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Botanical Journal of the Linnean Society, 2013, 171, 429–440. With 4 figures
The Scotch broom, Cytisus scoparius (Fabaceae),
a paradox in Denmark – an invasive plant or
endangered native species?
LARS ROSENMEIER, ERIK D. KJÆR and LENE R. NIELSEN*
Forest & Landscape, Faculty of Science, University of Copenhagen, Rolighedsvej 23, DK-1958
Frederiksberg C, Denmark
Received 6 February 2012; revised 19 June 2012; accepted for publication 7 September 2012
Scotch broom, Cytisus scoparius, spreads rapidly in parts of Denmark and is considered an invasive species by some
authors. However, the species has been present in the Danish flora for centuries and is therefore considered native
to Denmark. In the present study we explore whether Danish Scotch broom consists of one or two gene pools with
potential differences in phenotype and invasiveness. One plastid and five nuclear microsatellite markers were used
to reveal potential substructuring of Danish Scotch broom. Nine populations were included representing populations exhibiting invasive behaviour and populations showing non-invasive behaviour. An Italian population was
used as reference. Bayesian analysis based on genetic markers indicated that the sampled populations form two
distinct gene pools, and this pattern was supported by neighbour-joining trees. Measurements of height and width
of the analysed plants showed that the two gene pools correspond to populations exhibiting either a horizontal
habit and non-invasive behaviour or an erect habit and, in some cases, invasive behaviour. The Italian population
clustered with the erect ones. We discuss the origin and taxonomic status of the two gene pools and conclude that
Danish horizontal Scotch broom should be given a formal taxonomic status in order to initiate conservation
activities for its protection. © 2012 The Linnean Society of London, Botanical Journal of the Linnean Society,
2013, 171, 429–440.
ADDITIONAL KEYWORDS: morphology – native – nuclear microsatellites – plastid microsatellites.
INTRODUCTION
Recent meta-analyses have shown that there is a
clear impact of invasive species on species richness at
the local scale (Gaertner et al., 2009; Powell, Chase &
Knight, 2011). Direct ecological consequences of invasive species include changes in the function of the
ecosystem or simple outnumbering of the indigenous
species (Vilà, Weber & D’Antonio, 2000; Pauchard &
Shea, 2006; Suetsugu et al., 2012). Native species
may also be put at risk by hybridization and introgression by invading species, if these are closely
related with only weak reproductive barriers (Mooney
& Cleland, 2001). With intensified modern travel and
trade, the spread of alien organisms has rapidly
increased (Meyerson & Mooney, 2007; Pautasso et al.,
*Corresponding author. E-mail: [email protected]
2010; Gladieux et al., 2011), enhancing the risk of
interspecific interactions. To protect native species, it
is important to have operational taxonomic descriptions of the biota. However, incomplete taxonomic
knowledge is a general problem as many species
and species borders are not yet fully described
(May, 1988). This can hinder conservation initiatives,
because conservationists typically work from a
species list at the national level (Isaac, Mallet &
Mace, 2004). In the case of closely related native and
invasive species, the lack of operational taxonomic
units can become critical for conservation measures
as discussed here.
Scotch broom, Cytisus scoparius (L.) Link
(Fabaceae), is native to most of Europe. It has been
introduced to other parts of the World including
Africa, the Americas and Australasia (Potter et al.,
2009), where the shrub in many cases spreads rapidly
in open areas and often invades abandoned pastures
© 2012 The Linnean Society of London, Botanical Journal of the Linnean Society, 2013, 171, 429–440
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L. ROSENMEIER ET AL.
(Potter et al., 2009). In addition, invasive populations
of Scotch broom seem to have similar levels of
inbreeding, genetic diversity and structure as native
ones, suggesting that the species is a highly successful invader (Kang, Lowe & Buckley, 2007b).
In Denmark, C. scoparius also behaves as an invasive in many areas, and it is classified as a ‘landscape
weed’. Due to its invasive behaviour in some areas,
with negative effects on the biodiversity of semiheathlands and grasslands, the Danish Ministry of
the Environment included the species on the preliminary list of invasive species in May 2008 (Skov- og
Naturstyrelsen, 2008). However, treating the species
as invasive is controversial as it has been present
in the Danish flora for centuries and is therefore
regarded as native in the country. It was already
reported in Denmark in 1648 by Simon Paulli in
Flora Danica (Paulli, 1648) written for the King,
Christian IV, and it has been known in Scandinavia
since the Middle Ages (Buchwald, 2008). For this
reason, C. scoparius was left out of the final ‘Actionplan for Invasive Species’ (Skov- og Naturstyrelsen,
2009), because invasive species must by definition be
non-native.
In addition to the areas where the species is acting
as an invasive, there are sites where it is found in
relatively small populations and where it does not
show invasive behaviour. Because of these small and
potentially vulnerable populations the species was
included in the strategy for gene conservation of trees
and shrubs in Denmark in 1995 (Graudal, Kjær &
Canger, 1995). The unclear status of the species has,
however, hindered measures to protect potentially
native Danish populations of Scotch broom.
The taxonomic status of Danish Scotch broom has
been under discussion for many years. Böcher &
Larsen (1958) distinguished three major groups in
C. scoparius, each with one or several ecotypes. The
groups differed from each other mainly in height and
frost hardiness, i.e. traits related to vigour and
fitness. In Denmark, Böcher & Larsen, (1958) found
representatives from two of the major groups.
Members of one group were tall (150–250 cm) with a
low to medium degree of frost resistance, and
members of the other group were short (60–100 cm)
with a high degree of frost hardiness. Böcher &
Larsen (1958) believed that only the frost-tolerant low
type is native in Denmark and that it should have at
least varietal rank. According to Buchwald (2008),
however, historical records combined with measurements of herbarium material suggest that Danish
Scotch broom is a single variable species (although no
metric data were presented in the paper).
The main objective of the present study is to detect
whether invasive populations are genetically distinguishable from populations not showing invasive
behaviour. This is based on the hypothesis that C. scoparius in Denmark consists of two gene pools, one
representing a vulnerable native species with presumably relatively low phenotypes mainly confined to
protected semi-natural ecosystems, and the second
composed of potentially taller plants that may spread
rapidly in an invasive manner. We test the hypothesis
by genotyping individuals from Danish populations,
of which some are known to exhibit invasive behaviour whereas others are not, using one plastid and
five nuclear microsatellite markers. Based on the
marker data we determine the likely number of gene
pools in the material by applying a Bayesian analysis
of population structure. We measure the height and
width of each sampled plant and establish the correlation of morphological characters with genetic
markers. Finally, we discuss our results in relation to
the ongoing taxonomic debate of C. scoparius s.l. and
discuss the implications of the results for future management of Danish Scotch broom.
MATERIALS AND METHODS
THE SPECIES
Scotch broom is a shrub in Fabaceae with yellow
flowers, flowering in May–June. Scotch broom is
mainly outcrossing and pollinated by insects (Parker,
1997), and the fruits are pods. The primary dispersal
method is ballistic with dispersal lengths of up to 7 m
(Malo, 2004), but seeds can be transported short
distances by insects. The seeds are long-lived and
remain viable for many years (80 years) in the seed
bank (Turner, 1933).
SAMPLING
All collections took place from late January to March
2009. Nine populations of Scotch broom from
Denmark were included in the study. Populations of
non-invasive and short Scotch broom were identified
using the search facility in http://www.naturdata.dk
and four of these, all from Jutland, were selected and
used for sampling. Five Danish locations, with presumably taller and vigorously growing plants, were
located in Jutland (two locations) and on Funen (one
location) and Sealand (two locations). Scotch broom
has been reported to exhibit invasive behaviour and is
considered a landscape weed at two of these sites,
Mols Bjerge and Svanninge Bjerge. Sampling locations are indicated in supporting Table S1.
The populations were well-defined groups of plants
of 50–200 individuals. Approximately 25 plants were
randomly chosen at each site. Leaves and sprigs were
collected for DNA analyses and stored at -20 °C until
DNA extraction. The distances between the locations
were 0.6–266 km; due to the fact that the prostrate
© 2012 The Linnean Society of London, Botanical Journal of the Linnean Society, 2013, 171, 429–440
DANISH SCOTCH BROOM – INVASIVE OR ENDANGERED?
form is only present in a more limited area, the
maximum distance between the populations of the
prostrate form was 58 km.
For comparison, one population from Italy was
genotyped. This population was collected in the forest
Macchia Grande di Manziana (Province of Rome) and
documented by photographs that suggest that the
sampled individuals are of an erect, vigorously
growing form. This population was collected by Laura
Celesti-Grapow, Università La Sapienza, Rome, Italy,
and after collection the samples were dried in silica
gel and sent to Denmark.
DNA
EXTRACTION
DNA material was purified using the DNeasy 96
Plant Kit (Qiagen, Hilden, Germany), using approximately 45 mg of frozen plant material (or 10 mg of
the silica-dried samples from Italy) and following
the DNeasy 96 protocol. After DNA extraction the
genomic DNA was stored at 5 °C.
GENOTYPING
Eight nuclear primers developed for Scotch broom,
Cs03, Cs18, Cs34, Cs39, Cs40, Cs42, Cs63 and Cs65
(Kang, Buckley & Lowe, 2007a), and four universal
plastid primer pairs, ccmp2, ccmp5, ccmp7 and
ccmp10 (Weising & Gardner, 1999) previously found
to be polymorphic in Scotch broom (Kang et al.,
2007b), were tested on samples from the Melby,
Villingerød, Henne Strand, Nymindegab and Mols
Bjerge populations to check for amplification and
polymorphism. One plastid primer pair, ccmp5,
and five nuclear primers, Cs03, Cs18, Cs34, Cs39 and
Cs65, that amplified well and were polymorphic were
chosen for genotyping. PCR amplification was conducted with minor modifications following the procedure in the Qiagen Multiplex PCR handbook except
that the reaction volume was scaled down to 15 mL
(1¥ multiplex mastermix, 0.2 mmol L-1 of each primer,
approximately 60 ng template DNA and H2O to make
final volumes of 15 mL). PCR was performed in a
GeneAmp 2700 (Applied Biosystems) using the following profile: initial denaturing at 95 °C for 15 min,
followed by 30 cycles of denaturing for 30 s at 94 °C,
annealing for 90 s at 57 °C and extension for 60 s at
72 °C. The 30 cycles were followed by a final 30-min
step at 60 °C and then storage at 5 °C. Separation of
diluted (1:20) PCR products with GeneScan 500 LIZ
as an internal standard was achieved on an Applied
Biosystems ABI 3130XL.
MORPHOLOGICAL
DATA
Two size measures were assessed for all genotyped
individuals: maximum height was measured from soil
431
surface to height of plant (rounded to the nearest
5 cm), and maximum width was measured horizontally across the centre of the plant (rounded to the
nearest 5 cm).
DATA ANALYSIS
GENETIC DIVERSITY AND HARDY–
WEINBERG PROPORTIONS
According to Kang et al. (2007b), C. scoparius is polyploid, but we observed multiple peaks hinting at
polyploidy in only seven of the 214 genotyped individuals. For each of these samples, three peaks were
observed for loci Cs18 and Cs34. Two individuals were
from Villingerød and five from Melby. These seven
samples were not further used in the study. Cs65 did
not amplify in two individuals (from Idom Hede and
Mols Bjerge), which could be due to null alleles given
that the other markers amplified well. The two individuals were included in the study but with missing
data for Cs65.
The level of genetic diversity was compared by
calculating the mean number of alleles (Na) across loci
and for each population as well as Nac, mean number
of alleles corrected for differences in sample size,
using the rarefaction method with the software
HP-Rare (Kalinowski, 2005).
To test for the presence of Hardy–Weinberg (HW)
proportions, the observed (Ho) and expected (He) heterozygosity (Nei, 1987) and Fis per population were
calculated using GenAlEx 6.2 software (Peakall &
Smouse, 2006). Tests for statistical significance were
performed as an exact test using Markov chains as
implemented in the Arlequin (vers. 3.11) software
(Excoffier, Laval & Schneider, 2005). The probability
values were adjusted by table-wide sequential Bonferroni correction (Rice, 1989) using Holm’s method.
POPULATION
STRUCTURE
Assessment of population structure was done by the
use of the following two methods: population structure was estimated using the software STRUCTURE
version 2.3.1 (Pritchard, Stephens & Donnelly, 2000),
which uses a model-based Bayesian method to assign
distinct individuals to a number of clusters on the
basis of their multilocus genotypes. Both the plastid
and the nuclear markers were used for this clustering
analysis. The admixture method was used, as there
was no information of priors and admixture between
populations was a possibility. The length of the
burn-in period was 104 and the number of Markov
chain Monte Carlo replicates was 104. Runs were
repeated 20 times for the chosen number of clusters
(K). The most likely number of clusters present in the
data, K*, was estimated based on the DK statistics
© 2012 The Linnean Society of London, Botanical Journal of the Linnean Society, 2013, 171, 429–440
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L. ROSENMEIER ET AL.
Table 1. Sample size, N, allelic diversity, Na, before and after rarefaction, observed (Ho) and expected heterozygosity (He)
and inbreeding coefficient, Fis, for all populations across five nuclear loci
Population
N
Na
Nac
Ho*
He*
Fis*
Henne Strand
Lystbæk
Nr. Nebel
Nymindegab
Melby
Mols Bjerge
Villingerød
Svanninge Bjerge
Idom Hede
Italy
20
24
24
24
17
20
24
21
20
20
6.40 (1.66)
9.40 (2.23)
6.00 (1.64)
6.20 (2.04)
5.60 (1.03)
5.80 (1.88)
10.40 (1.50)
12.20 (1.77)
9.40 (1.66)
9.80 (1.20)
6.00 (1.48)
8.02 (1.76)
5.21 (1.35)
5.57 (1.68)
5.60 (1.0.3)
5.60 (1.77)
9.04 (1.23)
11.45 (1.54)
9.02 (1.66)
9.38 (1.16)
0.700 (0.087)
0.656 (0.046)
0.458 (0.059)
0.458 (0.105)
0.515 (0.139)
0.413 (0.155)
0.698 (0.079)
0.750 (0.023)
0.688 (0.063)
0.713 (0.031)
0.632 (0.069)
0.712 (0.073)
0.494 (0.066)
0.495 (0.094)
0.590 (0.100)
0.456 (0.113)
0.724 (0.080)
0.836 (0.030)
0.732 (0.081)
0.792 (0.027)
–0.102 (0.032)
0.055 (0.091)
0.064 (0.053)
0.050 (0.118)
0.197 (0.142)
0.190 (0.138)
0.028 (0.064)
0.099 (0.044)
0.040 (0.044)
0.094 (0.068)
Values are mean (SD).
*Calculations of Ho, He and Fis are based on four loci because of null alleles in CS 65.
(K = 1–6) as described by Evanno, Regnaut & Goudet
(2005). The results from STRUCTURE were compared with an unrooted neighbour-joining (NJ) tree
based on Nei’s genetic distance using the NJ method
of Saitou & Nei (1987) as implemented in MEGA
version 5 (Tamura et al., 2011). The level of genetic
differentiation for groups defined by STRUCTURE
was quantified with Wright’s F-statistics estimated
according to Weir (1990) as implemented in GenAlEx
6.2 (Peakall & Smouse, 2006).
ANALYSIS
OF HEIGHT–WIDTH RATIOS
To determine whether the genetically defined groups
reflected differences in growth habit, as described by
Böcher & Larsen (1958), the following approach was
taken: on the basis of the STRUCTURE assignment
of individuals to clusters (K = 2, i.e. an estimated
genetic structure consisting of two populations), we
compared the height–width relationship for individuals assigned with confidence to either of the two
different groups. Twenty individuals out of the 195
individuals had an admixture coefficient q1 between
0.20 and 0.80, and these were excluded from this
analysis because they might be results of gene flow
between two gene pools. For the remaining 175 individuals we applied a simple regression model
Hij = ai + bi Wij + eij, where Hij and Wij are height and
width of the jth individual from either of the two
groups (i = 1, 2), to test if the slopes (bi) were significantly different for the two clusters. Distinct differences in slope (bi) would separate erect (tall and
narrow) from low types (short and wide). Based on
fitted linear relationships between height and width
within groups, we estimated 95% confidence intervals
for the prediction of height (H) based on width (W) for
each group. The residuals eij were assumed to be
independent and normally distributed. We did not
include a population effect in this analysis because we
wanted variation between populations to be included
as noise in the model. We applied SAS ver. 9.2 procedure GLM (SAS, 2008) for these analyses.
GENETIC
RESULTS
HW
DIVERSITY AND
PROPORTIONS
Expectation of HW proportions for nuclear markers
was in general accepted with the exception of locus
Cs65 where genotypes differed significantly from HW
proportions in three (Melby, Villingerød and Idom
Hede) out of ten examined populations after sequential Bonferroni corrections. This locus had a surplus
of homozygous individuals, probably due to null
alleles, as also indicated by the lack of amplification
in two individuals. Locus Cs03 had genotypes that
deviated significantly from HW proportions in one
population (Melby); HW proportions were not rejected
after Bonferroni corrections for the remaining loci
and populations.
The mean number of alleles per locus for the
nuclear markers ranged from 5.60 (Melby) to 12.20
(Svanninge Bjerge) (Table 1). The numbers did not
differ much after sample size corrections, from 5.60
(Melby) to 11.45 (Svanninge Bjerge). The Italian
population had an average number of alleles
(Nac = 9.38) that was within the range observed in
Danish populations. The average inbreeding coefficient (Fis) did not differ significantly from 0 in the ten
populations when Cs65 was left out of the calculations (Table 1).
Allele frequencies for the plastid microsatellite
marker are shown in Table 2. The Danish populations
are in most cases either fixed or close to being fixed
for one allele (114 or 115 bp). The Svanninge Bjerge
© 2012 The Linnean Society of London, Botanical Journal of the Linnean Society, 2013, 171, 429–440
DANISH SCOTCH BROOM – INVASIVE OR ENDANGERED?
433
Table 2. Allele frequencies for the plastid marker Ccmp 5
Allele
length (bp)
Henne
Strand
Lystbæk
Nr.
Nebel
Nymindegab
Melby
Mols
Bjerge
Villinge-rød
Svanninge
Bjerge
Idom
Hede
Italy
110
112
113
114
115
116
117
118
0.000
0.000
0.000
0.000
1.000
0.000
0.000
0.000
0.000
0.000
0.000
0.042
0.917
0.000
0.042
0.000
0.000
0.000
0.000
0.000
1.000
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.917
0.000
0.083
0.000
0.000
0.000
0.000
0.941
0.000
0.059
0.000
0.000
0.000
0.000
0.000
0.850
0.150
0.000
0.000
0.000
0.000
0.000
0.000
1.000
0.000
0.000
0.000
0.000
0.048
0.000
0.095
0.714
0.095
0.000
0.000
0.048
0.000
0.000
0.000
1.000
0.000
0.000
0.000
0.000
0.000
0.050
0.000
0.200
0.750
0.000
0.000
0.000
Most frequent alleles per population are in bold type.
Figure 1. Bayesian analysis of the genetic structure of Scotch broom based on one plastid and five nuclear microsatellite
loci. The q1 admixture coefficient is presented in shaded colour, and q2 = 1 – q1 is in bright colour.
population is the most variable, showing five alleles
in the 21 genotyped individuals. The two most
common alleles in the Italian population are also 114
and 115 with frequencies of 0.2 and 0.75, respectively.
POPULATION
STRUCTURE
The STRUCTURE analysis based on nuclear and
plastid markers suggested the presence of two gene
pools (K = 2) based on the approach of Evanno et al.
(2005). Below we refer to these groups as ‘N’ (with q1
admixture coefficients close to 1) and ‘I’ (with q1
admixture coefficients close to 0). Admixture coefficients for all individuals are presented as a bar
diagram in Figure 1. In general, the grouping corresponded closely to populations. Individuals from the
non-invasive Henne Strand, Lystbæk, Nymindegab
and Nr. Nebel populations were mainly of the ‘N’ type
(admixture coefficient q1 above 0.8), whereas individuals from the remaining five populations (Idom Hede,
Melby, Mols Bjerge, Villingerød and Svanninge
Bjerge) were mainly of the ‘I’ type (admixture coefficient q1 below 0.2). However, most populations
included a few individuals that deviated from the
general pattern. This was especially the case for Mols
Table 3. Average admixture coefficient (q1) of individuals
from each of ten runs (K = 2)
Population
Nuclear
markers
Nuclear + plastid
markers
Henne Strand
Lystbæk
Nr. Nebel
Nymindegab
Melby
Mols Bjerge
Villingerød
Svanninge Bjerge
Idom Hede
Italy
0.91
0.83
0.95
0.92
0.04
0.30
0.11
0.08
0.09
0.03
0.95
0.91
0.96
0.91
0.02
0.20
0.05
0.10
0.06
0.09
Bjerge (Fig. 1). The Italian population grouped
together with the Danish ‘I’ type populations.
Running STRUCTURE without the locus with null
alleles (Cs65) did not change the signal (data not
shown). Table 3 presents the average admixture coefficient q1 of individuals from each of the populations.
The population from Nr. Nebel had the highest
© 2012 The Linnean Society of London, Botanical Journal of the Linnean Society, 2013, 171, 429–440
434
L. ROSENMEIER ET AL.
Figure 2. Map showing sampling locations and proportion of membership of each pre-defined population in each of the
two clusters based on five nuclear and one plastid microsatellite marker. Proportion of membership = mean admixture
coefficient of all individuals in population. Key: dark grey = group ‘N’, white = group ‘I’.
average q1 value of 0.95 (based on nuclear markers
only) which indicates that the individuals from this
population were almost purely identified as being of
‘N’ origin. In contrast, the Melby population had the
lowest average q1 value of 0.05 (nuclear markers only)
indicating that these individuals predominantly represent an ‘I’ origin. Other populations were slightly
more intermediate, but all showed clear membership
to one of the two groups. The inclusion of the plastid
marker increased this pattern with the exceptions of
Svanninge Bjerge and, in particular, Italy. In Figure 2
the geographical positions of the Danish populations
are shown as pie charts based on mean admixture
scores for each population.
An unrooted NJ-tree based on genetic distances
using all markers is presented in Figure 3. This supports the grouping of Danish populations found by the
STRUCTURE analysis. The main difference is the
Italian population, which is located on its own node
between the two Danish groups. However, when the
plastid microsatellite marker is excluded from the
analysis the Italian population is found among the
Danish vigorous ‘I’ type populations (results not
shown).
Individuals from predominantly ‘N’ type populations (Henne Strand, Lystbæk, Nymindegab, Nr.
Nebel) had slightly lower gene diversity than those
from predominantly ‘I’ type populations (Idom Hede,
Melby, Mols Bjerge, Villingerød, Svanninge Bjerge)
(Table 4). The level of population differentiation (Fst)
within groups was similar (‘N’: 0.11; ‘I’: 0.15), but
inbreeding coefficients (Fis) were in general slightly
higher for ‘I’ type populations (Fis = 0.08) than for ‘N’
type populations (Fis = 0.02).
HEIGHT–WIDTH RATIOS AMONG ‘N’
‘I’ TYPE INDIVIDUALS
AND
Based on the 85 samples with q1 values > 0.8 (type ‘N’
individuals) and the 90 samples with q1 values < 0.2
(type ‘I’ individuals), the height–width slopes (bi) were
found to be significantly different between ‘N and ‘I’
© 2012 The Linnean Society of London, Botanical Journal of the Linnean Society, 2013, 171, 429–440
DANISH SCOTCH BROOM – INVASIVE OR ENDANGERED?
Nymindegab
Lystbæk
435
the genetic groups. The 20 samples that were intermediate between groups (0.2 < q1 < 0.8) showed no
clear picture.
Henne strand
Nr. Nebel
DISCUSSION
Italy
RELATIONSHIP
BETWEEN INVASIVE BEHAVIOUR,
GROWTH FORM AND GENETIC GROUPS
Mols Bjerge
Idom Hede
Villingerød
Svanninge Bjerge
Melby
0.1
Figure 3. Unrooted neighbour-joining tree using data
from both the plastid marker and the nuclear markers,
based on Nei’s genetic distance.
type individuals (P < 0.001). The estimated slope was
bN = 0.196 (0.030) for the ‘N’ group, but four times
higher bI = 0.796 (0.060) for the ‘I’ group (see also
Fig. 4).
It is clear from Figure 4 that the ‘N’ type individuals in general correspond to a prostrate growth form,
and no members of this group were taller than 1.1 m,
whereas several individuals of the ‘I’ type were taller
than 2.5 m. The estimated 95% confidence intervals
for prediction of height from width for each group are
also shown in Figure 4. For individuals < 1.5 m wide
or < 1 m high, the upper 95% interval of type ‘N’
individuals overlaps with the lower 95% interval of
type ‘I’ individuals. However, for individuals > 1 m
high or > 1.5 m wide, the width–height ratio allows
quite precise prediction of type, with erect individuals
being ‘I’ type and low individuals being ‘N’ type. This
means that once the individuals have obtained some
age (size), they differentiate into one of the two
growth forms, and these growth forms correspond to
The genetic markers separated individuals into two
genetic groups, ‘N’ and ‘I’. The ‘N’ gene pool is characteristic for four of the studied populations from
central/western Jutland: Henne Strand, Nr. Nebel,
Nymindegab and Lystbæk. These sites were selected
a priori due to expected occurrence of non-invasive
populations of the species. The five populations dominated by ‘I’ type individuals are spread across the
country: Mols Bjerge, Svanninge Bjerge, Melby, Villingerød and Idom Hede. The Idom Hede population
is located near to the Lystbæk population from group
‘N’ (see Fig. 2), but is clearly of type ‘I’. The two
genetic types are characterized by different growth
form in terms of height/width ratio, with ‘I’ types
being the highest. The populations reported to show
invasive behaviour (Mols Bjerge and Svanninge
Bjerge) are dominated by individuals of the erect type
belonging to genetic group ‘I’. The results of the
present study thus support our initial hypothesis that
Danish populations of Scotch broom seem to consist of
two gene pools, of which only one is characteristic for
populations with invasive behaviour. Moreover, the
results suggest that the height/width ratio will allow
an easy assignment of individuals to a gene pool.
Exceptions will be small (young or browsed) plants
and plants assigned an intermediate status according
to the genetic structure.
TAXONOMIC
STATUS OF
DANISH SCOTCH
BROOM
Our results reject the suggestion by Buchwald (2008)
that Scotch broom in Denmark comprises a single
morphologically variable species. The two revealed
gene pools correspond to the presence of two taxonomic units of Scotch broom in Denmark as acknowledged by Böcher & Larsen (1958). They described one
group (with two ecotypes from Denmark) as more or
less erect, 150–250 cm high, where one ecotype had a
low degree of frost hardiness and the other an intermediate level. Böcher & Larsen (1958) believed that
both of these ecotypes had been introduced to
Denmark and then became naturalized. Our genetic
group ‘I’ fits well into this group because of its erect
growth habit and height up to 300 cm and its invasive
behaviour in some areas. Furthermore, our genetic
type ‘N’ corresponds well to ecotype B (2) of Böcher &
Larsen (1958), which measured 50–100 cm in height
and had a high degree of winter hardiness. The
© 2012 The Linnean Society of London, Botanical Journal of the Linnean Society, 2013, 171, 429–440
436
L. ROSENMEIER ET AL.
Table 4. F-statistics and gene diversity for the grouped Danish populations: total gene diversity, Ht, mean expected
heterozygosity across populations, He, mean observed heterozygosity across populations, Ho, inbreeding coefficient, Fis,
overall inbreeding coefficient, Fit, fixation index, Fst
Group ‘N’
Mean
Group ‘I’
Locus
Ht
He
Ho
Fis
Fit
Fst
Cs03
Cs18
Cs34
Cs39
Cs65
0.737
0.725
0.709
0.466
0.888
–
0.839
0.874
0.805
0.615
0.920
–
0.657
0.625
0.622
0.429
0.832
–
0.747
0.716
0.672
0.527
0.844
–
0.679
0.567
0.567
0.460
0.746
–
0.743
0.597
0.614
0.496
0.427
–
–0.033
0.093
0.088
–0.073
0.104
0.019 (0.042)
0.006
0.166
0.087
0.060
0.494
0.080 (0.033)
0.079
0.218
0.201
0.012
0.160
0.128 (0.049)
0.115
0.316
0.238
0.194
0.536
0.216 (0.042)
0.109
0.137
0.124
0.079
0.063
0.112 (0.012)
0.110
0.181
0.165
0.143
0.083
0.150 (0.015)
Cs03
Cs18
Cs34
Cs39
Cs65
Mean
Group‘N’: Henne Strand, Lystbæk, Nymindegab and Nr. Nebel. Group ‘I’: Melby, Mols Bjerge, Svanninge Bjerge, Idom
Hede, Villingerød. Cs65 is excluded from averages due to null alleles.
ecotype and geographical distribution of our populations from this ‘N’ gene pool also corresponds to the
description from Böcher & Larsen (1958), stating that
this low type is found on sandy hillsides in a
restricted area of central Jutland. They believed that
this type should have at least varietal status. Our
study confirms that some of the Scotch broom populations from Jutland represent a different genetic
stock than the remaining Danish populations. We
suggest that there are two taxa of Scotch broom in
Denmark and agree with Böcher & Larsen (1958)
that attention should be given to the populations of
Scotch broom with the characteristics of the ‘N’ type
described in the present study, today probably only
found in parts of Jutland.
ORIGIN
OF
DANISH SCOTCH
BROOM GENE POOLS
It is interesting that the genotyped Italian population
cluster together with the Danish type ‘I’ populations
with vigorous growth in the STRUCTURE analysis
(Fig. 1). This matches the frequently raised hypothesis that the vigorous populations of Danish Scotch
broom are likely to represent successors of plants
introduced from the Mediterranean area, whereas the
short types are native. However, Scotch broom can
show invasive behaviour even in areas where it is
considered to be native (Rousseau & Loiseau, 1982;
Prevosto et al., 2006), and management practices
such as grazing and burning seem to trigger this
invasiveness (Rousseau & Loiseau, 1982; Rees &
Paynter, 1997; Paynter et al., 1998). The short form
with the restricted distribution area is most probably
native to Denmark, but the status of the vigorously
growing form is still unclear.
Similarly to Kang et al. (2007b), we find high levels
of genetic diversity in the putative invasive populations (our ‘I’ type). In fact, gene diversity (He) is in
general a little higher at population level and at
group level compared with the ‘N’ gene pool. The ‘N’
type populations are relatively small and isolated and
may have lost genetic diversity due to genetic drift. If
the ‘I’ type does consist of introduced and naturalized
populations, the high levels of diversity would suggest
multiple independent introductions based on several
seed lots.
The relatively high level of Fis in the ‘I’ type
populations suggest to us that the invasive dynamics and corresponding recent colonization that have
taken place in these populations could have created
small-scale spatial structures which could have generated Wahlund effects (Wahlund, 1928). However,
our sampling design does not allow us to explore
this hypothesis further.
FUTURE
STATUS OF THE
DANISH SCOTCH
BROOM
The two gene pools of Scotch broom discovered here
are well separated based on a relatively limited
number of markers. There is only little indication of
gene transfer between the two groups. The lack of a
potential hybrid swarm may, however, be biased by
our sampling as we did not include localities in
between eastern and western Denmark.
In our study, we have only two geographically close
populations that were classified into different groups:
© 2012 The Linnean Society of London, Botanical Journal of the Linnean Society, 2013, 171, 429–440
DANISH SCOTCH BROOM – INVASIVE OR ENDANGERED?
437
400
350
300
Height (cm)
250
TYPE I
200
Type I, Lower 95%
150
100
Type N, Upper 95%
50
TYPE N
0
0
50
100
150
200
250
300
350
400
Width (cm)
Figure 4. Height–width relationship for individuals assigned with confidence to either of the two gene pools. Type N (䉱)
corresponds to individuals with admixture coefficients (q1) > 0.8 and Type I (䊐) to those with q1 < 0.2. The estimated 95%
confidence intervals for prediction of height from width for each group are shown by solid lines.
the ‘N’ type (putative native) Lystbæk population
located fairly close to the ‘I’ type Idom Hede population. Due to their proximity, the Lysbæk population
may have experienced gene flow from the vigorous ‘I’
type. It was the ‘N’ type population that was estimated to have the lowest degree of admixture with
q1 = 0.83 (Table 3). It also had the same average
number of alleles per locus (nuclear markers) as Idom
Hede, two or three more alleles per locus than the
other ‘N’ type populations (Table 1). In addition,
Lystbæk was the only population from the ‘N’ group
where we found the plastid marker allele 114, which
is the most common for ‘I’ type populations (Table 2).
As the plastid marker is maternally inherited in
Scotch broom as in most angiosperms (Aguinagalde
et al., 2005; Petit et al., 2005), some of the putative
gene flow is likely to have been via seed. Looking at
‘I’ type populations, Mols Bjerge has the highest estimated admixture with the ‘N’ types. As the seed of
Scotch broom is viable for a long time in the seed
bank (Turner, 1933) it is possible that the relatively
high admixture coefficient (‘N’ type) reflects the remnants of a former more common presence of short
Scotch broom in Mols Bjerge.
Crosses between C. scoparius (s.l.) and congeners
have been produced for ornamental reasons (Peterson
© 2012 The Linnean Society of London, Botanical Journal of the Linnean Society, 2013, 171, 429–440
438
L. ROSENMEIER ET AL.
& Prasad, 1998) and suggest that interspecific barriers are weak or non-existent. Furthermore, both types
flower during the same time period (May and June)
and are likely to share pollinators. The clear genetic
separation of the two taxa with only few indications of
gene exchange could be due to slight differences in
ecological preferences as reported by Citharel & Citharel (1986) for C. scoparius and C. scoparius subsp.
maritimus (Rouy) Heywood. Controlled transplant
experiments with Danish ‘I’ type seed and seedlings
into ‘N’ type populations could reveal if the case for
Danish Scotch broom is similar. The clear separation
of the two Danish gene pools could also be due to the
history with no or only little contact between the two
types. All but one of the studied ‘N’ type populations
were located at remote sites on the west coast of
Jutland. In contrast, the ‘I’ type populations, except
Idom Hede, were located further to the east. The
vigorous type may reach the isolated areas in western
Jutland relatively soon, because the species is known
for rapid expansion during early colonization (Parker,
2000; Magda et al., 2009). Whether the vigorous form
will swamp the non-vigorous one, as seen in other
native–invasive species pairs (Daehler & Strong,
1997; Prentis et al., 2007), is unknown, but the risk of
genetic erosion calls for intensified genetic conservation effort.
CONCLUSION
The original question for this study was whether
Scotch broom in Denmark is an invasive plant or a
vulnerable native species. Our results suggest both.
We find two distinct gene pools that are morphologically separable by plant size. Populations dominated
by the tall type sometimes show invasive behaviour,
whereas short individuals do not. We find that at
present it is still possible to identify populations that
are almost purely of the short type. However, given
the high multiplication rate of C. scoparius s.l., we
suspect that the tall, putatively introduced form will
expand to areas where the short form is currently the
dominant form and perhaps exclude it unless they
have clear divergent habitat preferences. Identification of populations for in situ conservation in a
network of genecological zones (as outlined in
Graudal et al., 1995) can be based on screening of
growth forms, and verified by the genetic tool box
applied in the present study to reveal and quantify
indications of introgression.
At present it is a conservation paradox that expansion of Scotch broom populations with invasive behaviour may present a genetic conservation risk to native
Scotch broom populations. It is important to give the
short form a formal taxonomic status as either a
subspecies or species in order to ensure the initiation
of conservation activities to protect the native taxon.
ACKNOWLEDGEMENTS
We thank Hans Peter Ravn and Laura CelestiGrapow for providing material of Scotch broom from
Italy. Rita Buttenschøn and Poul Hald-Mortensen
provided information on Scotch broom locations in
Denmark. Lea Vig McKinney is thanked for help with
the DNA extractions and Ida Hartvig Larsen for valuable comments on the manuscript.
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SUPPORTING INFORMATION
Additional Supporting Information may be found in the online version of this article:
Table S1. GPS coordinates for each locality.
© 2012 The Linnean Society of London, Botanical Journal of the Linnean Society, 2013, 171, 429–440