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Phylogenetics of tribe Sabiceeae (Ixoroideae, Rubiaceae)
revisited, with a new subgeneric classification for
Sabicea
LISE ZEMAGHO
1,2
, SIGRID LIEDE-SCHUMANN
1
*, BONAVENTURE SONK
E
2
, STEVEN
JANSSENS
3
, OLIVIER LACHENAUD
3
, BRECHT VERSTRAETE
3
and STEVEN DESSEIN
3
1
Department of Plant Systematics, University of Bayreuth, Bayreuth, Germany
2
Plant Systematic and Ecology Laboratory, Department of Biology, Higher Teachers’ Training College,
University of Yaound
e I, Yaound
e, Cameroon
3
Botanic Garden Meise, Meise, Belgium
Received 24 April 2015; revised 4 March 2016; accepted for publication 18 July 2016
Tribe Sabiceeae (Ixoroideae, Rubiaceae) has undergone recent taxonomical changes with the incorporation of the
related genera Ecpoma,Pseudosabicea and Stipularia into the type genus Sabicea. We use phylogenetic analysis
and morphological data to verify the relationships among members of the tribe, including the most
comprehensive taxon sampling of the tribe to date with 74 of 145 species. Sequence data from the nuclear
internal transcribed spacer (ITS) and three plastid markers (petD,rps16,trnT–F) were used to infer relationships
among the members of the tribe. Individual analyses using maximum likelihood, parsimony and Bayesian
approaches reveal several supported clades: the former genus Stipularia is resolved as a monophyletic unit, but
Ecpoma is monophyletic only if Sabicea urbaniana and Sabicea xanthotricha are included (corresponding to
Sabicea subgenus Stipulariopsis sensu Wernham). Pseudosabicea is biphyletic, with one clade corresponding to
section Anisophyllae of Hall
e (1964) and the other one to the other sections (Floribundae and Sphaericae) of the
genus. Eleven morphological characteristics were recorded for all species studied and seven have been mapped
onto the phylogenetic tree to study their evolution in the group and assess their value for the classification of
Sabicea s.l. Finally, our study shows that a combination of diagnostic characteristics should be used to
differentiate each group and we propose to recognise four subgenera in Sabicea. ©2016 The Linnean Society of
London, Botanical Journal of the Linnean Society, 2016, 182, 551–580
ADDITIONAL KEYWORDS: cpDNA – Ecpoma –Hekistocarpa – morphology – nrITS –
Pseudosabicea –Stipularia –Tamridaea –Virectaria.
INTRODUCTION
Tribe Sabiceeae (Ixoroideae, Rubiaceae) was pro-
posed for the first time by Bremekamp (1966) in sub-
family Cinchonoideae, to accomodate the genus
Sabicea Aubl. Bremekamp (1966) excluded Sabicea
from tribe Mussaendeae because of its simple (not
bifid) stipules, axillary (not terminal) inflorescences
and the narrow (not large) testa cells. Several Rubi-
aceae taxonomists did not accept Sabiceeae as a sep-
arate tribe (Verdcourt, 1958; Hall
e, 1961, 1963, 1966;
Kirkbride, 1979, 1982; Robbrecht, 1988, 1993) and
included Sabicea, with supposedly related genera, in
tribe Isertieae or Mussaendeae. Andersson (1996),
however, conducted a cladistic analysis using mor-
phological characteristics of tribe Isertieae sensu
Robbrecht (1988) and resurrected tribe Sabiceeae in
which he included Acranthera Arn. ex Meisn. (Meis-
ner, 1838), Amphidasya Standl. (Standley, 1936),
Ecpoma K.Schum. (Schumann, 1896), Pentaloncha
Hook.f. (Hooker, 1873), Pittierothamnus Steyerm.
(Steyermark, 1962), Pseudosabicea N.Hall
e (Hall
e,
1963), Sabicea Aubl. (Aublet, 1775), Schizostigma
Arn. ex. Meisn. (Meisner, 1838) and Temnopteryx
Hook.f. (Hooker, 1873); a tenth genus included in his
cladistic analysis, Stipularia P.Beauv. (Palisot-Beau-
vois, 1810), was erroneously omitted from the list.
He considered the tribe more closely related to
*Corresponding author. E-mail: sigrid.liede@uni-bayreuth.de
551©2016 The Linnean Society of London, Botanical Journal of the Linnean Society, 2016, 182, 551–580
Botanical Journal of the Linnean Society, 2016, 182, 551–580. With 9 figures
members of subfamily Ixoroideae than to tribe Iser-
tieae (subfamily Cinchonoideae). However, the broad
taxonomic delimitation of Sabiceeae as proposed by
Andersson (1996) is not supported by molecular data
(Bremer & Thulin, 1998; Alejandro, Razafimandim-
binson & Liede-Schumann, 2005; Smedmark et al.,
2008; Khan et al., 2008a; Rydin et al., 2009). On the
basis of rps16 intron sequences, Dessein et al.
(2001a) proposed that tribe Virectarieae, including
Hekistocarpa,Tamridaea and Virectaria, are sister
to Sabiceeae. Khan et al. (2008a) proposed a new tri-
bal delimitation for Sabiceeae, based on the phyloge-
netic signal of nrITS and plastid trnT-F sequences
and altered the generic delimitations in the tribe.
Virectarieae were merged with Sabiceeae and
Ecpoma,Pseudosabicea,Schizostigma and Stipularia
were included in Sabicea s.l. As a result, Sabiceeae,
as broadly delimited by Khan et al. (2008a), included
only four genera: Hekistocarpa,Sabicea s.l.,Tamri-
daea and Virectaria. This tribal concept was adopted
in later classifications (Bremer & Eriksson, 2009;
Kainulainen, Razafimandimbison & Bremer, 2013).
The genus Sabicea was originally described by
Aublet (1775) with two species: S. aspera Aubl. and
S. cinerea Aubl., the latter designated later as the
type species (Standley, 1921: 148). Wernham (1914)
revised the genus and recognised two subgenera
based mainly on the growth form: subgenus
Eusabicea Wernham [=subgenus Sabicea] with 96
species, having a twining, climbing or creeping habit,
and subgenus Stipulariopsis Wernham with nine
species, having an erect habit with large leaves and
stipules. He further subdivided subgenus Sabicea
into four sections based on the type of inflorescence:
sections Laxae Wernham, Sessiles Wernham [= sec-
tion Sabicea], Capitatae Wernham and Floribundae
Wernham. Wernham (1914) accepted the closely
related Stipularia as a distinct genus, based on its
peculiar involucre, whereas Hepper (1958) syn-
onymized Stipularia under Sabicea.
Hall
e (1963) reviewed all characteristics used by
Wernham (1914) in his classification of Sabicea [(1)
the indumentum, especially of the leaves; (2) the nat-
ure of the inflorescence –whether sessile or peduncu-
late, involucrate or ex-involucrate; (3) the length of
the calyx lobes and their shape; and (4) the shape
and relative size of the corolla tube and lobes] and
considered them as irrelevant for the generic classifi-
cation, except for the size of the corolla tube and
lobes. Instead, he emphasised the number of ovary
locules, a characteristic largely overlooked by Wern-
ham (1914). Hall
e (1963) restricted Sabicea to twin-
ing or creeping species with pentalocular ovaries. He
recognised Stipularia (with tri- to pentalocular ovar-
ies) as a distinct genus, based on the erect habit and
involucral characteristics. He transferred the species
with bilocular ovaries to Ecpoma (Schumann, 1896)
which grouped the erect species, and the new genus
Pseudosabicea, including sarmentose or creeping spe-
cies. Ecpoma had been overlooked since its descrip-
tion. In addition to the type species E. apocynaceum
K.Schum., Hall
e (1963) included in it five of the nine
original species of Sabicea subgenus Stipulariopsis
(S. bicarpellata K.Schum., S. cauliflora Hiern, S.
geantha Hiern, S. gigantostipula K.Schum., S. hierni-
ana Wernham). Sabicea bicarpellata proved to be
misplaced in the genus and was later transferred to
Bertiera Aubl. (Aublet, 1775) as B. bicarpellata
(K.Schum.) N.Hall
e (Hall
e, 1970). In Pseudosabicea,
Hall
e distinguished three sections: section Anisophyl-
lae N.Hall
e(S. batesii Wernham, S. mildbraedii
Wernham, S. medusula K.Schum. ex Wernham);
section Floribundae N.Hall
e[S. floribunda K.Schum.,
S. segregata Hiern, S. proselyta (N.Hall
e) Razafim.
et al.]; and section Sphaericae N.Hall
e [= section
Pseudosabicea](S. arborea K.Schum., S. nobilis
R.D.Good, S. pedicellata Wernham). The transfer of
S. pedicellata to Pseudosabicea was in fact erroneous,
as this species has a pentalocular ovary (L. Zemagho,
pers. observ.). Several new taxa of Pseudosabicea
were later described by Hall
e (1966, 1971).
The monotypic Schizostigma Arn. ex. Meisn. is
endemic to the forests of south-western Sri Lanka
and has often been associated with Sabicea and its
allies. Arnott (1839) presented a description in which
he stated, ‘may be placed near Sabicea’.
Hiern (1877) suggested a relationship between
the African genus Pentaloncha and Schizostigma,
whereas Baillon (1880, cited in Puff, Igersheim &
Buchner, 1998) included both Pentaloncha and Tem-
nopteryx in Schizostigma, stating that they repre-
sent ‘sections africaines’ of the genus with
multiflorous inflorescences. Puff et al. (1998) com-
pared Schizostigma and Sabicea and suggested that
there are no characteristic states by which the two
genera can be reliably separated and that the only
reason that could be brought forward in favour of
keeping them as separate entities would be
geographical distribution. Finally, Khan et al.
(2008a) clearly showed that Schizostigma belongs to
Sabicea s.l.
Although recent phylogenetic investigations have
improved our understanding of the tribe and of the
systematic position of its type genus Sabicea, the
relationships between Sabicea s.s. and its satellite
genera are still unclear. Three questions are perti-
nent in this context: (1) Are Sabiceeae monophyletic
as presently circumscribed? (2) Do the formal and
informal characteristics support the phylogenetic
classification? (3) Are Ecpoma,Pseudosabicea and
Stipularia nested in Sabicea or should they be con-
sidered as segregate genera?
©2016 The Linnean Society of London, Botanical Journal of the Linnean Society, 2016, 182, 551–580
552 L. ZEMAGHO ET AL.
This study presents the first analysis of morpho-
logical characteristics in a phylogenetic context in
Sabiceeae. In Sabicea s.l., we studied the suitability
of morphological characteristics for infrageneric clas-
sification purposes. Finding characteristics that sup-
port the phylogenetic classification will help to
position species of which sequenceable material is
not available. In addition, characteristic evolution
might give a clue to the understanding of an evolu-
tionarily successful lineage. The aims of this study
are (1) to re-assess the phylogenetic relationships
within the broadly circumscribed Sabiceeae adding
additional species and further sequence data from
the plastid regions petD and rps16 and (2) to study
the systematic value of morphological characteristics,
which is defined here as their potential to identify
monophyletic groups and to find morphological sup-
port for the major clades in Sabicea s.l.
MATERIAL AND METHODS
TAXON SAMPLING
We aimed at as broad a sampling as possible, includ-
ing all genera of Sabiceeae s.l., and as many species
as possible for each genus. For nrITS and plastid
DNA markers (trnT-F,petD and rps16) 74 species of
Sabiceeae were included in the study: 51 species of
Sabicea s.s.,12Pseudosabicea spp., two Stipularia
spp., two Ecpoma spp., four Virectaria spp. and the
monotypic genera Hekistocarpa,Schizostigma
(=Sabicea ceylanica Puff) and Tamridaea. Whenever
possible, the type species of the genera were
included, but our efforts to isolate DNA from herbar-
ium specimens of Ecpoma apocynaceum K.Schum.
(the type species of Ecpoma) were unsuccessful. Mus-
saenda erythrophylla Schumach. & Thonn., M. arcu-
ata Lam. ex Poir. and Pseudomussaenda flava Verdc.
were chosen as outgroups because Mussaendeae
have been identified as sister to Sabiceeae s.l.
(Kainulainen et al., 2013) (Appendix 3). Most DNA
samples were obtained from silica gel collections;
rarely leaf samples were taken from herbarium spec-
imens (BR, WAG) or from fresh leaf material col-
lected from the living collections of the Meise
Botanic Garden. We incorporated 78 previously pub-
lished sequences in our dataset (38 nrITS and 40
trnT-F; Khan et al., 2008a).
DNA EXTRACTION,AMPLIFICATION AND SEQUENCING
DNA was extracted using the 2 9cetyl trimethyl
ammonium bromide method (Doyle & Doyle, 1987)
and purified using caesium chloride/ethidium bro-
mide gradient centrifugation. Four molecular mark-
ers, ITS of nrDNA, the plastid introns petD and
rps16, and the trnT-F region of plastid DNA (includ-
ing the trnT-L and trnL-F spacers and the trnL
intron) were sequenced. Amplification and sequenc-
ing was carried out using the primers listed in
Table 1. Polymerase chain reactions (PCR) for nrITS,
petD,rps16 and trnT-F were carried out as 25-lL
reactions including 10.875 lLH
2
O, 2.5 lL reaction
buffer (109,59GC, 59AB), 2.5 lL 25 mM MgCL
2
,
2.5 lL 2 mM BSA, 0.25 lL each forward and reverse
primers (20 lM), 0.125 lLTaq DNA polymerase
(Kapa-TAQ, Dream-TAQ, PE-TAQ) and 2 lL DNA
template. PCR reactions of nrITS were run on a
Thermocycler Controller 2720 with initial denatura-
tion at 95 °C for 3 min, followed by ten cycles of 30 s
at 60 °C, 72 °C for 1 min 30 s, 95 °C for 30 s, fol-
lowed by 30 cycles of 30 s at 54 °C, 72 °C for 1 min,
72 °C for 7 min, finishing with 10 °C. All PCR and
sequencing reactions of trnT-F,petD and rps16 were
performed using a Programmable Biometra Personal
Cycler (Biometra, G€
ottingen, Germany) and were
carried out as follows: 96 °C for 2 min; 34 cycles at
94 °C for 1 min, 50 °C for 1 min, 72 °C for 1 min
30 s, and a final extension at 72 °C for 10 min. PCR
products were sent to Macrogen (Seoul, South Korea)
Table 1. Amplification primers used in this study
Region Primers Primers sequences (50
–30) References
nrITS Forward (P17F)
Reverse (26S–82R)
CTA CCG ATT GAA TGG TCC GGT GAA
TCC CGG TTC GCT CGC CGT TAC TA
Alejandro et al. (2005)
petD Forward (petB1365F)
Reverse (petD738 R)
TTG ACY CGT TTT TAT AGT TTA C
AAT TTA GCY CTT AAT ACA GG
L€
ohne & Borsch (2004)
rps16 Forward (rps16)
Reverse (rpsR2)
GTG GTA GAA AGC AAC GTG CGA CTT
TCG GGA TCG AAC ATC AAT TGC AAC
Oxelman, Lid
en & Berglund (1997)
trnT-FForward (c)
Reverse (f)
CGA AAT CGG TAG ACG CTA CG
AT TTG AAC TGG TGA CAC GAG
Razafimandimbison & Bremer (2002)
Forward (trnL-A1)
Reverse (trnL-I)
ACA AAT GCG ATG CTC TAA CC
CC AAC TCC ATT TGT TAG AAC
©2016 The Linnean Society of London, Botanical Journal of the Linnean Society, 2016, 182, 551–580
PHYLOGENETICS OF SABICEEAE (RUBIACEAE) 553
for sequencing. Two hundred and ten sequences are
newly generated (73 petD,69rps16, 37 ITS and 31
trnT-F) (see Appendix 1).
SEQUENCE ALIGNMENT
Forward and reverse sequences generated for nrITS,
petD,rps16 and trnT-F regions were assembled
using Geneious 6.1.6 (Biomatters Ltd, Auckland,
New Zealand). Each individual base position was
examined for agreement between the two strands.
Consensus sequences were aligned with the plugin
MAFFT implemented in Geneious 6.1.6 and the
resulting alignment was edited manually.
PHYLOGENETIC ANALYSIS
Before analysing the concatenated data matrix, each
marker was analysed separately to recognise puta-
tive phylogenetic differences visually between the
different plastid datasets and then, between the plas-
tid and ITS datasets. A partition homogeneity test
(implemented in PAUP*4.0b10a; Swofford, 2002)
was carried out to detect statistically whether the
data matrices were incongruent. The best performing
substitution model for maximum likelihood (ML) and
Bayesian inference (BI) was determined for each par-
tition using the Akaike information criterion (AIC)
as implemented in jModelTest 2.1.6 (Guindon & Gas-
cuel, 2003; Darriba et al., 2012). For all partitions,
the GTR +G model was suggested. Bayesian analy-
ses of the single markers or the concatenated dataset
were conducted with MrBayes 3.1 (Huelsenbeck &
Ronquist, 2001; Ronquist & Huelsenbeck, 2003). Two
runs of four chains (one cold, three heated), initiated
from a random starting tree, were monitored for two
million generations at which stationarity was
reached. Every 100 generations, a tree was sampled
from the chain for a total of 20 000 trees. Conver-
gence of the runs was checked with Tracer 1.5
(Drummond & Rambaut, 2007) resulting in a
removal of 5000 trees as burn-in. Geneious 6.1.6 was
used to compute the 50% majority rule consensus
tree. Maximum likelihood analyses were carried out
using the RAxML search algorithm (Stamatakis,
Ludwig & Meier, 2005) under the GTRGAMMA
approximation of rate heterogeneity for each gene
(Stamatakis, Ludwig & Meier, 2006) as implemented
in RAxML 7.2.8. Five hundred bootstrap trees were
inferred using the RAxML rapid bootstrap algorithm
to provide support values for the best-scoring ML
tree. Maximum parsimony (MP) analyses were con-
ducted using PAUP*4.0b10a (Swofford, 2002).
Heuristic searches were conducted applying tree
bisection-reconnection (TBR) branch swapping on
10 000 random addition sequence replicates, with
five trees held at each step. For each of the 1000
bootstrap replicates, a heuristic search was con-
ducted with identical settings as in the original
heuristic analysis. A clade was considered well sup-
ported if the support values for ML and MP was
>80% and the posterior probability was >0.95. A
clade was considered moderately supported if sup-
port values for both ML and MP were >70% and the
posterior probability between 0.90 and 0.95. A clade
was considered weakly supported if support values
for both ML and MP were <70% and the posterior
probability was <0.90.
MORPHOLOGICAL ANALYSIS
To identify synapomorphic characteristics supporting
monophyletic lineages, all specimens from BR,
BRLU, MO, NY, UPS and WAG (herbarium acro-
nyms according to Thiers, continuously updated)
were examined, totalling 3012 specimens. In addi-
tion, previous taxonomic works (Wernham, 1914;
Khan, 2007) and local taxonomic treatments of gen-
era of Sabiceeae were used (Hepper, 1963; Hall
e,
1966; Bridson & Verdcourt, 1988). Eleven morpholog-
ical characteristics, most of which had been used by
Wernham (1914) or Hall
e (1963) in their classifica-
tions of Sabicea s.l., are chosen and evaluated here
(Appendix 1), then compiled into a matrix (Appen-
dix 2). Variability of some characteristics used in the
analysis is illustrated in Figure 1. Evolution of
morphological characters was studied by plotting
each character on the phylogenetic tree based on the
Bayesian topology of the combined ITS and plastid
DNA data using a maximum parsimony approach.
We used the ‘Trace Character History’ option to
reconstruct evolution of each character with the soft-
ware package Mesquite 2.75 (Maddison & Maddison,
2001).
RESULTS
MOLECULAR STUDY
Sequence and alignment characteristics are given in
Table 2. Single locus phylogenetic tree revealed no
topological discrepancies and partition homogeneity
tests did not indicate conflict between the plastid
data sets or between plastid DNA and nrITS
(P>0.02). As a result, we combined all data sets for
further phylogenetic analyses. The concatenated data
set consisted of 81 taxa and 491 informative charac-
teristics. No topological conflict was found between
the ML tree, the MP tree and the Bayesian consen-
sus tree; however, support values were generally
lower for the MP-based phylogenetic reconstruction
(Fig. 2).
©2016 The Linnean Society of London, Botanical Journal of the Linnean Society, 2016, 182, 551–580
554 L. ZEMAGHO ET AL.
AB
B´ C
DEF
I
JHG
Figure 1. Variation of some morphological characters included in the phylogenetic analysis of Sabiceeae. Anisophylly
at nodes: unequal leaves: Sabicea aff. aurifodinae sp.nov. (A from OL1272)/equal leaves: Sabicea proselyta (B from
OL1531) –entire stipule: Sabicea proselyta (B0from OL1531) –bract shape: Sabicea bigerrica (C*from greenhouse
Meise), Sabicea africana (G*from BSO6239), Sabicea urniformis (H*from OL1532) –inflorescence type: branched: Sabi-
cea proselyta (B from OL1531)/unbranched and pedunculate: Sabicea bigerrica (C from greenhouse Meise)/unbranched
and sessile: Sabicea aff. aurifodinae sp.nov. (D from BSO5962) –first order bracts in fruiting stage: not splitting Sabicea
africana (G from BSO6239)/splitting Sabicea urniformis (H from OL1532) –fruit colour: Sabicea sp. (E from OL1498),
Sabicea sp. (F from BSO5004) –indumentum corolla throat: short trichomes: Sabicea dinklagei (I from BSO6109)/long
trichomes: Sabicea sp. –OL1447 (J from OL1447). A, B, E, H, J by O. Lachenaud; C, D, F, G, I by B. Sonk
e.
©2016 The Linnean Society of London, Botanical Journal of the Linnean Society, 2016, 182, 551–580
PHYLOGENETICS OF SABICEEAE (RUBIACEAE) 555
Relationships in Sabiceeae are highly congruent
with those described in previous phylogenetic studies
(Alejandro et al., 2005; Khan et al., 2008a; Kainu-
lainen et al., 2013). The present phylogenetic analy-
ses confirmed that Sabiceeae as currently
circumscribed are monophyletic. Several moderately
to well supported clades can be found in the tribe.
The Hekistocarpa–Tamridaea–Virectaria clade is
resolved with moderate support (ML: 69, BI: 0.82,
MP: 72) as sister to the clade containing all members
of Sabicea s.l. (Fig. 2). Hekistocarpa is sister to a
well supported (ML: 99, BI: 1.0, MP: 100) Tamri-
daea–Virectaria clade. In the strongly supported
(ML: 100, BI: 1.0, MP: 100) Sabicea s.l. clade, the
highly supported Sabicea africana–S. elliptica clade
(ML: 100, BI: 1.0, MP: 100) branches off first. Next,
a strongly supported clade (ML: 82, BI: 0.97, MP:
97), corresponding to Pseudosabicea section Aniso-
phyllae [comprising Sabicea aurifodinae (N.Hall
e)
Razafim., B.Bremer, Liede & Saleh A.Khan, S. bate-
sii,S. becquetii (N.Hall
e) Razafim., B.Bremer, Liede
& Saleh A.Khan, S. medusula,S. mildbraedii includ-
ing var. letestui (N. Hall
e), S. sthenula (N.Hall
e)
Razafim., B.Bremer, Liede & Saleh A.Khan, S. sp-
SD1865 and S. sp-SD2149], is retrieved as sister to
the rest of the group. Species previously referred to
Ecpoma form a moderately supported clade with
Sabicea xanthotricha Wernham and S. urbaniana
Wernham (ML: 56, BI: 0.99, MP: 66). The Ecpoma
clade is sister to a large, moderately supported clade
(ML: 88, BI: 0.99, MP: 58) with little internal struc-
ture, comprising all sequenced species of Sabicea s.s.
(sensu Hall
e, 1963, 1966), species of Pseudosabicea
sections Sphaericae and Floribundae and the mono-
typic Schizostigma. Resolution in this clade is poor,
but a few supported clades can be identified. Schizos-
tigma forms a moderately supported clade (ML: 70,
BI: 0.90, MP: 75) with the Central African Sabicea
speciosa K.Schum. Sabicea pilosa Hiern. and S. urce-
olata Hepper form a strongly supported clade (ML:
99, BI: 1.0, MP: 98). The Madagascan species S.
acuminata Baker and S. diversifolia Pers. form a
moderately supported clade (ML: 55, BI: 0.96, MP:
59). The Neotropical Sabicea spp. also form a
moderately supported clade (ML: 73, BI: 0.94, MP:
53) sister to a moderately supported (ML: 82, BI:
0:99, MP: 86) Continental African clade comprising
S. orientalis Wernham, S. venosa Benth. and S.laxa
Wernham, as well as S. mollis K.Schum. ex Wern-
ham. Sabicea discolor Stapf from west tropical Africa
is retrieved as moderately supported (ML: 51, BI:
0.82) sister to this alliance. The species from S~
ao
Tom
e, Sabicea exellii G.Taylor and S. ingrata
K.Schum. do not form a clade; the former groups
with S. rosea Hoyle from West Africa (ML: 97, BI:
1.0, MP: 93), whereas the position of the second in a
clade of mostly Central African species receives only
weak support in the Bayesian analysis (BI: 0.71) and
is unsupported in the ML and MP analyses. Thus,
Pseudosabicea spp. are resolved in two clades, the
first as sister to Ecpoma–Sabicea s.s. and the second
as a moderately supported clade (ML: 89, BI: 1.0,
MP: 68) comprising four species nested in Sabicea
s.s. Sister to the latter clade, with moderate support
(ML: 88, BI: 0.99, MP: 58) is Sabicea mabouniensis
O.Leachenaud & Zemagho from Gabon.
MORPHOLOGICAL STUDY
Patterns of character evolution of potentially taxo-
nomically important characters are shown in Figures
3–9. The morphological comparison of monophyletic
units in Sabicea s.l. allows them to be recognised by
a combination of characters (Table 3) and four sub-
genera can be delimited in Sabicea. Further details
are given in the discussion.
DISCUSSION
HEKISTOCARPA–TAMRIDAEA–VIRECTARIA CLADE
In our analysis, Hekistocarpa,Tamridaea and Virec-
taria form a moderately supported clade, sister to
Sabicea s.l. (Fig. 2). This is in agreement with Des-
sein et al. (2001a) and Bremer & Eriksson (2009),
but in conflict with Khan et al. (2008a, b), who
retrieved Hekistocarpa as sister to all other genera
of Sabiceeae, and Kainulainen et al. (2013), who
Table 2. Characteristics of sequences and the alignments used in the phylogenetic analyses for Sabiceeae
Markers nrITS rps16 petD trnT-F
Plastid
combined
nrITS +all
plastid
Number of taxa 79 79 79 77 80 81
Range of GC contents in Sabiceeae
s.l. sequences (%)
54 30.3 30.7 26.0 31.8 36.8
Potentially informative characters 60 98 90 243 248 491
Aligned length in Sabiceeae s.l. 625 819 875 1931 2813 4318
©2016 The Linnean Society of London, Botanical Journal of the Linnean Society, 2016, 182, 551–580
556 L. ZEMAGHO ET AL.
Sabicea amazonensis
Tamridaea capsulifera
Hekistocarpa minutiflora
Mussaenda erythrophylla
Mussaenda arcuata
Pseudomussaenda flava
Virectaria multiflora
Virectaria procumbens
Virectaria angustifolia
Sabicea elliptica
Sabicea africana
Sabicea becquetii
Sabicea segregata
Sabicea a floribunda
Sabicea aurifodinae
Sabicea sp. SD2149
Sabicea mildbraedii var. letestui
Sabicea mildbraedii
Sabicea medusula
Sabicea sthenula
Sabicea batesii
Sabicea proselyta
Sabicea urbaniana
Sabicea gigantostipula
Sabicea hierniana
Sabicea xanthotricha
Sabicea caminata
Sabicea laxa
Sabicea mollis
Sabicea discolor
Sabicea urniformis
Sabicea bigerrica
Sabicea rufa
Sabicea carbunica
Sabicea capitellata
Sabicea pedicellata
Sabicea johnstonii
Sabicea gracilis
Sabicea ingrata var. ingrata 2
Sabicea ingrata var. ingrata 1
Sabicea calycina
Sabicea ceylanica
Sabicea diversifolia
Sabicea speciosa
Sabicea acuminata
Sabicea venosa
Sabicea orientalis
Sabicea villosa
Sabicea cinerea
Sabicea aspera
Sabicea aspera var. glabrescens
Sabicea brachycalyx
Sabicea tayloriae
Sabicea mexicana
Sabicea erecta
Sabicea cuneata
Sabicea pyramidalis
Sabicea chocoana
Sabicea panamensis
Sabicea humilis
Sabicea mattogrossensis
Sabicea cana
Sabicea najatrix
Sabicea gabonica
Sabicea urceolata
Sabicea pilosa
Sabicea brevipes
Sabicea gilletii
Sabicea ferruginea
Sabicea tchapensis
Sabicea duparquetiana
Sabicea bullata
Sabicea dewevrei
Sabicea vogelii
Sabicea harleyae
Sabicea rosea
Sabicea exellii
Virectaria major
Sabicea sp. SD1865
Sabicea mabouniensis
Sabicea nobilis
0.67/65/59
0.94/73/53
0.99/58/56
0.94/-/-
0.98/90/68
0.63/-/-
0.99/88/58
0.89/76/74
1.0/89/68
0.86/-/-
0.90/70/75
0.86/79/88
1.0/76/89
0.82/51/-
0.99/82/86
0.89/64/-
0.98/59/68
1.0/76/78
0.81/-/-
1.0/98/97
0.99/56/66
0.71/-/-
1.0/97/65
1.0/71/60
1.0/99/98
1.0/99/98
0.96/55/59
1.0/97/93
1.0/100/86
0.97/82/97
0.94/64/57
1.0/87/84
1.0/91/98
1.0/100/96
1.0/100/100
1.0/100/100
1.0/99/100
1.0/100/100
0.95/76/93
0.91/81/57
0.82/69/72
0.95/52/57
1.0/85/81
1.0/97/87
1.0/92/93
Asia
Madagascar
São Tomé & Príncipe
São Tomé & Príncipe
Sabicea
subg. Sabicea
Sabicea
subg. Stipulariopsis
Sabicea
subg. Stipularia
Sabicea
subg. Anisophyllae
Virectaria
Tamridaea
Hekistocarpa
Outgroup
Neotropis
Figure 2. Strict consensus tree derived from maximum likelihood (ML), maximum parsimony (MP) and Bayesian anal-
ysis (BI) of ITS–PetD–rps16–trnT-F data of tribe Sabiceeae and three outgroups from Pseudomussaenda and Mussaenda.
The numbers to the left represent Bayesian posterior probabilities (>0.95), in the middle ML bootstrap support values
(>50%) and to the right MP bootstrap values.
©2016 The Linnean Society of London, Botanical Journal of the Linnean Society, 2016, 182, 551–580
PHYLOGENETICS OF SABICEEAE (RUBIACEAE) 557
Erect
Creeping
Twining
Sarmentose
Pseudomussaenda flava
Mussaenda arcuata
Mussaenda erythrophylla
Hekistocarpa minutiflora
Tamridaea capsulifera
Virectaria angustifolia
Virectaria procumbens
Virectaria multiflora
Virectaria major
Sabicea africana
Sabicea elliptica
Sabicea becquetii
Sabicea sthenula
Sabicea medusula
Sabicea sp SD2149
Sabicea aurifodinae
Sabicea batesii
Sabicea sp SD1865
Sabicea mildbraedii
Sabicea mildbraedii var letestui
Sabicea gigantostipula
Sabicea hierniana
Sabicea urbaniana
Sabicea xanthotricha
Sabicea caminata
Sabicea excellii
Sabicea rosea
Sabicea dewevrei
Sabicea duparquetiana
Sabicea ferruginea
Sabicea gabonica
Sabicea najatrix
Sabicea tchapensis
Sabicea brevipes
Sabicea gilletii
Sabicea harleyae
Sabicea vogelii
Sabicea speciosa
Sabicea ceylanica
Sabicea bullata
Sabicea pilosa
Sabicea urceolata
Sabicea acuminata
Sabicea diversifolia
Sabicea mabouniensis
Sabicea segregata
Sabicea floribunda
Sabicea nobilis
Sabicea proselyta
Sabicea calycina
Sabicea ingrata var ingrata2
Sabicea ingrata var ingrata1
Sabicea rufa
Sabicea bigerrica
Sabicea urniformis
Sabicea capitellata
Sabicea carbunica
Sabicea gracilis
Sabicea johnstonii
Sabicea pedicellata
Sabicea discolor
Sabicea mollis
Sabicea laxa
Sabicea orientalis
Sabicea venosa
Sabicea cuneata
Sabicea aspera
Sabicea brachycalyx
Sabicea cana
Sabicea chocoana
Sabicea cinerea
Sabicea erecta
Sabicea aspera var glabrescens
Sabicea mattogrossensis
Sabicea panamensis
Sabicea amazonensis
Sabicea mexicana
Sabicea tayloriae
Sabicea pyramidalis
Sabicea villosa
Sabicea humilis
Character: growth form
Outgroup
Virectaria
Sabicea
subg. Stipularia
Sabicea
subg. Anisophyllae
Sabicea
subg. Stipulariopsi
s
Sabicea
subg. Sabicea
Tamridaea
Hekistocarpa
Figure 3. Evolution of growth form in Sabiceeae.
©2016 The Linnean Society of London, Botanical Journal of the Linnean Society, 2016, 182, 551–580
558 L. ZEMAGHO ET AL.
Character: anisophylly
Leaves equal or subequal
Leaves strongly unequal
Pseudomussaenda flava
Mussaenda arcuata
Mussaenda erythrophylla
Hekistocarpa minutiflora
Tamridaea capsulifera
Virectaria angustifolia
Virectaria procumbens
Virectaria multiflora
Virectaria major
Sabicea africana
Sabicea elliptica
Sabicea becquetii
Sabicea sthenula
Sabicea medusula
Sabicea sp SD2149
Sabicea aurifodinae
Sabicea batesii
Sabicea sp SD1865
Sabicea mildbraedii
Sabicea mildbraedii var letestui
Sabicea gigantostipula
Sabicea hierniana
Sabicea urbaniana
Sabicea xanthotricha
Sabicea caminata
Sabicea excellii
Sabicea rosea
Sabicea dewevrei
Sabicea duparquetiana
Sabicea ferruginea
Sabicea gabonica
Sabicea najatrix
Sabicea tchapensis
Sabicea brevipes
Sabicea gilletii
Sabicea harleyae
Sabicea vogelii
Sabicea speciosa
Sabicea ceylanica
Sabicea bullata
Sabicea pilosa
Sabicea urceolata
Sabicea acuminata
Sabicea diversifolia
Sabicea mabouniensis
Sabicea segregata
Sabicea floribunda
Sabicea nobilis
Sabicea proselyta
Sabicea calycina
Sabicea ingrata var ingrata2
Sabicea ingrata var ingrata1
Sabicea rufa
Sabicea bigerrica
Sabicea urniformis
Sabicea capitellata
Sabicea carbunica
Sabicea gracilis
Sabicea johnstonii
Sabicea pedicellata
Sabicea discolor
Sabicea mollis
Sabicea laxa
Sabicea orientalis
Sabicea venosa
Sabicea cuneata
Sabicea aspera
Sabicea brachycalyx
Sabicea cana
Sabicea chocoana
Sabicea cinerea
Sabicea erecta
Sabicea aspera var glabrescens
Sabicea mattogrossensis
Sabicea panamensis
Sabicea amazonensis
Sabicea mexicana
Sabicea tayloriae
Sabicea pyramidalis
Sabicea villosa
Sabicea humilis
Outgroup
Virectaria
Sabicea
subg. Stipularia
Sabicea
subg. Anisophyllae
Sabicea
subg. Stipulariopsi
s
Sabicea
subg. Sabicea
Tamridaea
Hekistocarpa
Figure 4. Evolution of anisophylly in Sabiceeae.
©2016 The Linnean Society of London, Botanical Journal of the Linnean Society, 2016, 182, 551–580
PHYLOGENETICS OF SABICEEAE (RUBIACEAE) 559
Character: stipule type
Entire to bidentate
Multifid
Pseudomussaenda flava
Mussaenda arcuata
Mussaenda erythrophylla
Hekistocarpa minutiflora
Tamridaea capsulifera
Virectaria angustifolia
Virectaria procumbens
Virectaria multiflora
Virectaria major
Sabicea africana
Sabicea elliptica
Sabicea becquetii
Sabicea sthenula
Sabicea medusula
Sabicea sp SD2149
Sabicea aurifodinae
Sabicea batesii
Sabicea sp SD1865
Sabicea mildbraedii
Sabicea mildbraedii var letestui
Sabicea gigantostipula
Sabicea hierniana
Sabicea urbaniana
Sabicea xanthotricha
Sabicea caminata
Sabicea excellii
Sabicea rosea
Sabicea dewevrei
Sabicea duparquetiana
Sabicea ferruginea
Sabicea gabonica
Sabicea najatrix
Sabicea tchapensis
Sabicea brevipes
Sabicea gilletii
Sabicea harleyae
Sabicea vogelii
Sabicea speciosa
Sabicea ceylanica
Sabicea bullata
Sabicea pilosa
Sabicea urceolata
Sabicea acuminata
Sabicea diversifolia
Sabicea mabouniensis
Sabicea segregata
Sabicea floribunda
Sabicea nobilis
Sabicea proselyta
Sabicea calycina
Sabicea ingrata var ingrata2
Sabicea ingrata var ingrata1
Sabicea rufa
Sabicea bigerrica
Sabicea urniformis
Sabicea capitellata
Sabicea carbunica
Sabicea gracilis
Sabicea johnstonii
Sabicea pedicellata
Sabicea discolor
Sabicea mollis
Sabicea laxa
Sabicea orientalis
Sabicea venosa
Sabicea cuneata
Sabicea aspera
Sabicea brachycalyx
Sabicea cana
Sabicea chocoana
Sabicea cinerea
Sabicea erecta
Sabicea aspera var glabrescens
Sabicea mattogrossensis
Sabicea panamensis
Sabicea amazonensis
Sabicea mexicana
Sabicea tayloriae
Sabicea pyramidalis
Sabicea villosa
Sabicea humilis
Outgroup
Virectaria
Sabicea
subg. Stipularia
Sabicea
subg. Anisophyllae
Sabicea
subg. Stipulariopsi
s
Sabicea
subg. Sabicea
Tamridaea
Hekistocarpa
Figure 5. Evolution of stipule type in Sabiceeae.
©2016 The Linnean Society of London, Botanical Journal of the Linnean Society, 2016, 182, 551–580
560 L. ZEMAGHO ET AL.
Character: bracts
Free to shortly connate at the base
Fused and urceolate,
Pseudomussaenda flava
Mussaenda arcuata
Mussaenda erythrophylla
Hekistocarpa minutiflora
Tamridaea capsulifera
Virectaria angustifolia
Virectaria procumbens
Virectaria multiflora
Virectaria major
Sabicea africana
Sabicea elliptica
Sabicea becquetii
Sabicea sthenula
Sabicea medusula
Sabicea sp SD2149
Sabicea aurifodinae
Sabicea batesii
Sabicea sp SD1865
Sabicea mildbraedii
Sabicea mildbraedii var letestui
Sabicea gigantostipula
Sabicea hierniana
Sabicea urbaniana
Sabicea xanthotricha
Sabicea caminata
Sabicea excellii
Sabicea rosea
Sabicea dewevrei
Sabicea duparquetiana
Sabicea ferruginea
Sabicea gabonica
Sabicea najatrix
Sabicea tchapensis
Sabicea brevipes
Sabicea gilletii
Sabicea harleyae
Sabicea vogelii
Sabicea speciosa
Sabicea ceylanica
Sabicea bullata
Sabicea pilosa
Sabicea urceolata
Sabicea acuminata
Sabicea diversifolia
Sabicea mabouniensis
Sabicea segregata
Sabicea floribunda
Sabicea nobilis
Sabicea proselyta
Sabicea calycina
Sabicea ingrata var ingrata2
Sabicea ingrata var ingrata1
Sabicea rufa
Sabicea bigerrica
Sabicea urniformis
Sabicea capitellata
Sabicea carbunica
Sabicea gracilis
Sabicea johnstonii
Sabicea pedicellata
Sabicea discolor
Sabicea mollis
Sabicea laxa
Sabicea orientalis
Sabicea venosa
Sabicea cuneata
Sabicea aspera
Sabicea brachycalyx
Sabicea cana
Sabicea chocoana
Sabicea cinerea
Sabicea erecta
Sabicea aspera var glabrescens
Sabicea mattogrossensis
Sabicea panamensis
Sabicea amazonensis
Sabicea mexicana
Sabicea tayloriae
Sabicea pyramidalis
Sabicea villosa
Sabicea humilis
Fused and urceolate,
splitting in fruiting stage
not splitting in fruiting stage
Outgroup
Virectaria
Sabicea
subg. Stipularia
Sabicea
subg. Anisophyllae
Sabicea
subg. Stipulariopsi
s
Sabicea
subg. Sabicea
Tamridaea
Hekistocarpa
Figure 6. Evolution of bracts in Sabiceeae.
©2016 The Linnean Society of London, Botanical Journal of the Linnean Society, 2016, 182, 551–580
PHYLOGENETICS OF SABICEEAE (RUBIACEAE) 561
Character: inflorescence type
Branched
Unbranched and sessile
Unbranched and pedunculate
Pseudomussaenda flava
Mussaenda arcuata
Mussaenda erythrophylla
Hekistocarpa minutiflora
Tamridaea capsulifera
Virectaria angustifolia
Virectaria procumbens
Virectaria multiflora
Virectaria major
Sabicea africana
Sabicea elliptica
Sabicea becquetii
Sabicea sthenula
Sabicea medusula
Sabicea sp SD2149
Sabicea aurifodinae
Sabicea batesii
Sabicea sp SD1865
Sabicea mildbraedii
Sabicea mildbraedii var letestui
Sabicea gigantostipula
Sabicea hierniana
Sabicea urbaniana
Sabicea xanthotricha
Sabicea caminata
Sabicea excellii
Sabicea rosea
Sabicea dewevrei
Sabicea duparquetiana
Sabicea ferruginea
Sabicea gabonica
Sabicea najatrix
Sabicea tchapensis
Sabicea brevipes
Sabicea gilletii
Sabicea harleyae
Sabicea vogelii
Sabicea speciosa
Sabicea ceylanica
Sabicea bullata
Sabicea pilosa
Sabicea urceolata
Sabicea acuminata
Sabicea diversifolia
Sabicea mabouniensis
Sabicea segregata
Sabicea floribunda
Sabicea nobilis
Sabicea proselyta
Sabicea calycina
Sabicea ingrata var ingrata2
Sabicea ingrata var ingrata1
Sabicea rufa
Sabicea bigerrica
Sabicea urniformis
Sabicea capitellata
Sabicea carbunica
Sabicea gracilis
Sabicea johnstonii
Sabicea pedicellata
Sabicea discolor
Sabicea mollis
Sabicea laxa
Sabicea orientalis
Sabicea venosa
Sabicea cuneata
Sabicea aspera
Sabicea brachycalyx
Sabicea cana
Sabicea chocoana
Sabicea cinerea
Sabicea erecta
Sabicea aspera var glabrescens
Sabicea mattogrossensis
Sabicea panamensis
Sabicea amazonensis
Sabicea mexicana
Sabicea tayloriae
Sabicea pyramidalis
Sabicea villosa
Sabicea humilis
Outgroup
Virectaria
Sabicea
subg. Stipularia
Sabicea
subg. Anisophyllae
Sabicea
subg. Stipulariopsi
s
Sabicea
subg. Sabicea
Tamridaea
Hekistocarpa
Figure 7. Evolution of inflorescence type in Sabiceeae.
©2016 The Linnean Society of London, Botanical Journal of the Linnean Society, 2016, 182, 551–580
562 L. ZEMAGHO ET AL.
Short trichomes
Long trichomes
Pseudomussaenda flava
Mussaenda arcuata
Mussaenda erythrophylla
Hekistocarpa minutiflora
Tamridaea capsulifera
Virectaria angustifolia
Virectaria procumbens
Virectaria multiflora
Virectaria major
Sabicea africana
Sabicea elliptica
Sabicea becquetii
Sabicea sthenula
Sabicea medusula
Sabicea sp SD2149
Sabicea aurifodinae
Sabicea batesii
Sabicea sp SD1865
Sabicea mildbraedii
Sabicea mildbraedii var letestui
Sabicea gigantostipula
Sabicea hierniana
Sabicea urbaniana
Sabicea xanthotricha
Sabicea caminata
Sabicea excellii
Sabicea rosea
Sabicea dewevrei
Sabicea duparquetiana
Sabicea ferruginea
Sabicea gabonica
Sabicea najatrix
Sabicea tchapensis
Sabicea brevipes
Sabicea gilletii
Sabicea harleyae
Sabicea vogelii
Sabicea speciosa
Sabicea ceylanica
Sabicea bullata
Sabicea pilosa
Sabicea urceolata
Sabicea acuminata
Sabicea diversifolia
Sabicea mabouniensis
Sabicea segregata
Sabicea floribunda
Sabicea nobilis
Sabicea proselyta
Sabicea calycina
Sabicea ingrata var ingrata2
Sabicea ingrata var ingrata1
Sabicea rufa
Sabicea bigerrica
Sabicea urniformis
Sabicea capitellata
Sabicea carbunica
Sabicea gracilis
Sabicea johnstonii
Sabicea pedicellata
Sabicea discolor
Sabicea mollis
Sabicea laxa
Sabicea orientalis
Sabicea venosa
Sabicea cuneata
Sabicea aspera
Sabicea brachycalyx
Sabicea cana
Sabicea chocoana
Sabicea cinerea
Sabicea erecta
Sabicea aspera var glabrescens
Sabicea mattogrossensis
Sabicea panamensis
Sabicea amazonensis
Sabicea mexicana
Sabicea tayloriae
Sabicea pyramidalis
Sabicea villosa
Sabicea humilis
Character: corolla-throat indumentum
Outgroup
Virectaria
Sabicea
subg. Stipularia
Sabicea
subg. Anisophyllae
Sabicea
subg. Stipulariopsi
s
Sabicea
subg. Sabicea
Tamridaea
Hekistocarpa
Figure 8. Evolution of corolla throat indumentum in Sabiceeae.
©2016 The Linnean Society of London, Botanical Journal of the Linnean Society, 2016, 182, 551–580
PHYLOGENETICS OF SABICEEAE (RUBIACEAE) 563
Character: ovary
2-locular
(3)- 5-locular
Pseudomussaenda flava
Mussaenda arcuata
Mussaenda erythrophylla
Hekistocarpa minutiflora
Tamridaea capsulifera
Virectaria angustifolia
Virectaria procumbens
Virectaria multiflora
Virectaria major
Sabicea africana
Sabicea elliptica
Sabicea becquetii
Sabicea sthenula
Sabicea medusula
Sabicea sp SD2149
Sabicea aurifodinae
Sabicea batesii
Sabicea sp SD1865
Sabicea mildbraedii
Sabicea mildbraedii var letestui
Sabicea gigantostipula
Sabicea hierniana
Sabicea urbaniana
Sabicea xanthotricha
Sabicea caminata
Sabicea excellii
Sabicea rosea
Sabicea dewevrei
Sabicea duparquetiana
Sabicea ferruginea
Sabicea gabonica
Sabicea najatrix
Sabicea tchapensis
Sabicea brevipes
Sabicea gilletii
Sabicea harleyae
Sabicea vogelii
Sabicea speciosa
Sabicea ceylanica
Sabicea bullata
Sabicea pilosa
Sabicea urceolata
Sabicea acuminata
Sabicea diversifolia
Sabicea mabouniensis
Sabicea segregata
Sabicea floribunda
Sabicea nobilis
Sabicea proselyta
Sabicea calycina
Sabicea ingrata var ingrata2
Sabicea ingrata var ingrata1
Sabicea rufa
Sabicea bigerrica
Sabicea urniformis
Sabicea capitellata
Sabicea carbunica
Sabicea gracilis
Sabicea johnstonii
Sabicea pedicellata
Sabicea discolor
Sabicea mollis
Sabicea laxa
Sabicea orientalis
Sabicea venosa
Sabicea cuneata
Sabicea aspera
Sabicea brachycalyx
Sabicea cana
Sabicea chocoana
Sabicea cinerea
Sabicea erecta
Sabicea aspera var glabrescens
Sabicea mattogrossensis
Sabicea panamensis
Sabicea amazonensis
Sabicea mexicana
Sabicea tayloriae
Sabicea pyramidalis
Sabicea villosa
Sabicea humilis
Outgroup
Virectaria
Sabicea
subg. Stipularia
Sabicea
subg. Anisophyllae
Sabicea
subg. Stipulariopsi
s
Sabicea
subg. Sabicea
Tamridaea
Hekistocarpa
Figure 9. Evolution of ovary in Sabiceeae.
©2016 The Linnean Society of London, Botanical Journal of the Linnean Society, 2016, 182, 551–580
564 L. ZEMAGHO ET AL.
retrieved Tamridaea and Virectaria as sister to the
remaining Sabiceeae. The position of Hekistocarpa
seems to be ambiguous, pending further study. In
our study, the monophyly of the Hekistocarpa–Tam-
ridaea–Virectaria clade is entirely based on the
rps16 sequences and it should be confirmed by addi-
tional molecular data. Morphologically, this clade dif-
fers from Sabicea s.l. by its dry fruits. Fruit
characteristics, among others (see key to the genera),
can also be used to separate the three genera: fruits
indehiscent or tardily splitting into two mericarps in
Hekistocarpa, capsules with two persistent valves in
Tamridaea and capsules with one caducous valve
and the other remaining attached to the pedicel in
Virectaria (Dessein et al., 2001a, b). In this clade,
the African genus Virectaria is fully supported as sis-
ter to Tamridaea, a genus endemic to Socotra, which
is in agreement with previous molecular studies. The
two genera are characterized by terminal inflores-
cences, a characteristic not found in other members
of Sabiceeae.
SYSTEMATIC VALUE OF INDIVIDUAL MORPHOLOGICAL
CHARACTERISTICS
Evaluation of morphological characteristics
The results from our study of morphological charac-
ter evolution demonstrate that many characters used
to hypothesise relationships in Sabicea s.l. are labile
and do not delineate the genera as proposed by
earlier studies. Several characters were judged as
uninformative for generic and subgeneric delimita-
tion after preliminary morphological examination,
because they exhibited either little information (e.g.
pollen morphology, size and shape of the disc) or con-
siderable intraspecific variation (external indumen-
tum of vegetative parts, dimensions of leaves and
stipules). These were excluded from further analysis.
Our molecular phylogenetic tree for Sabiceeae pro-
vides an independent hypothesis to evaluate taxo-
nomically important morphological characters such
as growth form (Fig. 3), anisophylly (Fig. 4), stipule
type (Fig. 5), bracts (Fig. 6), inflorescence type
(Fig. 7), corolla-throat indumentum (Fig. 8) and the
number of ovary locules (Fig. 9) (see Appendices 1
and 2).
Growth form (Fig. 3)
Sabicea s.l. shows a remarkable range of variation
in growth habit. In the present study, we sum-
marised the habits in four major groups: Erect shrub
or herb (0); creeping (1); twining (2), i.e. with stems
climbing in spiral around a support; and sarmentose
(3), i.e. more or less lianescent, but not climbing in
spiral. In the creeping category we only included
the small species (<30 cm tall) with the stems
appressed to ground or shortly ascending at the
apex. There are some ambiguous cases in which the
stems are creeping at base and twining at apex (S.
pilosa,S. bullata) or creeping at base and sarmen-
tose at apex (S. sp.-SD2149); these were considered
as twining or sarmentose, respectively. Although the
outgroups are somewhat ambiguous with the two
Mussaenda spp. described as ‘shrub, scandent
shrub or climber’ (Bridson & Verdcourt, 1988), Pseu-
domussaenda flava is an erect shrub and the early
branching lineages in Sabiceeae s.l. are all erect
(Hekistocarpa,Tamridaea,Virectaria, and Sabicea
subgenus Stipularia). Thus, an erect habit is pre-
sumably ancestral in tribe Sabiceeae s.l., in contrast
to the findings of Razafimandimbison et al. (2012)
for Morindeae. Sabicea subgenus Anisophyllae com-
prises creeping and sarmentose growth forms, but
no twining ones. In contrast, most species of sub-
genus Sabicea are twining and this habit presum-
ably evolved a single time. In Africa, one clade (the
Malagasy S. acuminata and S. diversifolia, the for-
mer Pseudosabicea spp. has evolved a sarmentose
habit, also including the creeping S. mabouniensis.
The only Asian species, S. ceylanica, however, is a
creeper and, in the New World, several reversals to
an erect habit have taken place, supporting the view
that evolutionary changes between lianescent and
erect habits are reversable (Razafimandimbison
et al., 2012). Possibly a lianescent habit provided an
evolutionary advantage in the rain forests of West
Africa, the preferred habitat of Sabicea subgenus
Sabicea, in which there is no lack of supporting
trees, but intense competition for light, the ideal
combination for lianescent growth forms (Schnitzer
& Bongers, 2002).
Anisophylly (Fig. 4)
Some species of Sabicea s.l. are anisophyllous and
Hall
e (1963) used this character to diagnose
Pseudosabicea section Anisophyllae. There is a good
range of variation in the level of anisophylly: leaves
slightly unequal with one about twice larger than
the other (S. tersifolia), strongly unequal with one
normally developed and the other one vestigial (e.g.
S. aurifodinae), or, at the extreme, a single leaf per
node (S. batesii,S. sthenula). Our study shows that
unequal leaves appeared two or three times in Sabi-
ceeae: once or possibly twice in subgenus Sabicea (S.
mabouniensis from Gabon, S. acuminata and S.
diversifolia from Madagascar) and once in subgenus
Anisophyllae.Sabicea becquetii with equal or slightly
unequal leaves is sister to the remainder of the Anis-
ophyllae clade. Anisophylly is restricted to species
with creeping or sarmentose habit (Fig. 3), and can
be understood as an adaptation that maximizes light
capture in dimly lit understories (Dengler, 1999).
©2016 The Linnean Society of London, Botanical Journal of the Linnean Society, 2016, 182, 551–580
PHYLOGENETICS OF SABICEEAE (RUBIACEAE) 565
Anisophylly is therefore homoplasious and cannot be
used alone to delimit subgenus Anisophyllae.
Stipule type (Fig. 5)
Stipules are often useful in distinguishing genera of
Rubiaceae (Gentry, 1993). The stipules in Sabiceeae
are interpetiolar and persistent; in Sabicea subgenus
Anisophyllae they are often connate with the base of
the petioles. They vary in shape from entire or
shortly bidentate at apex, to multifid (divided in
many segments). Multifid stipules are found in some
(but not all) species of Sabicea subgenus Anisophyl-
lae, but also in the Malagasy taxa S. acuminata and
S. diversifolia. Consequently, this character has to
be considered homoplasious.
Bracts (Fig. 6)
The genus Stipularia has been separated from Sabi-
cea (e.g. Hall
e, 1966) based on its peculiar urn-
shaped involucre surrounding the inflorescence. This
involucre consists of two to four bracts that are fused
for most of their length, and either split in fruit
[S. elliptica(Schweinf. ex Hiern) Hepper or remain
intact (S. africana P.Beauv.). Although nothing is yet
known about pollination in subgenus Stipularia, the
conspicuous bracts are most likely detectable at long
distances by long-ranging pollinators, as has been
shown for the bracts of Mussaenda frondosa L.
(Borges, Gowda & Zacharias, 2003). As pointed out by
Hepper (1958), the bracts of Sabicea s.s. show consid-
erable variation: they can be free and inconspicuous
(e.g. S. exellii), free and conspicuously surrounding
the inflorescence (e.g. S. capitellata Benth.) or shortly
fused at the base (S. dewevrei De Wild. & T.Durand,
S. urniformis Zemagho, O.Lachenaud & Sonk
e; in the
latter species the bracts split in fruit); an exception is
S. urceolata that has bracts fused for most of their
length and splitting in fruit, thus much resembling
the involucre of Stipularia. This character is therefore
also homoplasious. In subgenus Anisophyllae bracts
are free, usually small and inconspicuous (in Sabicea
sp.-SD2149, they are larger and enclose the flowers
when young, but not the whole inflorescence). In sub-
genus Stipulariopsis, bracts are free and small, except
in S. gigantostipula, which has two large free bracts
surrounding the inflorescence.
Table 3. Comparison of morphological characters of the subgenera of Sabicea s.l. (as used in this paper)
Characteristics
Sabicea subgenus
Anisophyllae
Sabicea subgenus
Sabicea
Sabicea subgenus
Stipularia
Sabicea subgenus
Stipulariopsis
Growth habit Creeping or sarmentose Usually twining,
sometimes
creeping or
sarmentose
Erect Erect
Leaves Strongly anisophyllous
(except S. becquetii
and S. arborea)
Isophyllous, rarely
anisophyllous
Isophyllous Isophyllous
Stipule type Entire or multifid Entire or multifid Entire Entire
Bracts Free, usually small,
not surrounding
inflorescence
Free or shortly connate
at base,
rarely (S. urceolata)
connate for most of
their length into
an involucre
splitting laterally in fruit
Fused for most their
length into an
urceolate
involucre, splitting in
fruit (S. elliptica)or
remaining intact
(S. africana)
Free and
usually small
(large in
S. gigantostipula)
Inflorescence Unbranched,
sessile
Branched or unbranched,
sessile or pedunculate
Unbranched,
sessile
Unbranched,
sessile
Indumentum
of corolla throat
Long trichomes
forming a dense
beard (except
S. becquetii
and S. arborea)
Short trichomes, or,
rarely, long
trichomes forming
a dense beard
(S. acuminata,
S. diversifolia)
Short trichomes Short trichomes
Ovary Bilocular Bi- to pentalocular Tri- to pentalocular Bilocular
Distribution Central and
East Africa
Pantropical West and
Central Africa
Central Africa
(Lower Guinea
Domain)
©2016 The Linnean Society of London, Botanical Journal of the Linnean Society, 2016, 182, 551–580
566 L. ZEMAGHO ET AL.
Inflorescence type (Fig. 7)
In the present study, we summarised inflorescences
of Sabiceeae in three major types: (1) branched; (2)
unbranched and sessile; or (3) unbranched and
pedunculate. Our study shows that all inflorescence
types occur in subgenus Sabicea. The inflorescences
of subgenus Stipularia are always unbranched, but
vary from sessile to pedunculate. In subgenus Stipu-
lariopsis and Anisophyllae, the inflorescences are
always unbranched and sessile (with fruits pedicel-
late in the former, and usually sessile in the latter).
Unbranched and sessile inflorescences are also found
in some species of subgenus Sabicea (e.g. S. bullata,
S. diversifolia,S. speciosa), hence, this character is
also homoplasious.
Indumentum of corolla throat (Fig. 8)
An important character that has been overlooked in
the discussion of Sabiceeae is the nature of the
internal indumentum of the corolla. Verdcourt (1958)
examined the trichomes of many genera of Rubiaceae
and distinguished two groups: the internal indumen-
tum (inside the corolla tube) and the external indu-
mentum (other plant organs). Although Verdcourt
(1958) considered the trichome structure of external
indumentum as a useful secondary character, the
systematic importance of the inner indumentum has
not been extensively discussed in the literature. The
inner indumentum mostly consists of unicellular
thin-walled trichomes, which are flat and ribbon-like
or sometimes moniliform (Robbrecht, 1988). In Sabi-
ceeae s.l., the internal indumentum consists of thin
white multicellular trichomes of variable length. Of
particular systematic value are the trichomes occur-
ing around the corolla throat, which are short (c.
0.5 mm) and sparse in subgenera Stipulariopsis and
Stipularia and in most species of subgenus Sabicea.
In subgenus Anisophyllae (except for S. becquetii)
these trichomes are long (1–2 mm) and form a dense
beard around the throat, sometimes extending to the
basal part of the lobes. However, the Madagascan
species S. acuminata and S. diversifolia also display
a long and dense corolla throat indumentum, so this
character appears to be homoplasious. Dense beards
around the corolla throat are also found in the out-
group species of Mussaendeae. However, in that
tribe, the trichomes are usually red or yellow and
therefore different from the ones encountered in
Sabiceeae.
Number of ovary locules (Fig. 9)
Hall
e (1963) considered the number of ovary locules
as an useful character to separate Ecpoma and
Pseudosabicea (bilocular) from Sabicea pentalocular),
whereas Wernham (1914) judged this character
as quantitative and therefore of little taxonomic
significance. In subgenus Sabicea, most species are
pentalocular, however, the former Pseudosabicea
spp. nested in Sabicea and S. mabouniensis are
bilocular and so are the members of subgenera Stip-
ulariopsis and Anisophyllae. Thus, locule number is
homoplasious in Sabicea and cannot be used alone
for subgeneric delimitation. Similar homoplasy of
locule number has been demonstrated recently in
other genera of Rubiaceae, e.g. Pyrostria Comm. ex
Juss. (Vanguerieae; Razafimandimbison et al., 2009),
diminishing the value of this character for generic
delimitations.
NEW SUBGENERIC CLASSIFICATION FOR SABICEA S.L
The synonymization of Ecpoma,Pseudosabicea and
Stipularia under Sabicea as proposed by Khan et al.
(2008a) is accepted in the present study. However,
Khan et al. (2008a) did not propose a new subgeneric
classification for Sabicea s.l., indicating that a better
sampling was needed for taxa previously placed in
Ecpoma,Pseudosabicea and Stipularia. With the
sampling currently presented, several moderately to
well supported clades can be identified in Sabicea
s.l., allowing us to propose a new subgeneric classifi-
cation (see key to the subgenera).
We chose to use subgenera instead of sections,
since one of the clades (subgenus Sabicea) may have
to be further subdivided into sections in the future.
Moreover, although additional studies are needed to
delimitate monophyletic groups in Sabicea sensu
Hall
e, some previous sections could be clearly
found in this latter (sections Floribundae, Laxae,
Sphaericae).
The first clade branching off consists of S. afri-
cana and S. elliptica and is strongly supported.
This clade is characterized by the erect habit and
the peculiar urceolate involucre surrounding the
inflorescences. On account of the latter characteris-
tic, both species were previously referred to the
separate genus Stipularia (Palisot-Beauvois, 1810;
Hall
e, 1963, 1966). As the clade is sister to the
rest of Sabicea s.l., the generic status of Stipularia
could be retained. However, since none of its diag-
nostic characteristics is synapomorphic (the erect
habit also occurs in subgenus Stipulariopsis, and
the urceolate involucre in S. urceolata), we agree
with Hepper (1958) and Khan et al. (2008a) to
include Stipularia in Sabicea.
A second well supported lineage in Sabicea s.l.
corresponds to Pseudosabicea section Anisophyllae
(Hall
e, 1963). This group is well supported and
comprises six described species and one variety
(Table 4) and two still undescribed taxa (S. sp.-
SD1865, S. sp.-SD2149). Hall
e (1963) characterized
section Anisophyllae by anisophyllous nodes
©2016 The Linnean Society of London, Botanical Journal of the Linnean Society, 2016, 182, 551–580
PHYLOGENETICS OF SABICEEAE (RUBIACEAE) 567
combined with sessile inflorescences. However, both
characteristics are also found in the two Madagas-
can species of subgenus Sabicea, which differ by
their pentalocular ovaries. The present study shows
that this clade lacks a unique synapomorphy and
that it can only be circumscribed by a combination
of characters, i.e. bilocular ovaries and inflores-
cences constantly unbranched and sessile. Species
of this group also usually show strong anisophylly
and long trichomes in the corolla throat. However,
S. becquetii (and S. arborea, not sequenced here
and morphologically similar) are aberrant in having
leaves equal or only slightly unequal and a corolla
throat with short trichomes. A case could be made
for placing them in a separate subgenus (especially
if we consider the early branching position of S.
Table 4. Subgeneric composition and name changes in Sabicea
Pseudosabicea section Anisophyllae N.Hall
eSabicea subgenus Anisophyllae (N.Hall
e)
Zemagho, Sonk
e, Dessein & Liede
Pseudosabicea arborea (K.Schum.) N.Hall
eSabicea arborea K.Schum.
Pseudosabicea aurifodinae* N.Hall
eSabicea aurifodinae (N.Hall
e) Razafim.,
B.Bremer, Liede & Saleh A.Khan
Pseudosabicea batesii* (Wernham) N.Hall
eSabicea batesii Wernham
Pseudosabicea becquetii* N.Hall
eSabicea becquetii (N.Hall
e) Razafim.,
B.Bremer, Liede & Saleh A.Khan
Pseudosabicea medusula*
(K.Schum. ex Wernham) N.Hall
e
Sabicea medusula (K.Schum) Wernham
Pseudosabicea mildbraedii* (Wernham) N.Hall
eSabicea mildbraedii Wernham
Pseudosabicea sthenula* N. Hall
eSabicea sthenula (N.Hall
e) Razafim., B.Bremer,
Liede & Saleh A.Khan
Ecpoma K.Schum. Sabicea subgenus Stipulariopsis Wernham
Ecpoma apocynaceum K.Schum. Sabicea apocynacea (K.Schum.) Razafim.,
B.Bremer, Liede & Saleh A.Khan
Ecpoma cauliflorum (Hiern) N.Hall
eSabicea cauliflora Hiern
Ecpoma geanthum (Hiern) N.Hall
eSabicea geantha Hiern
Ecpoma gigantostipulum* (K.Schum.) N.Hall
eSabicea gigantostipula K.Schum.
Ecpoma hiernianum*
(Wernham) N.Hall
e & F.Hall
e
Sabicea hierniana Wernham
Sabicea subgenus Stipulariopsis Wernham
Sabicea urbaniana* Wernham Sabicea urbaniana Wernham
Sabicea xanthotricha* Wernham Sabicea xanthotricha Wernham
Stipularia P.Beauv. Sabicea subgenus Stipularia (P.Beauv.)
Zemagho, Sonke
´, Dessein & Liede
Stipularia africana*P.Beauv. Sabicea africana (P.Beauv.) Hepper
Stipularia elliptica* Schweinf. ex Hiern Sabicea elliptica (Schweinf. ex Hiern) Hepper
Stipularia mollis Wernham Sabicea lanata Hepper
Sabicea Aublet Sabicea subgenus Sabicea
43 species New World 43 species New World
c. 75 species Old World c. 75 species Old World
Schizostigma Arn. ex Meisn.
Schizostigma hirsutum* Arn. ex Meisn. Sabicea ceylanica Puff
Pseudosabicea N.Halle
´section Sphaericae N.Hall
e
Pseudosabicea nobilis* (R.D.Good) N.Hall
eSabicea nobilis R.D.Good
Pseudosabicea N.Halle
´section Floribundae Wernh. ex N.Hall
e
Pseudosabicea floribunda* (K.Schum.) N.Hall
eSabicea floribunda K.Schum.
Pseudosabicea proselyta* N.Hall
eSabicea proselyta (N.Hall
e) Razafim., B. Bremer,
Liede & Saleh A.Khan
Pseudosabicea sanguinosa N.Hall
eSabicea sanguinosa (N.Hall
e) Razafim.,
B.Bremer, Liede & Saleh A.Khan
Pseudosabicea segregata* (Hiern) N.Hall
eSabicea segregata Hiern
Type species are marked in bold.
*Species included in the present study.
©2016 The Linnean Society of London, Botanical Journal of the Linnean Society, 2016, 182, 551–580
568 L. ZEMAGHO ET AL.
becquetii in the group), but due to lack of sequence
data for S. arborea, we refrain from doing so at
this moment (Table 4).
A third moderately well supported lineage in Sabi-
cea s.l. groups the former Ecpoma species (E. gigan-
tostipula (K.Schum.) N.Hall
e and E. hierniana
(Wernham) N.Hall
e and F.Hall
e with Sabicea urba-
niana and S. xanthotricha. The latter two species
were not included in Ecpoma by Hall
e (1963) pre-
sumably because he had not seen the material. This
clade is morphologically homogeneous and can be
characterized by the erect habit, bilocular ovaries
and unbranched sessile inflorescences with free (and
usually small) bracts. It could be retained as a
genus, but this would render Sabicea paraphyletic,
unless subgenus Anisophyllae is also elevated to gen-
eric level. This group largely corresponds with Wern-
ham’s (1914) subgenus Stipulariopsis, so this
subgenus is here reinstated. Two aberrant species
placed in this subgenus by Wernham have already
been excluded from Sabicea:S. bicarpellata
K.Schum. is a Bertiera (Hall
e, 1970) and the
Neotropical S. umbrosa Wernham is now placed in
Amphidasya (Standley, 1936). Following Art. 11 of
the International Code of Botanical Nomenclature,
the name Stipulariopsis has priority over Ecpoma at
the infrageneric rank, because the latter was
described as a genus.
The fourth clade comprises all species of Sabicea
s.s., plus Schizostigma,Pseudosabicea section Flori-
bundae, and Sabicea nobilis (the type species of the
genus Pseudosabicea and of section Sphaericae).
Despite the generally low resolution, the three latter
groups are deeply and unambiguously nested in this
clade (Table 4). Because the type species of Sabicea,
S. cinerea Aubl., falls into this clade, it has to be
recognised as Sabicea subgenus Sabicea. It com-
prises c. 75 species in the Old World and 43 species
in the New World (Khan, 2007). With the deeper
nodes unresolved or only weakly supported, the rela-
tionships in this subgenus remain unclear and will
be subject of further phylogenetic studies including
more taxa and/or characteristics.
TAXONOMIC TREATMENT
TRIBE SABICEEAE Bremek., Rec. Trav. Bot. Neerl.
31: 253 (1934). Type: Sabicea Aubl.
Virectarieae Verdc., Kew Bull. 30: 366 (1975).
Type: Virectaria Bremek.
Plants annual or perennial, herbaceous to woody,
erect to variously climbing or creeping. Raphides
absent. Stipules persistent, interpetiolar, some-
times extensively fused to petiole, entire
to deeply laciniate. Inflorescences axillary or
terminal, branched or not, sessile or pedunculate.
Flowers (tetra-)penta-(hexa-)merous, heterostylous or
homostylous. Corolla tube cylindrical to narrowly fun-
nel-shaped, 3–35 mm long, villose inside; lobes
reduplicate or truly valvate, glabrous to villose inside.
Ovary bi- to pentalocular. Placentas elliptic or oblong,
rarely cordate, attached throughout their length or
rarely peltate (and then the stipe usually attached
above middle), each with numerous ovules. Fruits fle-
shy and indehiscent, dry and indehiscent or capsular
with two valves. Seeds numerous, angular; exotesta
cells without secondary thickening in outer tangential
wall, radial walls distinctly thickened, at least in
lower part, inner tangential wall with distinct sec-
ondary thickening in the shape of bands or an even
sheet perforated by minute pores. Pollen grains tri- or
tetracolporate or tri- or tetra- (penta-)pororate.
KEY TO THE GENERA OF TRIBE SABICEEAE
1. Inflorescences terminal; ovary bilocular; fruits
dry, capsular and bivalved. . . . . . . . . . . . . . . 2.
Inflorescences axillary; ovary bi- to pentalocular;
fruits dry or fleshy, indehiscent or tardily split-
tingintotwomericarps.................3.
2. Capsules with two valves remaining attached to
the pedicel; corolla tube much longer than lobes,
the latter unequally trilobed at apex; corolla aes-
tivation reduplicate valvate; anthers included;
style with two filiform lobes . . . . . . . .Tamridaea.
Capsules with one valve caducous and the other
remaining attached to the pedicel; corolla tube
not or only slightly longer than lobes, the latter
entire; corolla aestivation valvate, anthers long-
exserted; style linear, without stigmatic
branches . . . . . .................Virectaria.
3. Fruits dry, indehiscent or tardily splitting into
two mericarps; inflorescence scorpioid; corolla
aestivationreduplicatevalvate.............
..........................Hekistocarpa.
Fruits fleshy, indehiscent; inflorescence not scor-
pioid; corolla aestivation truly valvate ........
..............................Sabicea.
DIVERSITY AND GEOGRAPHICAL DISTRIBUTION OF TRIBE
SABICEEAE
Tribe Sabiceeae are most diverse in tropical Africa
and presumably have their origin there. The genera
Hekistocarpa and Virectaria are African endemics.
Virectaria is essentially Guineo-Congolian with pene-
tration into the Zambezian region and has its centre
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PHYLOGENETICS OF SABICEEAE (RUBIACEAE) 569
of diversity in the Lower Guinea subcentre of ende-
mism (Dessein et al., 2001b; phytochoria according to
White, 1979). Hekistocarpa is a monospecific genus
with a limited distribution in Lower Guinea, from
Nigeria to northern Congo-Brazzaville. The
monospecific Tamridaea is endemic to Socotra. Only
Sabicea, by far the largest genus, has a broad distri-
bution across the tropics, with its main centre of
diversity in tropical Africa (especially Lower Guinea)
and an important secondary centre in tropical Amer-
ica; it also includes a few representatives in Mada-
gascar and a single species in Sri Lanka.
DESCRIPTION OF SABICEA AUBL
Sabicea Aubl. Hist. Pl. Guiane Franc
ßoise 1: 192, t.
75. Jun-Dec 1775 –Lectotype: S. cinerea Aubl., des-
ignated by Standley (1921: 148).
Schwenkfelda Schreb. (Gen. Pl. 1: 123.
1789) Schwenkfeldia Willd. (Sp. Pl. 4 [post Reich-
ardianum quinta]: 982. 1798) –Illegitimate replace-
ment name for Sabicea.
Plants perennial, woody or, more rarely, herba-
ceous, usually lianescent, more rarely erect or creep-
ing, sometimes anisophyllous. Stipules persistent,
interpetiolar, free or rarely connate with the petioles
at base, entire to deeply laciniate. Inflorescences
axillary, branched or not, sessile to pedunculate.
Bracts conspicuous or not, often surrounding the
inflorescence, free to shortly connate, or rarely
connate for most of their length. Flowers (tetra-)
penta-(-hexa-)merous, heterostylous. Calyx campanu-
late to infundibuliform, usually green, lobes antrorse
to reflexed, usually lanceolate, subulate to subulate-
setaceous or setaceous. Corolla white to pale green
or occasionally bright red; tube cylindrical or nar-
rowly infundibuliform, 3–22 mm long, villose inside
in the upper part; lobes valvate, triangular, (sub-)
acute at apex, glabrous to villose inside. Stamens
(four–)five (to ten) per flower, included to fully
exserted; anthers dorsifixed near the middle by short
free part of filament, linear to narrowly oblong,
acute to rounded at base and apex. Pollen grains col-
porate to pororate, apertures three or four, exine
surface minutely reticulate, released as monads.
Style glabrous, included or exserted, divided into
two to five linear to broadly spathulate stigmatic
lobes. Ovary bi- to pentalocular with axile placenta-
tion and numerous ovules per locule. Fruits fleshy,
indehiscent and bacciform, white, red, purple or
black when mature, globose to ellipsoid or more
rarely fusiform. Seeds numerous and small, vari-
ously angular, usually with parallel striations or
more rarely reticulate; exotesta cells variable in
shape, with few to many rounded pits, radial wall
with verrucose thickenings.
Number of species: c. 150 species (c. 96 confined to
the African mainland, 43 to the Neotropics, six to
Madagascar, three to S~
ao Tom
e and Pr
ıncipe and
one to Sri Lanka).
KEY TO THE SUBGENERA OF SABICEA
1 Erectherbsorshrubs....................2.
Plants twining, creeping or sarmentose (never
erect).................................3.
2 Outer bracts fused into an urn-shaped involucre;
ovary tri- to pentalocular; calyx lobes shorter than
tube or at most equalling it; leaves strongly discol-
orous with dense felt of whitish trichomes
beneath . . . . . . . . . . . . . . . . subgenus Stipularia.
All bracts free, usually small; ovary bilocular;
calyx lobes much longer than tube; leaves green on
both sides . . . .........subgenus Stipulariopsis.
3 Ovary 5-locular; plants usually twining, rarely
creeping or sarmentose subgenus Sabicea
Ovary 2-locular; plants sarmentose or creeping,
nevertwining ......................... 4.
4 Inflorescences sessile, unbranched; corolla throat
with long trichomes (except S. becquetii); leaves
usually anisophyllous . . . . . subgenus Anisophyllae
Inflorescences pedunculate (sessile in flowering
stage in S. nobilis), usually branched; corolla
throat with short trichomes; leaves usually not
anisophyllous subgenus Sabicea.
DESCRIPTION OF THE SUBGENERA
1. SABICEA AUBL. SUBGENUS SABICEA
=Paiva Vell., Fl. Flum.: 104. 7 Sep–28 Nov 1829
(‘1825’) –Type: P. verticillata Vell. (=Sabicea villosa
Willd. ex Schult.).
=Pseudosabicea N.Hall
e, Adansonia ser 2, 3: 170.
1963 –Type: P. mitisphaera N. Hall
e(=Sabicea
nobilis R. Good).
=Pseudosabicea sect. Floribundae N.Hall
e, syn.
nov. (1963: 172) –Type: Pseudosabicea floribunda
(K.Schum.) N.Hall
e(Sabicea floribunda K.Schum.).
=Pseudosabicea sect. Sphaericae N.Hall
e (1963:
170) –Type: Pseudosabicea mitisphaera N.Hall
e
(=Sabicea nobilis R.D.Good), nom. illeg. acc. to
ICBN, Art. 22.2.
=Schizostigma Arn. ex Meisn., Pl. Vasc. Gen. 1:
164; 2: 115. 1838 –Type: S. hirsutum Arn. ex Meisn.
(Sabicea ceylanica Puff).
Lianas, usually twining, sometimes creeping or sar-
mentose. Stipules interpetiolar, persistent, erect to
©2016 The Linnean Society of London, Botanical Journal of the Linnean Society, 2016, 182, 551–580
570 L. ZEMAGHO ET AL.
reflexed, usually entire (but multifid in Madagascan
species). Nodes with two opposite leaves, leaf blades
usually equal in size (sometimes strongly unequal),
symmetrical to markedly asymmetrical at base, discol-
orous or not. Inflorescences axillary, one or two per
node, branched or not, sessile or pedunculate, one- to
many-flowered. Bracts surrounding the inflorescence
or not, usually free, sometimes shortly connate at
base (rarely connate for most of the length forming an
involucre splitting laterally at fruiting stage in S.
urceolata). Corolla with tube 5–25 (30) mm; corolla
throat inside with short multicellular trichomes (long
in Madagascan species). Ovary bi- to pentalocular.
2. SABICEA AUBL. SUBGENUS ANISOPHYLLAE
(N.HALLE) ZEMAGHO, SONK
E, DESSEIN &
LIEDE COMB. & STAT. NOV.
Pseudosabicea sect. Anisophyllae N.Hall
e, Adanso-
nia, s
er. 2, 3: 170 (1963). –Lectotype (designated
here): Sabicea mildbraedii Wernham
Sarmentose shrubs or creeping herbs. Stipules
interpetiolar, persistent, erect to reflexed, usually
shortly connate to the base of the petioles, entire to
multifid. Nodes usually with two strongly unequal
leaves, or sometimes with a single leaf opposed to a
stipule; rarely with two subequal leaves. Leaf blades
usually asymmetric at base, strongly discolorous, the
lower side covered with a dense felt of white to buff
woolly trichomes. Inflorescences axillary, sessile,
unbranched, usually many-flowered, one or two per
node, on leafy or leafless stems. Bracts and bracteoles
free and usually small, not enclosing the inflorescence,
sometimes enclosing the flowers when young. Corolla
with tube 3–11 mm long, throat usually densely pub-
escent with long multicellular trichomes sometimes
extending up to the base of the corolla lobes (tri-
chomes shorter in S. becquetii). Ovary bilocular.
Notes: Hall
e (1963) did not choose a type species
for Pseudosabicea section Anisophyllae among the
three species he originally placed in that section (S.
batesii,S. medusula,S. mildbraedii). The last spe-
cies is here chosen as lectotype since it is the most
widespread and its characteristics are representative
of the group.
3. SABICEA AUBL. SUBGENUS STIPULARIA
(P.BEAUV.) ZEMAGHO, SONKE, DESSEIN &
LIEDE COMB. & STAT. NOV.
=Stipularia P.Beauv., Fl. Owar. 2: 26. 1807 –
Type: S. africana P.Beauv. [Sabicea africana
(P.Beauv.) Hepper].
Erect shrubs, usually branched at base. Stipules
interpetiolar, persistent, erect, entire. Nodes with
two opposite leaves, leaf blades equal in size,
symmetrical at base, strongly discolorous (the lower
side covered with a dense felt of white to buff woolly
trichomes). Inflorescences axillary, sessile or subses-
sile, one or two per node, unbranched and many-
flowered. Involucre deeply urceolate and surrounding
the inflorescence, reddish, formed by the fusion of
two to four bracts, remaining intact or splitting in
the fruiting stage. Corolla tube 25–30 mm long; cor-
olla throat with short multicellular trichomes. Ovary
tri- to pentalocular.
4. SABICEA AUBL. SUBGENUS STIPULARIOP-
SIS WERNHAM (1914: 27).
Lectotype (designated here): Sabicea hierniana
Wernham (1914: 29).
Ecpoma K.Schum., Bot Jahrb. Syst. 23; 430 (1896),
syn. nov. –Type: E. apocynaceum K.Schum. (1896:
430).
Erect shrubs or herbs, usually single-stemmed or
more rarely ramose. Stipules interpetiolar, persistent,
erect, entire. Nodes with two opposite leaves, leaf
blades equal in size, symmetrical at base, green on
both sides. Inflorescence axillary, two per node, ses-
sile, unbranched, usually many-flowered. Bracts usu-
ally small, rarely (S. gigantostipula) two large free
bracts surrounding the inflorescence and resembling
stipules. Corolla tube 15–20 mm long; corolla throat
with short multicellular trichomes. Ovary bilocular.
Notes: Wernham (1914) did not choose a type spe-
cies for subgenus Stipulariopsis.Sabicea hierniana
is chosen here as lectotype, since it is the most wide-
spread and its characteristics are representative of
the group. The selection of the type of Ecpoma,E.
apocynaceum K.Schum., as lectotype is not possible,
because this species is mentioned anywhere by
Wernham (1914), even though it was published
before the publication of his treatment.
CONCLUSION
Based on nrITS and three plastid markers (petD,
rps16 and trnT-F), tribe Sabiceeae are revisited and
found to comprise four genera: Hekistocarpa,Sabi-
cea,Tamridaea and Virectaria. Our results show
that Hekistocarpa–Tamridaea–Virectaria are sister
to Sabicea s.l. so that re-establishment of Virectariae
for the dry-fruited genera in contrast to Sabiceeae
comprising taxa with fleshy fruits could be consid-
ered. However, support for Hekistocarpa–Tamri-
daea–Virectaria is only moderate and mainly based
on the rps16 sequences, so that further study is
needed to clarify the position of these genera. In
Sabicea s.l., a subgeneric concept is considered the
best option to reflect the polymorphism of the group,
©2016 The Linnean Society of London, Botanical Journal of the Linnean Society, 2016, 182, 551–580
PHYLOGENETICS OF SABICEEAE (RUBIACEAE) 571
resulting in four subgenera: subgenus Stipularia
(corresponding to the former genus Stipularia), sub-
genus Stipulariopsis (including all former Ecpoma
spp., S. urbaniana and S. xanthotricha), subgenus
Anisophyllae (previously Pseudosabicea section Anis-
ophyllae) and the large Sabicea subgenus Sabicea
(including all the remaining species).The larger spe-
cies number of S. subgenus Sabicea compared with
the other subgenera, and, in particular, to the other
genera of Sabiceeae might be attributable to the
combination of fleshy fruits and lianescent habit in
Sabicea subgenus Sabicea. Fleshy fruits are advanta-
geous in promoting long-distance dispersal and a
lianescent habit possibly opens low-light habitats not
available to other growth forms.
ACKNOWLEDGEMENTS
This study is a part of the dissertation of LZ who
holds a PhD research grant from the Deutscher
Akademischer Austausch Dienst (DAAD) for the
revision of Sabicea s.l. from continental Africa. We
express our gratitude to the Meise Botanic Garden
(Belgium) and the Departement of Plant Systematics
of the University of Bayreuth for financial and logis-
tic support offered to LZ. We wish to thank the
Moabi foundation (Wageningen, the Netherlands),
Nature+(Gembloux, Belgium) to enable the first and
the third author to visit Europe. We thank the cura-
tors of the following herbaria for their help while
working in their institutions and/or the loan of silica
gel and herbarium material: BM, BR, BRLU, GH,
MO, NY, P and WAG. Wim Baert and Angelika
T€
auber are gratefully acknowledged for the labora-
tory support. We warmly thank Petra De Block,
Ulrich Meve and Tariq St
evart for their useful com-
ments, help and suggestions. We also thank two
anonymous reviewers for their helpful and construc-
tive comments on a previous version of the paper.
The authors gratefully acknowledge the financial
support by International Association for Plant Tax-
onomy (IAPT).
REFERENCES
Alejandro GD, Razafimandimbinson SG, Liede-
Schumann S. 2005. Polyphyly of Mussaenda inferred from
ITS and trnT-F data and its implication for generic limits
in Mussaendeae (Rubiaceae). American Journal of Botany
92: 544–557.
Andersson L. 1996. Circumscription of the tribe Isertieae
(Rubiaceae). Opera Botanica Belgica 7: 139–164.
Arnott GAW. 1839. Descriptions of some new or rare Indian
plants. Annals of Natural History 3: 20–23.
Aublet JBCF. 1775. Sabicea, Pentandria, Monogynia. In
Histoire des plantes de la Guiane Franc
ßoise, Vol. 1. London
& Paris: P.-F. Didot jeune, 192–196.
Baillon H. 1880. Histoire des plantes, Vol. 7. Paris:
Hachette.
Borges RM, Gowda V, Zacharias M. 2003. Butterfly polli-
nation and high-contrast visual signals in a low-density dis-
tylous plant. Oecologia 136: 571–573.
Bremekamp CEB. 1966. Remarks on the position, the
delimitation and the subdivision of the Rubiaceae. Acta
Botanica Neerlandica 15: 1–33.
Bremer B, Eriksson T. 2009. Timetree of Rubiaceae: phy-
logeny dating the family, subfamily and tribes. Interna-
tional Journal of Plant Science 170: 766–793.
Bremer B, Thulin M. 1998. Collapse of Isertieae, re-estab-
lishment of Mussaendeae and a new genus of Sabiceeae
(Rubiaceae): phylogenetic relationships based on rbcL data.
Plant Systematics and Evolution 211: 71–92.
Bridson D, Verdcourt B. 1988. Rubiaceae (part2). In: Pol-
hill RM, ed. Flora of tropical East Africa. Rotterdam: Balk-
ema, 415–747.
Darriba D, Taboada GL, Doallo R, Posada D. 2012. jMo-
delTest 2: more models, new heuristics and parallel com-
puting. Nature Methods 9: 772.
Dengler NG. 1999. Anisophylly and dorsiventral shoot sym-
metry. International Journal of Plant Sciences 160(Suppl
6):S67–S80.
Dessein S, Andersson L, Robbrecht E, Smets E. 2001a.
Hekistocarpa (Rubiaceae): a member of an emended tribe
Virectarieae. Plant Systematics and Evolution 229:
59–78.
Dessein S, Janssens S, Huysmans S, Robbrecht E,
Smets E. 2001b. A morphological and anatomical survey of
Virectaria (African Rubiaceae), with a discussion of its taxo-
nomic position. Botanical Journal of the Linnean Society
137: 1–29.
Doyle JJ, Doyle JL. 1987. A rapid DNA isolation procedure
for small quantities of fresh leaf tissue. Phytochemical Bul-
letin, Botanical Society of America 19: 11–15.
Drummond AJ, Rambaut A. 2007. ‘BEAST’: Bayesian evo-
lutionary analysis by sampling trees. BMC Evolutionary
Biology 7: 1–214.
Gentry AH. 1993. Tropical forest biodiversity and the poten-
tial for new medicinal plants. In: Kinghorn AD, Balandrin
MF, eds. Human medicinal agents from plants. Washing-
ton: American Chemical Society, 13–24.
Guindon S, Gascuel O. 2003. A simple, fast and accurate
method to estimate large phylogenies by maximum-likeli-
hood. Systematic Biology 52: 696–704.
Hall
e F. 1961. Contribution
al’
etude biologique et tax-
onomique des Mussaendeae (Rubiaceae) d’Afrique tropicale.
Adansonia 1: 266–298.
Hall
e N. 1963. D
elimitation des genres Sabicea Aubl. et
Ecpoma K.Schum. en regard d’un genre nouveau: Pseudos-
abicea (Mussaendeae-Rubiaceae). Adansonia ser. 2 3: 168–
177.
Hall
e N. 1966. Famille des Rubiac
ees (1re partie). In: Aubr
e-
ville A, ed. Flore du Gabon, Vol. 12. Paris: Mus
eum
©2016 The Linnean Society of London, Botanical Journal of the Linnean Society, 2016, 182, 551–580
572 L. ZEMAGHO ET AL.
National d’Histoire Naturelle, Laboratoire de
Phan
erogamie, 1–277.
Hall
e N. 1970. Famille des Rubiaceae (2e partie). In: Aubr
e-
ville A, ed. Flore du Gabon, Vol. 17. Paris: Mus
eum
National d’Histoire Naturelle, Laboratoire de
Phan
erogamie, 1–335.
Hall
e N. 1971. Rubiaceae gabonaises nouvelles du genre
Pseudosabicea.Adansonia ser. 2 11: 313–317.
Hepper FN. 1958. Sabicea Aubl. and Stipularia Beauv.
(Rubiaceae-Mussaendeae) in tropical Africa. Kew Bulletin
13: 289–294.
Hepper FN. 1963. Sabicea. In: Hutchinson J, Dalziel JM, eds.
Flora of West tropical Africa, Vol. 2, 2nd edn. London: Crown
Agents for Overseas Governments, 104–224.
Hiern WP. 1877. Ordo LXX. Rubiaceae. In: Oliver D, Dyer
WTT, Prain D, Hill AW, eds. Flora of tropical Africa, Vol.
3. London: Reeve & Co., 33–82.
Hooker JD. 1873. Ordo LIV, Rubiaceae. In: Bentham G,
Hooker JD, eds. Genera plantarum, Vol. 2. London: L.
Reeve & Co., 7–151.
Huelsenbeck JP, Ronquist F. 2001. MrBayes: Bayesian
inference of phylogeny. Bioinformatics 17: 754–755.
Kainulainen K, Razafimandimbison SG, Bremer B.
2013. Phylogenetic relationships and new tribal delimita-
tions in subfamily Ixoroideae (Rubiaceae). Botanical Jour-
nal of the Linnean Society 173: 387–406.
Khan SA. 2007. New delimitations and phylogenetic rela-
tionships of Sabiceeae (Ixoroideae, Rubiaceae) and revision
of the Neotropical species of Sabicea Aubl. Unpublished
D.Phil. Thesis, University of Bayreuth.
Khan SA, Razafimandimbison SG, Bremer B, Liede-
Schumann S. 2008a. Sabiceeae and Virectarieae (Rubi-
aceae, Ixoroideae): one or two tribes? New tribal and gen-
eric circumscriptions of Sabiceeae and biogeography of
Sabicea s.l.Taxon 57: 1–17.
Khan SA, Razafimandimbison SG, Bremer B, Liede-
Schumann S. 2008b. Phylogeny and biogeography of the
African genus Virectaria Bremek. (Sabiceeae s.l., Ixoroi-
deae, Rubiaceae). Plant Systematics and Evolution 275: 43–
58.
Kirkbride MCG. 1979. Review of the neotropical Isertieae
(Rubiaceae). Brittonia 31: 313–332.
Kirkbride MCG. 1982. A preliminary phylogeny for neotrop-
ical Rubiaceae. Plant Systematics and Evolution 141: 115–
121.
L€
ohne C, Borsch T. 2004 [2005].Molecular evolution and
phylogenetic utility of the petD group II intron: a case
study in basal angiosperms. Molecular Biology and Evolu-
tion 22: 317–32.
Maddison DR, Maddison WP. 2001. MacClade 4: analysis
of phylogeny and character evolution. Version 4.03. Sunder-
land: Sinaur Associates.
Meisner CDF. 1838. Plantarum vascularium genera secun-
dum ordines naturals naturales digesta, Vol. 1. Leipzig:
Liberia Weidmannia.
Oxelman B, Lid
en M, Berglund D. 1997. Chloroplast
rps16 intron phylogeny of the tribe Sileneae (Caryophyl-
laceae). Plant Systematics and Evolution 206: 393–410.
Palisot-Beauvois AMFJ. 1810 [1807].Stipulaire. Stipular-
ia, Fam. des Rubiac
ees. In: Flore d’Oware et de B
enin.
Paris: Imprimerie de Fain et compagnie, 26, t.75.
Puff C, Igersheim A, Buchner R. 1998. Character states
and taxonomic position of the monotypic Sri Lankan Schi-
zostigma (Rubiaceae–Isertieae). In: Dransfield J, Coode
MJE, Simpson DA, eds. Plant diversity in Malesia III. Kew:
Royal Botanic Gardens, 187–203.
Razafimandimbison SG, Bremer B. 2002. Phylogeny and
classification of Naucleeae s.l. (Rubiaceae) inferred from
molecular (ITS, rbcl, and trnT-F) and morphological data.
American Journal of Botany 89: 1027–1041.
Razafimandimbison SG, Ekman S, McDowell TD, Bre-
mer B. 2012. Evolution of growth habit, inflorescence
architecture, flower size, and fruit type in Rubiaceae: its
ecological and evolutionary implications. PLoS ONE 7: 10.
Razafimandimbison SG, Lantz H, Mouly A, Bremer B.
2009. Evolutionary trends, major lineages, and new gen-
eric limits in the dioecious group of the tribe Vanguerieae
(Rubiaceae): insights into the evolution of functional
dioecy. Annals of the Missouri Botanical Garden 96:
161–181.
Robbrecht E. 1988. Tropical woody Rubiaceae. Opera
Botanica Belgica 1: 1–271.
Robbrecht E. 1993. Supplement to the 1988 outline of the
classification of the Rubiaceae, index to genera. In: Rob-
brecht E, ed. Advances in Rubiaceae macrosystematics.
Opera Botanica Belgica 6: 173–196.
Ronquist F, Huelsenbeck J. 2003. MrBayes 3: Bayesian
phylogenetic inference under mixed models. Bioinformatics
19: 1572–1574.
Rydin C, Kainulainen K, Razafimandimbinson SG,
Smedmark EEJ, Bremer B. 2009. Deep divergences in
the coffee family and the systematic position of Acranthera.
Plant Systematics and Evolution 278: 101–123.
Schnitzer SA, Bongers F. 2002. The ecology of lianas and
their role in forests. Trends in Ecology and Evolution 17:
223–229.
Schumann K. 1896 [1897].Beitr€
age zur Flora von Afrika,
XIII, Rubiaceae africanae. Botanische Jahrb€
ucher f €
ur Sys-
tematik, Pflanzengeschichte und Pflanzengeographie 23:
412–470.
Smedmark EEJ, Rydin C, Razafimandimbinson SG,
Khan SA, Liede-Schumann S, Bremer B. 2008. A phy-
logeny of Urophylleae (Rubiaceae) based on rps16 intron
data. Taxon 57: 24–32.
Stamatakis A, Ludwig T, Meier H. 2005. RAxML-III: a
fast program for maximum likelihood-based inference of
large phylogenetic trees. Bioinformatics 21: 456–463.
Stamatakis A, Ludwig T, Meier H. 2006. RAxML-VI-HPC:
maximum likelihood-based phylogenetic analyses with
thousands of taxa and mixed models. Bioinformatics 22:
2688–2690.
Standley PC. 1921. Rubiaceae (pars). North American Flora
32: 87–156.
Standley PC. 1936. Rubiaceae. In: Macbride JF, ed. Flora of
Peru, part 6. Publications of the Field Museum of Natural
History, Botanical Series 13: 3–261.
©2016 The Linnean Society of London, Botanical Journal of the Linnean Society, 2016, 182, 551–580
PHYLOGENETICS OF SABICEEAE (RUBIACEAE) 573
Swofford DL. 2002. PAUP*. Phylogenetic analysis using
parsimony (*and other methods). Version 4. Sunderland:
Sinauer Associates.
Thiers B. continuously updated. Index herbariorum: a
global directory of public herbaria and associated staff,
New York Botanical Garden’s Virtual Herbarium. Available
at: http://sweetgum.nybg.org/science/ih/
Verdcourt B. 1958. Remarks on the classification of the
Rubiaceae. Bulletin du Jardin Botanique de l’
Etat, Brux-
elles 28: 209–314.
Wernham HF. 1914. A monograph of the genus Sabicea.
London: British Museum (Natural History).
White F. 1979. The Guineo-Congolian Region and its rela-
tionships to other phytochoria. Bulletin du Jardin Botani-
que National de Belgique 49: 11–55.
APPENDIX 1: MORPHOLOGICAL
CHARACTERISTICS USED FOR
OPTIMIZATION
CHARACTERISTICS AND CHARACTER STATES
1. Growth habit: erect (0), creeping (1), twining
(2), sarmentose (3).
2. Anisophylly: leaves equal to subequal (0), leaves
strongly unequal (1).
3. Stipule size: < 15 mm high (0), more than
16 mm (1).
4. Stipule type: entire to bidentate (0), multifid
(1).
5. Bracts: free to shortly connate (0), fused and
urceolate, splitting in fruiting stage (1), fused
and urceolate, not splitting in fruiting stage
(2).
6. Inflorescence type: branched (0), unbranched
and sessile (1), unbranched and pedunculate
(2).
7. Calyx lobes length: shorter than 3 mm long (0),
between 4–9 mm long (1), longer than 13 mm
long (2).
8. Corolla-throat indumentum: short trichomes
(0), long trichomes (1).
9. Ovary: two-locular (0), (three to)five-locular (1).
10. Pedicels length in fruit: sessile (0), between
0.5–4 mm (1), more than 5 mm (2).
11. Fruit colour: white (0), purple to black (1), red
(2), yellow (3), fruits dry and brown (4).
©2016 The Linnean Society of London, Botanical Journal of the Linnean Society, 2016, 182, 551–580
574 L. ZEMAGHO ET AL.
Appendix 2: Morphological matrix for Sabiceeae and outgroups
Taxa\characters 1 2 3 4 5 6 7 8 9 10 11
1Mussaenda arcuata 0&300 000 0 00? 3
2Mussaenda erythrophylla 0&300 000 1&200? 3
3Pseudomussaenda flava 0 00 000 0&1000 4
4Hekistocarpa minutiflora 0 00 000 0 000 4
5Sabicea acuminata 3 10 101 1 110 0
6Sabicea africana 0 01 021 0 010 2
7Sabicea amazonensis 2 00 001&21 010 2
8Sabicea aspera 2 00 001&20 011 1
9Sabicea aspera var. glabrescens 2 00 001 0 011 1
10 Sabicea aurifodinae 3 1 0&1 0 0 1 0&1 1 0 0 ?
11 Sabicea batesii 3 10&1001 0 100 ?
12 Sabicea becquetii 3 00 001 1 000 2
13 Sabicea bigerrica 2 00 002 1 012 0
14 Sabicea brachycalyx 2 00 000 0 010 2
15 Sabicea brevipes 2 00 002 2 010 2
16 Sabicea bullata 2 00 001 1 010 2
17 Sabicea calycina 2 00 002 2 012 1
18 Sabicea caminata 2 00 002 0 010 0
19 Sabicea cana 2 00 001&20 011 1
20 Sabicea capitellata 2 00 002 0&1010 0
21 Sabicea carbunica 2 00 002 0 010 2
22 Sabicea ceylanica 1 00 001 1 012 0
23 Sabicea chocoana 2 00 000 0 011&21
24 Sabicea cinerea 0 00 001 1 010 2
25 Sabicea cuneata 2 00 000 1 011&2?
26 Sabicea dewevrei 2 00&1002 0 010 2
27 Sabicea discolor 2 00 000 0 012 0
28 Sabicea diversifolia 3 10 101 1 110 2
29 Sabicea duparquetiana 2 00 002 1&2010 2
30 Sabicea elliptica 0 0 0&1 0 1 1&2 0 0 1 0 2
31 Sabicea erecta 0 00 000 1 011 ?
32 Sabicea exellii 2 00 001 0 011 1
33 Sabicea ferruginea 2 01 001&22 012 2
34 Sabicea floribunda 3 00 000 0 001 0
35 Sabicea gabonica 2 00 002 2 011 2
36 Sabicea gigantostipula 0 01 001 2 001 1
37 Sabicea gilletii 2 00 001&22 011 2
38 Sabicea gracilis 2 00 002 1 012 0
39 Sabicea harleyae 2 00 002 2 012 2
40 Sabicea hierniana 0 01 001 1&2001 1
41 Sabicea humilis 0 00 002 1 011 ?
42 Sabicea ingrata var. ingrata1 2 00 002 1 010 0
43 Sabicea ingrata var. ingrata2 2 00 002 1 010 0
44 Sabicea johnstonii 2 00 002 1 012 0
45 Sabicea laxa 2 00 000 0 011 0
46 Sabicea mabouniensis 1 10&1000 1 001 ?
47 Sabicea mattogrossensis 2 00 002 2 012 1
48 Sabicea medusula 1 10 001 1 100 2
49 Sabicea mexicana 2 00 000 1 012 1
50 Sabicea mildbraedii 1 10 101 0 100 2
51 Sabicea mildbraedii var. letestui 1 10 101 0 100 ?
52 Sabicea mollis 2 00 000 0&1011 0
©2016 The Linnean Society of London, Botanical Journal of the Linnean Society, 2016, 182, 551–580
PHYLOGENETICS OF SABICEEAE (RUBIACEAE) 575
Appendix 2. Continued
Taxa\characters 1 2 3 4 5 6 7 8 9 10 11
53 Sabicea najatrix 2 00 002 1 010 2
54 Sabicea nobilis 3 01 001&20 001 0
55 Sabicea orientalis 2 00 000 1 011 0
56 Sabicea panamensis 2 00 000 1 012 2
57 Sabicea pedicellata 2 00 002 0 012 0
58 Sabicea pilosa 2 00 002 2 010 2
59 Sabicea proselyta 3 00 000 0 001 0
60 Sabicea pyramidalis 2 00 000 0 012 1
61 Sabicea rosea 2 00 001 1 011 2
62 Sabicea rufa 2 00 002 0 010 0
63 Sabicea segregata 3 00 000 1 001 1
64 Sabicea speciosa 2 00 002 2 011 2
65 Sabicea sthenula 1 10 101 1 100 ?
66 Sabicea tayloriae 2 00 001&21 010 1
67 Sabicea tchapensis 2 00 002 2 011 2
68 Sabicea urbaniana 0 01 001 2 001 ?
69 Sabicea urceolata 2 00 011 0 010 2
70 Sabicea urniformis 2 00 011&21 010 0
71 Sabicea venosa 2 00 000 1 011 0
72 Sabicea villos 2 00 000 1 011 1
73 Sabicea vogelii 2 00 002 2 012 2
74 Sabicea xanthotricha 0 01 001 2 001 ?
75 Sabicea sp.nov.-SD1865 1 10&1101 1 100 ?
76 Sabicea sp.nov-SD2149 3 10 001 1 100 ?
77 Tamridaea capsulifera 0 00 000 1 002 4
78 Virectaria angustifolia 0 00 000 0 001 4
79 Virectaria major 0 00 000 1 000 4
80 Virectaria multiflora 0 00 000 0&1000 4
81 Virectaria procumbens 0 00 000&10 001 4
? means unknown.
©2016 The Linnean Society of London, Botanical Journal of the Linnean Society, 2016, 182, 551–580
576 L. ZEMAGHO ET AL.
Appendix 3: Species, vouchers and GenBank accession numbers
Species Voucher Herbarium Country ITS petD rps16
trnT-F
region
Outgroup
Mussaenda arcuata Poir Bremer 3080 UPS Tanzania AJ846860 –FM204721 AJ847380
Mussaenda erythrophylla
Schum. & Thonn.
Alejandro 129 USTH Philippines AJ846858 –FJ884628 AJ847372
Pseudomussaenda flava Verdc. Bidgood 8030 BR Ethiopia AJ846877 JQ958026 FM204729 AJ847385
Hekistocarpa
H. minutiflora Hook.f. Sonke 5603 BR Cameroon LN878254 LN878092 LN878213 LN878285
Sabicea subgenus Anisophyllae
Sabicea aurifodinae (N.Hall
e)
Razafim., B.Bremer, Liede
& Saleh A.Khan
Wieringa 5026 WAG Gabon AM409046 LN878025 LN878150 AM409162
Sabicea batesii Wernham Van Valkenburg
2569
WAG Gabon AM409048 LN878026 LN878151 AM409139
Sabicea becquetii
(N.Hall
e) Razafim.,
B.Bremer, Liede
& Saleh A.Khan
Reekmans 11116 K Burundi AM409050 LN878027 LN878152 AM409138
Sabicea medusula K.Schum.
ex Wernham
Van Andel 3555 WAG Cameroon AM409047 LN878028 LN878153 AM409163
Sabicea mildbraedii
var. letestui N.Hall
e
Dessein 1909 BR Gabon LN878219 LN878029 LN878154 LN878257
Sabicea mildbraedii Wernham Wieringa 5032 BR Gabon AM409051 LN878030 LN878155 AM409137
Sabicea sp.nov1 Dessein 2149 BR Gabon LN878220 LN878031 LN878156 LN878258
Sabicea sp.nov2 Dessein 1865 BR Gabon LN878221 LN878032 LN878157 LN878259
Sabicea sthenula (N.Hall
e)
Razafim., B.Bremer, Liede
& Saleh A.Khan
Dessein 1829 BR Gabon LN878222 LN878033 LN878158 LN878260
Sabicea subgenus Sabicea
Sabicea acuminata Baker Luckow 4448 BR Madagascar LN878255 LN878034 LN878159 LN878261
Sabicea amazonensis Wernham Campbell P22037 MO Brazil AM409007 –– AM409157
Sabicea aspera Aubl. Andersson 2003 NY French Guiana AM409008 LN878035 –AM409143
Sabicea aspera var. glabrescens
(Benth.) K.Schum.
Gillespie 825 NY Guyana AM409024 –– AM409147
Sabicea bigerrica N.Hall
eDessein 2498 BR Gabon LN878223 LN878036 LN878160 –
Sabicea brachycalyx Steyerm. Beck 10056 NY Bolivia LN878240 LN878067 LN878188 –
Sabicea brevipes Wernham Jongkind 2793 WAG Ghana AM409178 LN878094 LN878216 AM409178
©2016 The Linnean Society of London, Botanical Journal of the Linnean Society, 2016, 182, 551–580
PHYLOGENETICS OF SABICEEAE (RUBIACEAE) 577
Appendix 3. Continued
Species Voucher Herbarium Country ITS petD rps16
trnT-F
region
Sabicea bullata Zemagho,
O.Lachenaud &Sonk
e
Dessein 2572 BR Cameroon LN878224 LN878037 LN878161 LN878262
Sabicea calycina Benth. Dessein 3147 BR Cameroon LN878225 LN878038 LN878162 LN878263
Sabicea caminata N.Hall
eDe Wilde 10311 WAG Gabon AM409010 LN878039 –AM409118
Sabicea capitellata Benth. Carvalho 3201 BR Cameroon LN878226 LN878040 LN878163 LN878264
Sabicea carbunica N.Hall
eWieringa 5151 WAG Gabon LN878227 LN878041 LN878164 –
Sabicea ceylanica Puff Iwarsson 576 UPS Sri Lanka AM409057 LN878093 LN878214 AM409168
Sabicea chocoana C.M.Taylor Delprete 6342 NY Colombia AM409013 LN878042 LN878165 AM409144
Sabicea cinerea Aubl. Andersson 1903 MO French Guinea AM409014 –– AM409120
Sabicea cuneata Rusby Solomon 17654 NY Bolivia LN878228 LN878043 LN878166 –
Sabicea dewevrei De Wild.
& T.Durand
Lemaire 1393 BR Republic of
the Congo
AM409016 LN878095 LN878215 AM409121
Sabicea discolor Stapf Jongkind 4880 WAG Ivory Coast AM409018 LN878044 –AM409145
Sabicea diversifolia Pers. De Block 901 BR Madagascar LN878229 LN878045 LN878167 LN878265
Sabicea duparquetiana Baill.
ex Wernham
Dessein 1725 BR Gabon LN878230 LN878046 LN878168 LN878266
Sabicea erecta Rusby Nee 43237 MO Bolivia LN878231 LN878047 LN878169 –
Sabicea exellii G.Taylor Joffroy 188 BRLU S~
ao Tom
e&
Pr
ıncipe
AM409020 LN878048 –AM409124
Sabicea ferruginea Benth. Jongkind 5683 WAG Liberia AM409021 LN878049 LN878170 AM409125
Sabicea gabonica Hutch. Van Andel 3417 WAG Cameroon AM409019 LN878050 LN878171 AM409123
Sabicea floribunda K.Schum. Wieringa 5440 WAG Gabon LN878232 LN878051 LN878172 LN878267
Sabicea gilletii De Wild. Lejoly 82/903 BR Democratic
Republic
of the Congo
AM409023 LN878052 LN878173 AM409154
Sabicea gracilis Wernham Dessein 1418 BR Cameroon LN878233 LN878053 LN878174 –
Sabicea harleyae Hepper Jongkind 4867 WAG Ivory Coast AM409025 LN878054 LN878175 AM409152
Sabicea humilis S.Moore Malme 2684 S Brazil AM409026 LN878055 –AM409148
Sabicea ingrata K.Schum.
var. ingrata (1)
Joffroy 150 BR S~
ao Tom
e&
Pr
ıncipe
LN878234 LN878056 LN878176 LN878268
Sabicea ingrata K.Schum.
var. ingrata (2)
Ogonovsky 10 BRLU S~
ao Tom
e&
Pr
ıncipe
AM409027 LN878057 LN878177 AM409149
Sabicea johnstonii K.Schum.
ex Wernham
Boyekoli Ebale Congo
2010 Expedition 847
BR Gabon LN878235 LN878058 LN878178 LN878269
Sabicea laxa Wernham Lachenaud 867 BR Cameroon LN878236 LN878059 LN878179 LN878270
©2016 The Linnean Society of London, Botanical Journal of the Linnean Society, 2016, 182, 551–580
578 L. ZEMAGHO ET AL.
Appendix 3. Continued
Species Voucher Herbarium Country ITS petD rps16
trnT-F
region
Sabicea longipetiolata De Wild. Boyekoli Ebale Congo
2010 Expedition 640
BR Gabon LN878237 LN878060 LN878180 LN878271
Sabicea mabouniensis
O.Lachenaud
&. Zemagho
Lachenaud 1446 BRLU Gabon LN878249 LN878082 LN878203 LN878286
Sabicea mattogrossensis
Wernham
Beck 9986 NY Bolivia AM409029 –LN878181 AM409127
Sabicea mexicana Wernham Hahn 639 NY Mexico AM409030 LN878061 LN878182 AM409153
Sabicea mollis K.Schum. ex
Wernham
Dessein 2258 BR Gabon LN878238 LN878062 LN878183 LN878272
Sabicea najatrix N.Hall
eWieringa 4653 WAG Gabon AM409031 LN878063 LN878184 AM409128
Sabicea nobilis (R.D.Good)
Razafim., B.Bremer, Liede
& Saleh A.Khan
Wieringa 7673 WAG Gabon LN878239 LN878064 LN878185 LN878273
Sabicea orientalis Wernham Mhoro 443 UPS Tanzania AM409032 LN878065 LN878186 AM409155
Sabicea panamensis Wernham Harling 26896 S Ecuador AM409033 LN878066 LN878187 AM409156
Sabicea pedicellata Wernham Sonk
e 5567 BRLU Cameroon LN878241 LN878068 LN878189 LN878274
Sabicea pilosa Hiern Dessein 2814 BR Gabon LN878242 LN878069 LN878190 LN878275
Sabicea proselyta (N.Hall
e)
Razafim., B.Bremer, Liede
& Saleh A.Khan
Van Valkenburg
2646
WAG Gabon AM409053 LN878070 LN878191 AM409166
Sabicea pyramidalis
L.Andersson
Burnham 1455 F Ecuador AM409034 LN878071 LN878192 AM409129
Sabicea rosea Hoyle Jongkind 10096 WAG Liberia LN878243 LN878072 LN878193 LN878276
Sabicea rufa Wernham Sonk
e 5082 BR Cameroon LN878244 LN878073 LN878194 –
Sabicea segregata Hiern Wieringa 5025 WAG Gabon AM409054 LN878074 LN878195 AM409164
Sabicea speciosa K.Schum. Van Meer 1623 WAG Nigeria AM409037 LN878075 LN878196 AM409131
Sabicea tayloriae Rusby ex
Liede, Meve & S.A.Khan
Nee 46014 MO Bolivia –LN878076 LN878197 LN878277
Sabicea tchapensis K.Krause Dessein 3017 BR Cameroon LN878245 LN878077 LN878198 LN878278
Sabicea urceolata Hepper Dessein 1803 BR Gabon LN878246 LN878078 LN878199 LN878279
Sabicea urniformis Zemagho,
O.Lachenaud & Sonk
e
Wieringa 5584 WAG Gabon LN878247 LN878079 LN878200 LN878280
Sabicea venosa Benth. Sonke 2797 WAG Central African
Republic
AM409041 LN878096 LN878218 AM409134
©2016 The Linnean Society of London, Botanical Journal of the Linnean Society, 2016, 182, 551–580
PHYLOGENETICS OF SABICEEAE (RUBIACEAE) 579
Appendix 3. Continued
Species Voucher Herbarium Country ITS petD rps16
trnT-F
region
Sabicea villosa Willd.
ex. Roem. & Schult.
Delprete 5102 NY Costa Rica AM409042 LN878080 LN878201 AM409160
Sabicea vogelii Benth. Jongkind 4859 WAG Ivory Coast AM409044 LN878081 LN878202 AM409136
Sabicea subgenus Stipularia
Sabicea africana P.Beauv. Dessein 1824 BR Gabon LN878250 LN878083 LN878204 LN878281
Sabicea elliptica Schweinf.
ex Hiern
Lisowski 56663 BR Congo AM409058 LN878084 LN878205 AM409169
Sabicea subgenus
Stipulariopsis
Sabicea gigantostipula
K.Schum.
Dessein 1650 BR Cameroon LN878251 LN878085 LN878206 LN878282
Sabicea hierniana
Wernham
Dessein 1955 BR Gabon LN878252 LN878086 LN878207 LN878283
Sabicea urbaniana
Wernham
Chatrou 488 BR Cameroon LN878253 LN878087 LN878208 LN878284
Sabicea xanthotricha
Wernham
Sonke 1082 BR Cameroon AM409045 LN878088 LN878209 AM409151
Tamridaea
Tamridaea capsulifera
(Balf.f.) Thulin &
B.Bremer
Miller 10087 UPS Yemen AM409170 LN878089 LN878210 AM409059
Virectaria
Virectaria angustifolia
(Hiern.) Bremek.
Lisowski 56663 BR Republic
of the Congo
LN878256 LN878097 LN878217 LN878287
Virectaria multiflora
(Sm.) Bremek.
Leeuwenberg 2295 UPS Ivory Coast AM409060 LN878090 LN878211 AM409171
Virectaria major
(K.Schum.) Verdc.
Lejoly 2934 BRLU Democratic Republic
of the Congo
AM981282 –– AM982741
Virectaria procumbens
(Sm.) Bremek.
Adams 453 UPS Liberia AM409061 LN878091 LN878212 AM409172
©2016 The Linnean Society of London, Botanical Journal of the Linnean Society, 2016, 182, 551–580
580 L. ZEMAGHO ET AL.
