Tragopogon lainzii, a New Species of Tragopogon (Asteraceae) Segregated from T.
dubius: Evidence from Morphological and Molecular Data
Author(s): Víctor N. Suárez-Santiago, Consuelo Díaz de la Guardia, Douglas E. Soltis, Pamela S. Soltis,
and Gabriel Blanca
Source: Systematic Botany, 36(2):470-480. 2011.
Published By: The American Society of Plant Taxonomists
URL: http://www.bioone.org/doi/full/10.1600/036364411X569651
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Systematic Botany (2011), 36(2): pp. 470–480
© Copyright 2011 by the American Society of Plant Taxonomists
DOI 10.1600/036364411X569651
Tragopogon lainzii, a New Species of Tragopogon (Asteraceae) Segregated from
T. dubius: Evidence from Morphological and Molecular Data
Víctor N. Suárez-Santiago,1,4 Consuelo Díaz de la Guardia,1
Douglas E. Soltis,2 Pamela S. Soltis,3 and Gabriel Blanca1
1
Departamento de Botánica, Facultad de Ciencias, Universidad de Granada, c/ Severo Ochoa, s/n, 18071 Granada, Spain.
2
Department of Biology, University of Florida, Gainesville, Florida 32611, U. S. A.
3
Florida Museum of Natural History, University of Florida, Gainesville, Florida 32611, U. S. A.
4
Author for correspondence (vsuarez@ugr.es)
Communicating Editor: Andrew Hipp
Abstract—Tragopogon dubius is one of the most widespread species of Tragopogon, extending across much of Eurasia. Traditionally, T. dubius
has been considered a morphologically homogeneous species that includes all Tragopogon collections with yellow flowers and swollen peduncles under capitula. Here we describe a new species of Tragopogon from the Iberian Peninsula, T. lainzii, which has heretofore been included in
T. dubius. To this end, we performed comparative morphological, cytogenetic, and molecular analyses on many populations of both species. Our results show that T. dubius is not a homogeneous species and that different lineages exist across its broad geographic distribution.
Moreover, we show that hybridization has occurred in the wild between sympatric populations of T. dubius, T. lainzii, and T. porrifolius.
Keywords—Cytogenetics, ETS, hybridization, ITS, morphology, rpl16.
species. Jacquin (1773), using material from Austria, proposed
T. major Jacq. to include those plants like T. dubius but with
wide leaves, large heads, and 10–12(–18) involucral bracts;
Chaubard (1837) and Lindemann (1881) described new varieties for T. major (var. decipiens Chaub. ex Noulet; var. desertorum
Lindem.). However, most authors later considered T. major a
synonym of T. dubius, because the diagnostic characters for
T. major are not sufficiently distinct to warrant its recognition.
Thus, Vollmann (1914) considered T. major as a subspecies of
T. dubius [subsp. major (Jacq.) Vollmann], while Bolòs and Vigo
(1989) treated it as a variety of T. dubius [var. major (Jacq.) Bolòs
& Vigo; var. decipiens (Chaub. ex Noulet) Bolòs & Vigo]. Tzvelev
(1985) combined the variety described by Lindemann for
T. major as a subspecies of T. dubius [subsp. desertorum (Lindem.)
Tzvelev]. Richardson (1976), in his review for Flora Europaea,
did not recognize any infraspecific taxonomic units within
T. dubius and considered T. major a synonym of T. dubius.
Recent evidence from molecular analyses seems to refute
the supposed homogeneity for T. dubius across its geographic
distribution. For example, what has been called T. dubius from
India seems to be genetically, as well as morphologically, distinct from the European T. dubius and appears to warrant recognition as a new species (Mavrodiev et al. 2008a).
Thus, specimens referred to as T. dubius from different geographic areas may represent distinct, previously unrecognized
species. Observations of collections in nature first suggested
to us that plants recognized as T. dubius from southern Spain
likely represented a separate entity, morphologically distinct
from other collections of T. dubius. In this study, we describe
a new species of Tragopogon from the Iberian Peninsula,
T. lainzii, which has traditionally been considered as T. dubius.
To this end, we performed comparative morphological, cytogenetic, and molecular analyses on multiple populations
of both species. Moreover, we show that hybridization has
occurred in the Iberian Peninsula between sympatric species
of Tragopogon, involving both T. dubius and T. lainzii, as well as
more distantly related species such as T. porrifolius.
Tragopogon L. includes about 150 species; the genus has a
Eurasian distribution with a center of diversification in the eastern Mediterranean basin. On the Iberian Peninsula, Tragopogon
is well represented with nine recognized species (Blanca and
Díaz de la Guardia 1996; Díaz de la Guardia and Blanca 2004).
Tragopogon comprises annual, biennial, and mostly perennial herbs, with leaves entire and parallel-veined, involucral bracts in one row, and achenes muricate or scabrous
and almost always with a long beak. The main morphological characters used to distinguish species involve fruit morphology, ligule color, ratio of ligule/involucral bract lengths,
number of involucral bracts, and the thickness of the peduncle below the heads. Tragopogon is a taxonomically complex
genus, and the morphological variation of the species has
resulted in different interpretations by various authors; as
a result, extensive taxonomic and nomenclatural confusion has occurred (reviewed in Mavrodiev et al. 2005, 2007).
Hybridization is a frequent process that also increases taxonomic difficulty in the genus (Ownbey 1950; Krahulec et al.
2005); intermediate forms are commonly found in the wild
where species occur sympatrically. However, hybridization
is also an important mechanism of speciation in Tragopogon,
especially when associated with polyploidy (Ownbey 1950;
Ownbey and McCollum 1953; Díaz de la Guardia and Blanca
1990; 2004; Soltis et al. 2004; Mavrodiev et al. 2008a, b).
Tragopogon dubius Scop. (section Majores Kuth.) is one of
the most widespread species of Tragopogon, extending across
much of Eurasia. The species is mainly characterized by its
pale yellow ligules, all shorter than the involucral bracts, with
commonly eight to 12 involucral bracts, and strongly inflated
peduncles below the heads. However, the recognition of this
species is usually restricted to those plants that are yellowflowered and have swollen peduncles. In this delineation, other
important characters (e.g. achene morphology and ratio of
ligule/involucral bract lengths) are underutilized. As a result,
the true extent of intraspecific morphological variability has
been underestimated, which may even mask the presence of
other yellow-flowered and swollen-peduncled species under
the name T. dubius. In fact, unlike T. porrifolius L. (another widespread Tragopogon species; see Mavrodiev et al. 2007), T. dubius
has often been considered a morphologically homogeneous
species, and few nomenclatural problems have involved this
Materials and Methods
Morphology—A comparative analysis of morphological characteristics was performed on 17 populations (eight of “typical” T. dubius and
470
2011]
SUÁREZ-SANTIAGO ET AL.: A NEW TRAGOPOGON SEGREGATED FROM T. DUBIUS
nine of the putative new species) across their distribution in the Iberian
Peninsula (Appendix 1, Fig. 1). Two of the 17 populations analyzed were
not included in the statistical analyses because they were determined to
be hybrid populations (see Results) and the morphology of these individuals was influenced by the species involved in the hybridizations.
Characters used in the morphological analysis (quantitative and qualitative) were those exhibiting the greatest variation between the two putative
species (18 quantitative and five qualitative characters; Table 1). Flower
characters were measured during anthesis of middle-inflorescence flowers, while achene characters were measured on mature achenes from the
peripheral flowers of the capitulum. The achenes were photographed
using a variable-pressure LEO 1430 VP scanning electronic microscope
(SEM) in conventional mode, after gold–palladium coating (Thornill
et al. 1965). The terminology used basically follows Font Quer (1979) and
Stearn (1980).
The statistical program used for analyses was SPSS version 15.0.1 (SPSS,
Inc., Chicago, Illinois). Because our data are not normally distributed, we
applied a Mann-Whitney U test for each of the quantitative variables studied to test for differences between T. dubius and the putative unrecognized
species. To check for intraspecific and interpopulation homogeneity of the
quantitative morphological characters a data matrix was produced from
the 18 characters studied and subjected to a clustering analysis (the qualitative characters were constant within each putative species and hence
not included in this analysis). To construct the data matrix, the population
average value for each continuous quantitative character was calculated.
A logarithmic transformation of these average values was performed [log
(x + 1)]. The clustering method used was UPGMA (Unweighted PairGroups Method using Arithmetic averages), using a dissimilarity matrix
generated with Euclidean distances. We used this hierarchical clustering
method because it permitted us to check for structure in the morphological diversity between populations.
Fig. 1.
codes.
471
Karyological Analysis—Chromosome numbers were counted at
metaphase in root-tip meristems taken from germinating seeds. Roots
were pretreated with 8-hydroxy-quinoline, fixed in ethyl alcohol–acetic
acid (3:1), hydrolysed in 1 N HCl, stained in acetic orcein solution, and
then flattened for light microscopy (Darlington and La Cour 1969).
Chromosomes were matched and paired based on centromere position and size, and the karyotypes were represented by their chromosomal
formula. For chromosome nomenclature we followed that proposed by
Levan et al. (1964), and for quantification of karyotype symmetry we followed the classification of Stebbins (1971).
Molecular Analysis—The internal transcribed spacer region (ITS-1,
5.8S, ITS-2; hereafter ITS) and the 3′portion of the external transcribed
spacer (Lee et al. 2002; hereafter ETS) of the nuclear ribosomal (nr) DNA,
as well as the first intron of the rpl16 chloroplast gene, were used as molecular markers. Intraspecific variability of T. dubius and the putative segregate was explored by sequencing individuals from different populations.
In addition to the populations studied from a morphological standpoint,
in the molecular analysis we included other populations of T. dubius and
the putative unrecognized species (Appendix 1). To facilitate placement
of the latter samples with their closest relatives, we included sequences
of representative species of the Majores s. l. clade (following the phylogeny of Tragopogon by Mavrodiev et al. 2005). These sequences were either
taken from GenBank or generated as part of this study from Iberian taxa.
Tragopogon olympicus Boiss. was selected as an outgroup based on its position as sister to the remainder of the Majores clade in the aforementioned
phylogeny. EMBL accession numbers for the new sequences are given in
Appendix 1, while the GenBank accession numbers for the remainder are
given on the phylogenetic trees.
Total genomic DNA was extracted, using the CTAB method (Doyle
and Doyle 1987), from fresh leaves collected in the wild. The entire ITS
region (ITS-1, 5.8S and ITS-2), the 3′portion of the ETS and the plastid
Map of the Iberian Peninsula showing the localities sampled of T. dubius (asterisk) and T. lainzii (triangle). See Appendix 1 for population
472
SYSTEMATIC BOTANY
[Volume 36
Table 1. Principal morphological characters distinguishing T. dubius and T. lainzii. Results of biometric analysis are shown: interval of extreme values,
mean (± 1 s. e.), N = number of samples analysed and p values obtained by Mann-Whitney U test (significant p values are marked with an asterisk).
Character
T. dubius
T. lainzii
Stem color
Leaf margin
Peduncle width at anthesis (mm)
Peduncle width in fruit (mm)
Bud shape
No. of involucral bracts
Involucral bract length at anthesis (mm)
Involucral bract length in fruit (mm)
Involucral bract width at anthesis (mm)
Involucral bract width in fruit (mm)
Ligule length (mm)
Ligule length/ involucral bract length
No. of achenes per head
Total achene length (mm)
Achene-body length (mm)
Achene-body width (mm)
Achene-body length/achene-body width
Achene-beak length (mm)
Achene-beak length/achene-body length
Achene ornamentation
Apex-beak shape
Pappus length (mm)
Achene length/pappus length
Greenish
Not-undulate
6–12 [8.80 ± 0.13 (N = 95)]
12–20 [14.85 ± 0.14 (N = 97)]
Triangular
7–14 [9.64 ± 0.15 (N = 155)]
29–45 [37.99 ± 0.24 (N = 210)]
55–85 [63.40 ± 0.37 (N = 180)]
4–7 [5.53 ± 0.04 (N = 210)]
7–11 [8.65 ± 0.06 (N = 180)]
16–33 [23.25 ± 0.27 (N = 210)]
0.45–0.82 [0.61 ± 0.01 (N = 210)]
49–134 [91.75 ± 0.02 (N = 28)]
23.5–33.5 [29.52 ± 0.13 (N = 210)]
11–14 [12.12 ± 0.04 (N = 210)]
1.5–2.3 [1.90 ± 0.01 (N = 210)]
5–8.13 [6.41 ± 0.04 (N = 210)]
10.5–21.5 [17.40 ± 0.13 (N = 210)]
0.78–1.87 [1.44 ± 0.01 (N = 210)]
Coarse-ornamented
Bulbous
18–27 [22.86 ± 0.14 (N = 210)]
1–1.67 [1.30 ± 0.01 (N = 210)]
Reddish
Undulate
7–13 [10.05 ± 0.13 (N = 110)]
12–20 [14.88 ± 0.15 (N = 102)]
Ovate-oblong
9–16 [13.00 ± 0.02 (N = 245)]
27–45 [34.32 ± 0.21 (N = 282)]
44–66 [54.79 ± 0.29 (N = 252)]
4–9 [5.7 ± 0.06 (N = 282)]
5–11 [7.92 ± 0.05 (N = 252)]
30–48 [37.29 ± 0.20 (N = 282)]
0.93–1.43 [1.09 ± 0.00 (N = 282)]
115–263 [186.30 ± 0.02 (N = 23)]
25–38.5 [31.50 ± 0.19 (N = 252)]
7.5–11.5 [9.96 ± 0.05 (N = 252)]
1–1.7 [1.31 ± 0.01 (N = 252)]
6–11 [7.66 ± 0.05 (N = 252)]
15.5–27.5 [21.53 ± 0.17 (N = 252)]
1.45–2.89 [2.17 ± 0.02 (N = 252)]
Fine-ornamented
Obpyramidal
20–30 [23.98 ± 0.12 (N = 252)]
1.06–1.65 [1.31 ± 0.01 (N = 252)]
rpl16 intron were amplified by PCR, using primers N-nc18S10 and C26A
(Wen and Zimmer 1996) for the ITS region, L-ETS (Lee et al. 2002) and 18SETS (Baldwin and Markos 1998) for ETS, and rpL16F71 (Jordan et al. 1996)
and rpL16R1516 (Small et al. 1998) for the rpl16 intron. Amplification reactions were performed in a volume of 50 µl, under standard conditions
(Innis et al. 1990) for ITS and ETS, and following Small et al. (1998) for the
rpl16 intron. Automated sequencing of the purified PCR products was performed in both directions using the amplification primers on a 3100-Avant
Genetic Analyzer (Applied Biosystems, Inc., Foster City, California).
Nucleotide sequences were edited and aligned with the SEQMAN II v.
3.61 and MEGALIN v. 3.18 programs, respectively, from the DNAstar software package (DNASTAR, Inc., Madison, Wisconsin) and then adjusted
by eye. To estimate the extent of sequence identity (intra and interspecific),
we calculated the divergence (uncorrected p-distance) between sequences
using MEGA v. 4 (Tamura et al. 2007).
Phylogenetic analyses were performed using two optimality criteria:
maximum parsimony (MP) as implemented in PAUP* v. 4.0b10 (Swofford
2003) and Bayesian inference using MrBayes v. 3.1.2 (Ronquist and
Huelsenbeck 2003). Separate analyses were conducted on the ITS, ETS
and rpl16 data sets, with an additional analysis on the combined ITS +
ETS matrix. The data matrices are available from TreeBASE (study number S10410).
Parsimony analyses involved heuristic searches. The data matrices
were subjected to 1,000 replicates of random sequence additions using
tree bisection-reconnection (TBR) branch-swapping under the Fitch criterion (unordered states and equal weights). Gaps were treated as missing data. Potentially phylogenetically informative and unambiguously
aligned gaps (there were no ambiguous regions in the alignment) were
coded as individual characters and subjected to specific step matrices following the method proposed by Lutzoni et al. (2000) and using the program INAASE v. 3 (Lutzoni et al. 2000). Only ten trees were held at each
step, to minimize the time spent searching for trees on suboptimal islands.
The starting tree was obtained by stepwise addition. Finally, 1,000 bootstrap replicates (BS: Felsenstein 1985) with 10 heuristic searches, as above,
were performed to assess internal support for nodes. Statistics reflecting
the amount of phylogenetic signal in the parsimony analysis were provided by the consistency index (CI: Kluge and Farris 1969) and the retention index (RI: Swofford 1993).
Bayesian analyses were implemented using the best-fit nucleotide
substitution model for each data set [HKY (nst = 2; rates = equal; statefreqpr = dirichlet) for ETS, K80 + G (nst = 2; rates = gamma; statefreqpr =
fixed) for ITS, and F81 + I (nst = 1; rates = propinv; statefreqpr = dirichlet)
for rpl16]. These models were selected using MrModeltest 2.3 (Nylander
2004) and the Akaike information criterion (Akaike 1973). For the ETS +
ITS combined analysis a partitioned model was used, which included the
p value
0.000*
0.932
0.000*
0.000*
0.000*
0.239
0.000*
0.000*
0.000*
0.000*
0.000*
0.000*
0.000*
0.000*
0.000*
0.000*
0.000*
0.090
selected models for the independent data sets. The analyses were based on
5,000,000 generations with four simultaneous runs (sixteen Markov chain
Monte Carlo, MCMC, chains) starting from random trees that were sampled every 100 generations. To determine apparent stationarity of the runs
the variation in log-likelihood scores was examined graphically. The initial
25% of the samples obtained were discarded as burn-in. Convergence was
assessed checking the standard deviation of split frequencies for the four
independent runs and also graphically using the slide command (it shows
the posterior probabilities of clades for nonoverlapping samples of trees
in the sample) and compare command (it plots pairwise split frequencies
for a series of independent MCMC runs) of the program AWTY online
(Wilgenbusch et al. 2004). As for the parsimony analyses, those potentially
informative gaps were treated as individual characters, and the standard
discrete model was specified for them.
Results
Morphology—The results of the biometric analysis, as
well as the main qualitative morphological characters, differentiate between T. dubius and the putative new species
(Table 1, Fig. 2). Because the data support the segregation of
a new species from T. dubius, we will henceforth refer to it
as T. lainzii; a formal description appears in the Taxonomic
Treatment. The main differences between the species involve
both floral and achene characters, as well as leaf features.
Tragopogon lainzii always possesses longer ligules than does
T. dubius, with the ligules as long as or longer than the involucral bracts in T. lainzii (Fig. 3), while in T. dubius the involucral
bracts always exceed the ligules. The number of involucral
bracts is almost constant (13) in T. lainzii, while in T. dubius
the number frequently varies from (7–)8–12(–14). Although
the achene length is similar in both species, T. lainzii differs
from T. dubius in having an achene-beak that is longer and an
achene-body that is shorter and thinner. Moreover, T. lainzii
characteristically possesses a fine-ornamented achene with an
obpyramid-shaped apex-beak, while T. dubius has a coarseornamented achene with a bulbous-shaped apex-beak (Fig. 4).
Furthermore, T. lainzii always has undulate leaves, while the
leaves are not undulate in T. dubius (Fig. 3), and the number of
2011]
SUÁREZ-SANTIAGO ET AL.: A NEW TRAGOPOGON SEGREGATED FROM T. DUBIUS
Fig. 2. Box plots of the distributions of the 18 morphological quantitative characters for T. dubius (DU) compared with T. lainzii (LAI).
Measurements are expressed in mm. *: extreme values.
achenes per head is higher in T. lainzii than in T. dubius (Table 1).
Statistically significant differences were found (p < 0.001) in
almost all quantitative characters, except for involucral-bract
width during flowering, ratio between the lengths of the
achene and pappus, and the peduncle width during ripening
(Table 1). UPGMA analysis likewise supports the morphological separation of T. lainzii and T. dubius (Fig. 5).
Karyological Analysis—All populations of T. lainzii studied exhibited the diploid chromosome number for the genus
(2n = 12). Figure 6 shows both a metaphase plate as well as the
karyotypes for T. lainzii and T. dubius. Both species possess a
similar bimodal karyotype, having three chromosome pairs
each with a submedian centromere and three chromosomal
pairs each with a median centromere.
A secondary constriction is always found in the first
chromosome pair, constituting a satellite in its short arm.
Chromosomal formula: 2n = 12 = 2sm sat + 4sm + 6m.
Karyotype asymmetry is of the 2B type (2: 51–99% of chromosomes with an arm ratio < 2:1; B: ratio between the largest and the smallest chromosome of 2:1–4:1). Neither species
exhibited any intraspecific chromosomal variation. There are
no obvious karyotypic differences between the two species.
Molecular Analysis—Alignment of all 46 ETS sequences
resulted in a 535-bp data matrix. The ITS-5.8S matrix is 728 bp
in length and consists of 49 sequences, and the rpl16 matrix
is 1,087 bp in length and includes 33 sequences. The ETS +
ITS combined matrix includes all ETS and ITS sequences (51)
and is 1,263 bp in length; in this matrix six accessions were
only represented by one of the two regions (only by ETS: collections of T. porrifolius from population CAZ ; only by ITS:
473
T. dubius from Germany, AJ633503, and from Switzerland,
AJ633500, T. porrifolius from Germany, AJ633494, and from
Italy, EF374206, and T. krascheninnikovii, AY645821). To check
the possible impact of including these six accessions on the
resulting topology, the analysis was repeated including only
those taxa for which both regions were available. The resulting topology was concordant with that obtained including
taxa with only one type of sequence and the support of the
clades did not vary substantially (data not shown).
Parsimony analysis of the ITS sequences yielded 5,730
most-parsimonious trees (length: 102, CI: 0.912, RI: 0.940),
while 54,216 trees were obtained for the ETS matrix (length:
71, CI: 0.958, RI: 0.990), and 23,967 trees were obtained for the
combined ETS + ITS matrix (length: 180, CI: 0.894, RI: 0.960).
Parsimony analysis of the rpl16 data set reached the maximum
number of trees permitted in memory (length: 69, CI: 0.928,
RI: 0.951). The resulting strict consensus topologies were all
similar to those obtained via Bayesian inference. The analyses
of the nrDNA regions (ETS, ITS) resulted in no contradictory
topologies, because the main differences between them were
due to the higher resolution shown by the ETS sequences to
resolve the phylogeny at internal nodes (data not shown). The
combined analysis of these two regions yielded an almost
completely resolved phylogeny (Fig. 7).
The ETS + ITS tree shows three highly supported clades
that correspond to the three main subclades (Majores s. s.,
Chromopappus, and Hebecarpus) within the Majores s. l.
clade. Sequences of the different populations of T. lainzii are
identical for both ETS and ITS and form a strongly supported
clade (BS: 98%, PP: 1.00); these populations are included in
the core of the Majores s. s. clade and are clearly separated
from the clade formed by the Spanish populations of T. dubius.
Sequences from one population of T. lainzii (RIO) are included
in the T. dubius clade (Fig. 7), showing a high identity with
the sequences of the latter (ETS: 100%, ITS: 99.9–100%). Two
additional sequences were also placed in clades that contradicted their morphology (Fig. 7): one of them was amplified
from an individual of T. porrifolius (population CAZ), but was
placed in the Spanish T. dubius clade (99% identity); the other
was obtained from a T. dubius individual (population CH)
and is related to T. porrifolius in the Hebecarpus clade (100%
identity).
The phylogenetic tree places populations of T. dubius in
different clades (Fig. 7). Sequences of T. dubius from Turkey
(including T. major) and from Switzerland are included in
the core of the Majores s. s. clade, while T. dubius from Spain
appears as a sister group to the latter. Finally, T. dubius from
India and one collection from Germany form a group not
related to the Majores s. l. clade.
The rpl16 intron yielded trees (Fig. 8) less resolved than
those obtained with the nrDNA markers. Like the ETS + ITS
tree, the rpl16 tree shows a strongly supported T. lainzii clade.
However, unlike the nrDNA tree, the chloroplast tree shows a
close relationship between T. lainzii and T. dubius from Spain
[even more closely related than the sequence of T. dubius from
Turkey, EU392002 (from the same individual as ETS sequence
AY645859 and ITS sequence AY645813; see Fig. 7)]. The
sequence of T. lainzii from population RIO and the T. dubius
sequence from population CH show the same relationships
as those shown in the ETS + ITS tree (Figs. 7, 8); however, the
sample of T. porrifolius from the CAZ population is related
to the other T. porrifolius sequences (Fig. 8) instead of to the
T. dubius ones (unlike in the ETS + ITS tree; Fig. 7).
474
SYSTEMATIC BOTANY
Fig. 3A. Tragopogon lainzii (GDA 52778, holotype), a: general appearance, b: flowering head, c: ripening head, d: achene.
Divergent Populations—In the morphological analysis we
detected one population of T. lainzii and one of T. dubius in
which the individuals showed anomalous character states
with regard to the other populations. These populations are
the same ones that did not group by taxonomy in the molecular trees (RIO of T. lainzii, and CH of T. dubius; Figs. 7, 8).
Individuals from RIO always showed the floral morphology typical of T. lainzii. Moreover, the individuals from this
population always showed undulate leaves like T. lainzii.
However, the features of the achenes were variable within
this population; we could find achenes typical of T. lainzii, but
also achenes typical of T. dubius (coarse-ornamented with an
achene-beak shorter and an achene-body longer and thicker,
respectively, than typical of T. lainzii). Also an intermediate
gradation between species was found for achene characters.
Despite the presence of yellow flowers in the individuals
from the CH population, these plants showed different values for almost all other morphological characters compared
to those typical (Table 1) of the individuals from other populations of T. dubius (e.g. mean values: ligule length at anthesis: 29.23 mm, ratio ligule length/bract length: 0.757, total
achene length: 33.35 mm, achene-beak length: 21.7 mm, ratio
beak length/body length: 1.87). The qualitative characters of
individuals from the CH population (e.g. undulate leaves, the
fine-ornamented achenes, and a (sub)cylindrical apex-beak
shape) were also different compared to other populations of
T. dubius.
[Volume 36
Fig. 3B. Tragopogon dubius (GDA 54996), a: general appearance,
b: flowering head, c: ripening head, d: achene.
Finally, after obtaining the molecular results, we checked
the morphology for individuals of T. porrifolius from population CAZ (another population that did not group by taxonomy in the molecular trees; Fig. 7). In this case we found
variability in achene morphology; in the extreme cases the
achenes were similar to those of T. dubius.
All sampled individuals from these three populations were
diploids and showed the same karyotype found in T. dubius
and T. lainzii.
Discussion
Taxonomic Characterization of T. lainzii—Tragopogon
lainzii is a newly described diploid species of Tragopogon (see
Taxonomic Treatment) that can be distinguished morphologically from T. dubius by the following: the tinge color on the
stems, margin of the leaves, shape of the capitula, number of
involucral bracts, length of the ligules as a proportion of the
length of the involucral bracts, ornamentation of the achene
surface and the shape of the apex of the beak (Table 1; Figs.
2, 3). With regard to the number of involucral bracts, except
for two cases, T. lainzii always showed a constant number of
13, while in T. dubius this number was more variable, ranging from (7–)8–12(–14) with eight the most common number. Among the aforementioned characters some of the most
important morphological characters to distinguish the species
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SUÁREZ-SANTIAGO ET AL.: A NEW TRAGOPOGON SEGREGATED FROM T. DUBIUS
475
Fig. 4. Scanning electron micrographs (SEM) of achenes of T. lainzii,
GDA 52778, holotype (A, body of achene; B, apex of beak) and T. dubius,
GDA 54996 (C, body of achene; D, apex of beak). Scale bar = 100 µm.
of Tragopogon are the length of the ligules relative to the length
of the involucral bracts, the number of involucral bracts and
the characters derived from the achenes (Blanca and Díaz
de la Guardia 1997). Moreover, there are other important
characters that, in spite of some overlap in their measurements, nonetheless distinguish T. lainzii from T. dubius. Thus,
T. lainzii in general shows shorter involucral bracts during
achene ripening, a shorter and more slender achene body and
more achenes per head than does T. dubius (Table 1).
Both the quantitative analysis and UPGMA phenogram
support the taxonomic recognition and characterization of
T. lainzii (Table 1; Fig. 5). The phenogram shows that the populations of each species group together by morphological
similarity, indicating the interpopulational homogeneity of
the morphological characters of this new species.
Karyologically, no differences were found between the
populations of T. lainzii and T. dubius; all studied individuals
showed the same chromosomal formula: 2n = 12 = 2sm sat +
4sm + 6m (Fig. 6).
Other Iberian species of Tragopogon with yellow flowers are
T. lamottei Rouy, T. pratensis L., and T. pseudocastellanus Blanca
& C. Díaz. However, T. lainzii can be easily distinguished
from them based on morphological (see key to yellowflowered species in the Iberian Peninsula) and, in the case of
T. pseudocastellanus, which is polyploid with 2n = 24, cytological features (Blanca and Díaz de la Guardia 1996).
Fig. 5. UPGMA phenogram obtained from the Euclidean distances
calculated for 18 morphological quantitative characters in 15 populations
of T. lainzii and T. dubius of the Iberian Peninsula.
Fig. 6. Metaphase plates and karyotypes of T. lainzii (A, C) and
T. dubius (B, D). Scale bar = 2 µm.
Phylogenetic Analysis—All phylogenetic analyses support
the recognition of T. lainzii as an independent species from
the morphologically similar species, T. dubius. Importantly, all
sequences of T. lainzii (exceptions discussed below) formed
a strongly supported clade separated from T. dubius, for all
molecular markers used.
Contrary to the meaning of the specific epithet (dubius =
doubtful), T. dubius has long been considered morphologically homogeneous by taxonomists, with invariant morphological characters. Thus, the traditional concept of T. dubius
has included all Tragopogon species with yellow flowers and
swollen peduncles under the capitula (e.g. Vollmann 1914;
Richardson 1976; Tzvelev 1985; Bolòs and Vigo 1989). Our
results show that T. dubius is not morphologically homogeneous and the specific epithet is well deserved. Tragopogon
lainzii has been considered part of T. dubius, and it illustrates
that not all yellow-flowered Tragopogon with swollen peduncles are T. dubius. In fact, distinct lineages, to date unrecognized as species, may be included under this broad concept
of T. dubius. Mavrodiev et al. (2008a), studying the parentage of the tetraploid T. kashmirianus G. Singh, suggested that
T. dubius from India and neighboring countries may be a different species from the European T. dubius, on the bases of
molecular and morphological data. Our phylogenetic analyses agree with Mavrodiev et al. (2008a) and show T. dubius
as a nonmonophyletic species, with different lineages existing across its geographic distribution. Thus, in addition to the
distinctiveness of T. lainzii, other samples of T. dubius from the
Iberian Peninsula are not part of the same subclade as other
sampled European (Switzerland, AJ633500) and Turkish
(including T. major) T. dubius. While the latter are included
in the core of the Majores s. s. clade (sensu Mavrodiev et al.
2005), the sequences of the Iberian populations form a sister
clade to the Majores s. s. clade. Finally, the sequence of the
T. dubius collection from India used by Mavrodiev et al.
(2008a) appeared closely related to a sequence from Germany
and clearly is not related to the species of the Majores s. l. clade
[in Mavrodiev et al. (2008a), T. dubius from India occurred in
a clade of native species from Kashmir, which is sister to the
Tragopogon clade sensu Mavrodiev et al. (2005)]. This relationship between T. dubius from India and one collection from
Germany would increase the range of the putative new Indian
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SYSTEMATIC BOTANY
[Volume 36
Fig. 7. Bayesian consensus tree for analysis of the ETS + ITS data set for studied species of Tragopogon. Numbers above branches are posterior probabilities, and numbers below branches are bootstrap values ≥ 50% obtained in the parsimony analysis, which generated the identical topology. Specific
epithets for sequences from hybrid populations are in quotes. Country codes (in parentheses) for non-Spanish T. dubius populations: CH: Switzerland;
DE: Germany; IN: India; TR: Turkey; see Appendix 1 for Spanish population codes of the sequences generated in this study. Names for clades within
Majores s. l. are taken from Mavrodiev et al. (2005).
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SUÁREZ-SANTIAGO ET AL.: A NEW TRAGOPOGON SEGREGATED FROM T. DUBIUS
477
Fig. 8. Bayesian consensus tree for analysis of the plastid rpl16 sequences of the studied Tragopogon species, showing the same topology as the 50%
majority tree obtained in the parsimony analysis. Numbers above branches are posterior probabilities, and numbers below branches are bootstrap values ≥ 50%. Specific epithets for sequences from hybrid populations are in quotes. Country codes (in parentheses) for non-Spanish T. dubius populations:
TR: Turkey; see Appendix 1 for Spanish population codes of the sequences generated in this study.
478
SYSTEMATIC BOTANY
species (suggested by Mavrodiev et al. 2008a). Clearly more
sampling and study of this material is needed.
Karyological studies of T. dubius by Dvořák et al. (1978)
showed two different karyotypes in different populations.
Thus, one of these two karyotypes was the same as that
reported here in the Iberian populations of T. dubius (6sm +
6m), while the other ones differed in having 8, instead of 6,
chromosomes with the centromere in median position (4sm
+ 8m). Thus, other unrecognized cryptic species may exist
within what is now called T. dubius.
The molecular differentiation of plants of T. dubius from
Spain compared to the taxa of the Majores s. s. clade, as well
as the nonmonophyletic nature of T. dubius throughout its
geographic distribution and the reported variation in karyotypes, merit further studies involving both morphological
and molecular data.
Hybridization in Populations Studied—Natural hybridization between individuals of wild populations from Tragopogon
has been reported in many studies on the basis of both morphology and molecular data (e.g. cf. Ownbey 1950; Díaz de la
Guardia and Blanca 2004; Mavrodiev et al. 2008b). The combination of morphological and molecular data is a powerful
tool for detecting instances of hybridization, especially recent
hybridization. In the present study, we detected several putative hybrid populations involving T. dubius, T. lainzii, and
T. porrifolius on the basis of the morphological and molecular
features. Moreover, the inclusion in the molecular analyses of
a chloroplast region has allowed us to suggest which species
are the maternal parents. Ownbey (1950), in his study of natural allopolyploid formation involving introduced Tragopogon
species in North America, stated that natural hybrids can be
expected wherever any of these species grow together. For
each of the hybrids we detected, the two parental species
were growing together (RIO: T. dubius and T. lainzii; CH and
CAZ: T. dubius and T. porrifolius).
In all three hybrid populations, the individuals showed
a combination of the morphological features typical of the
parental species, and few characters showed intermediate
forms. Thus, in the population where T. dubius and T. lainzii occur together, the hybrids detected always show the floral and leaf features of T. lainzii, and therefore they look like
T. lainzii. However, achene characters were more variable.
In this population, some individuals had the achene type of
T. dubius and others the achene type of T. lainzii, and intermediate forms between them were detected. Our results obtained
from the chloroplast rpl16 region suggest that in this population T. lainzii acts as the maternal parent.
With regard to the hybridization detected in those populations where T. dubius and T. porrifolius grow together, the
results suggest T. porrifolius as the maternal parent (consistent with the maternal parentage of the allotetraploid T. mirus,
which is derived from T. dubius and T. porrifolius; Soltis and
Soltis 1989). Contrary to Ownbey’s observations (1950), nonbicolored ligules were detected in the hybrids, but they look like
either parental species (yellow ligules in the CH population,
and purple ligules in the CAZ population). Unlike ligule color,
the achene characters are good markers to distinguish hybrids
between T. dubius and T. porrifolius. Tragopogon porrifolius has
a characteristic subcylindric beak-apex shape (club-shaped
in T. dubius; Blanca and Díaz de la Guardia 1997), which is
a good hybridization marker when it appears in T. dubiuslike hybrids. The club-shaped beak-apex is also a good hybridization marker when it appears in T. porrifolius-like hybrids.
[Volume 36
The diploid hybrids studied by Ownbey (1950) were highly
sterile, as their heads did not continue to develop normally
after flowering, and almost all ovaries aborted at the flowering stage or shortly thereafter. Based on this evidence, together
with the morphological uniformity of the hybrids, Ownbey
inferred that these hybrids were F1 individuals. Although we
did not carry out an exhaustive analysis of hybrid sterility, all
individuals sampled from the hybrid populations were fertile;
they showed normal heads, and their seeds germinated normally (pers. obs.). These observations, together with the floral
features (flower like T. lainzii in T. dubius × T. lainzii plants,
and nonbicolored flowers in T. dubius × T. porrifolius individuals) and the range of variation in the achene characters, would
suggest that the hybrids of these populations are not F1 plants,
but may represent later-generation backcrosses to one parent.
Finally, hybridization seems to be a frequent phenomenon
in T. dubius populations where it grows together with other
species (e.g. cf. Ownbey 1950). Hybridization should be considered as a possible source of variation when studies of the
genus Tragopogon are conducted, especially in those studies
including molecular markers, in which the use of a “hybrid”
plant that is not recognized as such can yield mistaken conclusions. Thus, the combination of morphology and molecular markers is a useful approach for detecting hybridization.
Taxonomic Treatment
Tragopogon lainzii V. N. Suárez-Santiago, P. S. Soltis, D. E.
Soltis, C. Díaz de la Guardia & G. Blanca sp. nov.—TYPE:
SPAIN. Granada: ctra. Orce-María, 950 m, 13 May 2006,
V. N. Suárez-Santiago and I. López-Flores 52778 (holotype:
GDA). See Appendix 1 for additional specimens.
Differt a plus minusve simili T. dubius Scop. caulibus rubellis, foliis margine undulatis, bracteis involucralibus 13 atque
post anthesin accrescentibus [ita ut 44(54.8 ± 0.3)66 mm longae fiant], per anthesin autem aequilongis aut brevioribus
quam ligulis externis [hae quidem 30(37.3 ± 0.2)48 mm longae sunt] et achaeniis 115(186.3 ± 0.02)263 in capitulo unoquoque [rostrum achaenii uniuscuiusque 15.5(21.5 ± 0.2)27.5
mm longum, corpus autem 7.5(10.0 ± 0.1)11.5 item longum].
Biennial plants. Stems 30–80(–100) cm, ramose in the basal
zone, glabrous, woolly-floccose pubescent in the leaf axils, reddish. Leaves linear, broadening at the base to form a sheath,
margin entire and undulate; the basal leaves numerous, rosulate; the cauline leaves (10–)15–30(–35) × 1.5–2.2 cm, decreasing in size towards the apex, alternate, semiamplexicaul,
largely subulate. Capitula homogamous, terminal, solitary;
peduncles cylindrical, markedly swollen at the apex, 7–13 mm
wide at anthesis, 12–20 mm in fruit; involucre with 13 bracts
in one row; involucral bracts 27–45 × 4–9 mm at anthesis, 44–66
× 5–11 mm in fruit, lanceolate, subkeeled, glabrous, greenish. Ligules 30–48 mm long, as long as or longer than involucral bracts, pale-yellow, with dark veins on the back and teeth
greenish-brown; anthers dark-brown and style greenish-brown.
Achenes 25–38.5 mm long; body 7.5–11.5 mm, sligthly curved,
scabrous, brown, gradually tapering towards the beak, beak
15.5–27.5 mm long, obpyramidal at the tip. Pappus 20–30 mm,
with plumose hairs. 2n = 12. (IV)V-VI(VII). Figure 3.
Etymology—The epithet honors our friend and colleague,
the botanist P. Manuel Laínz, S. J.
Habitat—Grassy areas of nitrified lands, usually in road
taluses and ditches, on carbonate-loamy substrate, 500–1,500 m
SE of Iberian Peninsula.
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SUÁREZ-SANTIAGO ET AL.: A NEW TRAGOPOGON SEGREGATED FROM T. DUBIUS
479
Key to Yellow-Flowered Species of TRAGOPOGON in the Iberian Peninsula
1.
Involucral bracts 5(–7); achenes 17–25 mm long . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2
2. Cauline leaves subulate, 0.3–0.5 cm wide; ligules 4/5 length of involucral bracts or almost the same; achenes 17–22 mm long . . . . . . . T. pratensis
2. Cauline leaves linear-lanceolate, 0.6–0.8 cm wide; ligules 2/3 length of involucral bracts; achenes 20–25 mm long . . . . . . . . . T. pseudocastellanus
1. Involucral bracts (7–)8–13(–16); achenes 23–39 mm long . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3
3. Peduncles contracted below the capitulum during fruiting; involucral bracts with reddish or blackish margin; florets yellow,
with reddish-orange dorsal veins . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . T. lamottei
3. Peduncles gradually swollen towards the capitulum, involucral bracts greenish; florets yellow . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 4
4. Leaf margin not undulate; ligules 2/3–1/2 length of involucral bracts . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . T. dubius
4. Leaf margin undulate; ligules as long as or longer than involucral bracts . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . T. lainzii
Acknowledgments. The authors would like to thank Padre M. Laínz
for assistance with the text in Latin. We are also grateful to the staff of
the herbarium of the University of Granada, to the staff of the Marion
Ownbey Herbarium, Washington State University, to Dr. K. Pistrick of
the Leibniz-Institut für Pflanzengenetik und Kulturpflanzenforschung in
Gatersleben, and to Dr. B. Gemeinholzer of the Botanischer Garten und
Botanisches Museum Berlin-Dahlem, Freie Universität Berlin. We thank
Dr. F. Alba-Sánchez and Dr. D. Nieto-Lugilde for assistance with Fig. 1.
Dr. Suárez-Santiago was funded by a postdoctoral grant from the Spanish
Ministry of Education and Science. This work was supported in part by
NSF grants MCB-0346437 and DEB-06144421. We also thank two anonymous reviewers and Dr. Andrew Hipp for their comments.
Literature Cited
Akaike, H. 1973. Information theory and an extension of the maximum
likelihood principle. Pp. 267–281 in Second international symposium on information theory, eds. B. N. Petran and F. Csaki. Budapest:
Akademiai Kiado.
Baldwin, B. G. and S. Markos. 1998. Phylogenetic utility of the external
transcribed spacers (ETS) of 18S–26S rDNA: congruence of ETS and
ITS trees of Calycadenia (Compositae). Molecular Phylogenetics and
Evolution 10: 449–463.
Blanca, G. and C. Díaz de la Guardia. 1996. Sinopsis del Género Tragopogon
L. (Asteraceae) en la Península Ibérica. Anales del Jardin Botanico de
Madrid 54: 358–363.
Blanca, G. and C. Díaz de la Guardia. 1997. Fruit morphology in Tragopogon
L. (Compositae: Lactuceae) from the Iberian Peninsula. Botanical
Journal of the Linnean Society 135: 319–329.
Bolòs, O. and J. Vigo. 1989. Notes sobre taxonomia i nomenclatura de
plantes, IV. Folia Botanica Miscellanea 6: 85–86.
Chaubard, L. A. 1837. Tragopogon L. Pp. 395–397 in Flore du bassin souspyrénéen, ed. J. B. Noulet. Toulouse.
Darlington, C. D. and L. F. La Cour. 1969. The handling of chromosomes.
London: Allen and Unwin.
Díaz de la Guardia, C and G. Blanca. 1990. Tragopogon castellanus Levier =
T. crocifolius subsp. badalii Willk. Anales del Jardin Botanico de Madrid
47: 253–256.
Díaz de la Guardia, C and G. Blanca. 2004. A new Spanish species of
Tragopogon (Asteraceae: Lactuceae). Botanical Journal of the Linnean
Society 146: 505–511.
Doyle, J. J. and J. L. Doyle. 1987. A rapid DNA isolation procedure for
small quantities of fresh leaf tissue. Phytochemical Bulletin 19: 11–15.
Dvořák, F., P. Trnka, and B. Dadáková. 1978. Cytotaxonomic study of
Tragopogon L. in Czechoslovakia. Folia Geobotanica et Phytotaxonomica
13: 305–330.
Felsenstein, J. 1985. Confidence limits on phylogenies: an approach using
the bootstrap. Evolution 39: 783–791.
Font Quer, P. 1979. Diccionario de Botánica. Barcelona: Labor.
Innis, M. A., D. H. Gelfand, J. J. Sninsky, and T. J. White. 1990. PCR protocols: a guide to methods and applications. New York: Academic Press.
Jacquin, N. J. 1773. Florae Austriacae vol. 1. Vienna: Viennae Austriae, Typis
Leopoldi Joannis Kaliwoda, Aulae imperialis typographi.
Jordan, W. C., M. W. Courtney, and J. E. Neigel. 1996. Low levels of
intraspecific genetic variation at a rapidly evolving chloroplast DNA
locus in North American duckweeds (Lemnaceae). American Journal
of Botany 83: 430–439.
Kluge, A. G. and J. S. Farris. 1969. Quantitative phyletics and the evolution
of anurans. Systematic Zoology 40: 315–328.
Krahulec, F., Z. Kaplan, and J. Novák. 2005. Tragopogon porrifolius ×
T. pratensis: the present state of an old hybrid population in Central
Bohemia, the Czech Republic. Preslia 77: 297–306.
Lee, J., B. G. Baldwin, and L. D. Gottlieb. 2002. Phylogeny of Stephanomeria
and related genera (Compositae–Lactuceae) based on analysis of
18S–26S nuclear rDNA ITS and ETS sequences. American Journal of
Botany 89: 160–168.
Levan, A., K. Fredga, and A. A. Sandberg. 1964. Nomenclature for centromeric position on chromosomes. Hereditas 52: 201–220.
Lindemann, E. E. 1881. Flora Chersonensis vol. 1. Stockholm: Odessae.
Lutzoni, F., P. Wagner, V. Reeb, and S. Zoller. 2000. Integrating ambiguously
aligned regions of DNA sequences in phylogenetic analyses without
violating positional homology. Systematic Biology 49: 628–651.
Mavrodiev, E. V., I. Nawchoo, P. S. Soltis, and D. E. Soltis. 2008a. Molecular
data reveal that the tetraploid Tragopogon kashmirianus (Asteraceae:
Lactuceae) is distinct from the North American T. mirus. Botanical
Journal of the Linnean Society 158: 391–398.
Mavrodiev, E. V., P. S. Soltis, M. A. Gitzendanner, R. M. Baldini, and
D. E. Soltis. 2007. Polyphyly of Tragopogon porrifolius (Asteraceae), a
Eurasian native with intercontinental disjuncts. International Journal
of Plant Sciences 168: 889–904.
Mavrodiev, E. V., P. S. Soltis, and D. E. Soltis. 2008b. Putative parentage of
six old world polyploids in Tragopogon L. (Asteraceae: Scorzonerinae)
based on ITS, ETS, and plastid sequence data. Taxon 57: 1215–1232.
Mavrodiev, E. V., M. Tancig, A. M. Sherwood, M. A. Gitzendanner,
J. Rocca, P. S. Soltis, and D. E. Soltis. 2005. Phylogeny of Tragopogon
L. (Asteraceae) based on internal and external transcribed spacer
sequence data. International Journal of Plant Sciences 166: 117–133.
Nylander, J. A. A. 2004. MrModeltest v. 2. Program distributed by the
author. Evolutionary Biology Centre, Uppsala University.
Ownbey, M. 1950. Natural hybridization and amphiploidy in the genus
Tragopogon. American Journal of Botany 37: 487–499.
Ownbey, M. and G. D. McCollum. 1953. Cytoplasmic inheritance and
reciprocal amphiploidy in Tragopogon. American Journal of Botany 40:
788–796.
Richardson, I. B. K. 1976. Tragopogon L. Pp. 322–325 in Flora Europaea
vol. 4, eds. T. G. Tutin, V. H. Heywood, N. A. Burges, D. M. Moore,
D. H. Valentine, S. M. Walters, and D. A. Webb. Cambridge:
Cambridge University Press.
Ronquist, F. and J. P. Huelsenbeck. 2003. MRBAYES 3: Bayesian phylogenetic inference under mixed models. Bioinformatics 19: 1572–1574.
Small, R. L., J. A. Ryburn, R. C. Cronn, T. Seelanan, and J. F. Wendel. 1998.
The tortoise and the hare: choosing between noncoding plastome and
nuclear Adh sequences for phylogenetic reconstruction in a recently
diverged plant group. American Journal of Botany 85: 1301–1315.
Soltis, D. E. and P. S. Soltis. 1989. Allopolyploid speciation in Tragopogon:
Insights from chloroplast DNA. American Journal of Botany 76:
1119–1124.
Soltis, D. E., P. S. Soltis, J. C. Pires, A. Kovarik, and J. Tate. 2004. Recent and
recurrent polyploidy in Tragopogon (Asteraceae): genetic, genomic,
and cytogenetic comparisons. Biological Journal of the Linnean Society.
Linnean Society of London 82: 485–501.
Stearn, W. T. 1980. Botanical Latin. London: David and Charles.
Stebbins, G. L. 1971. Chromosomal evolution in higher plants. London:
Edward Arnold.
Swofford, D. L. 1993. PAUP: phylogenetic analysis using parsimony,
v. 3.1.1. Champaign: Illinois Natural History Survey.
Swofford, D. L. 2003. PAUP*. Phylogenetic analysis using parsimony
(*and other methods), v. 4.0. Sunderland: Sinauer Associates.
Tamura, K., J. Dudley, M. Nei, and S. Kumar. 2007. MEGA4: Molecular
Evolutionary Genetics Analysis (MEGA) software version 4.0.
Molecular Biology and Evolution 24: 1596–1599.
Thornill, J. W., R. K. Matta, and W. H. Wood. 1965. Examining three dimensional microstructures with the scanning electron microscope. Grana
Palynologica 6: 3–6.
480
SYSTEMATIC BOTANY
Tzvelev, N. N. 1985. Genus Tragopogon L. (Asteraceae) in the European
part of the USSR. Pp. 238–250 in News in higher plant systematics
vol. 22, ed. T. Egorova. Leningrad: Nauka.
Vollmann, F. 1914. Flora von Bayern. Stuttgart.
Wilgenbusch, J. C., D. L. Warren, and D. L. Swofford. 2004. AWTY: A system for graphical exploration of MCMC convergence in Bayesian
phylogenetic inference. http://ceb.csit.fsu.edu/awty.
Wen, J. and E. A. Zimmer. 1996. Phylogeny and biogeography of Panax
L. (the ginseng genus, Araliaceae): inference from ITS sequences of
the nuclear ribosomal DNA. Molecular Phylogenetics and Evolution 6:
167–177.
Appendix 1. Population data, voucher information and type of analysis performed for taxa used in this study. Information is listed as follows:
species: country: population code; locality; voucher data; type of analysis
(M: morphological analysis, C: cytogenetical analysis, I: molecular analysis); EMBL accession number (in parenthesis). All specimens are deposited at GDA herbarium (University of Granada).
Tragopogon angustifolius: Spain: A-92; Granada, Autovía A-92, en
la salida 303 a Hernán Valle; V. N. Suárez-Santiago and I. López-Flores,
13/V/2006, 52757 (GDA); I (ETS: FN675685, ITS: FN675712, rpl16:
FN687182). ORC; Granada, ctra. a Orce desde la ctra. comarcal 330; V.
N. Suárez-Santiago and I. López-Flores, 13/V/2006, 52722 (GDA); I (ETS:
FN675684, ITS: FN675711, rpl16: FN687181). T. dubius: Spain: AD; Burgos,
Aranda de Duero, Salida Sur, junto gasolinera CEPSA; V. N. Suárez-Santiago
and I. López-Flores, 28/V/2006, 52768 (GDA); M, C, I (ETS: FN675675, ITS:
FN675703, rpl16: FN687175). ALAR; Palencia, Alar del Rey, junto vía del
tren; V. N. Suárez-Santiago and C. Abellán-López, 6/VII/2006, 52776 (GDA);
C, I (ETS: FN675669, ITS: FN675697). AT; Guadalajara, Atienza, monte
detrás del pueblo; V. N. Suárez-Santiago and C. Abellán-López, 30/VI/2006,
52767 (GDA); M, C, I (rpl16: FN687170). CAZ; Jaén, Ctra. Peal de BecerroCazorla (A-319), prop. Cazorla, antes del cruce con A-322, 750 m; V. N.
Suárez-Santiago et al. 4/VI/2008, 55948 (GDA); C, I (ETS: FN675668, ITS:
FN675698, rpl16: FN687168).CH; Granada, Sierra Nevada, La Cortichuela,
Jardín Botánico, 1,650 m; G. Blanca et al. 27/VI/2006, 52773 (GDA); M, C,
I (ETS: FN675677, ITS: FN675705, rpl16: FN687177). LEN; Burgos, nacional 114 entre Sigüenza y Aranda de Duero, 1 km pasado Aldealengua,
a 30 km de Aranda de Duero; V. N. Suárez-Santiago and C. Abellán-López,
30/VI/2006, 52770 (GDA); M, C, I (ETS: FN675671, ITS: FN675701, rpl16:
FN687171). IZN; Granada, Ctra Iznalloz-Guadahortuna (A-323) Km 2,
entre las Encebras y el Navazuelo, 1,000 m; V. N. Suárez-Santiago et al.,
4/VI/2008, 54994 (GDA); M, C, I (ETS: FN675670, ITS: FN675696, rpl16:
FN687169). OAS; Jaén, Ctra. Jaén-Granada (A-44), estación de servicio el Oasis, junto gasolinera; V. N. Suárez-Santiago and F. López-Soriano,
25/V/2008, 54996 (GDA); M, C, I (ETS: FN675672, ITS: FN675702, rpl16:
FN687172). SOM; Madrid, Pto. Somosierra, Salida 79 hacia Horcajo
de la Sierra, Ctra. entre Horcajo de la Sierra y Montejo de la Sierra
(M-141), 1,300 m; V. N. Suárez-Santiago and S. Schiaffino, 21/VI/2008,
54997 (GDA); M, C, I (ETS: FN675673, ITS: FN675699, rpl16: FN687173).
[Volume 36
SOR; Soria, Nacional 122 entre el Burgo de Osma y Soria, km 206-205;
V. N. Suárez-Santiago and C. Abellán-López, 1/VII/2006, 52777 (GDA); C, I
(ETS: FN675676, ITS: FN675704, rpl16: FN687176). TOB; Ctra. OcañaAlbacete (N-301), Villatobas, 1km pasado el pueblo, 600 m; V. N. SuárezSantiago and S. Schiaffino, 20/VI/2008, 54998 (GDA); M, C, I (ETS: FN675674,
ITS: FN675700, rpl16: FN687174). T. lainzii: Spain: CASTR; Granada,
Sierra de Castril, río Castril, 950 m; V. N. Suárez-Santiago et al., 19/VI/2008,
54992 (GDA); M, C, I (ETS: FN675663, ITS: FN675691, rpl16: FN687159).
CAZ; Jaén, Ctra. Peal de Becerro-Cazorla (A-319), prop. Cazorla, antes del
cruce con A-322, 750 m; V. N. Suárez-Santiago et al., 54993 (GDA); M, C, I
(ETS: FN675666, ITS: FN675694, rpl16: FN687167). FAD; Granada, entre
la Cañada de Cañepla y la Puebla de Don Fadrique; ctra. A-317 (prop. La
Puebla); V. N. Suárez-Santiago and I. López-Flores, 13/V/2006, 52775 (GDA);
M, C, I (ETS: FN675658, ITS: FN675686, rpl16: FN687160). JOD; Jaén, Ctra.
Jódar-Peal de Becerro (A-322), 450 m; V. N. Suárez-Santiago et al., 4/VI/2008,
54986 (GDA); M, C, I (ETS: FN675662, ITS: FN675690, rpl16: FN687163).
ORC; Granada, ctra. Orce-María, 950 m; V. N. Suárez-Santiago and I. LópezFlores, 13/V/2006, 52778 (GDA); M, C, I (ETS: FN675664, ITS: FN675692,
rpl16: FN687161). PUCAST; Granada, Sierra de Castril, Ctra. Pozo AlcónCastril (A-326), prop. embalse de la Bolera. 950 m; V. N. Suárez-Santiago et
al., 19/VI/2008, 54990 (GDA); M, C, I (rpl16: FN687164). PZ; Jaén, Sierra del
Pozo, Loma de Cagasebo; V. N. Suárez-Santiago and G. Blanca, 14/VI/2006,
52774 (GDA); C, I (ETS: FN675665, ITS: FN675693, rpl16: FN687162). RIO;
Albacete, Srra de Alcaraz; Ctra. entre Salobre y Riópar (CM-412), prop.
Riópar, junto cruce con Ctra. C-415 hacia ptos. Crucetas y Crucetillas,
1,100 m; V. N. Suárez-Santiago et al., 4/VI/2008, 54989 (GDA); M, C, I
(ETS: FN675667, ITS: FN675695, rpl16: FN687165). SAL; Albacete, Sierra
de Alcaraz, ctra. entre Salobre y Riópar (CM-412), entre Las Parideras y
Las Dehesas, 1,000 m; V. N. Suárez-Santiago et al., 4/VI/2008, 55104 (GDA);
M, C, I (ETS: FN675660, ITS: FN675688). SAG; Granada, Sierra de la Sagra,
Cortijo Casa de la Virgen. 1,350 m; V. N. Suárez-Santiago et al., 4/VI/2008,
55001 (GDA); M, C, I (ETS: FN675661, ITS: FN675689). SR; Albacete,
Sierra de Alcaraz, ctra entre Siles y Riópar; V. N. Suárez-Santiago, and
G. Blanca, 15/VI/2006, 52772 (GDA); C, I (ETS: FN675659, ITS: FN675687,
rpl16: FN687166). T. porrifolius: Spain: CARO; Jaén, La Carolina, ctra. al
Centenillo; V. N. Suárez-Santiago and I. López-Flores, 19/V/2006, 52796 (GDA);
I (ETS: FN675678, ITS: FN675709, rpl16: FN687179). CAZ; Jaén, Ctra. Peal
de Becerro-Cazorla (A-319), prop. Cazorla, antes del cruce con A-322,
750 m; V. N. Suárez-Santiago and S. López-Vinyallonga, 4/VI/2009, 55103
(GDA); M, C, I (ETS: FN675683, rpl16: FN687180). CH; Granada, Sierra
Nevada, La Cortichuela, Jardín Botánico, 1,650 m; V. N. Suárez-Santiago
et al., 12/VI/2008, 54961 (GDA); C, I (ETS: FN675679, ITS: FN675706, rpl16:
FN687178). IZN; Granada, Ctra Iznalloz-Guadahortuna (A-323) Km 2,
entre las Encebras y el Navazuelo, 1,000 m; V. N. Suárez-Santiago et al.,
4/VI/2008, 54962 (GDA); C, I (ETS: FN675680, ITS: FN675707). OAS; Jaén,
ctra. Jaén-Granada (A-44), estación de servicio el Oasis, junto gasolinera;
V. N. Suárez-Santiago and F. López-Soriano, 25/V/2008, 54954 (GDA); C, I
(ETS: FN675681, ITS: FN675710). ORC; Granada, carretera Cúllar-Orce,
dirección Orce desde comarcal 330; V. N. Suárez-Santiago et al., 14/V/2008,
54963 (GDA); C, I (ETS: FN675682, ITS: FN675708).