenrecently proposed based upon these clades with lowsupport (Long et al., 2016a,b). For example, giventhat the genera Exormotheca Mitt. and StephensoniellaKashyap were found to be the sister taxa of CronisiaBerk., Exormothecaceae was merged with Corsiniaceae(Long et al., 2016a). However, the node constituted byCronisia, Exormotheca and Stephensoniella lacked sig-nificant support values (fig. 2 in Villarreal et al., 2016).On the other hand, taxonomic proposals based onnodes with high support were controversial from amorphological perspective (Forrest et al., 2015; Longet al., 2016b). Even if the average support at the fam-ily level was high in recent molecular studies (Forrestet al., 2006; Villarreal et al., 2016), deep nodesremained ambiguous (Wheeler, 2000; Boisselier-Dubayle et al., 2002; Forrest et al., 2006; Villarrealet al., 2016). A large number of the inclusive nodeswithin Marchantiales, the crown group of the subclass(Crandall-Stotler et al., 2009), still had low supportvalues (Villarreal et al., 2016). In particular, derivednodes (Ricciaceae, Oxymitraceae, Monocleaceae, etc.)were hard to resolve even in combined analyses (Bois-selier-Dubayle et al., 2002). Therefore, with few excep-tions, interfamily relationships persisted as ataxonomic challenge (Crandall-Stotler et al., 2009).The strategies to overcome environmental stress and

their significance to bryophytes evolution were regularissues of interest in previous studies (Bischler andJovet-Ast, 1981; Longton, 1988a; Hedderson andLongton, 1996; Bischler, 1998). Bischler (1998) definedfour different groups based on different morphologicaland ecological characteristics, postulating a strong cor-respondence of those groups with morphological char-acters and taxonomic/phylogenetic membership.Similar morphologies were explained as consequencesof common ancestry rather than similar selective forces(Bischler, 1998; Bischler et al., 2005). Concerningmosses, Longton (1988b) stated that the basic lifestrategy was being perennial, considering ephemeralstrategies as derived. In contrast, Bischler (1998) con-sidered ephemeral life traits as ancestral for complexthalloid liverworts. Although character mapping hasbeen considered to elucidate the evolutionary patternof some traditional characters (Villarreal et al., 2016),Bischler’s hypothesis on the concerted evolution oflife-history traits has not been challenged analytically.The recent modifications within the phylogeny of

Marchantiidae have involved both new scenarios ofevolutionary trends and changes in morphologicaldiagnosis of different clades within this group. Herewe present the results of the largest combined phyloge-netic analysis in Marchantiidae in terms of taxon andcharacter sampling. This study allows the solving ofprevious uncertainties in some nodes, updating group

2 Flores et al. / Cladistics 0 (2017) 1–25

diagnoses and testing previous ideas about characterevolution. In addition, the contribution to and conflictbetween morphological characters from the differentlife phases were evaluated in depth.

Materials and methods

Taxonomic and morphological character sampling

Despite taxonomic changes having been recentlyproposed for some groups, the main taxonomicscheme followed throughout this study was that ofCrandall-Stotler et al. (2009). After Long et al. (2016a,b), the genera Exormotheca and Aitchisoniella Kayshapwere considered members of Corsiniaceae and Cle-veaceae, respectively. However, in order to testrecently proposed synonyms (Long et al., 2016a), thefollowing genera were scored as independent entities:Stephensoniella and Exormotheca, and Preissia Corda,Bucegia Radian and Marchantia L. In the combineddataset, ingroup sampling consisted of 37 genera ofMarchantiopsida (35 genera of Marchantiidae plustwo genera of Blasiidae), which represented all of theorders, families and genera within the class. Outgroupsincluded 15 species of Jungermanniopsida, with thethree major subclasses of this group being represented(Jungermaniidae, Pelliidae and Metzgeriidae). In addi-tion, the dataset included species from the three generaof Haplomitriopsida. Takakia ceratophylla (Mitt.)Grolle (Bryophyta) was employed to root the tree.Five genera were absent in the morphological parti-tion: Cavicularia Steph. (Blasiidae), Riella Mont.(Marchantiidae), Apotreubia S. Hatt. & Mizut.(Haplomitriopsida), Pleurozia Dumort. and Pallavici-nia Gray (Jungermanniopsida). All but five specieswere scored mainly from observed specimens: Athala-mia pinguis Falc., Austroriella salta J. Milne & Cargill,Geothallus tuberosus Campb. (Marchantiidae), Pelliaepiphylla L. Corda and Lejeunea cavifolia (Ehrh.)Lindb. (Jungermanniopsida). In total, the combineddataset comprised 56 species; 20 more taxa than thelast combined analysis (Boisselier-Dubayle et al.,2002). Taxa and vouchers are listed on Table 1.The morphological dataset comprised 126 characters

from different sources. The present morphologicalmatrix was initially constructed by extending that of Bis-chler (1998). Some characters scored at the phylum levelby Crandall-Stotler and Stotler (2000) were alsoincluded. The original coding of nine of these “tradi-tional” characters were modified (see Character Defini-tion in the Supplementary material). Along with thesetraits, 66 completely novel characters were incorporated.Pegged rhizoids, for instance, were recently studied indepth by Duckett et al. (2014). Most of these characters,especially those that were now re-interpreted, were

included in a phylogenetic matrix for the first time. Simi-larly, characters derived from spores have been studiedextensively in several taxa (Gupta and Udar, 1986) butinfrequently used for phylogenetic analyses within thisgroup; novel data from spores of Marchantiidae wereconsidered in this study. Features representing continu-ous traits have seldom been considered in phylogeneticanalyses of bryophytes. In cases where this sort of infor-mation was included, characters were commonly dis-cretized losing valuable phylogenetic information(Farris, 1990; Goloboff et al., 2006). In the presentstudy, 10 different variables were analysed as continuouscharacters (Goloboff et al., 2006). In order to incorpo-rate the morphological variation among populations, aminimum of 10 specimens per species was examined.The final range of variation for each character was estab-lished as the mean value � standard deviation (SD).Ninety-two characters were scored from the gameto-phytic phase and 34 from the sporophyte. In conjunc-tion, these included 12 cytological characters, ninedevelopmental features, 18 sexual traits and 87 macro-scopic characters. The morphological dataset presented4540 more cells than the most extensive morphologicaldataset at the subclass level to date (1886 cells; Bischler,1998). A detailed description of character, state defini-tion and images can be found in the Supplementarymaterial. The final dataset is available at Morphobank(Project: 2674; morphobank.org/permalink/?P2674).

Molecular data

The previous most comprehensive molecular sam-pling was carried out by Villarreal et al. (2016). Thepresent study focused on extending outgroup samplingand filling gaps within the former analysis. Therefore,psbA and rbcL markers were sequenced for three spe-cies (Plagiochila sp. (Dumort.) Dumort.; Radula volutaTaylor ex Gottsche, Lindenb. & Nees and Riccia sp.L.). Amplifications were carried out at the Real Jard�ınBot�anico de Madrid (Spain) following the conditionsdescribed by Forrest and Crandall-Stotler (2004; pri-mers described in Table S1). Sequencing was conductedby Macrogen Inc. (Seoul). Sequences by Villarreal et al.(2016) were downloaded from GenBank. Nucleotideswere aligned with MAFFT (Katoh and Toh, 2008)using default penalty settings. Ambiguously alignedpositions were excluded from the analysis. Data werecompiled using GB2TNT (Goloboff and Catalano,2012), a pipeline to build molecular datasets from Gen-Bank.

Phylogenetic analyses, constrained searches andtopology comparisons

Although molecular data are commonly analysed byfollowing model-based methods, the study of

Flores et al. / Cladistics 0 (2017) 1–25 3

Table

1Taxaincluded

intheanalysis

Species

Locality:Voucher

Support

literature

Genbankaccessionnumber

26S

rbcL

rps4

psbA

psbT

atpB

rpoC1

cpIT

Snad1

nad5

rps3

Aitchisoniellla

him

alayensis

Kashyap

India:Ahmad346(PC)

(Bischleret

al.,1994;

Longet

al.,2016b)

––

––

–KT793364

––

––

–

Aneura

pinguis(L.)

Dumort.

Argentina:JR

Flores43

AY608195

AY688744

DQ983852

AY607921

JF513424

AY507346

–AF033631

JF513358

AY688744

–

Apotreubia

nana

(S.H

att.&

Inoue)

S.H

att.&

Mizut.

(Hattoriet

al.,1966)

DQ460030

AB476552

DQ268983

AY877397

KJ590878

KJ590840

KT793586

–KJ590799

DQ268907

KJ590920

Asterella

tenella

(L.)

P.Beauv.

Mexico:

Ortcutt1562

(Evans,1920;Little,

1936;Sharp,1939;

Wittlake,

1954;

Bischler,1998)

KT793344

KT793556

KT793706

KT793458

KT793496

KT793374

KT793594

KT793739

–KT793799

KT793650

Athalamia

pinguis

Falc.

(SokhiandMehra,1973;

Bischler,1998;

Boisselier-D

ubayle

etal.,2002;Crandall-

Stotler

etal.,2009;

Rubasinghe,

2011)

DQ265774

DQ286003

DQ220677

DQ265747

KJ590880

KJ590842

KT793596

KT793741

KJ590801

DQ268912

KJ590922

Austroriella

salta

Milne&

Cargill

(CargillandMilne,

2013)

KT356898

KT356969

KT356979

KT356959

KT793498

KT356898

KT356990

KT356911

KT356946

––

BlasiapusillaL.

France:Fesolowicz123;

Pierrot24057;Cuynet

18486;Poirions/n;

Gardet

s/n;Coppey

s/n.

(DuckettandRenzaglia,

1993)

AY688683

AF536232

AY507436

AY507477

KJ590881

AY507349

–KT793742

KJ590802

AY688746

KJ590923

Bucegia

romanica

Radian

Hungary:Boross/n

Poland:Szw

eykowski&

Chudzinska1002

KJ590831

KJ590910

KJ590953

KJ590867

KJ590882

KJ590843

KT793598

KT793744

KJ590803

KT793801

KJ590924

Caviculariadensa

Steph.

–DQ268968

DQ268984

AY507479

KJ590883

KJ590844

KT356991

KT356914

KJ590804

DQ268913

KJ590925

Cleveanana

(=

Athalamia

hyalina)

France:Bischleret

Leclerc

93614;Bonot&

Pierrot604;Pierrot,

3410;Skrzypczak&

Skrzypczak1226

DQ265773.1

DQ286002

DQ220676

DQ265746

KJ590884

KJ590845

KT356992

KT356915

KJ590805

DQ268911

KJ590926

Conocephalum

conicum

(L.)

Dumort.

France:Bischleret

Baudoin

75137,75175;

Jovet-Ast

etBischler

71734,71738,71264,

71747,71882;Castelli

s/n;Vicqs/n;Dismier

s/n

KT356888

KT356971

KT356981

KT356961

KT793502

KT356899

KT356993

KT356916

KT356948

KT793804

KT793654


Table

1(Continued)

Species

Locality:Voucher

Support

literature


26S

rbcL

rps4

psbA

psbT

atpB

rpoC1

cpIT

Snad1

nad5

rps3

Corsinia

coriandrina

(Spreng.)Lindb.

France:Jovet-Ast

et

Bischler,71136-7,

71155,71377,71343,

71354,71387,71422-3,

71454,71492,71499,

71545,74328;Bischler

74058,74067,74089,

74106,74108,74113,

74124,74170,74142,

74148,74162,74178

(Hassel

deMen� en

dez,

1963;Bischler,1998;

Bischleret

al.,2005)

KT793348

KT793560

KT793710

KT793462

KT793506

KT793381

KT793604

KT793750

KT793423

KT793806

KT793658

Cronisia

fimbriata

(Nees)

Whittem.&

Bischl.

Bolivia:

Fuentes2841

(Bischleret

al.,2005)

–KT793562

–KT793464

–KT793383

–KT793752

––

–

Cryptomitrium

tenerum

(Hook.)

Underw.

Argentina:Hassel

de

Men� endez

(Hassel

deMen� en

dez,

1963)

KT356889

KT356972

KT356982

KT356962

KT793507

KT356900

FJ173789

KT356918

KT356949

KT793807

KT793659

Cyathodium

cavernarum

Kunze

CaboVerde:

Bolles/n

(Bischleret

al.,2005)

KT793349

KT793565

KT793712

KT793467

KT793509

–KT793607

KT793756

KT793425

KT793809

KT793661

Dumortiera

hirsuta

(Sw.)Nees

Argentina:JR

Flores

(Hassel

deMen� en

dez,

1963;Bischleret

al.,

2005)

DQ265780

DQ286009

DQ220683

DQ265753

KJ590888

KJ590849

KT356996

KT356920

KJ590809

DQ268920

KJ590931

Exorm

otheca

pustulosa

Mitt.

France:Bischler74077;

Jovet-Ast

&Bischler

74609

(Bischleret

al.,2005)

DQ265781

DQ286010

DQ220684

DQ265754

KJ590889

KJ590850

KT356997

KT356921

KJ590810

DQ268921

KJ590932

Fossombronia

foveolata

Lindb.

Argentina:JR

Floress/n

(Haupt,1920;Renzaglia,

1982;Crandall-Stotler

etal.,2005)

AF226029

HQ446985

HQ447037

AY507482

–AY507354

––

––

–

Frullania

asagrayana

Mont.

Bolivia:Curchillet

al.

23373(m

ixed

withF.

beauverdii)

(Schuster,1984a,b,1992)

KX493338

FJ380822

–FJ380657

––

––

––

–

Geothallustuberosus

Campb.

(Campbell,1896)

KJ590833

KJ59091

KJ590955

KJ590869

KJ590890

KJ590851

KT356998

KT356922

KJ590811

KT793817

KJ590933

Haplomitrium

hookeri(Sm.)Nees

German:Lehmans/n

(Grubb,1970;Crandall-

Stotler

etal.,2005)

DQ268882

–AY608068

––

HQ412996

KT793615

–KT793431

––

Jungermannia

sp.L.

Argentina:JR

Flores58

(Crandall-Stotler

etal.,

2005)

AY316351

AY507409

AY507493

AY149816

––

–X00667

KF852564

AY688756

KF851659

Lejeunea

cavifolia

(Ehrh.)Lindb.

(Schuster,1980;

Crandall-Stotler

and

Stotler,2000)

DQ026545

–DQ787470

AM396279

AY312077

DQ646043

––

–AJ000701

–

Lepidoziareptans

(L.)Dumort.

Indonesia:Richards,P.

s/n

(Schuster,1969;

Crandall-Stotler

and

Stotler,2000)

AY608229

U87075

AY608083

AY607962

–DQ646025

––

JF513380

AY608293

JF316404

Lophocolea

heterophylla

(Schrad.)Dumort.

Spain:Mu~ nozet

al.s/n

DQ268889

DQ268932

DQ268987

DQ268999

–DQ646034

–AF033625

AY354959

DQ268932

KF942740

Lunulariacruciata

(L.)Lindb.

Argentina:JR

Flores11

(Hassel

deMen� en

dez,

1963)

DQ265782

DQ286011

DQ220685

DQ265755

KT793519

KT356902

KT357000

KT356923

KT356951

DQ268933

KT793671


Table

1(Continued)

Species

Locality:Voucher

Support

literature


26S

rbcL

rps4

psbA

psbT

atpB

rpoC1

cpIT

Snad1

nad5

rps3

Makinoacrispate

(Steph.)Miyake

Japan:U.Fauries/n

AY877374

AY877390

AY877393

AY877399

KF852178

AY877395

––

JF513370

AY877386

KF851588

Mannia

californica

(Gottsche)

L.C.

Wheeler

Mexico:McG

regor&

Rosarios/n

GQ910726

KJ590913

KJ590956

KJ590870

KJ590893

KJ590852

KT357001

KT356924

KJ590814

KT793818

KJ590936

Marchantia

polymorphaL.

Argentina:JR

Flores18,

28

KT356891

KT356974

KT356984

KT356964

KT793528

KT356903

KT357002

KT356925

KT356952

KT793822

KT793677

Metzgeria

furcata

(L.)Corda

Bolivia:MarkoLew

iss/n

(Renzaglia,1982;

Kuwahara,1986;

Crandall-Stotler

and

Stotler,2000)

DQ268876

DQ268975.1

AM396271

JF513436

––

––

JF513371

DQ268940

–

Monocarpus

sphaerocarpus

Carr.

Australia:Fawcetts/n

KT356886

KT356975

KT356985

KT356965

–KT356904

KT357003

KT356926

KT356953

––

Monoclea

gottschei

Lindb.

Argentina:JR

Flores8,

23

(Hassel

deMen� en

dez,

1963)

DQ265787

DQ286016

DQ220690

DQ265760

KJ590895

KJ590854

HQ026346

KT793777

KJ590816

DQ268943

KJ590938

Monosolenium

tenerum

Griff.

Japan:Inoue915;

Deguchi962

(Crandall-Stotler,1980;

Bischler,1998)

DQ265788

DQ286017

DQ220691

KJ590871

KJ590896

KJ590855

KT357004

KT356928

KJ590817

DQ268944

KJ590939

Neohodgsonia

mirabilis(Perss.)

Perss.

New

Zealand:Streimann,

H51072

(Bischler,1998)

AY688692

AY507415

AY507456

AY507499

–AY877396

–KT793778

–AY688759

–

Oxymitra

incrassata

(Brot.)S� ergio

&

Sim

-Sim

France:Sotiauxet

Sotiaux1234

(Hassel

deMen� en

dez,

1963;Bischler,1998;

Bischleret

al.,2005)

KJ590834

KJ590914

KJ590957

KJ590872

KJ590898

KJ590856

KT357005

KT356930

KJ590819

KT793826

KJ590941

Pallavicinia

lyellii

(Hook.)Gray

AY734742

AY507416.1

AY688798

AY507501

KF852181

DQ646049

––

JF513374

AY688761

KF851591

PelliaepiphyllaL.

Corda

(Renzaglia,1982;

Crandall-Stotler

and

Stotler,2000;Crandall-

Stotler

etal.,2005)

AY688693

AY688787

DQ463120

AY507502

–DQ646048

––

JF513375

AY688764

–

Peltolepisquadrata

(Saut.)M

€ ull.Frib.

France:Castelli65001a,

66001,67001

AY688693

AY688787

AY507457

AY507502

KT793534

AY688823

–JF

513375

––

–

Plagiochasm

a

rupestre(J.R

.

Forst.&

G.Forst.)Steph.

Argentina:JR

Flores27;

Schiavone,

633

KJ590835

KJ590915

KJ590958

KJ590873

KJ590900

KJ590858

FJ173796

KT793779

KJ590821

KT793828

KJ590943

Plagiochilasp.

(Dumort.)

Dumort.

Argentina:JR

Flores54

–KY985367

–MF036173

––

––

––

–

Pleuroziapurpurea

Lindb.

DQ268899

AY877391

AY608100

AY877401

–HQ412997

––

AY607889

DQ268951

–

Preissiaquadrata

(Scop.)Nees

France:Allorges/n;

Jeanperts/n;Durands/

n

DQ265793

KJ590916

KJ590959

KJ590874

KJ590901

KJ590859

KT357008

KT356933

KJ590822

DQ268953

KJ590944

Radula

voluta

Taylorex

Gottsche,

Lindenb.&

Nees

Argentina:JR

Flores51

––

HQ447037

MF036172

––

––

––

–


Table

1(Continued)

Species

Locality:Voucher

Support

literature


26S

rbcL

rps4

psbA

psbT

atpB

rpoC1

cpIT

Snad1

nad5

rps3

Reboulia

hem

isphaerica(L.)

Raddi

France:Bischler74132,

74128,74110,74626;

Jovet-Ast

etBischler

74562

KJ590836

KJ590917

KJ590960

KJ590875

KJ590902

KJ590860

KT357010

KT356935

KJ590823

–KJ590945

Ricciasp.(cf.

paraguayensis)

L.

Argentina:JR

Flores49

(Hassel

deMen� en

dez,

1963)

–KY985366

–MF036171

––

––

––

–

RicciafluitansL.

France:

DGabriel

s/n

(Underwood,1894;

Sharp,1939;Jovet-A

st,

1986;Singh,2014)

DQ265795

DQ286023

DQ220696

DQ265766

KJ590903

KJ590861

KT793634

KT356936

KJ590824

DQ268956

KJ590946

Ricciocarposnatans

(L.)Corda

Uruguay:

Suarezet

al.1317

(Hassel

deMen� en

dez,

1963)

DQ265796

DQ286024

DQ220698

DQ265767

KJ590904

KJ590862

KT793637

KT356937

KJ590825

DQ268958

KJ590947

Riellahelicophylla

(Bory

&Mont.)

Mont.

Algeria:Trabuts/n;

Durieu

deMaisonneuve

s/n

(Montagne,

1852)

KJ590837

KJ590918

KJ590961

KJ590876

KJ590905

KJ590863

KT357011

KT356938

KJ590826

KT793832

KJ590948

Sauteriaalpina

(Nees)

Nees

Austria:PGeissler17087

Italia:Bischler866

Poland:Szw

eykowski&

Chudizinska1001

(Rubasinghe,

2011;

Borovichev

etal.,2012)

DQ265797

DQ286025

DQ220699

DQ265768

KJ590906

KJ590864

KT357012

KT356939

KJ590827

DQ268960

KJ590949

Sphaerocarpos

texanusAust.

Morocco:Jovet-Ast

a,b

s/n

(Haynes,1910)

AY688697

AY507425

AY507466

AY507511

KT793545

AY507382

KT357015

KT356942

KT356956

AY688771

KT793696

Stephensoniella

brevipedunculata

Kashyap

Him

alaya:Chopra

500

(Kashyap,1914a,b,

1915)

–KT793584

–KT793486

–KT793410

–KT793789

––

–

Symphyogyna

undulata

Colenso

Chile:

JR

Flores26,50

(Crandall-Stotler

etal.,

2005)

AY877380

AY688790

AY688804

AY688835

–AY688825

––

–AY688773

–

Takakia

ceratophylla

(Mitt.)Grolle

(Grubb,1970;Renzaglia

etal.,1997)

HM751509

GU295867

AY908023

JF513404

AB254135

DQ646093

–AF426188

AY354937.1

DQ268963

–

Targionia

hypophyllaL.

Argentina:JR

Flores56,

48;Schiavone2417

(Hassel

deMen� en

dez,

1963;Bischleret

al.,

2005)

KJ590839

AY507427

AY688805

AY507514

KJ590908

AY507384

FJ173802

KT793790

KJ590829

DQ268965

KJ590951

Treubia

sp.K.I.

Goebel

Samoa:F.Reinecke17

(Schuster

andScott,

1969;Schuster,1984a,

b,a)

AY688699

AY507428

AY507468

AY507515

KT793547

HQ412995

KT793639

–JF

973315

AY688774

JF973315

Wiesnerella

denudata

(Mitt.)Stephani

Japan:Akiyama&

Hiraoka1;Akiyama

13013,Inoue875,Suire

&Toyota

3

Indonesia,Java:

Verdoorn

s/n

Nepal:Long17247

LaReuni� on:Bischler

89021;Gim

alac1703,

1706,12569

DQ265799

DQ286027

DQ220701

–KJ590909

KJ590866

KT793640

KT356944

KJ590830

DQ268966

KJ590952

Voucher

andsupportingliterature

referto

thetaxaincluded

inthemorphologicaldataset.New

sequencesare

marked

inbold

type.


morphology has been based largely on parsimony. Incontrast with molecular data, the use of probabilisticmodels to evaluate morphology has continually led toundesirable results (Goloboff and Pol, 2005; Beck andLee, 2014; Goloboff et al., 2017). For instance, Mkmodels have recently been shown to be particularlysensitive to the high heterogeneity rate present in mor-phological datasets, resulting in spurious groupingsand imprecise dating (Steiper and Seiffert, 2012; Beckand Lee, 2014). These outcomes still generate a greatreluctance amongst many authors to adopt models asa mean to analyse morphological data. Weighted par-simony, however, has been shown to improve the anal-ysis of morphological data (Goloboff et al., 2008a,b)and has acquired a key role in combined phylogeneticanalyses (e.g. Mirande, 2017). As the principal premiseof this paper is to achieve a phylogenetic hypothesisbased on a total-evidence approach, searches were per-formed under weighted parsimony as the optimalitycriterion.Therefore, the main systematic and taxonomic con-

clusions of the present study are derived from theanalysis of the combined dataset under extendedimplied weighing, weighting blocks in accordance totheir average homoplasy (Goloboff, 2014). Theblocks represented each gene and the morphologicaldataset.

Group sensitivity and searches. In order to assessgroup stability, searches were conducted across severalweighting schemes for individual partitions and thecombined dataset. In addition to the searches underblock weighting (BW), an alternative search strategyinvolved weighting each character according to itshomoplasy but taking into account the proportion ofmissing entries (IWM). In IWM, each missing entrywas assumed to have half of the homoplasy of theobserved entries (P = 0.5). In searches under bothIWM and BW, the TNT default concavity value wasused (K = 3; K3). Three concavity values for standardimplied weighting were also explored (K3, K5 and K7;Goloboff, 1993). Finally, the data also were analysedunder equal weighting (EqW). All weighting settingswere applied to the individual partitions and thecombined data (except BW in the case of themorphological dataset). Hence, a total of 17 differentanalyses were conducted (six for combined data, sixfor the molecular dataset and five for morphologicaldataset). In all cases, analyses were carried out inTNT 1.5 (Goloboff et al., 2008a,b; Goloboff andCatalano, 2016) with new technology searches using 10RAS as starting point. For each replicate, 10 iterationsof Ratchet (Nixon, 1999) and Tree Drifting (Goloboff,1999) were performed, ending the search when afterthe best score was hit seven times. Support values wereestimated by Symmetric Resampling (SR) considering

the difference between the most frequent group andthe most frequent contradictory group (GC; Goloboffet al., 2003). Additional estimates of support(Bootstrapping and Bremer) were also calculated andare available in Fig. S1.

Topological comparisons. In order to assess thecontribution of each source of information to the finalresult, the topologies recovered from different datatypes were compared in terms of (i) SPR (subtreepruning and regrafting) distance (Goloboff, 2008) and(ii) number of shared nodes. As a single SPRrearrangement may involve a movement of a few orseveral nodes, each movement was weighted by aconstant value of 5 (Goloboff, 2008). Themorphological trees obtained under each weightingcondition were compared to all the molecular trees. Inturn, the molecular trees were compared to thecombined tree. The number of SPR moves and thenumber of shared nodes were also calculated forthe morphological trees obtained by Bischler (1998) andBoisselier-Dubayle et al. (2002). In addition, the samecomparison was performed to discern the relativecontribution of gametophyte and sporophyte characters.To compare our morphological tree with Bischler (1998)and Boisselier-Dubayle et al. (2002) trees, evaluationsinvolved two different trees as reference: (a) ourmolecular tree obtained under BW and (b) Villarrealet al.’s (2016) molecular tree. Regarding our two sets ofmorphological characters, comparisons only used ourmolecular tree as reference.

Constrained searches. In order to quantify thesuboptimality of the groups not retrieved in the besttrees, constrained searches were carried out in thecombined data under BW. Understandingsuboptimality in terms of fit (F) is hardly intuitive.Therefore, the fit difference between the constrainedand the unconstrained trees was translated into thenumber of mean homoplastic characters with an extrastep required to explain the suboptimal tree (�x+1;Carrizo and Catalano, 2015). The estimation ofcharacter with mean homoplasy (�x was obtained as thetree length divided by the number of characters.The ratio between the topology fit differences(Funconstrained, Fconstrained) and mean character fitdifferences (�x, �x+1) allowed suboptimality to beconceived in terms of the number of �x+1

[(Funconstrained – Fconstrained)/(F�x � F�xþ1Þ ].

Character reconstruction, synapomorphies and diagnosis

Group diagnoses were determined by mapping mor-phological characters onto the combined tree obtainedunder BW. In addition, the role of different sets ofcharacters to diagnose specific clades was addressed.


Because sporophytic characters have commonly beenviewed as having no taxonomic information at lowerlevels (Schuster, 1984a,b; Crandall-Stotler and Stotler,2000; Crandall-Stotler et al., 2009), special attentionwas paid to those characters. In particular, the propor-tion of synapomorphies derived from sporophyticcharacters was quantified for each node. To assesswhether sporophytic characters diagnose taxonomicgroups below the class level, the number of sporo-phytic synapomorphies per taxonomic category wasalso measured.Phylogenetic searches, sensitivity assessment and diag-

nosis evaluation were implemented in scripts using TNTmacro language. These are available upon request.

Life history patterns and taxonomic membership

Bischler (1998) hypothesized that life-history traitswere phylogenetically correlated in Marchantiidae and,consequently, morphological similarity was caused bycommon ancestry. In order to test these hypotheses inlight of the new data, the four patterns found by Bis-chler found by Bischler (named Groups I–IV) and theputative associated characters were mapped onto thefinal tree. Given their relevance in liverwort biology(Bischler et al., 2005), four features were optimized:branch length, number of spores per capsule, sporesize and capsule dehiscence. Branch length and sporesize were scored as continuous characters; capsuledehiscence was scored as a binary character (cleistocar-pous/noncleistocarpous). The number of spores per cap-sule was scored as a multistate character: less than 500spores (0), more or equal to 1000 and less than 10000(1) and more than 10000 spores (2).

Results

Combined Tree under BW and Constrained Searches

The combined dataset analysed under extendedimplied weighting (BW) produced a single fullyresolved tree (Fig. 1). It was largely congruent withrecent phylogenetic analyses (Forrest et al., 2015; Longet al., 2016a,b).The major groups (Haplomitriopsida, Jungerman-

niopsida andMarchantiopsida) were recovered as mono-phyletic with high support values (�100; Fig. 1), as wellas the sister relationship among the four orders withinMarchantiidae. Some of these nodes had already beenfound in previous studies but have remained without aformal recognition (Crandall-Stotler et al., 2009; Forrestet al., 2015). To facilitate descriptions, some of thesenodes are referred to as clades A–F. Clade A was ren-dered as the association between the genus Monocarpusand the order Sphaerocarpales (Sphaerocarpus,

Geothallus, Austroriella and Riella) with strong evidence(SR 98, Fig. 1); the monophyly of Sphaerocarpales s.s.received more moderate support (SR 72). As in recentmolecular phylogenies (Forrest et al., 2015; Villarrealet al., 2016), the previous definition of Marchantialeswas not supported. Clade B was established upon thenode constituted by Lunularia (Lunulariales) and theremaining genera of Marchantiales expect Monocarpus.This clade with a moderate (to high) support value (SR88), was recovered as the sister group to Clade A.Mid-level nodes within Clade B were recovered in

most cases with low support. The node F (Riccia +Ricciocarpos + Oxymitra) + E (Monoclea + Cono-cephalum) was the sister clade to the monophyleticAytoniaceae (Asterella + Cryptomitrium + Mannia +Plagiochasma + Reboulia). The poorly supported cladeMonosolenium + Cyathodium was related to the highlysupported Clade D (Exormotheca + Aitchisoniella +Stephensoniella + Corsinia + Cronisia). Clade C (Wies-nerella + Targionia) was sister to both the remainingmorphologically simple groups and Aytoniaceae(Fig. 1). In contrast, the less inclusive nodes betweenfamilies tended to be well supported. Clades C, D, Fand E received support values from 82 to 100 (Fig. 1).At the family level, Cleveaceae and Corsiniaceae (Long

et al., 2016a) were both nonmonophyletic becauseAitch-isoniella was nested within the latter group. The generaCronisia and Corsinia formed a moderately supportedclade (SR 76; Fig. 1) with Exormotheca as their sistertaxa (SR 64; Fig. 1). Stephensoniella constituted a highlysupported clade with Aitchisoniella (SR 100; Fig. 1). Theclade formed by Sauteria, Clevea, Peltolepis and Athala-mia was also highly supported (SR 100; Fig. 1). Theother nonmonotypic families were retrieved as mono-phyletic with general high support.Phylogenetic searches forcing the monophyly of

groups absent from the best trees led to widely differ-ent suboptimality values depending on the consideredtaxa. On the one hand, ingroup families were highlysuboptimal regarding the BW combined tree. Themonophyly of Corsiniaceae entailed a difference of167.1 mean homoplastic characters gaining a furtherstep. As for Cleveaceae, its monophyly involved 172.2mean characters gaining an extra step. Comparatively,outgroup taxonomic groups implied less than one stepin a character with mean homoplasy (0.2–0.6). There-fore, there was no evidence to discard the monophy-letic status of outgroup taxa whereas the monophylyof the ingroup families was considerably questioned.

Partitioned analyses

Molecular dataset. The strict consensus of themolecular trees recovered across the entire analyticalconditions had 34 nodes (out of 54; Fig. 2).Molecular trees recovered Jungermanniopsida and


Fig. 1. (a) Combined tree under extended implied weighting (Block weighting). Symmetric Resampling values are indicated above branches (only> 50 values are shown; see additional support measures in the Fig. S1 in the Supplementary material). Sensitivity plots for ingroup nodes mono-phyly are shown below branches. (b) Sensitivity plots for Cleveaceae, Corsiniaceae and Marchantiales [as defined by Long et al. (2016a,b) andCrandall-Stotler et al. (2009)], two groups that were not monophyletic in the combined analysis under block weighting. Sensitivity plot reference:coloured squares represents monophyly. C = combined data, MOL = molecular data, MOR = morphological data. Extended implied weightingsettings: block weighting (BW) and character weighted considering their homoplasy and number of missing entries (IWM). K3, K5 and K7 areconcavities values explored under standard implied weighting. EqW refers to equal weighting.


Marchantiopsida as monophyletic. InsideMarchantiopsida, relationships were poorly resolved.The association between the orders Lunulariales orNeohodgsoniales, and the remaining orders wasunresolved. The family Marchantiaceae was sister tothe remaining Marchantiales. The rest of the familieswithin Marchantiales were placed in a nine-waypolytomy. Clades with high support values in thecombined tree under BW were also recovered in theanalysis of the molecular partition. The genera

Aitchisoniella and Stephensoniella were sister taxawithin a clade constituted by the remaining genera ofCorsiniaceae. Cleveaceae was not recovered asmonophyletic. Searches under BW concluded in asingle fully resolved tree, highly similar to thecombined tree (see below under Relative datacontribution).

Morphological dataset. The strict consensus of themorphological trees obtained across the entire

Fig. 2. Strict consensus of the molecular (left) and morphological trees (right) obtained under the different weighting schemes.


analytical conditions had 29 nodes (out of 49; Fig. 2).The order Marchantiales (in the sense of Crandall-Stotler et al., 2009) was not recovered asmonophyletic. Monoclea was found to be closelyrelated to Sphaerocarpales and Lunularia was nestedwithin Marchantiales. Monocarpus was the sister taxonof the remaining Marchantiales. Neohodgsonia was notrecovered inside Marchantiaceae. The rest of the taxawere placed within a 16-way polytomy. The genusAitchisoniella was not grouped with the rest of thefamily Cleveaceae although its association with otherfamilies was unclear. The other members ofCleveaceae did not constitute a monophyletic group.Corsiniaceae was also nonmonophyletic. The generaExormotheca and Stephensoniella were sister taxawhereas Corsinia + Cronisia were related to Ricciaceaeand Oxymitraceae. Searches under standard impliedweighting (K7) led to a single fully resolved tree whichmaximized congruence with both the combined dataand the molecular data (see below under Relative datacontribution).

Sensitivity analysis

About two-thirds of the ingroup clades (22 of 36)were not affected by different weighting schemes(Fig. 1). Although it was not a strict pattern, cladeswith high support values tended to be less sensitive toparameter variation. Unsupported nodes (SR < 50)were nonmonophyletic in 11 to 15 weighting schemes.Weakly to moderately supported clades (SR 50–90)were not monophyletic in two to seven weightingschemes. Highly supported groups (SR > 90) were notmonophyletic in five weighting conditions or less. Twoexceptions to this pattern were Sphaerocarpales (SR72) and Oxymitraceae + Ricciaceae (SR 100), whichwere nonmonophyletic in nine analytical conditions.The genus Exormotheca behaved as a wildcard taxon,alternating its position with members of Corsiniaceae.The most inclusive nodes within Marchantiales (i.e.excluding Marchantiaceae and Dumortieraceae) wereextremely sensitive. Moreover, such nodes were onlyrecovered in two analytical conditions (molecular andcombined data under BW; SR < 50). However,lower level clades within Marchantiales were recoveredunder different weighting schemes. The cladesMonocleaceae + Conocephalaceae, Exormothecaceae +Corsiniaceae and Monosoleniaceae + Cyathodiaceaewere markedly stable (10 to 12 schemes). The genusAitchisoniella constituted a stable clade with Stephen-soniella in 12 weighting schemes.Nodes of the morphological trees were more sensi-

tive to parameter variation than molecular trees (sensi-tivity plots in Fig. 1). Five families, and relationshipsinside them, were resolved by the morphological data(Ricciaceae, Aytoniaceae, Corsiniaceae, Cleveaceae

and Marchantiaceae). Taxa relationships inside Sphae-rocarpales were considerably more stable in the molec-ular data.

Relative data type contribution

Trees derived from molecular data were highly simi-lar to the BW combined tree. In particular, the BWmolecular tree was the most similar to the BW com-bined cladogram (1.2 SPR; Fig. 3). The molecular treeobtained under BW also shared 48 nodes with thecombined tree. Among morphological trees, the treeobtained considering a concavity value of seven (K7)was the most similar to the BW molecular tree (8.1SPR moves and 17 shared nodes).Regardless of the reference tree (our BW molecular

tree or Villarreal et al.’s (2016) tree), the comparisonsbetween the K7 morphological tree and Bischler(1998) and Boisselier-Dubayle et al. (2002) treesyielded lower SPR values for our morphological tree(Table 2). Bischler’s (1998) morphological treeinvolved 9.1 and 9.2 SPR moves to the BW moleculartree and Villarreal et al.’s (2016) tree, respectively.Boisselier-Dubayle et al.’s (2002) tree entailed 9.5 SPRmoves to the BW molecular tree and 10.7 SPR movesto Villarreal et al.’s (2016) tree.In terms of shared nodes, values varied depending

on the reference tree (Table 2). The number of sharednodes was slightly higher for Bischler’s tree when ourBW molecular tree was used as reference (18 sharednodes for Bischler’s tree; 17 shared nodes for the K7tree and Boisselier-Dubayle et al.’s tree). In contrast,the number of shared nodes was higher for the K7morphological tree when Villarreal et al.’s tree wasused as reference (18 shared nodes for our K7 tree; 17shared nodes for Bischler’s and Boisselier-Dubayleet al.’s trees).The gametophyte tree was more similar to the BW

molecular tree than the sporophyte tree. In terms ofshared nodes, the single gametophytic tree had 15common nodes with the molecular tree and an SPRdistance of 8.38 weighted movements (Fig. 4). Theanalysis including only sporophytic characters pro-duced 100 optimal trees which shared eight nodes withthe molecular tree and had an average SPR distanceof 12.8 weighted movements.Conflict and contribution of data sources were

also reflected in the support values. Despite the widedifferences among molecular trees and morphologicaltrees, average support values were improved in thecombined analysis in comparison with the molecular-only analysis. In particular, five clades of theingroup had higher support values (Figs 2 and 4).The mean SR support value was 70 for the BWcombined tree, whereas it was 65.7 for the BWmolecular topology.


Synapomorphy mapping and diagnosis assessment

Mapping morphological characters on the com-bined tree allowed determination of synapomorphiesfor 26 out of 35 ingroup nodes. The class Marchan-tiopsida and seven high-level clades inside it had highto moderate support values and morphological apo-morphic characters (Fig. 5; Table 3). Marchantiopsidaand Marchantiidae were both supported by nine

synapomorphies whereas node B had 10 synapomor-phic characters (being diagnosed for the first time).Clade F was supported by seven synapomorphies;two continuous characters and five discrete charac-ters. The Clade E was supported by a single sporetrait, germ tube absent. Two characters were diagnos-tic for Clade D. Three characters were synapomor-phies for both clades C and A. Descriptions of thesespecific groups are in Table 3, a detailed list of

Fig. 3. Molecular tree under extended implied weighting (Block weighting) and morphological tree under implied weighting (K = 7) which max-imised similarity among data types. Weighted SPR movements to the reference tree and shared nodes are indicated below each topology.


Table 2Comparison among different morphology-based trees (our morphological tree obtained under implied weighting, and Bischler’s and Boisselier-Dubayle et al.’s trees). The tree obtained in the present study under K7 implied weighting maximized similarity with reference trees in three outof four cases (bold). Bischler’s tree maximized the number of shared nodes with the BW molecular tree (bold)

SPR moves to theBW molecular tree

SPR moves toVillarreal et al.’s tree

Shared nodes with theBW molecular tree

Shared nodes withVillarreal et al.’s tree

K7 morphological tree 8.1 7.8 17 18

Bischler’s morphological tree 9.1 9.2 18 17Boisselier-Dubayle et al.’smorphological tree

9.5 10.7 17 17

BW, block weighting; SPR, subtree pruning and regrafting.

Fig. 4. Morphological trees derived from gametophytic characters and sporophytic characters under implied weighting (K = 7). Families withinMarchantiales are highlighted. SPR values and shared nodes with the molecular tree are indicated below each tree. For sporophytic tree, theaverage SPR value is provided (SD in parenthesis).


Fig. 5. Groups with improved diagnoses (Combined tree under BW). A detailed diagnosis for each group is in Table 3. The selected inter-familylevel nodes were those groups simultaneously well supported and diagnosed.


synapomorphies for each node in the phylogeny is inTable S2).Within the entire phylogeny, continuous charac-

ters diagnosed 18 clades (16 within Marchantiop-sida). Additionally, 18 clades were diagnosed by atleast one sporophytic feature. In ten out of these18 nodes, 50% or more of the diagnostic traitswere derived from the sporophyte (Fig. 6; Fig. S2).

The distribution of this proportion per taxonomicrange was observed to be higher for orders thanfor classes or subclasses. The inclusion of sporo-phytic characters into the diagnosis at the interfam-ily level was also considerably high. In fact, orderand interfamily nodes received the highest sporo-phyte contribution in terms of synapomorphies(Fig. 6; Fig. S2).

Table 3Synapomorphies for mid- and high-level groups within Marchantiidae and Marchantiopsida

Clade name Taxa included Character number: synapomorphies

F Ricciaceae [Riccia + Ricciocarpos]+Oxymitraceae [Oxymitra]

2: ratio apex width/base width: 1.1–1.5 -> 1.8–2.23: pegged rhizoids/smooth rhizoids rate: 0.6–1.0 -> 0.35–0.45108: capsule dehiscence: irregular -> cleistocarpous111: elaters: present -> absent53: perigonia position: embedded in receptacles -> in anacrogynousclusters

95: embryo ontogeny: foot and seta from hypobasal cell ->sporangium from all of it

69: antheridia stalk anatomy: four or more seriate -> uniseriateE Monocleaceae [Monoclea] +

Conocephalaceae [Conocephalum]125: germ tube: present -> absent

C Wiesnerellaceae [Wiesnerella] +Targioniaceae [Targionia]

2: ratio apex width/base width: 1.0–1.1 -> 0.7–0.97: diameter pegged rhizoids/smooth rhizoids: 0.7 -> 0.76–0.77105: capsule internal wall thickenings: annular -> semi-annular

D Corsiniaceae [Cronisia + Corsinia] +Exormothecaceae [Exormotheca +Aitchisoniella + Stephensoniella]

53: perigonia position: embedded in receptacles -> in anacrogynousclusters

27: drought tolerance: no -> yesA Monocarpaceae [Monocarpus] +

Sphaerocarpales [Sphaerocarpus +Austroriella + Riella + Geothallus]

108: capsule dehiscence: into four valves -> cleistocarpous111: elaters: present -> absent120: spore, proximal gross morphology: as in distal face -> differentfrom distal face

Marchantiopsida Blasiidae [Blasia + Cavicularia] +Marchantiidae [Neohodgsoniales +Sphaerocarpales + Lunulariales +Marchantiales]

17: phyllotaxi: one-third/one half -> none34: ventral appendages: absent -> two rows117: polarized spores: no -> yes89: outer perichaetium: leaf-like scales -> marchantioid involucre92: calyptra: shoot calyptra -> true calyptra90: Position of involucre: none -> dorsal75: spermatid spline aperture: 1-tubule aperture- > 3-tubule apertureor more

77: spermatid, basal body dimorphism: no -> yes82: spermatid, notch presence: no -> yes

Marchantiidae Sphaerocarpales(including Monocarpus)+ Lunulariales + Marchantiales +Neohodgsoniales

104: capsule wall: multistratose -> unistratose116: spore mother cell lobed: present -> absent125: germ tube: absent -> present11: protonema: globose -> flattened28: air chambers: absent -> Marchantia type60: archegoniophore, stalk anatomy: none -> homogeneous62: female receptacle anatomy: none -> homogeneous106: capsule wall thickenings: absent -> present64: archegonia neck cells (CT): five-cells row -> six-cells row

B Lunulariales [Lunularia] + Marchantiales 9: female branch length: 0.4–0.5 -> 1.236: ventral appendage shape: acute -> lanceolate43: rhizoids type: unicellular, smooth only -> unicellular, dimorphic44: typed of pegged rhizoids: none -> slightly pegged rhizoids only45: pegs: nonpegged -> blunt to pointed47: pegged rhizoids, distribution throughout the plant: none-pegged -> ventrally distributed

94: embryo type: filamentous -> octant38: scales appendage number: without scales -> one39: scales margin: no scales -> entire42:appendage cell form: no scale -> hexagonal or pentagonal


Mapping life-history traits

None of the ecological groups defined by Bischler(1998) had a single origin (Fig. 7). Group II was thebasal group along Marchantiidae, whereas groups IIIand IV characterized distant nodes to Sphaerocarpales.Group I was polyphyletic in three different clades(Fig. 7). Capsule dehiscence was reconstructed with 13steps; the cleistocarpous state diagnosed Sphaero-carpales + Monocarpus, Oxymitraceae + Ricciaceaeand Corsinia + Cronisia. The optimization of the num-ber of spores per capsule required nine steps. The con-tinuous characters (spore size and branch length), had1.5 and 2.15 steps each. Spore size was largest amongmorphologically simple groups (Oxymitraceae + Ric-ciaceae + Aytoniaceae + Conocephalaceae + Mono-cleaceae), whereas smaller sizes were concentrated inthe basal clades. Conversely, gametophyte branchlength showed lower values in younger clades andhigher values among deep nodes. The number ofspores per capsule tended to be lower at distant groupsand in the basally placed group A (Fig. 7).

Discussion

In the current work, we present the results of thelargest combined analysis for the complex thalloid liv-erworts. Novel sources of morphological informationwere evaluated including continuous and structuralcharacters; features used in previous studies were re-interpreted. The results obtained from the combineddata corroborate many of the recent proposals butcast doubt on the monophyly of some families. The

relative contribution of different types of morphologi-cal characters indicate that sporophytic characters pro-vide considerable information for low taxonomiclevels. The exhaustive morphological character sam-pling allows improvement of the analysis in terms ofdata congruence, sheds light on phylogenetic relation-ships among families, and improves the diagnoses ofhigh-level groups. The main taxonomic and systemat-ics conclusions presented below are derived from thecombined tree under BW. A discussion on group sta-bility and character contribution derives from theresults of the partitioned analyses.

Monophyly and stability of groups

Most of the deep relationships within the subclassMarchantiidae are in agreement with previous studies(Wheeler, 2000; Boisselier-Dubayle et al., 2002; Forrestet al., 2006; Villarreal et al., 2016). The orderMarchantiales, as conceived by Crandall-Stotler et al.(2009) or Bischler (1998), is not retrieved as mono-phyletic in any of our analyses (Fig. 1). After finding aclose association between Monocarpus and Sphaero-carpales, Forrest et al. (2015) and Villarreal et al.(2016) rejected Marchantiales sensu Crandall-Stotleret al. (2009), which agrees with our finding of a well-diagnosed Clade A (Monocarpus + Sphaerocarpales;Figs 1 and 5).The present analyses recovered several interesting

groups above the family level (Figs 1 and 5). The ear-lier removal of the genus Lunularia from Marchan-tiales led to the proposal of the order Lunulariales(Long, 2006; Crandall-Stotler et al., 2009). Relation-ships among high-level taxa remained unresolved

0.00

0.25

0.50

0.75

1.00

Family Inter−family Order Subclass Class

PhaseGametophyteSporophyte

Fig. 6. Percentage of synapomorphies per ontogenetic phase per taxonomic category. C = class; SC = subclass; O = order; F = familyand X = inter-family. Proportion of sporophytic synapomorphies mapped along the combined tree is included as Fig. S2 in the Supplementarymaterial.


Fig. 7. Character mapping of selected adaptive features across ingroup (Combined tree under BW). Branches coloured according Bischler’s life-history groups. Selected characters shown as continuous characters below branches [branch length (cm)/spore size (mm)] and coloured symbols.Grey symbols and dotted branches represents ambiguous optimisations.


throughout different studies (Crandall-Stotler et al.,2009), and thus the proposal of four different orderswithin Marchantiidae seemed appropriate (Sphaero-carpales, Neohodgsoniales, Lunulariales and Marchan-tiales). However, Villarreal et al. (2016) found a closeassociation between Lunulariales and Marchantiales(except Monocarpus). Our analyses recover the samegroup, Clade B (Figs 1 and 5), which differs fromothers definitions of Marchantiales (Bischler, 1998;Crandall-Stotler and Stotler, 2000; Long, 2006; Cran-dall-Stotler et al., 2009) in excluding Monocarpus andincluding Lunularia (as Lunulariales). Although thedistinction of two different orders (Lunulariales andMarchantiales) is not necessarily contradicted, it doesnot seem to be completely suitable given the presentresults. This is strengthened by the fact that the nodecomprising Marchantiales (except Monocarpus) lacksdiagnostic characters and Lunularia only shows a lownumber of autapomorphic changes (Fig S1 andTable S2).Many of the clades recovered at mid-taxonomic

levels by Villarreal et al. (2016; i.e. those whichgrouped two families) are also found in the presentstudy. The Clade F (Riccia + Ricciocarpos + Oxymitra;Fig 5) was formerly considered as an order (Crandall-Stotler and Stotler, 2000) or suborder (Bischler, 1998).This clade was previously recovered with high supportvalue (Villarreal et al., 2016), and our results agreed inthe three analyses (combined, molecular and morpho-logical data; Fig. 1). The rest of the interfamilialgroups have not been recognized formally in modernclassifications. Clades E (Conocephalum + Monoclea)and C (Wiesnerella + Targionia; Fig. 5) are both highlysupported and stable in the combined and moleculardata. Clade D (Stephensoniella + Aitchisoniella + Exor-motheca + Corsinia + Cronisia) is exclusively recoveredby our combined and molecular data (Figs 1 and 5),contradicting recent changes (see below).The outcomes of the present study show discrepan-

cies with the recently made nomenclatural changes atthe family level and below (Long and Crandall-Stotler,2016; Long et al., 2016a,b). The genus Aitchisoniellawas recently transferred to Cleveaceae (Long et al.,2016b) on the basis of its sister relationship with theremaining genera of the family (Villarreal et al., 2016).In contrast to the results of Villarreal et al. (2016), theposition of Aitchisoniella within Cleveaceae is not sup-ported by our data (Fig. 1; see Sensitivity plots). Inour analyses, on the one hand, Aitchisoniella wasresolved as sister to Stephensoniella with high support(SR 100) and clearly nested in Corsiniaceae (Fig. 1;Clade D in Fig. 5). Exormotheca, on the other hand,was recovered as sister to Corsinia and Cronisia in theanalyses based on both the combined and moleculardata (Fig. 1). Morphology, nonetheless, produces puz-zling results regarding Corsiniaceae. As in Villarreal

et al. (2016), morphological trees recovered Exormoth-eca and Stephensoniella as sister taxa but these wereunrelated to the remaining Corsiniaceae (Fig. 3).Hence, the nomenclatural changes proposed to bothCleveaceae and the genus Stephensoniella (Long et al.,2016a,b) are not supported by the results of our analy-ses. Instead, our results suggest that Corsiniaceaeshould be reviewed to accommodate Aitchisoniella,and that Stephensoniella and Exormotheca are actuallyindependent taxa.The genera Corsinia and Cronisia were found to be

sister taxa by the first time in a combined analysis(Fig. 1). Although this clade was resolved here with amoderate support value, it was found in most of theanalytical conditions (13 out of 17); showing a highstability (Fig. 1). The fact that Villarreal et al. (2016)recovered a clade consisting of paraphyletic “tradi-tional” families (Corsiniaceae and Exormothecaceae;fig. 2 in Villarreal et al., 2016), led to a logical rear-rangement of these groups (Long et al., 2016a,b).Compared to previous studies (Boisselier-Dubayleet al., 2002; Forrest et al., 2006; Villarreal et al.,2016), the inclusion of extensive morphological dataturned over many relationships at the genus level.Unlike Stephensoniella and Exormotheca (Villarrealet al., 2016), the novel link between Cronisia and Cor-sinia is supported by a large number of morphologicalcharacters. In addition, both genera have few autapo-morphic characters (Table S2). Altogether, this makesCronisia and Corsinia suitable taxa for being mergedunder a single generic name.As Crandall-Stotler et al. (2009) pointed out, trying

to solve the deep relationships inside Marchantiidaehas commonly challenged researchers. Our resultsshow Clade B (Marchantiales and Lunulariales; Fig. 5)is an unstable clade in the molecular dataset andabsent in the morphological trees (sensitivity plots inFig. 1). By contrast, the Clade B showed up as astable group in the combined analysis. These outcomessuggest that adding morphological data to moleculardatasets improves the stability of the clade regardlessof the overall incongruence between partitions. Addi-tionally, Clade B received a higher support value whenmorphological data was included. Similarly, supportvalues of other less-inclusive groups (Sphaerocarpales,Ricciaceae and Corsiniaceae) were significantlyimproved after the addition of morphology (Table S3).These results highlight the fact that extensive morpho-logical data can successfully capture similar patterns tothose obtained with molecular data. Even more, unsta-ble or poorly supported results derived from moleculardata can be significantly improved after the additionof morphology.The differences in the results as compared with pre-

vious studies may be a consequence of both the addi-tion of new data and the different methods employed


to derive the phylogenetic hypotheses. It has beenpointed out that nodes with low support may be sensi-tive to changes in analytical conditions (Giribet, 2003;Miller and Hormiga, 2004). Under this reasoning, aclade founded on scarce evidence could be unstable tothe addition of new conflicting characters. The differ-ences between our results and previous phylogeniesfocused mainly on Corsiniaceae and Cleveaceae. InVillarreal et al.’s (2016) phylogeny, the groups Corsini-aceae + Exormotheca and Exormotheca + Stephen-soniella had low to moderate support (BS <50 and 79,respectively). In our study, these taxa are highly unsta-ble, especially Exormotheca (see sensitivity plots inFig. 1). Additionally, recent phylogenies were mostlyconducted under maximum-likelihood or Bayesiananalyses (Forrest et al., 2015; Villarreal et al., 2016),but implied weighting was never used for analysingdata of this group. Thus, the differences in the resultsmight be at least partially attributed to the differentanalytical methods.

Contribution of data, conflict and agreement

Boisselier-Dubayle et al. (2002) and Crandall-Stotleret al. (2005) have previously analysed the incongruencein results based on different data types in Marchan-tiales. Even if exhaustive, such comparisons did notquantify the degree of conflict and contribution ofspecific groups of morphological characters to recoverclades. The topological comparisons (SPR distanceand shared nodes) carried out in the present analysisshowed the high similarity of the molecular trees withthe combined tree. Boisselier-Dubayle et al. (2002) sta-ted that given a large amount of molecular data, suchan outcome should be expected. However, theextension to which morphological results depart frommolecular trees has been rarely evaluated. In thisstudy, the SPR measures indicated that our morpho-logical trees were similar to the molecular trees (8.1SPR movements; Fig. 3; Table 2). The comparisonsbetween the molecular trees and the morphologicaltrees of Bischler (1998) and Boisselier-Dubayle et al.(2002) yielded SPR values above 9 (Table 2). In termsof shared nodes, Bischler’s tree is similar to our BWmolecular tree (Table 2). This similarity was explainedby the monophyly of the traditional Exormothecaceaein both Bischler’s tree and our BW molecular tree.The alternative comparison, using Villarreal et al.’s(2016) tree as reference, retrieved the K7 morphologi-cal tree as the most similar (Table 2). Consequently,our K7 morphological tree maximized similarity withmolecular hypotheses in three out of four compar-isons.It has been formerly stressed that morphology-based

trees and molecular-based trees rendered markedly dif-ferent topologies (Boisselier-Dubayle et al., 2002).

Although this conflict is also reflected in our trees(Fig. 3), the K7 morphology-based tree has both dis-similarities with previous morphological trees andcommon aspects with molecular trees (Bischler, 1998;Crandall-Stotler and Stotler, 2000; Boisselier-Dubayleet al., 2002). As previous authors have pointed out,morphological trees tended to place morphologicallycomplex species in more distant positions regardingSphaerocarpales (Boisselier-Dubayle et al., 2002; For-rest et al., 2006; Crandall-Stotler et al., 2009). In con-trast, our morphological trees somewhat resemble thetopology of molecular trees (Fig. 3; see online supple-mental material). In the K7 morphological tree, themorphologically simple clade F was recovered in a dis-tant position regarding Sphaerocarpales (Fig. 3); con-trasting with earlier morphological phylogenies(Boisselier-Dubayle et al., 1997, 2002; Bischler, 1998;Crandall-Stotler and Stotler, 2000). The genera Corsi-nia and Cronisia were closely related to group F, as inForrest et al. (2006). The “traditional” Exormothe-caceae was rejected (Aitchisoniella + Stephen-soniella + Exormotheca; as in Villarreal et al., 2016)and Neohodgsonia was excluded from Marchantiaceae(as in Forrest et al., 2006). Nonetheless, our morpho-logical trees also have some coincidences with formermorphological trees. For example, they exclude Mono-clea from Marchantiales and do not support Lunulari-ales (as in Crandall-Stotler and Stotler, 2000). In thissense, results from our morphological data can beinterpreted as intermediate between the equally-weighted morphological trees of previous studies (Bis-chler, 1998; Crandall-Stotler and Stotler, 2000; Boisse-lier-Dubayle et al., 2002) and various molecularhypotheses (Boisselier-Dubayle et al., 2002; Forrestet al., 2006; Villarreal et al., 2016; and this study).The most in-depth analysis to elucidate the exten-

sion of data conflict between sets of morphologicalcharacters was performed by Crandall-Stotler et al.(2005). They concluded that sporophytic characters arenot more informative than gametophytic characters. Inour study, the topological similarity with the moleculartree is maximized by the tree derived from the gameto-phytic characters (Fig. 4). Nevertheless, this does notimply that sporophytic characters are uninformative.Although some groups are in conflict compared withtrees derived from other kind of characters (e.g.Sphaerocarpales’ nested position into Marchantiales),several taxonomic groups in the molecular tree arealso found in the sporophyte tree but not in the game-tophyte tree (Fig. 4). Clade F was found in the com-bined analysis and also in the sporophytic tree, inagreement with previous phylogenies (Bischler, 1998;Crandall-Stotler and Stotler, 2000; Wheeler, 2000;Boisselier-Dubayle et al., 2002; Forrest et al., 2006;Villarreal et al., 2016; Fig. 4). Similarly, the familyAytoniaceae is supported by the sporophytic data but


rejected by gametophytic characters (Fig. 4). Sporo-phytic traits also resolved two additional clades inaccordance with molecular hypotheses (Fig. 5): groupsC (Wiesnerella and Targionia) and A (Monocarpus andSphaerocarpales; Wheeler, 2000; Boisselier-Dubayleet al., 2002; Forrest et al., 2006; Villarreal et al.,2016).Our data show that trees based on morphological

data have a certain level of agreement with the molec-ular topology in terms of SPR distance, shared nodesand groups placement. The increased similarity com-pared to previous morphological datasets is probablyrelated to the inclusion of new information and the re-interpretation of previous characters. Until now, mor-phological datasets were relatively small (43 discretecharacters) and none of the larger matrices was con-centrated on Marchantiidae (Crandall-Stotler and Sto-tler, 2000; He-Nygr�en et al., 2006). In addition, it isworth noting that Boisselier-Dubayle et al. (1997) pre-viously suggested using weighting approaches fordiminishing the effect of incongruence between datatypes. Homoplasy reported by earlier works (Boisse-lier-Dubayle et al., 2002; Crandall-Stotler et al., 2005)was now downweighted by the use of implied weight-ing (Goloboff, 1993). Altogether, these factors can beconsidered as improvements in the analysis of morpho-logical data.

Synapomorphies and Diagnosis

Crandall-Stotler et al. (2009) remarked on the factthat some characters could not be confidently assignedto several groups within the current classification. Manyof the groups diagnosed for the first time in the presentstudy were already recovered in previous analyses (For-rest et al., 2006; Villarreal et al., 2016), yet they couldnot be fully evaluated on the basis of morphologicalcharacters. Consequently, their morphological definitionremained dubious. In the present combined study, thediagnosis of several groups was clarified.In the contemporary classification (Crandall-Stotler

et al., 2009), Marchantiopsida and Marchantiidae weredescribed in general terms. That is, some characterswere actually apomorphic (e.g. cuneate apical cell oruni-stratose capsule wall; Crandall-Stotler et al., 2005)whereas others were non-apomorphic traits (e.g. plantsthalloid, rarely leafy or dehiscence by valves, lid or cleis-tocarpous). In our BW combined tree, these groups aresupported by nine morphological characters each,mainly gametophytic features (Fig. 6; Table 3;Fig. S2). Clade B (Marchantiales and Lunulariales;Fig. 5) has not been diagnosed since its original recov-ery (Wheeler, 2000; Boisselier-Dubayle et al., 2002;Forrest et al., 2006). In the present combined analysis,such a node is delimited by 15 synapomorphies, mostof these scored from the gametophytic phase.

Hexagonal/pentagonal scale cells, lanceolate shapedscales and thin pegged rhizoids are examples of diag-nostic characters for this group.Most of the previous analyses (Wheeler, 1998, 2000;

Forrest et al., 2006; He-Nygr�en et al., 2006; Villarrealet al., 2016) recovered Clade F (Oxymitra, Ricciocarposand Riccia; Fig. 5). However, it was not recognizedwithin the contemporary classification (Crandall-Stotleret al., 2009). This group was defined almost exclusivelyby sporophytic characters in previous morphologicalanalyses (Bischler, 1998; Boisselier-Dubayle et al.,2002). In our study, this node is diagnosed by seven mor-phological synapomorphies, four of them being gameto-phytic traits. Thus, the number of synapomorphies notonly increased but also new gametophyte-related charac-ters are added. A distinctive diagnostic trait is the ratiobetween apex and base width of 1.8–2.2; indicating anobcordate thallus shape. A 0.3–0.4 proportion of peggedrhizoids/smooth rhizoids is likewise diagnostic for thisclade. Antheridia (male gametangia) primarily posi-tioned in anacrogynous clusters (not derived from anapical cell; as opposed to being gathered in receptacles orspecialized branches) is a further gametophytic synapo-morphy of this node.Molecular evidence has consistently recovered both

Clades E (Monoclea and Conocephalum; Forrest et al.,2015; Villarreal et al., 2016) and C (Wiesnerella andTargionia; Boisselier-Dubayle et al., 2002; Forrestet al., 2006; Villarreal et al., 2016); but none of thesestudies could provide synapomorphic morphologicalcharacters diagnosing both clades, as found here.Clade E is diagnosed by the absence of a germinaltube, whereas C is defined by three synapomorphies: aratio of 0.7–0.9 between apex and base width, a 0.76–0.77 proportion of pegged rhizoids and semi-annularcapsule thickenings (as opposed to annular thickeningsin remaining clades).The results obtained in the present analysis share

many nodes with previous phylogenetic hypotheses(Forrest et al., 2006, 2015; Villarreal et al., 2016)although the diagnoses of some nodes differ from thatpreviously proposed. Many characters were potentiallylinked to the recently proposed Corsiniaceae (Exor-motheca + Corsinia + Cronisia; Long et al., 2016a).However, only two characters are synapomorphies ofthis clade: pegged rhizoids originating near the thallusapex and a multiple-of-8 chromosome number. Nospore-related character supported the link betweenExormotheca and Cronisia + Corsinia as suggested(Long et al., 2016a). The novel Clade D, whichincludes Aitchisoniella and Corsiniaceae (Exormotheca,Stephensoniella, Cronisia and Corsinia), was diagnosedby two vegetative traits: antheridia in anacrogynousclusters and drought tolerance. Nevertheless, it mustbe noted that the diagnosis of Clade D is not compa-rable to the new Corsiniaceae (Long et al., 2016a)


because it comprises different taxa (i.e., includes Aitch-isoniella). Clade A (Monocarpus and Sphaerocarpales;Fig. 5) was recently found by Forrest et al. (2015)who proposed that this group could be described onthe basis of sporophytic trait reductions. Our resultsconfirmed Forrest et al.’s proposal by mapping rever-sions as synapomorphic character changes (cleistocar-pous capsules, no elaters and distinctiveornamentation of the spore proximal face).Diagnoses of the remaining families with more than

one genus were also modified relative to Crandall-Stotleret al.’s (2009) classification, as a result off Aytoniaceaeand Cleveaceae (excluding Aitchisoniella) constrainingtheir original delimitations. Nevertheless, 10 and eightsynapomorphies are found for each taxon, respectively.Many of these apomorphies were already proposed byCrandall-Stotler et al. (2009) whereas others are com-pletely new characters. For instance, a 1.09–1.1 peggedrhizoids/smooth rhizoids proportion and simple peggedrhizoids distributed in stalk furrows are new synapomor-phies for Aytoniaceae. Cleveaceae (excluding Aitchiso-niella) is fairly well diagnosed by a high peg density (9.4–11.0), mid to long pegs (4.5–4.6) and tuber presence,among others. Marchantiaceae rendered as apomorphiesmany of the characters previously proposed by Crandall-Stotler et al. (2009). However, novel characters relatedto pegged rhizoids are also added. Ricciaceae, as was thecase in several classifications (Bischler, 1998; Crandall-Stotler et al., 2009), is circumscribed mainly by reversals(absences).Two additional features characterize the new diag-

noses presented in this study: a high proportion of sporo-phytic characters and input of continuous characters.Bischler (1998) documented an important number ofsynapomorphies related to sporophytic characters.Namely, 52% of her sampled sporophytic characters pre-sented apomorphic states (Bischler, 1998). That percent-age was informative neither on the ubiquity of changesnor the proportion of sporophytic apomorphies pernode. After a scrutiny of the synapomorphies reportedby Bischler (1998), it turned out that the sporophyticphase contributed 46 out of 126 changes (36%; mainlyconcentrated at deep nodes). In our analysis, synapo-morphies are dominated by gametophytic features at thefamily level (Fig. 6; Fig. S2). These quantities may coin-cide with the notion that sporophytic traits providescarce information at low taxonomic levels (Schuster,1984a,b; Crandall-Stotler and Stotler, 2000). However,this idea is not supported when the synapomorphies pre-sented at the level of orders and nodes that grouped fami-lies are considered. These nodes were diagnosed by alarge number of sporophytic traits (Fig. 6; Fig. S2).Conversely, sporophytic features are poorly representedin the diagnosis at the level of subclasses and classes. Therelatively high number of sporophytic synapomorphiesat low taxonomic levels compared to that obtained in

Bischler (1998) could be related to the higher number ofsporophytic characters included in our dataset. Indeed,Bischler’s (1998) dataset included 12 sporophytic charac-ters whereas our matrix scored 34 sporophytic traits.These new findings provide a basis upon which newgroups could be proposed relying on both high supportvalues and a clearly stated diagnosis.Several synapomorphies retrieved in the present

analysis involve continuous characters, a novel resultgiven that previous phylogenetic analyses in this groupdid not include this type of character. The lack of con-tinuous characters in earlier studies (Bischler, 1998;Boisselier-Dubayle et al., 2002) can be explained bythe absence of a method that could deal with continu-ous variation, because Goloboff et al. published theirapproach later, in 2006. However, recent studies haveexplicitly suggested that continuous characters beexcluded. Oyston et al. (2016) argued that at highertaxonomic categories (� deeper nodes), taxa tend todiffer more contrastingly than at lower levels (� shal-low nodes). Consequently, Oyston et al. claimed thatclassical morphometrics (continuous characters) arenot suitable for deep levels. Discretized characterswere considered better because they exhibit a gap inthe continuous trait being evaluated (Oyston et al.,2016). The analysis of continuous characters analysedas such (Goloboff et al., 2006) led us to diagnose 16out of 35 ingroup nodes using this type of data.Indeed, even if the consensus is poorly resolved, somenodes at the family level and at middle level are recov-ered when continuous characters are used to infer phy-logenetic relationships (Fig. S4). Thus, our findingscontradict the notion that continuous traits are appro-priate only for shallow taxonomic levels. Further sur-veys on continuous characters should be conducted inorder to completely elucidate their relevance in liver-wort phylogeny.

Life history traits, morphological resemblance andevolutionary scenarios

Although some morphological traits were slightlydecoupled regarding life-history groups, a general cor-related pattern is clearly recovered (Fig. 7). Bischler(1998) established such life-history groups on thegrounds of a statistical analysis of environmental fac-tors and morphological features. By mapping thesegroups onto her morphological tree, she suggested thatmorphology and life strategies co-variation is phyloge-netically structured.1 Therefore, morphological

1

Note that Bischler (1998) changed group labelling in her phylo-

genetic mapping (fig. 15 in Bischler, 1998). However, the groups

remained the same (e.g. Marchantiaceae and Conocephalaceae were

first placed in group 2 (Bischler, 1998, p. 135) and then in group 4

(p. 136)).


resemblance was argued to be caused by ancestryrather than by environmental pressure. The characterreconstruction in our combined tree is in partial agree-ment with Bischler’s hypothesis. Explaining the mor-phological diversity in terms of common ancestrywould require each life-history group to be mono-phyletic or paraphyletic. Because groups II–IV had aunique origin, morphological resemblance within eachgroup could be attributed to common ancestry. Simi-larly, within each clade of Group I taxa tended to behighly similar with regard to morphological characters(Fig. 7). Environmental pressure could be consideredto explain morphological diversity among unrelatedtaxa. Unlike members of the same clade, similaritybetween unrelated taxa could not be caused by com-mon ancestry. The resemblance between, say, the gen-era Riccia and Monocarpus is easier to explain interms of similar environmental pressure (Fig. 7). Ourfindings, therefore, allow Bischler’s hypothesis to bereformulated. That is, based on the current phyloge-netic patterns, morphological resemblance is partiallyexplained by common ancestry (inside each clade) andpartially by convergence (between unrelated membersof the group I).Multiple independent character reductions occurred

in taxa of Group I, commonly limited to open dryhabitats (cleistocarpous capsules, small gametophytes,few and large spores). These features are favouredunder environmental stress (Bischler, 1998; Bischleret al., 2005). Conversely, members of groups II–IVtended to develop morphologically complex features(larger gametophytes, noncleistocarpous capsules,numerous and small spores), appropriate for mesichabitats (Bischler, 1998). Thus, the correlation betweenmorphological traits and life strategies found by Bis-chler (1998) is confirmed.Deviations from the expected evolutionary trends

(Bischler, 1998) were a consequence of both dissimilartopologies and methodological issues. The statisticalsurvey of Bischler (1998) was conducted without con-sidering phylogeny as a source of variation constraint.Rather, species were treated as independent statisticalentities. In order to evaluate the concerted evolutionof potentially adaptive characters, the effect of thephylogeny should be eliminated. By doing so, speciescould be treated as statistically independent units(Felsenstein, 1985). An exhaustive quantitative evalua-tion of putative adaptive characters is still needed. Atthe moment, the adaptive value of morphologyremains a poorly investigated area.

Final remarks

The first in-depth combined analysis for the complexthalloid liverworts was conducted. An improved

character sampling led to the construction of the lar-gest morphological dataset. Key findings were achievedregarding morphological contribution and diagnosisimprovement. In sum, the well-established assumptionthat morphology may produce completely incongruentpatterns with molecular data was discredited. Indeed,many nodes recovered with low support values in pre-vious studies were retrieved by our combined datawith increased support. This suggests that the combi-nation of apparently conflicting data types may revealhidden support for most of the groups.It is argued that common weaknesses of morpholog-

ical data are challenging character circumscriptionand, for this specific group, no informative changes(Boisselier-Dubayle et al., 2002). Morphological dataare often described as highly homoplastic in plants(Buck et al., 2000; Ranker et al., 2004; Liu et al.,2012; Yu et al., 2013; Wu et al., 2015; among others).Such a characterization became a widespread notionfor different groups of organisms. Scotland et al.(2003) even suggested that, regardless of the taxonomicgroup, morphological characters should not be evalu-ated given their problematic nature. Our results, aswell as others’ (e.g. Wahlberg et al., 2005), stronglyreject such a statement. The inclusion of more andnovel characters produced topologies which differfrom small-matrix-derived morphological phylogeniesand are more similar to molecular trees. Likewise, itwas shown not only that the number of diagnosedgroups increased, but also that most of the previouslyproposed diagnoses were imprecise.Several synapomorphies at the interfamily level were

found for the first time and some unsupported taxo-nomic changes were questioned. Although deep rela-tionships among derived families are still dubious,diagnoses and support values of many interfamilynodes were improved. Subsequently, this could betranslated into new groups gathering derived families.The taxonomy of the families Cleveaceae and Corsini-aceae, as well as the genera Aitchisoniella and Stephen-soniella, shall be reviewed. At the present, finding aproper set of synapomorphies for the order Marchan-tiales is still a major problem. A reasonable approachwould be to redefine Marchantiales so as to include itssister taxon Lunulariales. By doing this, a supportedand accurately diagnosed category could be proposed.The fact that morphology-derived trees improved theircongruence with molecular evidence (increased valuesof shared nodes and SPR distance) will encourage thesurvey of more and new morphological characters.

Acknowledgments

We thank Emilio Cano (Real Jard�ın Bot�anico) forhis technical assistance in extracting and amplifying


plant material. We are also indebted to the curators ofthe PC herbarium for providing invaluable specimens.Several colleagues also contributed fresh material col-lected in northern Argentina. J.R.F. is supported by aDoctoral Fellowship of the National Scientific andTechnical Research Council (CONICET). This studywas funded by FONCyT (PICT-1838, PICT-1930 andPICT -0810), the National University of Tucuman(PIUNT-G524) and CONICET (PUE 0070). TNT isfreely available thanks to the Willi Hennig Society.

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Supporting Information

Additional Supporting Information may be found inthe online version of this article:Fig. S1. Additional support measures.Fig. S2. Proportion of synapomorphies derived from

the sporophyte at each diagnosed branch.Fig. S3. Strict consensus of trees derived from the

ten continuous characters under K3.Table S1. Primers for sequenced genes.Table S2. Synapomorphies for each node of the

combined tree under block weighting (node numbersin Figure S1).Table S3. Support values for selected groups.


Character definition

Characters used in the present study. Characters already analysed by previous authors

(Bischler, 1998; Crandall-Stotler and Stotler, 2000) were modified; they are also

described as such. “*” and “**” refer to additive and continuous characters,

respectively. A minimum of 7-10 replicates for each continuous measure were

quantified. When possible, each of such replicates were measured from a different

specimen:

0. Plant branches length (cm)**: liverworts usually exhibit a broad range of sizes.

Sometimes discriminating a single non-clonal individual from a ramet is not

straightforward. Then, branch length (and not gametophyte length) was

considered as a more reliable measure. Longitude of branches was measured (in

cm) by taking the linear distance from the thallus apex down up to the first

dichotomy point.

1. Thallus width, apex/base ratio (ct)**: thallus width was measured as the ratio

between the width at the apex and base (apexw/basew) in cross section. “Base”

is understood as the dichotomy (ramification) point.

2. Thallus broadness, apex/base ratio (plane)**: thallus width, in surface view, was

measured by taking the distance among margins of the thallus at the apex and

the ramification point. Posteriorly, the ratio among such values were calculated.

A value below 1 indicated a cordate shape (as in Ricciaceae or Oxymitraceae)

whereas a value higher than 1 designated an obocordate layout. Linear forms

were about a value of 1.

3. Pegged/Smooth rhizoids, rate**: the proportion of pegged rhizoids was

computed by counting the number of pegged and smooth rhizoids within an area

of 150x150 µm (in 40x).

4. Pegged rhizoids, pegs density**: pegs density was calculated by counting the

number of pegs per 25 µm of pegged rhizoids longitude.

5. Peg length (µm)**: longitude of pegs was measured in 100x, from pegs tip up to

rhizoid wall beginning.

6. Pegged/Smooth rhizoids, spatio-temporal development pattern (mm) **:

according to Duckett et al. (2014) pegged rhizoids develop earlier than smooth

rhizoids. As a consequence, smooth rhizoids are far from the thallus apex.

Distance between thallus tip and smooth rhizoids might reflect heterochronic

patterns associated with adaptive strategies. The distance between the thallus

apex and the firsts smooth rhizoids were measured as a linear distance (mm).

The appearance of smooth rhizoids was considered when the proportion of

observed smooth rhizoids was about 5% of the proportion observed in old

portions of the thallus.

7. Pegged rhizoids, diameter of pegged/smooth rhizoids **: the ratio between the

diameter of pegged and smooth rhizoids calculated plane view (40x).

8. Male branch length (cm) **: the longitude of the branch carrying antheridia was

measured from its base up to the apex. For non-autoicous species, branches are

considered female and a missing entry is scored for the species.

Gametangiophores and ordinary branches are homologous structures; so they

are scored as different organs. In strict autoicous species, sexual branches are

considered different. When a female branch is associated with antheridia (as in

Asterella), both branches are considered independently. If the species develops

gametangiophore; the antheridiophore is considered a male branch.

Adventitious branches are also measured if carrying antheridia.

9. Female branch length (cm) **: as for character eight, branches are measured

from the base up to the apex. Archegoniophores and innovations are also

considered.

10. Thallus-Stem, gametophyte growth form: (0) leafy, mainly uni-stratose; (1) costa

and winged thallus; (2) multi-stratose thallus; (3) leaf, mainly pluri-stratose.

Gametophyte could be arranged in two different groups (leafy and thallus).

Gametophytes developing leaf-like expansions, were scored as uni- (0) or multi-

stratose (3) when leaf-like expansions are almost completely uni-stratose

(excepting their base) or are almost completely multi-stratose (excepting the

apex of the expansions). Thallus not producing a distinctive costa were scored as

multi-stratose (2).

11. Thallus-Stem, protonema: (0) filamentous, not heterotrichous; (1) globose; (2)

flattened plate; (3) long, cylindrical.

12. Thallus-Stem, apical cell geometry: (0) tetrahedral, dorsal; (1) tetrahedral,

ventral; (2) lenticular; (3) cuneate; (4) hemidiscoid.

13. Thallus-Stem, apical cell ventral face development: (0) broad ventral face; (1)

narrow ventral face.

14. Thallus-Stem, epidermal cells mucilage secretion: (0) absent; (1) present.

Epidermal cells mucilage-like secretion (as opposed to the parenchymatic cells

secretion). Typical of Haplomitriopsida.

15. Thallus-Stem, apical cell division plane: (0) parallel to apical cell wall (1) oblique

to cell wall.

16. Thallus-Stem, apical cell derivate first division: (0) periclinal (1) anticlinal.

17. Thallus-Stem, phyllotaxy at apex: (0) one third; (1) two fifths; (2) one half; (3)

none

18. Thallus-Stem, apical cell tilt: (0) absent; (1) dorsal; (2) ventral. Generally, apical

cells exhibit a tilt towards the dorsal or ventral surface. Cell tilt is usually reflected

in the plant bending.

19. Thallus-Stem, pitted parenchymatic cell (water conducting cell): (0) absent; (1)

thin walled, simple perforation; (2) thick walled, simple perforation; (3) thick

walled, pit fields. Parenchymatic cells often develop simple perforation (1, 2)

regardless of the cell walls thickness. However, sometimes cells can develop pit

fields in remarkably thickened cells (3).

20. Thallus-Stem, oil body occurrence: (0) absent; (1) many, in all cells; (2) one, in all

cells; (3) in idioblastic cells.

21. Thallus-Stem, schizogenous mucilage cavities: (0) absent; (1) present.

22. Thallus-Stem, food conducting cells: (0) absent; (1) specialised parenchyma.

23. Thallus-Stem, gemmae: (0) absent; (1) blastic; (2) discoid cups; (3) crescent shape

cups. Modified from Crandall-Stotler and Stotler (2000) to account for gemmae

types of Lunularia (3) and Marchantia (2).

24. Thallus-Stem, tubers presence: (0) absent; (1) present, apically; (2) present,

elsewhere.

25. Thallus-Stem, chromosome number: (0) multiple of 9; (1) multiple of 8; (2)

multiple of 5.

26. Thallus-Stem, Nostoc symbionts: (0) absent; (1) exogenous, in domatia. Domatia

are modified sheltered projections; in Blasia these structures host Nostoc

symbionts.

27. Thallus-Stem, drought tolerance: (0) no; (1) yes.

28. Thallus-Stem, air chambers: (0) absent; (1) vestigial; (2) Marchantia type; (3)

Riccia type; (4) Reboulia type; (5) pseudo-Reboulia type. Hassel de Menéndez

(1963) recognised that many Riccia species develop Reboulia-like air chambers

but are ontogenetically different from those of Reboulia. This pseudo-Reboulia

type primarily originates as uni-stratose chambers; following in the ontogeny, it

turns backwards so that air chambers superpose. The development can be

observed in a longitudinal section at the apex level.

29. Thallus-Stem, air pores on thallus: (0) absent; (1) simple, not elevated; (2) simple,

elevated; (3) compound. Two types of simple pores are differentiated. The first

involves a simple opening between epidermal cells (e.g. Plagiochasma rupestre;

1). However, some species shows elevated simple pores (e.g. Lunularia; 2).

30. Thallus-Stem, leaf-wing origin: (0) one central initial; (1) anodic and kathodic

initials; (2) two acroscopic initials. A central initial is simply a parenchymatic cell

which recovers its division capacity (0). Anodic and kathodic initials divide

towards or against the segmentation spiral of the apical cell (1). Acroscopic refers

to initials located next to the apical cell region (2).

31. Thallus-Stem, leaf morphology: (0) undivided; (1) two lobed, simple; (2) two

lobed, complicate and dorsal; (3) more than two lobes; (4) two lobed, complicate

and ventral; (5) none. Leaf laminae could be differentiated into several classes

according to their morphology. Many are completely entire (undivided; 0), or

widely divided. In addition, dorsal (2) or ventral (4) lobes may develop complex

structures.

32. Thallus-Stem, lateral branching: (0) absent; (1) dichotomous; (2) axillary, moss

type; (3) terminal, leaf modified; (4) terminal, leaf unmodified; (5) collared,

Lejeunea type; (6) endogenous; (7) exogenous, intercalary. Branches may be

produced from an internal unique cell within a thallus (1), from an axillary cell

(2), from a leafy stem apex and modifying leaves (3) or not (4), internally from a

leafy stem and producing a collar after protruding (5), developing internally or

externally between leaves but never producing collars (6, 7).

33. Thallus-Stem, ventral branching: (0) absent; (1) terminal, leaf modified; (2)

exogenous, delayed; (3) endogenous.

34. Thallus-Stem, mature ventral appendage: (0) absent; (1) one row; (2) two rows;

(3) more than two rows. Ventral appendages refer to both amphigastria or

scales. These appendages may be arranged in one row (1), two rows (2) or more

than two rows (well-organised or not; 3).

35. Thallus-Stem, ventral ventral appendage constitution: (0) short papillae; (1) long

cilia; (2) foliose, with chlorophyllous cells; (3) foliose, without chlorophyllous

cells. The term “form” refers to the appendage general aspect. Ventral

appendages may be reduced to short inconspicuous papillae (0) or cilia (1). They

may be conspicuously developed into foliose-like structures; carrying

photosynthetic cells or not (2, 3).

36. Thallus-Stem, ventral appendage shape: (0) acute throughout, inconspicuously

rounded or absent; (1) bifid apex, cordate ;(2) bifid apex, rounded; (3) Entire or

almost non-bifid; (4) lanceolate (wider base, thinner apex). Ventral appendages

may express different shapes. An acute appendage differs from a lanceolate

appendage in that its widest part usually has less than three cells. Bifid

appendages may have a cordate or rounded outline (2, 3).

37. Thallus-Stem, scales appendage form: (0) none; (1) filiform (few than three cell

width); (2) somewhat triangular (lanceolate, acute); (3) somewhat ovate or

obtuse (obovate, oblong); (4) reniform; (5) orbicular. Scales usually carry

appendages at their tips. The shape of these appendages can differ between

species or genera.

38. Thallus-Stem, scales appendage number: (0) without scale; (1) zero; (2) one; (3)

more than one. The number of appendages per scale also varies between taxa.

39. Thallus-Stem, medial scales margin: (0) no scales; (1) entire; (2) crenate; (3)

serrulate. Cells of the medial scales margin can vary in shape. Thus, resulting in

different margins shape.

40. Thallus-Stem, scale appendage margin: (0) no appendage; (1) entire; (2) crenate;

(3) serrulate. Similar to 39 but refers to appendage margin.

41. Thallus-Stem, appendage basal constriction: (0) no scale; (1) no appendage; (2)

no constricted; (3) constricted. The insertion point between the appendages and

the scales can be constricted or not.

42. Thallus-Stem, appendage cell form: (0) no scale; (1) without appendage; (2)

somewhat globose; (3) elongate rectangular; (4) hexagonal/pentagonal.

Appendages cells are seen at the middle of the appendage.

43. Thallus-Stem, rhizoids, development type: (0) absent; (1) unicellular, smooth

only; (2) unicellular, dimorphic. Two different types of rhizoids are commonly

present in liverworts. That is, rhizoids with smooth wall or with projections

(pegs). A single species may bear both types (2) or just the smooth one (1).

44. Thallus-Stem, rhizoids, types of pegged rhizoids along thallus: (0) slightly

developed pegs; (1) highly developed pegs (curved shape, non-protruding

papillae union); (2) highly developed pegs (with protruding papillae union); (3)

none. In general, Marchantiales bear pegged rhizoids with relatively small pegs

(e.g. most Aytoniaceae, Cleveaceae; 0). However, larger pegs may be developed

within thicker curved rhizoids. In this case, pegs tend to be large and a peg union

(“papillae union”) may be present. A papillae union is an internal projection

which links opposite pegs (e.g. in Marchantia; 2).

45. Thallus-Stem, pegged rhizoids, pegs internal structure: (0) blunt to pointed; (1)

blunt to branched; (2) hooked; (3) pointed; (4) blunt (5) none.

46. Thallus-Stem, Pegged rhizoids, orientation: (0) none; (1) parallel to thallus

surface; (2) at right angles. Rhizoids associated with water uptake are often

oriented parallel to thallus surface. If such parallel rhizoids were pegged rhizoids,

then species were scored as 1.

47. Thallus-Stem, pegged rhizoids, distribution along the mature plant: (0) none; (1)

ventral or randomly distributed; (2) rhizoids zonation. Pegged rhizoids may be

present only in the ventral surface of the thallus (1) or concentrated in particular

organs (receptacles, gametangiophores, etc.; 2).

48. Thallus-Stem, development related to positional segregation: (0) non-

segregated, pegs equally developed in the whole thallus; (1) segregated, female

stalk pegs more developed; (2) none. If segregated, pegged rhizoids may be

conspicuously larger within the archegoniophore stalk (1).

49. Thallus-Stem, parallel rhizoids/perpendicular rhizoids wall width: (0) equal; (1)

parallel rhizoids thicker. Regardless of bearing pegs, parallel rhizoids can develop

thicker walls than perpendicular rhizoids (1). This is a consequence of being

related to water uptake.

50. Thallus-Stem, rhizoids, pigmentation: (0) absent; (1) present.

51. Thallus-Stem, sexual condition: (0) dioicous, dimorphic; (1) dioicous,

monomorphic; (2) monoicous.

52. Thallus-Stem, perigonial position: (0) randomly scattered, dorsal; (1) in bracts on

main stem; (2) in acrogynous clusters; (3) embedded in receptacles; (4) on

specialised branches; (5) in anacrogynous clusters; (6) on antheridiophores.

53. Thallus-Stem, perichaetial position: (0) randomly scattered, dorsal; (1)

anacrogynous, clustered; (2) acrogynous on main stem; (2) acrogynous on short

branches; (3) on archegoniophores. Archegonia development may imply the loss

of the apical cell function (acrogynous) or not (anacrogynous). In the former

case, archegonia may develop on the main axis (2) or on a short branch (3). If

anacrogynous, archegonia are usually clustered in receptacles or in specialised

branches (archegoniophores). Otherwise, archegonia may be scattered along the

thallus in different types of groups.

54. Thallus-Stem, paraphyses in perigonium: (0) absent; (1) present. Usually,

paraphyses are no larger than sexual organs. They are easily observed within the

same receptacle of the organ sex.

55. Thallus-Stem, paraphyses in perichaetium: (0) absent; (1) present.

56. Thallus-Stem, Archegonia, distal-cell early development: (0) axial cell exposed;

(1) axial not exposed. Approximately at the stage III1 of the archegonial

ontogeny, the distal cell suffers transverse divisions. The new cells may cover the

ovum mother cell (1) or not (2).

1 Stage numbers arbitrarily fixed with Schuster (1984, pp. 859) illustrations as reference.

57. Thallus-Stem, archegonia, jacket cell early divisions: (0) division in two out of

three; (1) division in all the three cells. The three jacket cells initials can suffer

successive division (giving rise to a six-row-cell neck), or only two of them divide.

58. Thallus-Stem, number of archegonia per perichaetium: (0) one; (1) more than

one.

59. Thallus-Stem, archegoniophore, number of rhizoid furrows: (0) without

archegoniophore; (1) zero; (2) one; (3) two. Archegoniophore stalks may carry

several or none furrow. These are easily observed as depressions in the stalk

cross section.

60. Thallus-Stem, archegoniophore, stalk anatomy: (0) none; (1) homogeneous (fully

parenchymatous); (2) heterogeneous (with air chambers). Stalks may develop

photosynthetic air chambers (2) or not (1). This character is visualised in cross

section.

61. Thallus-Stem, archegoniophore stalk elongation: (0) absent stalk; (1) after

fertilization, before spore maturity; (2) after fertilization, at time of spore

maturity; (3) before fertilization.

62. Thallus-Stem, female receptacle anatomy: (0) not stalked receptacle; (1)

homogeneous (parenchymatous); (2) heterogeneous (with air chambers). This

character resembles that character 79 but it is strictly referred to the receptacle

(the structure bearing archegonia).

63. Thallus-Stem, female stalk pore/female receptacle pore: (0) none; (1)

simple/simple; (2) simple/compound; (3) compound/compound. Both stalks and

receptacles can develop pores. As in the thallus surface, these pores may be

simple or compound. The relationship between stalk and receptacle pore types

are observed in cross sections.

64. Thallus-Stem, archegonia, neck cells (CT): (0) four cell rows; (1) five cell rows; (2)

six cell rows.

65. Thallus-Stem, antheridia receptacles limit: (0) not bounded; (1) bounded by

scales; (2) bounded by membrane. Antheridia may be surrounded by scales or

membranes.

66. Thallus-Stem, antheridia development, oblique divisions of the distal cell: (0)

two, obliquely; (1) one, obliquely. During the antheridia development, at the

filamentous stage, the distal cell generates the androgonial initial(s) and the

jackets cell initials. Before producing the androgonial initial, the distal cell

undergoes mitotical divisions. These may be two (0) or one (1) diagonal division.

67. Thallus-Stem, antheridia development, filamentous-stage cell number: (0) four;

(1) 5< x < 7; (2) less than four. At the filamentous stage, each row of the filament

may be comprised by several cells. Typically, Haplomitriopsida have less than

four cells in each row (2) while the other groups are more variable.

68. Thallus-Stem, antheridia development, temporal origin of the bi-seriate stalk: (0)

never; (1) early, after the basal cell differentiation (2) later, after or

simultaneously to the androgonial differentiation. At the filamentous stage, the

stalk may suffer vertical divisions so that it becomes a bi-seriate stalk. These

divisions may occur early in the ontogeny (1), relatively later (2) or never (0).

69. Thallus-Stem, antheridia, stalk anatomy: (0) uni-seriate; (1) bi-seriate; (2) four or

more seriate. At maturity (after the androgonial and jacket cells settlement), the

stalk may suffer additional division. These divisions can produce a multi-seriate

stalk (up to 8 in foliose liverworts).

70. Thallus-Stem, anatomy of stalked male receptacle: (0) not stalked; (1)

homogeneous (fully parenchymatous); (2) heterogeneous (with air chamber).

71. Thallus-Stem, antheridial origin: (0) epidermal; (1) non-epidermal. Antheridia

develop from an epidermal cell while in hornworts antheridia are exclusively

derived from a sub-epidermal cell. However, in Takakia the antheridial origin is

unknown. Therefore, both states were included in this dataset. Takakia was

scored as ambiguous.

72. Thallus-Stem, apical cell in antheridium: (0) absent; (1) present.

73. Thallus-Stem, antheridia development, young antheridia (androgonial cells): (0)

four celled; (1) two celled one celled. After distal cell suffered transversal

divisions [approximately at the stage IV highlighted by Schuster (1984)2],

multiple androgonial initials can be produced. In Marchantiidae four androgonial

cells are typically produced.

2 Stage numbers arbitrarily fixed with Schuster (1984, pp. 853) illustrations as reference.

74. Thallus-Stem, antheridia, jacket cells: (0) irregular, isodiametric; (1) tiered,

elongate. Antheridia jacket cells may be regularly arranged in tiers or irregularly

arranged and being isodiametric.

75. Thallus-Stem, spermatid, spline aperture: (0) three tubules or more; (1) one

tubule, exceptionally two. Spermatid spline exhibits an aperture as a

consequence of tubules shortening.

76. Thallus-Stem, spermatid, LS tapering orientation: (0) left; (1) right. Spline is

usually shortened on its left or right as a consequence of tubules reduction.

77. Thallus-Stem, spermatid, basal body dimorphism: (0) yes; (1) no. Basal bodies are

structurally different between most of the Marchantiidae and remaining

liverworts.

78. Thallus-Stem, spermatid, spline width: (0) 30 or less; (1) more than 50. Spline

width varies among liverworts. Generally, Haplomitriopsida shows a wider spline

79. Thallus-Stem, spermatid, spline coiling: (0) almost non-coiled (one gyre); (1)

scarcely coiled (mora than 1 and less than 2 gyres); (2) pronouncedly coiled (3

gyres or more). Spline is proximally coiled; following the spermatid body outline.

80. Thallus-Stem, spermatid, BB overlapping degree: (0) highly overlapped, almost

totally superimposed; (1) considerably overlapped, nearly the half; (2) almost no

overlapped, less than the half. Basal bodies overlapping refers to a positional

staggering along the longest axis. Overlapping is seen in surface view.

81. Thallus-Stem, spermatid, spline associated with LS: (0) yes; (1) no. Spline may be

decoupled from the Layered Strip (LS) or not.

82. Thallus-Stem, spermatid, notch presence: (0) no; (1) yes. A notch is typically

associated with the spline aperture in marchantioid taxa.

83. Thallus-Stem, spermatid, AM extension at mature: (0) extended; (1) not

extended. The mitochondrion tends to extend its anterior region during

spermatid maturity.

84. Thallus-Stem, spermatid, spline geometry: (0) wider at the anterior region; (1)

wider at the posterior region. Tubules can be added to right region of the spline

anteriorly. This is typical of Haplomitrium.

85. Thallus-Stem, spermatid, ABBs position: (0) on the left; (1) on the right. The

anterior basal body can be located towards the left or right regarding the spline

axis.

86. Thallus-Stem, spermatid, LS/ABB length: (0) less than 1.5; (1) 1.5> x <3.0; (2)

more than 3.5. The ratio between the length of the Lamellar Strip and the

anterior basal body measured from the surface view.

87. Thallus-Stem, transfer cell, gametophyte: (0) absent; (1) present. Transfer cells

are observed as digitated cell walls around the foot.

88. Thallus-Stem, transfer cell, sporophyte: (0) absent; (1) present. Similar to

previous but projections extend towards gametophytic tissue.

89. Thallus-Stem, embryo, protection structure (outer perichaetium): (0) absent; (1)

leaf-like scales; (2) bracts and bracteoles; (3) marchantioid involucre.

90. Thallus-Stem, embryo protection strucutre, position of marchantioid involucre:

(0) dorsal related to thallus; (1) ventral, regarding thallus; (2) no marchantioid

involucre. Involucres can be located dorsally (e.g. Oxymitra; 0) or ventrally (e.g.

Targionia; 1) regarding thallus surface. Monoclea carries involucres at the apex;

this genus was scored as a polymorphism.

91. Thallus-Stem, embryo protection structure (inner perichaetium): (0) absent; (1)

pseudoperianth; (2) perianth; (3) hollow perigynium; (4) marchantioid

pseudoperianth.

92. Thallus-Stem, embryo protection structure (calyptra): (0) absent; (1) true

calyptra; (2) shoot calyptra; (3) vestigial, solid perigynium; (4) epigonium.

93. Thallus-Stem, embryo protection structure (chlorophyllous outer perichaetium):

(0) non-chlorophyllous; (1) chlorophyllous, multi-stratose; (2) chlorophyllous,

uni-stratose. This character refers to the outer perichaetium envelope capacity

to photosynthesise. Multi-stratose envelopes are typical of marchantioid taxa.

Uni-stratose envelopes are generally produced in foliose liverworts.

94. Sporophyte, embryo, development type: (0) filamentous; (1) octant; (2) free

nuclear. In early embryogenesis, cells can be concatenated (0) or within

quadrants (1). In few occasions, protoplasm divides without a correlated

membrane division. Then, the embryo is observed as a conjunction of nuclei

within a single cell (2).

95. Sporophyte, embryo, ontogeny: (0) foot and seta from hypobasal cell; (1) foot

from hypobasal cell and sporangium from epibasal; (2) sporangium from all of it.

In general, the foot and seta of the sporophyte derives from the hypobasal cell

(0). However, seta and capsule (sporangium) may differentiate from the epibasal

cell. The complete sporophyte may be derived from the unique fertilised cell (2).

96. Sporophyte, embryo, apical cell: (0) present; (1) absent.

97. Sporophyte, embryo, foot form: (0) small, bulbous; (1) conical; (2) collared, cup-

shaped; (3) irregular, with palisade layer; (4) irregular, no palisade layer; (5)

absent

98. Sporophyte, sporangial development: (0) determinant; (1) non-determinant.

99. Sporophyte, seta development: (0) absent; (1) from meristem; (2) synchronous,

elongation; (3) synchronous, no elongation. Meristematic-like regions, which

actively divide into new cells, are commonly differentiated in mosses and

hornworts. In liverworts, seta elongation is achieved by synchronic elongation of

seta cells. Elongation may be significant (producing a relatively massive seta) or

almost absent (slightly elongated seta).

100. Sporophyte, seta cell arrangement: (0) non-articulate; (1) articulate. Seta

cells arranged in well-defined tiers are commonly regarded as articulated.

101. Sporophyte, hydroids in sporophyte: (0) present; (1) absent.

102. Sporophyte, leptoids in sporophyte: (0) present; (1) absent.

103. Sporophyte, cruciate seta pattern: (0) absent; (1) present. Seta cells

arranged in a cruciate pattern in cross section.

104. Sporophyte, capsule wall: (0) uni-stratose; (1) bi-stratose; (2) multi-

stratose. Capsule wall layers were observed between the apex and the bottom

of capsule.

105. Sporophyte, capsule internal wall thickening: (0) none; (1) semi-annular;

(2) annular. Internal wall thickenings are observed in the middle of the capsule,

between the bottom and the apex.

106. Sporophyte, capsule external wall thickening: (0) absent; (1) present. At

the apex the capsules, additional wall layers may develop different thickenings

from that of inner layer (Hassel de Menéndez, 1963).

107. Sporophyte, capsule form: (0) cylindrical; (1) ellipsoidal; (2) globose.

108. Sporophyte, capsule dehiscence: (0) irregular; (1) along one suture; (2)

into two valves; (3) into four valves; (4) apically with an operculum; (5)

cleistocarpous. Since reduced sporophytes lack a clear dehiscence mechanism,

capsules must disintegrate before spore releasing. Therefore, they were

considered as cleistocarpous and not as absent [as opposed to Crandall-Stotler

and Stotler (2000)].

109. Sporophyte, capsule stomata: (0) absent; (1) present.

110. Sporophyte, columella: (0) absent; (1) present.

111. Sporophyte, elaters: (0) absent; (1) present, unicellular. The state

“present, multi-cellular” was omitted (Crandall-Stotler and Stotler, 2000).

112. Sporophyte, elaters thickening bands: (0) absent; (1) present.

113. Sporophyte, elaterophores: (0) absent; (1) basal; (2) apical. Groups of

elaters (elaterophore) may be disposed basally or apically onto a capsule valve.

114. Sporophyte, spore, perine layers on spores: (0) absent; (1) present.

115. Sporophyte, spore:elater division ratio: (0) 4:1 (no division); (1) > 4:1

(sporocites divide); (2) < 4:1 (elaterocites divide).

116. Sporophyte, spores mother cell lobes: (0) absent; (1) present.

117. Sporophyte, spore, polarised spores: (0) no; (1) yes. Spores of complex

thalloid liverworts usually have two differentiated faces. The proximal face bears

a trilete scar and shows a conic-like shape while the distal face is distinctly

ornamented.

118. Sporophyte, spore, germination: (0) exosporic; (1) endosporic. Spore may

germinate after wall rupture (1) or before (0).

119. Sporophyte, spore, distal gross morphology: (0) verruculate-like; (1)

baculate or pilate-like; (2) areolate-like, ill-defined muri (open reticulum); (3)

areolate, well-defined muri (closed reticulum); (4) domes-like.

120. Sporophyte, spore, proximal gross morphology: (0) as in distal face; (1)

differentiated from distal face. In general, spore proximal face develops the

same ornamentation pattern as the distal face (0). However, it may show a

striking different pattern (1).

121. Sporophyte, spore, equatorial cingulum: (0) not developed; (1) slightly

developed; (2) highly developed. A cingulum is a projection extending along the

equatorial line and separates the distal face from the proximal face. A cingulum

can be observed as a small linear projection almost empty of ornamentation (1);

or as a large projection carrying well-ornamented walls (2).

122. Sporophyte, spore, equatorial zone: (0) not developed; (1) distally

present. As opposed to the cingulum, a “zone” is a negative ornamentation. That

is, it is not an outgrowth but an invagination of the wall. Usually, it is produced

in the distal face immediately after the equatorial line (1).

123. Sporophyte, spore distal face, basal ornamentation: (0) none (smooth);

(1) rounded projections (verrucose, orbiculate, etc); (2) acute projections

(spines, bacules, etc.); (3) reticulum. The basal ornamentation (base

ornamentation) involves few wall layers than the ornamentation itself. Normally,

it is observed as a small ornamentation pattern within the larger walls (e.g. small

spines above reticulum muri). This base ornamentation encompasses rounded

projections (1), acute projections (2) or even reticulum (3).

124. Sporophyte, spore proximal face, basal ornamentation: (0) none

(smooth); (1) rounded projections (verrucose, orbiculate, etc.); (2) reticulum.

Similar to previous.

125. Sporophyte, spore, germ tube: (0) absent; (1) present.

Table S1. Primers for sequenced genes

Primers for amplified genes. Amplification followed Forrest and Crandall-Stotler (2004).

Gene Primer Direction Sequence

rbcL M007 Forward 5’-CCACAAACGGAGACTAAAGC

rbcL M28 Forward 5’-GTTGTTGGATTTAAAGCTGGTGTT

rbcL MtmRR Reverse 5’-GCTCTA TCCACTGAGCTAC

rbcL M636 Reverse 5’-GCGTTGGAGAGATCGTTTCT

psbA 501F Forward 5’-TTTCTCAGACGGTATGCC

psba trnHR Reverse 5’-GAACGACGGGAATTGAAC

Figure S1. Additional support measures. Additional support measures are shown above

branches (Bootstrapping > 50) and below (Bremer > 0.0). Node numbers are beneath

Bremer values.

Table S2. Synapomorphies for each node of the combined tree under block weighting

(node numbers in Figure S1).

Synapomorphic changes at inner nodes Autapomorphic changes

Node 57 :

No synapomorphies

Node 58 :

Char. 50: 0 --> 1

Node 59 :

Char. 80: 0 --> 2

Node 60 :

Char. 12: 0 --> 3

Char. 14: 1 --> 0

Char. 56: 1 --> 0

Char. 85: 1 --> 0

Char. 86: 0 --> 1

Node 61 :

No synapomorphies

Node 62 :

Char. 105: 2 --> 1

Node 63 :

No synapomorphies

Node 64 :

No synapomorphies

Node 65 :

Char. 2: 1.025-1.100 --> 0.930-1.010

Char. 105: 2 --> 1

Node 66 :

No synapomorphies

Node 67 :

Char. 69: 0 --> 1

Takakia_ceratophylla

No autapomorphies

Pellia_epiphylla

Char. 12: 3 --> 4

Char. 54: 0 --> 1

Char. 55: 0 --> 1

Char. 89: 1 --> 0

Char. 98: 01 --> 2

Char. 104: 0 --> 1

Char. 116: 1 --> 2

Char. 119: 0 --> 1

Fossombronia_foveolata

Char. 10: 2 --> 0

Char. 12: 3 --> 2

Char. 13: 1 --> 0

Char. 18: 0 --> 1

Char. 31: 5 --> 0

Char. 58: 1 --> 0

Char. 75: 0 --> 1

Makinoa_crispata

Char. 34: 0 --> 3

Char. 93: 2 --> 3

Char. 108: 2 --> 0

Metzgeria_furcata

Char. 8: 0.093-0.147 --> 0.020-0.030

Char. 9: 0.400-0.500 --> 0.050-0.130

Char. 11: 1 --> 0

Char. 20: 1 --> 0

Char. 33: 0 --> 3

Char. 34: 0 --> 3

Char. 51: 0 --> 1

Char. 90: 1 --> 0

Node 68 :

Char. 98: 0 --> 2

Node 69 :

Char. 32: 4 --> 3

Node 70 :

Char. 2: 0.930-1.010 --> 0.870-0.890

Char. 10: 12 --> 0

Char. 12: 23 --> 1

Char. 30: 0 --> 1

Char. 90: 1 --> 2

Char. 92: 0 --> 2

Char. 94: 0 --> 2

Node 71 :

Char. 18: 0 --> 2

Node 72 :

Char. 98: 2 --> 1

Node 73 :

Char. 18: 0 --> 1

Char. 54: 0 --> 1

Char. 55: 0 --> 1

Char. 70: 0 --> 2

Char. 80: 0 --> 1

Char. 82: 0 --> 1

Node 74 :

No synapomorphies

Node 75 :

No synapomorphies

Node 76 :

Char. 17: 0 --> 3

Char. 34: 0 --> 2

Char. 76: 1 --> 0

Char. 77: 1 --> 0

Aneura_pinguis

Char. 1: 0.900-0.950 --> 0.790-0.880

Char. 25: 0 --> 2

Char. 70: 0 --> 1

Char. 114: 0 --> 2

Symphyogyna_undulata

No autapomorphies

Jungermannia_sp

Char. 0: 1.000 --> 0.400-0.800

Char. 1: 0.900-0.950 --> 0.970-1.000

Char. 17: 0 --> 2

Char. 23: 0 --> 1

Char. 32: 3 --> 6

Lejeunea_cavifolia

Char. 0: 1.000 --> 0.500-0.800

Char. 11: 1 --> 2

Char. 32: 3 --> 5

Char. 58: 1 --> 0

Char. 75: 0 --> 1

Char. 101: 0 --> 1

Char. 104: 0 --> 1

Frullania_asagrayana

Char. 8: 0.400 --> 0.500-1.500

Char. 13: 1 --> 0

Char. 51: 2 --> 1

Radula_voluta

Char. 1: 0.900-0.950 --> 0.314-0.361

Char. 2: 0.870-0.890 --> 0.591-0.670

Char. 48: 2 --> 0

Char. 78: 1 --> 0

Char. 80: 2 --> 1

Char. 86: 1 --> 0

Char. 91: 2 --> 0

Lepidozia_reptans

Char. 18: 0 --> 2

Char. 83: 0 --> 1

Char. 87: 2 --> 1

Char. 90: 1 --> 3

Char. 118: 0 --> 1

Node 77 :

No synapomorphies

Node 78 :

Char. 10: 2 --> 1

Char. 11: 2 --> 3

Char. 28: 2 --> 0

Char. 53: 4 --> 0

Char. 58: 1 --> 0

Char. 60: 1 --> 0

Char. 96: 0 --> 1

Char. 101: 0 --> 1

Char. 104: 0 --> 1

Node 79 :

Char. 109: 3 --> 5

Char. 112: 1 --> 0

Char. 121: 0 --> 1

Node 80 :

No synapomorphies

Node 81 :

Char. 11: 1 --> 2

Char. 28: 0 --> 2

Char. 60: 0 --> 1

Char. 64: 1 --> 2

Char. 105: 2 --> 0

Char. 107: 1 --> 0

Char. 117: 1 --> 0

Char. 126: 0 --> 1

Node 82 :

Char. 91: 0 --> 2

Char. 92: 0 --> 4

Char. 31: 0 --> 3

Char. 33: 0 --> 3

Char. 52: 1 --> 4

Char. 53: 2 --> 3

Char. 108: 2 --> 1

Lophocolea_heterophylla

Char. 18: 0 --> 1

Char. 33: 0 --> 3

Char. 108: 2 --> 1

Plagiochila_species1

Char. 20: 1 --> 0

Char. 32: 3 --> 4

Char. 51: 2 --> 1

Char. 126: 0 --> 1

Haplomitrium_hookeri

Char. 19: 0 --> 1

Char. 33: 0 --> 2

Char. 43: 1 --> 0

Char. 53: 1 --> 0

Char. 64: 1 --> 0

Char. 90: 1 --> 2

Char. 94: 0 --> 2

Char. 105: 2 --> 0

Char. 107: 1 --> 0

Char. 109: 3 --> 1

Treubia_sp

No autapomorphies

Blasia_pusilla

No autapomorphies

Austroriella_salta

No autapomorphies

Sphaerocarpos_texanus

No autapomorphies

Node 83 :

No synapomorphies

Node 84 :

Char. 0: 0.900-1.700 --> 3.500-4.000

Char. 3: 0.730-1.060 --> 2.890-2.900

Char. 4: 8.272-10.052 --> 26.262-

29.545

Char. 5: 3.920-4.470 --> 6.230

Char. 8: 0.275-0.300 --> 1.200-1.300

Char. 9: 1.250-1.400 --> 3.000

Char. 29: 02 --> 3

Char. 37: 04 --> 5

Char. 44: 0 --> 2

Char. 47: 1 --> 2

Char. 48: 2 --> 1

Char. 62: 1 --> 2

Char. 92: 0 --> 4

Node 85 :

No synapomorphies

Node 86 :

Char. 9: 0.400-0.500 --> 1.250

Char. 36: 0 --> 4

Char. 38: 0 --> 2

Char. 39: 0 --> 1

Char. 42: 0 --> 4

Char. 43: 1 --> 2

Char. 44: 3 --> 0

Char. 45: 5 --> 0

Char. 47: 0 --> 1

Node 87 :

Char. 3: 0.730-1.030 --> 1.090-1.120

Char. 8: 0.275-0.300 --> 0.400

Char. 9: 0.300-0.700 --> 0.800

Char. 28: 2 --> 4

Char. 38: 2 --> 3

Char. 47: 1 --> 2

Char. 48: 2 --> 0

Geothallus_tuberosus

Char. 9: 0.400-0.500 --> 0.250-0.350

Char. 24: 0 --> 1

Char. 51: 0 --> 1

Char. 111: 0 --> 1

Bucegia_romanica

No autapomorphies

Marchantia_polymorpha

Char. 23: 0 --> 2

Char. 34: 2 --> 3

Char. 45: 0 --> 1

Char. 60: 1 --> 2

Char. 71: 1 --> 2

Char. 98: 0 --> 2

Preissia_quadrata

No autapomorphies

Cryptomitrium_tenerum

Char. 24: 0 --> 1

Char. 64: 2 --> 0

Char. 96: 0 --> 1

Char. 123: 0 --> 1

Char. 125: 1 --> 0

Reboulia_hemisphaerica

Char. 8: 0.400 --> 0.000

Char. 63: 0 --> 2

Plagiochasma_rupestre

Char. 4: 9.073-13.091 --> 13.223-

16.216

Char. 8: 0.400 --> 0.500-1.000

Char. 13: 1 --> 0

Char. 18: 0 --> 1

Char. 47: 2 --> 1

Char. 59: 2 --> 1

Mannia_californica

Char. 54: 0 --> 1

Char. 59: 0 --> 2

Char. 62: 1 --> 2

Char. 109: 0 --> 4

Node 88 :

No synapomorphies

Node 89 :

Char. 5: 3.420-4.470 --> 2.950-3.000

Node 90 :

Char. 2: 1.025-1.110 --> 1.160-1.420

Node 91 :

Char. 9: 1.250-1.400 --> 0.307-1.200

Node 92 :

Char. 6: 1.750-2.000 --> 1.000-1.570

Node 93 :

Char. 4: 8.272-8.371 --> 9.073-13.091

Char. 42: 4 --> 3

Char. 45: 02 --> 1

Char. 66: 0 --> 1

Node 94 :

Char. 27: 0 --> 1

Node 95 :

No synapomorphies

Node 96 :

Char. 28: 2 --> 5

Char. 38: 2 --> 1

Char. 41: 2 --> 1

Char. 69: 2 --> 0

Char. 90: 3 --> 0

Char. 91: 0 --> 2

Node 97 :

Char. 64: 2 --> 0

Char. 96: 0 --> 1

Asterella_tenella

Char. 22: 0 --> 1

Char. 63: 0 --> 2

Char. 123: 0 --> 1

Char. 124: 1 --> 3

Char. 125: 1 --> 2

Ricciocarpos_natans

Char. 1: 1.700-1.850 --> 3.220-4.710

Char. 39: 1 --> 3

Char. 43: 2 --> 0

Char. 44: 0 --> 3

Char. 45: 0 --> 5

Char. 47: 1 --> 0

Char. 51: 1 --> 2

Char. 120: 3 --> 2

Riccia_fluitans

Char. 28: 5 --> 3

Riccia_species1

No autapomorphies

Athalamia_pinguis

Char. 120: 4 --> 2

Char. 121: 0 --> 1

Char. 122: 0 --> 2

Char. 125: 1 --> 0

Clevea_nana_=Athalamia_hyalina

Char. 4: 9.415-11.010 --> 11.896-

13.702

Peltolepis_quadrata

Char. 6: 1.000-1.570 --> 2.310-3.560

Char. 44: 0 --> 2

Char. 46: 1 --> 2

Char. 2: 1.190-1.510 --> 1.840-2.280

Char. 3: 0.730-1.030 --> 0.350-0.450

Char. 52: 3 --> 5

Char. 70: 2 --> 0

Char. 96: 0 --> 2

Char. 109: 0 --> 5

Char. 112: 1 --> 0

Node 98 :

Char. 1: 1.000-1.010 --> 1.700-1.850

Char. 33: 2 --> 0

Char. 51: 2 --> 1

Node 99 :

Char. 25: 0 --> 1

Char. 52: 5 --> 0

Char. 53: 1 --> 0

Node 100 :

No synapomorphies

Node 101 :

Char. 1: 0.960-1.010 --> 1.020-1.170

Char. 5: 3.420-4.470 --> 4.570-4.600

Char. 24: 0 --> 1

Char. 28: 2 --> 4

Char. 34: 2 --> 1

Char. 45: 0 --> 3

Char. 62: 1 --> 2

Node 102 :

Char. 3: 0.730-0.900 --> 0.470-0.650

Node 103 :

Char. 126: 1 --> 0

Node 104 :

Char. 0: 0.640-0.800 --> 1.060

Char. 1: 0.960 --> 1.210-1.250

Char. 8: 0.275-0.300 --> 0.640-0.650

Char. 9: 0.317 --> 0.640-0.650

Sauteria_alpina

Char. 37: 1 --> 2

Char. 46: 1 --> 2

Char. 47: 1 --> 2

Char. 48: 2 --> 0

Char. 59: 1 --> 2

Conocephalum_conicum

Char. 11: 2 --> 1

Char. 21: 0 --> 1

Char. 22: 0 --> 1

Char. 59: 0 --> 2

Char. 62: 01 --> 2

Char. 63: 0 --> 2

Char. 66: 0 --> 2

Char. 119: 0 --> 1

Char. 120: 3 --> 0

Corsinia_coriandrina

Char. 29: 2 --> 1

Char. 34: 2 --> 3

Char. 40: 1 --> 2

Cronisia_fimbriata

No autapomorphies

Cyathodium_cavernarum

Char. 2: 1.190-1.600 --> 1.720-1.800

Char. 9: 0.200-0.317 --> 0.000

Char. 24: 0 --> 2

Char. 45: 0 --> 3

Char. 60: 1 --> 0

Char. 61: 1 --> 0

Char. 62: 1 --> 0

Char. 91: 0 --> 1

Char. 109: 0 --> 4

Dumortiera_hirsuta

Char. 4: 8.272-10.052 --> 3.322-5.863

Char. 5: 3.420-4.470 --> 1.170-2.270

Char. 24: 0 --> 2

Char. 33: 2 --> 0

Char. 45: 0 --> 4

Char. 53: 4 --> 1

Char. 60: 1 --> 0

Char. 61: 1 --> 0

Char. 62: 1 --> 0

Char. 66: 0 --> 1

Char. 90: 3 --> 1

Char. 109: 0 --> 5

Char. 112: 1 --> 0

Char. 113: 1 --> 0

Char. 125: 1 --> 0

Node 105 :

Char. 6: 0.500-0.875 --> 0.300-0.325

Char. 25: 0 --> 1

Node 106 :

Char. 27: 0 --> 1

Char. 52: 3 --> 5

Node 107 :

No synapomorphies

Node 108 :

Char. 8: 0.275-0.300 --> 0.000

Node 109 :

Char. 2: 1.025-1.110 --> 0.790-0.970

Node 110 :

No synapomorphies

Char. 28: 2 --> 1

Char. 35: 3 --> 0

Char. 38: 2 --> 1

Char. 41: 23 --> 1

Char. 42: 4 --> 1

Char. 98: 0 --> 1

Lunularia_cruciata

Char. 2: 1.025-1.110 --> 1.180-1.550

Char. 8: 0.275-0.300 --> 0.000

Char. 13: 1 --> 0

Char. 23: 0 --> 3

Char. 27: 0 --> 1

Char. 98: 0 --> 1

Monoclea_gottschei

Char. 28: 2 --> 0

Char. 29: 1 --> 0

Char. 34: 2 --> 0

Char. 36: 4 --> 0

Char. 38: 2 --> 0

Char. 43: 2 --> 1

Char. 45: 0 --> 5

Char. 46: 1 --> 0

Char. 51: 1 --> 0

Char. 54: 0 --> 1

Char. 55: 0 --> 1

Char. 56: 0 --> 1

Char. 94: 0 --> 1

Char. 95: 1 --> 2

Char. 108: 2 --> 1

Char. 109: 0 --> 1

Char. 123: 0 --> 1

Targionia_hypophylla

Char. 0: 0.900-1.700 --> 0.100-0.210

Char. 2: 0.790-0.970 --> 0.500-0.640

Char. 4: 8.272-10.052 --> 11.404-

16.742

Char. 9: 0.307-1.200 --> 0.000

Char. 13: 1 --> 0

Char. 18: 0 --> 1

Char. 21: 0 --> 1

Char. 27: 0 --> 1

Char. 53: 4 --> 2

Char. 60: 1 --> 0

Char. 61: 1 --> 0

Char. 62: 1 --> 0

Char. 91: 0 --> 1

Wiesnerella_denudata

Char. 0: 0.900-1.700 --> 2.000-5.000

Char. 1: 0.750-0.940 --> 0.530-0.600

Char. 4: 8.272-10.052 --> 6.637-7.806

Char. 20: 3 --> 0

Char. 33: 2 --> 0

Char. 44: 0 --> 1

Char. 47: 1 --> 2

Char. 48: 2 --> 0

Oxymitra_incrassata

Char. 0: 0.400-0.800 --> 0.130-0.150

Char. 4: 8.272-8.371 --> 12.507-

14.739

Char. 8: 0.275-0.300 --> 0.000

Char. 13: 1 --> 0

Char. 66: 0 --> 1

Char. 94: 0 --> 1

Char. 121: 0 --> 1

Exormotheca_pustulosa

Char. 1: 0.960 --> 0.768-0.852

Char. 8: 0.275-0.300 --> 0.000

Char. 21: 0 --> 1

Char. 24: 0 --> 1

Char. 47: 1 --> 2

Char. 48: 2 --> 1

Char. 59: 0 --> 2

Char. 62: 1 --> 2

Char. 100: 3 --> 2

Monocarpus_sphaerocarpus

Char. 0: 0.700-0.800 --> 0.150-0.300

Char. 27: 0 --> 1

Char. 69: 2 --> 0

Monosolenium_tenerum

Char. 2: 1.190-1.600 --> 0.860-1.090

Char. 28: 2 --> 0

Char. 29: 2 --> 0

Char. 48: 2 --> 0

Char. 59: 0 --> 3

Char. 122: 0 --> 2

Aitchisoniella_himalayensis

Char. 4: 7.386-7.905 --> 3.302-5.252

Char. 5: 2.950-3.000 --> 2.020-2.570

Char. 122: 0 --> 2

Stephensoniella_brevipedunculata

Char. 1: 0.960 --> 0.670-0.710

Char. 9: 0.300-0.317 --> 0.120-0.220

Char. 24: 0 --> 1

Char. 29: 2 --> 1

Char. 48: 2 --> 0

Char. 59: 0 --> 1

Neohodgsonia_mirabilis

Char. 29: 0 --> 3

Char. 38: 0 --> 1

Char. 41: 0 --> 1

Char. 42: 0 --> 1

Char. 60: 1 --> 2

Char. 62: 01 --> 2

Char. 63: 0 --> 3

Char. 71: 0 --> 2

Char. 92: 0 --> 4

Char. 102: 1 --> 0

Char. 103: 1 --> 0

Char. 122: 0 --> 1

Apotreubia_nana

No autapomorphies

Riella_helicophylla

No autapomorphies

Pallavicinia_lyellii

No autapomorphies

Pleurozia_purpurea

No autapomorphies

Cavicularia_densa

No autapomorphies

Figure S2. Proportion of synapomorphies derived from the sporophyte at each

diagnosed branch. C = classes, SC = subclasses, O = orders, X = inter-family, F = family.

Grey branches indicate synapomorphy absence.

Table S3. Support values for selected groups. Comparison between support values

when morphology is excluded. Symmetric Resampling values were compared under

IWM. Run settings: 250 replicates, each consisting of 10 RAS + TBR.

Groups Morphological data excluded Morphological data included

B SR: 51 SR: 94

Sphaerocarpales SR: 60 SR: 85

Ricciaceae SR: <50 SR: 65

Corsiniaceae SR: <50 SR: 91

Figure S3. Strict consensus of trees derived from the ten continuous characters under

K3.

Literature

Bischler, H.,. 1998. Systematics and evolution of the genera of the Marchantiales. Bryophyt. Bibl. 51, 1–201.

Crandall-Stotler, B., Stotler, R.,. 2000. Morphology and classification of the Marchantiophyta, in: Shaw, A., Goffinet, B. (Eds.), Bryophyte Biology. Cambridge University Press, Cambridge, pp. 21–70.

Duckett, J., Ligrone, R., Renzaglia, K.,. 2014. Pegged and smooth rhizoids in complex thalloid liverworts (Marchantiopsida): structure, function and evolution. Bot. J. Linn. Soc.

Hassel de Menéndez, G.,. 1963. Estudio de las Anthocerotales y Marchantiales de la Argentina. Opera Lilloana. 7, 1–297.

Documents

Cladistics · 2018. 1. 10. · 2 Flores et al. / Cladistics 0 (2017) 1–25. diagnoses and testing previous ideas about character evolution. In addition, the contribution to and conﬂict