Alejandro et al. Botanical Studies 2014, 55:65
http://www.as-botanicalstudies.com/content/55/1/65
RESEARCH
Open Access
Generic affiliations of Canthium species placed
under Pyrostria group B sensu Bridson
(Vanguerieae, Rubiaceae) inferred from
morphology and molecular data
Grecebio Jonathan D Alejandro1,2,3, Carizza Marie M Magdaleno1, Joseph Alvin T Pacia1, Lyn D Paraguison3,
Kim Karlo C Quiogue1, Annie Eliza D Wong1, Krysten Marie R Yayen1 and Axel H Arriola3,4*
Abstract
Background: Pyrostria sensu lato (s.l.) is regarded as one of the polyphyletic group within Vanguerieae formerly
comprising of Pyrostria sensu stricto (s.s.), Pyrostria group A and Pyrostria group B delineated by the number of
locules and geographical occurrence. Recent molecular phylogenetic studies within the genus have narrowed its
circumscription that resulted in the merging of Pyrostria group A and Pyrostria s.s. Although some species of
Pyrostria group B were already transferred to Pyrostria s.s. and Psydrax based on morphology, other representatives
of the group remain unsettled.
Results: Bayesian and parsimony analysis of the combined ITS (nrDNA) and trnL-F (cpDNA) datasets showed a
well-supported clade of the whole Vanguerieae containing four Philippine endemic representatives of Pyrostria
group B. The placement of Canthium oligophlebium, Canthium obovatifolium and Canthium ramosii within Pyrostria
s.s. (PP = 0.99; BS = 85%) is robustly supported likewise the affiliation of C. gynochthodes with Psydrax (PP = 0.94;
BS = 85%). Morphological features shared by our species with Pyrostria s.s. and Psydrax further supports our
molecular data.
Conclusion: Our study supports the earlier hypothesis that Canthium oligophlebium, C. obovatifolium and C. ramosii
should be placed under Pyrostria s.s. except for C. gynochthodes that grouped with Psydrax. Four new combinations
are proposed in this study. The generic affiliations of other species of Pyrostria group B should be reinvestigated
towards a more natural classfication in Vanguerieae.
Keywords: Bayesian analysis; ITS; Psydrax; Pyrostria; trnL-F; Vanguerieae
Background
The tribe Vanguerieae has long been regarded as monophyletic since it is easily delineated from the rest of
Rubiaceae by the presence of a unique type of pollen
presenter above the style (Verdcourt 1987; Bridson
1992). Recent molecular phylogenetic studies of the tribe
(Lantz et al. 2002; Lantz and Bremer 2004, 2005)
* Correspondence: arriolaaxel@yahoo.com
3
The Graduate School, University of Santo Tomas, España, Manila 1015,
Philippines
4
Department of Biological Sciences, College of Arts and Sciences, University
of the East, 2219 C.M. Recto Ave, Manila, Philippines
Full list of author information is available at the end of the article
support previous classifications of Bridson (1985, 1986,
1992) in reinstating Keetia E. Phillips and Psydrax Gaertn.
and raising Canthium subg. Afrocanthium Bridson to
a genus level. Meanwhile, Vangueria Juss. was recircumscribed and is distinguished from its close relatives by
having inflorescences borne from where leaves have fallen,
small bracteoles in secondary branch and large fruit with
2 to 5 locules. The genus Canthium was regarded as
highly polyphyletic and redelimited to include species with
supraaxillary spines.
Although major evolutionary lineages and new generic
limits have been established within Vanguerieae utilizing
© 2014 Alejandro et al.; licensee Springer. This is an Open Access article distributed under the terms of the Creative Commons
Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction
in any medium, provided the original work is properly credited.
Alejandro et al. Botanical Studies 2014, 55:65
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morphology and molecular data, there are still understudied species. When Bridson (1987) reinstated
the genus Pyrostria Comm. ex A. Juss., she lumped all
Pyrostria species and representatives of Canthium with
pair of persistent connate bracts (bracteate species)
under Pyrostria s.l. and suggested informal groups
(Pyrostria s.s., Pyrostria group A and Pyrostria group B)
based on the number of locules and geographical occurrence. Both Pyrostria s.s. (pluri-locular ovary) and
Pyrostria group A (bilocular ovary) have unisexual
flowers and well represented in Madagascar, the latter
extends from Africa to Arabia. Meanwhile, Pyrostria
group B shares characters with Pyrostria group A in
having unisexual flowers, bilocular ovaries, broad corolla tube, and 4–5 corolla lobes but the former radiates
as far as SE Asia. The polyphyly of Pyrostria s.l. was initially addressed by Lantz and Bremer (2004) in recovering a clade composed mainly of dioecious species but
failed to discussed further this group due to the poor internal support. Razafimandimbison et al. (2009) established a new generic delimitations using molecular data
within the dioecious Vanguerieae by lumping species
from Pyrostria group A to Pyrostria s.s. Their study,
however, failed to include any species under Pyrostria
group B and made an assumption that this group belongs to Pyrostria s.s. due to the presence of paired
bracts which is a typical character for this genus. In an
earlier study based on morphology, Utteridge and Davis
(2009) transferred two SE Asian Canthium species belonging to Pyrostria group B (C. brunnescens Craib and
C. cochinchinense Pierre ex Pit. in H.Lecomte) to Pyrostria s.s. Recently, Alejandro et al. (2013) transferred the
Philippine endemic C. subsessifolium (Merr.) Merr. to
Pyrostria. In contrast, Plectronia amplifolia Elmer informally placed under Pyrostria group B was transferred to
Psydrax (Ruhsam et al. 2008).
There are still left unresolved endemic Philippine species placed under Pyrostria group B probably associated
with Pyrostria such as C. brunneum (Merr.) Merr., C.
ellipticum (Merr.) Merr., C. gynochthodes Baill., C. megacarpum (Merr.) Merr., C. obovatifolium (Merr.) Merr., C.
oligophlebium (Merr.) Merr., C. ramosii (Merr.) Merr.,
and C. subcapitatum (Merr.) Merr. Available herbarium
materials of these species are scarce and lack reproductive parts for confirmation. In this study, four species of
Canthium informally placed under Pyrostria group B:
C. gynochthodes, C. obovatifolium, C. oligophlebium, and
C. ramosii were collected and challenged their phylogenetic positions within Vanguerieae utilizing molecular
sequence data. Furthermore, type specimens were meticulously examined to confirm our molecular results.
The present study is a good contribution in understanding a more robust phylogenetic evolutionary trends and
lineages within the tribe.
Page 2 of 8
Methods
Taxon sampling
This study is based on the examination of herbarium
sheets from various herbaria as well as field observation.
Canthium gynochthodes, C. obovatifolium, C. oligophlebium and C. ramosii were collected based on their type
protologues. Collected samples (herbarium specimens and
preserved reproductive structures in 70% ethanol) were
deposited at the USTH for accessioning. Leaf samples
were dried in silica-gel for DNA extraction (Chase and
Hills 1991).
Molecular methods
Genomic DNA was extracted from silica gel-dried leaf samples using the DNeasy Plant Minikit (Qiagen, Germany).
The entire ITS region (including the 5.8S gene) was amplified and sequenced using the primer pair P17F/26-82R
and P16F/P25R (Popp and Oxelman 2001). Meanwhile,
primer pair c/f were used for both amplification and sequencing of the trnL-F region (Taberlet et al. 1991). DNA
amplification was carried out following the work of
Alejandro et al. (2005, 2011). Amplified DNA was purified
using the QIA-quick Purification Kit (Qiagen, Germany).
Purified DNA was sent to MACROGEN Inc., Seoul, South
Korea for sequencing.
Phylogenetic analysis
The ITS and trnL-F sequences were assembled and edited using the Codon Code Aligner version 3.0.1. Novel
sequences of the four Philippine Canthium from each of
the markers used were incorporated with several related
sequences from the work of Lantz and Bremer (2004)
taken from the GenBank (Table 1). Ixora coccinea L. and
Mussaenda erythrophylla Schumach. & Thonn., considered as closely related to Vanguerieae were used as the
outgroups. Sequences were aligned manually using Se-Al
v.1.0al (Rambaut 1996).
Bayesian inference (BI) was used to estimate phylogenetic positions of the Philippine endemic Canthium species.
The analysis was carried out using the MrBayes v.3.1.2p
software (Huelsenbeck and Ronquist 2001; Ronquist and
Huelsenbeck 2003; Altekar et al. 2004). Model selection
for the best-performing evolutionary models were determined under three model selection criteria: a) Akaike
Information Criterion (AIC) (Akaike 1974), b) AICc
(seconder order criterion of AIC, necessary for smaller
samples) and c) the Bayesian Information Criterion (BIC)
(Schwartz 1978). The selected models were HKY and
GTR + G for the ITS and trnL-F, respectively. In analyzing
single marker, the best performing model was selected
and one million generation was considered with a sample
frequency of 1000 and four parallel chains. For combined
analyses, model selection as well as the settings is similar
with that of the single-marker analysis, however there
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Table 1 Nucleotide sequence database accession numbers of taxa used in this study
Taxon
GenBank/EMBL Accession Number
ITS
trnL-F
Afrocanthium burttii (Bullock) Lantz
AJ617749
AJ620120
Afrocanthium gilfillanii (N.E. Br.) Lantz
AJ617751
AJ620123
Afrocanthium keniense (Bullock) Lantz
AJ617753
AJ620126
Afrocanthium lactescens (Hiern) Lantz
AJ617754
AJ620127
Afrocanthium mundianum (Cham. & Schltdl.) Lantz
AJ315107
AJ620128
Afrocanthium parasiebenlistii (Bridson) Lantz
AJ617756
AJ620130
Afrocanthium pseudoverticillatum (S. Moore) Lantz
AJ617758
AJ620132
Afrocanthium siebenlistii (K. Krause) Lantz
AJ617759
AJ620133
Canthium ciliatum (D. Dietr.) Kuntze
AJ617750
AJ620121
Canthium coromandelicum (Brum. f.) Alston
AJ315081
AJ620122
Canthium glaucum Hiern ssp. glaucum
AJ617752
AJ620124
Canthium gynochthodes1
HG937666
HG937663
Canthium inerme (L.f.) Kuntze
AJ315120
AJ620125
Canthium mrimaense (Verdc.) Lantz
AJ617775
AJ620174
Canthium obovatifolium (Merr.) Merr.2
HG937664
HG937661
Canthium oligocarpum Hiern ssp. Captum (Bullock) Bridson
AJ617755
AJ620129
Canthium oligophlebium (Merr.) Merr.3
HG937665
HG937660
Canthium ramosii (Merr.) Merr.4
HG937667
HG937662
Fadogia ancylantha Schweinf.
AJ315103
AJ620136
Fadogia arenicola K.Schum. & K.Krause
AJ874981
AJ874943
Fadogia tetraquetra K. Schum. & K. Krause
AJ315099
AJ620139
Fadogia triphylla Baker
AJ874982
AJ874944
Keetia gueinzii (Sond.) Bridson
AJ315117
AJ620143
Keetia lukei Bridson
AJ617761
AJ620144
Keetia venosa (Oliv.) Bridson
AJ617762
AJ620145
Keetia zanzibarica (Klotzsch) Bridson ssp. zanzibarica
AJ315105
AJ620138
Psydrax kraussioides (Hiern) Bridson
AJ617786
AJ620157
Psydrax livida (Hiern) Bridson
AJ617769
AJ620158
Psydrax locuples (K. Schum.) Bridson
Aj617770
Aj620159
Psydrax parviflora (Afzel.) Bridson
Aj315110
AJ620162
Pyrostria ampijoroense (Arènes) Razafim., Lantz & B. Bremer
AJ617766
AJ719194
Pyrostria hystrix (Bremek.) Bridson
AJ315114
AJ620168
Pyrostria major (A. Rich. ex DC.) Cavaco
EU 584304
FN386344
Pyrostria orbicularis A. Rich. ex DC.
EU584285
FN386347
Pyrostria phyllantoidea (Baillon) Bridson
AJ315115
AJ620169
Pyrostria revoluta (Balf. f.) Razafim., Lantz & B. Bremer
AJ617776
AJ620176
Pyrostria sarodranensis Cavaco
EU584280
FN386366
Pyrostria serpentina Lantz, Klack. & Razafim.
EU584283
FN386350
Vangueria infausta Burchell
AJ617777
AJ620180
Vangueria proschii Briq.
AJ875009
AJ874975
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Table 1 Nucleotide sequence database accession numbers of taxa used in this study (Continued)
Vangueria parvifolia Sond.
AJ315092
AJ620181
Ixora coccinea L.
AJ224826
AJ620117
Mussaenda erythrophylla Schumach. & Thonn.
AJ224823
AJ620116
Since vouchers of most taxa included in the study were published only the voucher information of the Philippine Canthium included in the study are provided
as footnotes.
1
Philippines, Province of Palawan, Arriola and Alejandro 12442 (USTH).
2
Philippines, Province of Davao, Lemana and Alejandro BL10014 (USTH).
3
Philippines, Province of Isabela, Lemana and Alejandro BA10017 (USTH).
4
Philippines, Province of Quezon, Arriola and Alejandro S001 (USTH).
were a total of three million running generations. Clades
with posterior probability (PP) exceeding 0.95 were
regarded as strongly supported.
Parsimony analysis was conducted using PAUP version4.0b (Swofford 2000). Heuristic search was carried out
to determine the most parsimonious trees utilizing a treebisection reconnection (TBR) branch swapping using
10,000 random addition sequences, with MULTREES option on. Consistency index (Kluge and Farris 1969) and retention index (Farris 1989) were calculated to determine if
the data is far from being homoplasious. Bootstrapping
was determined using 10,000 replicates, MULTREES option off, TBR branch swapping, and five random addition
sequences. Clades receiving greater than 90% were considered strongly supported.
Results
Sequence characteristics
Table 2 shows the matrix characteristics of the separate
and combined ITS and trnL-F data sets. The aligned
matrix of the 43 taxa of the ITS region includes a total
of 691 positions, 190 base pairs (bp) of which are phylogenetically informative. The 43 sequences of trnL-F, have
a total of 1,002 positions, 43 bp of which are informative. The combined ITS/trnL-F of the 43 taxa with
1,693 characters generated a total of 233 bp informative
characters.
Phylogenetic analysis
The tree topologies of the separate ITS (PP = 1.00; BS =
100%) and trnL-F (PP = 0.89; BS = 90%) analyses (trees
not shown) revealed a monophyletic Vanguerieae. Both
trees resolved the phylogenetic positions of C. obovatifolium, C. oligophlebium and C. ramosii in Pyrostria clade
Table 2 Matrix characteristics of separate and combined
datasets
Number of taxa
ITS
trnL-F
Combined
43
43
43
Number of included characters
691
1,002
1,693
Number of informative characters
190
43
233
Consistency index
0.56
0.93
0.64
Retention index
0.74
0.93
0.76
with high support in ITS (PP = 0.96; BS = 89%) and trnLF (PP = 1.00; BS = 85%). However, both separate analyses
failed to resolve the placement of C. gynochthodes within
the tribe and polytomies were observed for members of
Canthium s.s and Psydrax.
Bayesian and parsimony analyses of the combined ITS/
trnL-F data sets (Figure 1) shows a robustly supported
Vanguerieae (PP = 1.00; BS = 99%) (Figure 1). The majority
rule consensus tree of the combined ITS/trnL-F data sets
(Figure 1) supports the monophyly of the included genera
and recovered tree topologies similar with Lantz and
Bremer (2004). For instance, the monophyly of Canthium
s.s. is supported (PP = 0.96; BS = 67%) and is closely related to the large flowered group (PP = 1.00; BS = 85%);
Keetia (PP = 1.00; BS = 64%) and Afrocanthium (PP = 1.00;
BS = 89%) as sister taxa is likewise supported (PP = 0.98;
BS = 72%); and the monophyly of Psydrax (PP = 0.94;
BS = 85%) and Pyrostria s.s. (PP = 0.99; BS = 85%) were
also sustained. The combined data analysis agrees with
single marker analyses on the close relatedness of C. obovatifolium, C. oligophlebium and C. ramosii with Pyrostria
(PP = 0.99; BS = 85%). Meanwhile, C. gynochthodes is
finally resolved within Psydrax (PP = 0.94; BS = 85%).
Discussion
Generic affiliations of species under Pyrostria group B
The results presented above clearly shows the polyphyly
of Canthium as earlier observed by Lantz et al. (2002),
Lantz and Bremer (2004, 2005) and Razafimandimbison
et al. (2009). The four Canthium (C. gynochthodes, C.
obovatifolium, C. oligophlebium and C. ramosii) should
be excluded from Canthium s.s. since these species are
spineless.
The phylogenetic position of C. obovatifolium, C. oligophlebium and C. ramosii within the Pyrostria clade was
already anticipated due to the presence of a persistent, basally paired, connate to acuminate bracts as observed in
our recent collections and available herbarium sheets. The
synapomorphic characters of Pyrostria such as dioecious
sexuality and fleshy corolla with trichomes in the throat
(Lantz and Bremer 2004) were also observed in our sampled Canthium species. The placement of these three
Canthium species in Pyrostria s.l. was already suggested
by Bridson (1987) but she was unsure of the placement in
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Page 5 of 8
Figure 1 Majority-rule consensus tree inferred from the Bayesian analysis of the combined ITS/trnL-F datasets of the 43 included taxa.
The results are congruent with the results of parsimony analysis except for the nodes marked with asterisks. Numbers above branches indicates
Bayesian posterior probabilities and those below branches are parsimony bootstrap values. Species under study are highlighted in grey.
the genus due to their geographical occurrence falling outside the known range, i.e. at that time Pyrostria was considered to be a predominately Afro-Madagascan genus.
Although Pyrostria is mostly represented in Africa,
Ruhsam et al. (2008) mentioned that the presence of SE
Asian Pyrostria could probably be a disjunct part of their
African relatives. There is a possibility that species under
this genus may have undergone long range dispersal
from Africa to Asia as in the case of Mussaenda L.
(Alejandro et al. 2005).
Meanwhile, the phylogenetic placement of C. gynochthodes within Psydrax does not support the earlier suggestion of Bridson (1985) in placing the species under
Pyrostria s.l.. Bridson may have assigned this SE Asian
species under Pyrostria group B due to the presence of
bracts which resembles that of Pyrostria although Baillon
(1879) did not mention the occurrence of this character.
However, we examined herbarium sheets of C. gynochthodes [Gaerlan, F.J.M 0542753 (L, PNH); Romero, E.M.
0542751 (L, PNH); Soejarto, D.D. 0219674 (L, PNH);
Arriola and Alejandro, 12442 (PNH, USTH); Arriola and
Alejandro, 11057 (PNH, USTH)] and revealed that bracts
exist in younger inflorescences but totally absent in older
ones. The presence of bracts on young inflorescences of
C. gynochthodes will not affect its close relatedness with
Psydrax. According to Bridson (1987) bracts may be
present in some representatives of Psydrax, however, it is
distinctive from the paired connate bracts of Pyrostria
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which are rare in Vanguerieae. The presence of bracts is
not a cardinal character to delimit Psydrax. For instance,
Bridson (1985) mentioned of the occurrence of bracts in
the Indian Psydrax umbellata (Whit.) Bridson and unnamed Malayan species. Furthermore, examination of C.
gynochthodes revealed that it posseses other diagnostic
features of Psydrax such as coriaceous leaf blades, keeled
stipules with truncate to triangular stipular base and falcate stipular apex, reflexed anthers, long style always exceeding the corolla tube, longer than wide stigmatic knob,
cartilaginous seed and a very shallow to nearly inconspicuous apical crest without a lid-like area in the pyrene
(Bridson 1985; Cheek and Sonke 2004). Additionally, the
occurrence of a unique insertion of 40 bp in the trnL-F
region of Psydrax that is non-alignable with other species
of Vanguerieae (Lantz and Bremer 2004) exists in
C. gynochthodes.
The close relatedness of species placed under Pyrostria
group B with Pyrostria s.s. and Psydrax are supported by
morphology and molecular data. Therefore, it is necessary
to recollect the remaining species of Pyrostria group B to
determine their correct generic affiliations within the tribe.
Taxonomic treatment
We present here novel combinations of four species that
were included in our study (Figure 2).
Pyrostria obovatifolia (Merr.) Wong, Magdaleno &
Alejandro, comb. nov. Basionym: Canthium obovatifolium
(Merr.) Merr., Philipp. J. Sci. 35 (1928) 8. Plectronia obovatifolia Merr., Philipp. J. Sci. C12 (1917) 167. Philippines,
Luzon, Tayabas Prov. Mount Dalindingan, Sept. 1916,
Ramos and Edano 26526 (holotype: PNH destroyed;
lectotype: designated here K!; isolectotypes: US, HUH)
(Figure 2A).
Shrub to small tree less than 3 m high; branches terete
to a more or less quadrangular and glabrous. Leaves obovate, 3.5–7.5 × 1.0–3.0 cm, glabrous on both sides; apex
rounded; base acute to acuminate; visible lateral nerves
3 to 4 on each side of the midrib; petiole 2.5–8.0 mm,
glabrous. Stipules triangular to broadly ovate, 5.0–6.0 ×
1.0 mm, glabrous on both sides. Female inflorescences
axillary on 3.0–5.5 mm long peduncles, 6-flowered;
Page 6 of 8
peduncular bracts present, 3.0–5.5 mm long, triangular to
broadly triangular, glabrous on both sides, enclosing the
young inflorescence; pedicels erect, 3.0–4.0 mm long at
flowering. Female flowers: calyx limb glabrous; tube
1.2- 2.5 mm long; lobes acuminate, 0.2 × 0.4 mm. Corolla
5-merous, white, glabrous outside; tube tubular, 0.8–
1.2 mm long, hairs present at the throat; lobes broadly
triangular, 2.0–2.5 × 1.0–1.2 mm, recurved. Stamens
exerted, attached to corolla tube; anthers narrowly ovate
to ovate, 0.3 mm long, exserted. Style including stigmatic
knob 3.0–3.9 mm long; stigmatic knob 1 mm long, with a
shallow cleft above, style not recessed into the stigmatic
head; disk glabrous. Ovary 2-locular. Male flower unknown. Fruits ovoid 8.5–10.5 mm, glabrous.
Distribution:—Luzon Island: Ilocos Norte, Quezon
Habitat:—In secondary forest; 200–350 m altitude.
Phenology:—Flowering from March to June; Fruiting
May to December
Taxonomic notes: This species approaches P. subsessifolia by its elliptic to ovate leaf shape but differs by its
less conspicuous lateral nerves, shorter bracts and nonkeeled fruits.
Pyrostria oligophlebia (Merr.) Pacia, Quiogue &
Alejandro, comb. nov. Basionym: Canthium oligophlebium
(Merr.) Merr., Philipp. J. Sci. 35 (1928) 8. Plectronia oligophlebia Merr., Philipp. J. Sci. 17 (1921) 442. Philippines,
Luzon, Rizal Prov. Mount Susong Dalaga, Aug. 1917,
Ramos and Edano 29342 (holotype, PNH destroyed;
lectotype: designated here US; isolectotype: HUH, K !)
(Figure 2B)
Shrub to small tree, less than 5.0 m high; branches
quadrangular to more or less terete and glabrous. Leaves
broadly lanceolate to oblong, 3.0–5.5 × 1.0–2.0 cm, glabrous on both sides; apex acute; base acute; visible lateral
nerves 2 to 4 on each side of the midrib; petiole 0.7–
1.0 mm, glabrous. Stipules broadly triangular to ovate,
1.0–2.0 × 1.0 mm, glabrous on both sides. Female inflorescences axillary on a glabrous peduncle less than
2 mm long, 7–12 flowered; peduncular bracts present,
2.5–3.0 mm long, triangular to broadly triangular, glabrous on both sides, enclosing the young inflorescence;
pedicels erect, 4.0–5.0 mm long at flowering, persistent.
Figure 2 Images of the four plant species included in the study. A. Pyrostria obovatifolia fruiting branch; B. Pyrostria oligophlebia infructiscences;
C. Pyrostria ramosii fruiting branch; D. Psydrax gynochthodes flowering branch.
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Female flowers: calyx limb glabrous 2–2.7 mm long;
lobes shortly toothed, 0.1 × 0.3 mm. Corolla 4-merous,
white, glabrous outside; tube tubular, 0.8–1.2 mm long,
hairs present at the throat; lobes broadly ovate, 2.0–
2.5 × 1.0–1.2 mm. Stamens attached to corolla tube adjacent to the throat; anthers ovate, 0.3 mm long, exserted.
Style including stigmatic knob 1.0–2.5 mm long; stigmatic
knob 1 mm long, with a shallow cleft above; disk glabrous.
Ovary 2-locular. Male flower unknown. Fruits ovoid 6.0–
6.5 mm, glabrous with distinct indentation when dry.
Distribution:—Luzon Island: Rizal;
Mindanao Island: Davao
Habitat:—In secondary forest; 500–900 m altitude.
Phenology:—Flowering from March to December;
Fruiting from Septmber to February
Taxonomic notes: The smaller and fewer nerved
leaves of P. oligophlebia approaches P. gynochthodes.
However, P. oligophlebia differs from the latter by having
persistent pair of bracts, many-flowered inflorescences
and a longer petioles, peduncles and pedicel.
Pyrostria ramosii (Merr.) Arriola, Paraguison &
Alejandro, comb. nov.
Basionym: Canthium ramosii (Merr.) Merr., Philipp. J.
Sci. 35 (1928) 9. Plectronia ramosii Merr., Philipp. J. Sci.
7 (1921) 443. Philippines, Luzon, Tayabas Prov., Mount
Umiray, June 1917, Ramos and Edano 28973 (holotype,
PNH destroyed; lectotype: designated here NY!; isolectotype: K!) (Figure 2C)
Tree less than 9 m high; branches terete, glabrous.
Leaves broadly lanceolate to oblong, 2.5–9.5 × 1.5–3.9 cm,
glabrous on both sides; apex triangular to acuminate; base
attenuate; visible lateral nerves 3 to 4 on each side of the
midrib; petiole 1.5–2.0 mm, glabrous. Stipules triangular,
3.5–4.0 × 1.5–2.0 mm, glabrous on both sides. Inflorescences 4 flowered umbels; pedicels erect, 6.5–7.0 mm,
persistent glabrous, peduncular bracts present, 0.5–
1.5 mm long, triangular, glabrous on both sides. Infructiscence glabrous; stalk glabrous. Fruits ovoid to didymous
6.5–8.5 mm, 2 –celled, glabrous.
Distribution:—Luzon Island: Quezon
Habitat:—In secondary forest; 200–350 m altitude.
Phenology:— Fruiting August to December
Taxonomic notes: Pyrostria oligophlebia and P. ramosii closely resemble each other due to their oblong shape
leaf with acuminate apex. However, the latter have longer peduncles and few (2–4) flowered umbellate inflorescences as compared to the numerous (7–12) flowered
inflorescence of the former.
Psydrax gynochthodes (Baill.) Arriola, Yayen &
Alejandro, comb. nov.
Basionym: Plectronia gynochthodes (Baill.) Merr., Enum.
Philipp. Fl. Pl. 3 (1923) 536. Canthium gynochthodes Baill.,
Adansonia 12 (1879) 199. Philippines, Luzon, Batangas
Prov., 1917, Cuming 1848 (holotype, K!) (Figure 2D)
Page 7 of 8
Tree, less than 7 m tall; branches flattened to subterete, glabrous. Stipules glabrous on both sides, triangular
to ovate, 2–4.5 mm, keel prominent on the abaxial side.
Petioles 1.0–5.0 × 0.3–0.5 mm, glabrous; leaf blades leathery, elliptic to elliptic-oblong or obovate, 3–8.5 × 1.5–
5 cm, base acute to broadly acute, apex obtuse, glabrous
throughout, glossy; lateral nerves 3–4 on each side of
the midrib, domatia present. Inflorescences axillary,
10–12(−several) flowered, glabrous; peduncle 0.8–2.5 mm,
glabrous; pedicel 4.0–10.0 mm, puberulent. Calyx tube
infundibuliform, 2–3 × 2.0–2.5 mm, glabrous; lobes triangular 1.5–2.0 mm. Corolla tube infundibuliform, 1.0–
1.5 mm, glabrous outside, with ring of white hairs inside;
lobes 4, triangular, glabrous outside and inside. Anthers 4,
reflexed. Style 2–6 mm, exceeding corolla tube, stigmaticknob longer than wide, 0.3 × 0.2 mm, bifid. Ovary bilocular, 1–ovule per locule. Fruits ovoid to didymous,
distinctly broader than long, 7.5–8.0 × 9–10 mm, green
when young, glabrous, calyx limb persistent; Seeds 2,
obliquely ovoid, ventrally flattened, 2–5.5 × 3 mm, apical
crest very shallow to nearly inconspicuous, pyrene
cartilaginous.
Distribution:—Philippines, Taiwan
Habitat:—In secondary forest; 500–900 m altitude.
Phenology:—Flowering from March to June; Fruiting
from June to December
Taxonomic notes: Psydrax gynochthodes is comparable with P. obovatifolia, P. oligophlebia and P. ramosii.
However it is delineated from the three species due to
the absence of persistent acuminate-connate bracts. For
the above reason, we do not agree with the observation
of Bridson (1987) that P. gyncochthodes is closely associated with P. villarii Vidal and that the two species
should be synonymize. Moreover, P. gynochthodes can
be distinguised from the Philippine P. amplifolia by its
smaller, thicker and darker glossy green leaves, umbellate to cymose inflorescences, longer peduncles and
smaller fruits.
Conclusion
The generic affiliations of four species previously hypothesized under Pyrostria group B have been resolved based
on morphology and molecular sequence data. We formally proposed three novel combinations in Pyrostria and
a Psydrax. Other species of Pyrostria group B should be
reinvestigated towards a more natural classfication in
Vanguerieae. Furthermore, large number of Pyrostria and
bracteate species temporarily placed under Canthium s.l
should be sampled to fully understand the evolutionary
dispersal of the genus.
Competing interests
The authors declare that they have no competing interests.
Alejandro et al. Botanical Studies 2014, 55:65
http://www.as-botanicalstudies.com/content/55/1/65
Authors’ contributions
GJDA and AHA drafted the manuscript; All authors participated in sample
collection, molecular work and taxonomic treatment. All authors read and
approved the final manuscript.
Acknowledgements
We thank the following herbaria and staff for providing loans and/or access
to collections: K, L, NY, P, PNH, US and USTH. The authors thank the Research
Center for Natural & Applied Sciences (RCNAS), University of Santo Tomas
and the Commission on Higher Education, Philippine Higher Education
Research Network (CHED-PHERNEt) for the funding.
Author details
1
College of Science, University of Santo Tomas, España, Manila 1015,
Philippines. 2Research Center for the Natural & Applied Sciences, University
of Santo Tomas, España, Manila 1015, Philippines. 3The Graduate School,
University of Santo Tomas, España, Manila 1015, Philippines. 4Department of
Biological Sciences, College of Arts and Sciences, University of the East, 2219
C.M. Recto Ave, Manila, Philippines.
Received: 21 November 2013 Accepted: 5 August 2014
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doi:10.1186/s40529-014-0065-3
Cite this article as: Alejandro et al.: Generic affiliations of Canthium
species placed under Pyrostria group B sensu Bridson (Vanguerieae,
Rubiaceae) inferred from morphology and molecular data. Botanical
Studies 2014 55:65.
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