Download (Primulaceae), a Wild

Annals of Botany 89: 559±562, 2002
doi:10.1093/aob/mcf083, available online at www.aob.oupjournals.org
Seed Germination and Reproductive Features of Lysimachia minoricensis
(Primulaceae), a Wild-extinct Plant
 1 , * and MA R IA MA Y O L 2
JOS E P A . R OSS E L L O
1JardõÂn Bota
 nico de Valencia, Universidad de Valencia c/Quart 80, E-46008, Valencia, Spain and 2Centre de Recerca
EcoloÁgica i Aplicacions Forestals (CREAF), Universitat AutoÁnoma de Barcelona, E-08193 Bellaterra, Barcelona,
Spain
Received: 25 May 2001 Returned for revision: 2 September 2001 Accepted: 3 December 2001
Lysimachia minoricensis is one of the few Mediterranean endemic plants (Minorca, Balearic islands) that has
gone extinct in the wild but which persists as extant germplasm or cultivated plants in several botanical gardens.
Reproductive features (seed set, number of seeds per capsule, seed weight) and germination responses to
constant temperatures, sea water and dry-heat pre-treatments were investigated to determine the extent to which
they may have in¯uenced the extinction of the species. Seed set in Lysimachia is not dependent on pollinators,
suggesting a functional selfer breeding system. Most plants produced a large mean number of fruits (23´2) and
seeds (46´6), and the mean production of seeds per individual was estimated to be almost 1100. Overall, no
highly speci®c requirements were observed for seed germination. Seed germination was not inhibited in the
dark, and a high germinability (over 87 % in all cases) was recorded in most experiments, with the exception of
those performed at low temperatures (5 and 10 °C). These data suggest that fertility and seed viability were not
the major causes of extinction. The high reproductive performance of L. minoricensis is in striking contrast to
its status as a wild-extinct plant, suggesting that extrinsic factors were responsible for its extinction.
ã 2002 Annals of Botany Company
Key words: Seed germination, Mediterranean ¯ora, extinct taxa.
INTRODUCTION
Islands are fragile ecosystems that often have a
remarkably high number of endemic taxa, most of
them being con®ned to a few geographically very
restricted populations (Carlquist, 1974). The risks of
extinction for plant and animal island species are usually
higher than those for other taxa growing in continental
areas (Alcover et al., 1999; Hylton-Taylor, 2000).
Amongst the 30-plus species of vascular plants to
have gone extinct in the Mediterranean basin in the
last two centuries (Greuter, 1994) were three insular
endemics: Diplotaxis siettiana Maire (Alboran islet),
Dianthus multinervis Vis. (Jabuka islet) and Lysimachia
minoricensis J.J. Rodr. (Minorca). Fortunately, two of
these wild-extinct taxa (D. siettiana and L. minoricensis)
persist in cultivation and/or seed banks (Greuter, 1994).
The reasons for the extinction of L. minoricensis are not
clear. The plant was believed to be very rare since its
description (RodrõÂguez, 1868), and it was assumed to have
disappeared from the single known location between 1926
and 1950 (IbaÂnÄez et al., 1999). Molecular screening (using
isozyme and RAPD markers) of germplasm accessions of
L. minoricensis from 12 European botanical gardens has
shown that the remaining living stocks of the species are
genetically depauperate since no variation was detected
(Calero et al., 1999; IbaÂnÄez et al., 1999). Several attempts
aimed at reintroducing L. minoricensis in the wild have
* For correspondence. Fax: +34 963156826, e-mail [email protected]
failed (GoÂmez Campo, 1987; Mayol, 1997), but whether or
not this is due to some underlying biological factor is
unclear.
Molecular analysis of extant Lysimachia accessions
cannot determine whether the amount of genetic variability
played any role in the decline of the species in the wild.
Survival of endangered species may not necessarily be
compromised by the absence of signi®cant levels of genetic
diversity, and low levels of genetic variation are not
necessarily associated with low population viability
(Arden and Lambert, 1997).
The low genetic variability (and thus reduced potential to
adapt to changing environmental conditions) and/or
inbreeding due to sel®ng or a limited number of mating
partners could have decreased the ®tness of the ex situ
cultured populations of L. minoricensis, lowering plant
performance through a reduction in growth, survivorship,
seed set, total mass of seeds, average seed mass and/or
percentage of germination. Germination requirements and
germination rate are amongst the most important seed traits
associated with plant ®tness (Harper, 1977), and are key
components in the ecology and evolution of plant life
histories. Knowledge of the performance of these stages
may help ascertain whether failure in germination success
has contributed to population decline in endangered plant
species. Unfortunately, there are no data concerning
any critical elements of the reproductive biology of
L. minoricensis. These data are essential, not only to
improve recovery programmes, but also to determine the
ã 2002 Annals of Botany Company
Rossello and Mayol РSeed Germination in Lysimachia
560
extent to which they may have in¯uenced the wild
extinction of this species.
MA TE R IA L S A N D ME T H O D S
Seeds of L. minoricensis were obtained from ten botanic
gardens (Cambridge, Brest, Berlin, Paris, Belgium, Nancy,
CoÂrdoba, SoÂller, Copenhagen and Edinburgh) and grown in
a glasshouse at Valencia University, Spain. Ultimately, all
accessions probably derived from a single source (Calero
et al., 1999; IbaÂnÄez et al., 1999). Several reproductive
features (number of capsules per scape, number of seeds per
capsule, seed weight) were recorded at the fruiting stage. To
avoid any bias concerning the origin of the samples in the
germination experiments, seeds were collected from plants
belonging to all accessions and pooled together. Seeds were
air-dried immediately upon collection and then stored at
room temperature (20 °C) until they were used in the
germination tests. Seeds were placed on wet ®lter paper in
Petri dishes and then placed in a germination chamber. First,
the responses of seeds to various constant temperatures
(5, 10, 15, 20, 25 and 30 °C) were investigated. The
in¯uence of ®re as a possible inhibitor of germination in
Lysimachia was investigated by pre-treating seeds at 80 °C
for 10, 60 and 90 min and recording subsequent germination
success at 25 °C. For salt tolerance tests, seeds were imbibed
in sea water for 1, 3 or 10 d before being allowed to
germinate at 25 °C. Given the high percentage of seed
germination, no further treatments involving lower salt
concentrations were attempted. For each germination test,
two batches of 50 seeds were used. Seeds were considered to
have germinated when the radicle penetrated the seed coat.
Germination was recorded every day for 30 d.
Data analysis
Germination data were arcsine-transformed and analysed
using ANOVA. Tukey's LSD multiple comparison was
used to test for signi®cant differences among treatment
means. The following parameters were determined: germinability (G), or percentage of germinating seeds; time to ®rst
observed germinant (T); number of days required for 50 %
of the total number of seeds to have germinated (T50);
number of days for 50 % of the total number of seeds
germinated (T¢50); time to maximum germination (T100);
germination rate (GR) or germinants per day, calculated as
the maximum number of germinants/(days to maximum
germination ± days to initial germination); index of
germination rate (IGR) = (100/n) S gi/ti, where n is the
number of seeds incubated, and gi is the number of seeds
germinated at time ti.
R E SU L T S
Reproductive features
Plants of L. minoricensis are biennial and usually have a
single ¯owering stem per individual. However, departures from this pattern are sometimes observed. Some
plants ¯owered in the ®rst year; exceptionally, some did
not die after ¯owering and so attained a second
¯owering season; lastly, up to two or three scapes
were observed in some large individuals. Isolated plants
(grown in an insect-free glasshouse) regularly developed
fruits in all ¯owers of the scape (Table 1), suggesting
that within-¯ower spontaneous sel®ng (autogamy) is the
main breeding system of L. minoricensis. Anthers and
stigmas close together at maturity, and anther dehiscence
occurs in unopened ¯ower buds, but we have failed to
verify pollen tube growth occurring at this stage. No
experiments were conducted to test whether or not the
high fruit set observed in this species was due to
apomixis. However, this seems unlikely since, as far as
we know, no apomictic taxa of Primulaceae have been
detected. Reproductive features concerning fruit set, seed
set, fruit number and seed weight are shown in Table 1.
Morphologically, the seeds conformed to earlier descriptions, and the mean weight of batches of 50 seeds
indicated that a single seed weighs as little as 0´20 mg
on average. From the values shown in Table 1 it is
estimated that the mean seed production per individual
is approx. 1100.
Germination rates
Results of the germination experiments are shown in
Table 2. Good germination success was obtained for most
treatments. Germination of Lysimachia seeds was neither
inhibited nor signi®cantly reduced when seeds were incubated in darkness. High germination rates were also
observed when additional temperature tests were conducted
under light (data not shown), suggesting that germination is
indifferent to light. Germinability was poor at the lower
temperatures tested (5 and 10 °C). No signs of germination
were observed after 30 d at 5 °C, but at 10 °C germination
started after 28 d. When the seeds were incubated with a
thermogradient between 15 and 30 °C germinability values
were over 98 %, and germination started between 5 and 12
d, with T50 between 9 and 14 d, and T100 within 22 d.
Germination rates were also high at 15 and 20 °C; at these
temperatures maximum germination occurred 6±8 d after
sowing.
Incubation of Lysimachia seeds in sea water for up to 10 d
had only a minor effect on germination rates. Statistically
non-signi®cant lower germination percentages were obtained for pre-soaked seeds (mean 88´3 % compared with
99 % for control seeds germinated at 25 °C). The lowest
values were recorded when seeds were immersed in sea
water for 24 h only.
The three pre-treatments did not signi®cantly decrease
the germination rate when the test was conducted without
immersion in sea water. On the contrary, the IGR index
increased in all pre-treatments, which indicates faster
germination regimes. Pre-heating the seeds at 80 °C resulted
in high germinability values (over 90 %) which were not
signi®cantly different from control values, irrespective of
the time of exposure to high temperatures. Pre-heating also
increased the speed of germination, as shown in Table 2.
The results after dry-heat treatments were similar to those
obtained when the seeds were imbibed in sea water.
Rossello and Mayol РSeed Germination in Lysimachia
D IS C U S SIO N
dra; Sytsma and Smith, 1992). Some ¯oral features
displayed by L. minoricensis seem to represent derived
traits (such as small corolla lobes, inconspicuous petal
pigmentation and a related autogamous breeding system),
which may indicate a young lineage. However, no
phylogenetic analysis of sect. Ephemerum (Reichb.) Endl,
in which L. minoricensis is currently included (Pax and
Knuth, 1905), has yet been performed.
Our reproductive and germination results do not help
explain the decline of L. minoricensis or its limited range.
Seed production in this species is high (best estimates
suggest that a single plant can produce up to 3300 seeds) and
should be free of year-to-year ¯uctuations (in the absence of
¯ower or fruit predators) through its sel®ng breeding
system. The high germinability obtained under different
temperature, salt and dry-heat regimes is unexpected for
such a narrowly restricted plant. The germination requirements of Lysimachia species have been studied rarely and,
as far as we know, are available for just a single annual herb
growing in central Africa and Afghanistan, L. ruhmeriana
Vatke (Teketay, 1998). These results are in striking contrast
with those for L. minoricensis. Germinability was less than
50 % in L. ruhmeriana at the different constant temperatures
assayed (between 10 and 30 °C), but better results were
achieved when diurnally alternating temperatures (20/12 °C
and 30/12 °C) were used. Furthermore, L. ruhmeriana did
not germinate in darkness, and when seeds previously
incubated in the dark were exposed to light germinability
increased very little, suggesting the induction of secondary
dormancy in this species (Teketay, 1998). The ability to
germinate over a wide range of temperatures, as is the case
for both Lysimachia taxa, has been associated with species
in which water supply is the main determinant of the timing
of germination in the ®eld (Grime et al., 1981). Since our
There is no evidence that a drastic decline in the distribution
range and population size of L. minoricensis has occurred in
recent times since its discovery. The species is known from
only a single location (Sa Vall Valley, Minorca), where it
was reported as very rare by RodrõÂguez (1878), and later by
other botanists (Porta, 1886; Knoche, 1921) who saw and
collected the plant. Although collecting individuals for
scienti®c collections was probably a major cause of the
extinction of L. minoricensis in the wild, it is likely that this
merely accelerated the inevitable decline of this endemic
lineage. All authors dealing with the taxonomic relationships of L. minoricensis have stated that it belongs to an old
lineage (Knoche, 1921; Contandriopoulos and Cardona,
1984) on the basis that no close relatives are known.
However, evolutionary changes leading to rapidly derived
morphological divergences could have obscured sister
relationships, as demonstrated by taxa showing a high
level of morphological change (e.g. Heterogaura heteran-
TA B L E 1. Reproductive features in L. minoricensis
Reproductive feature
Mean 6 s.d. (range)
Fruits/¯ower (n = 100)
Fruits/scape (n = 30)
Seeds/fruit (n = 50)
Seed weight (n = 18)
561
160
23´23 6 11´53 (6±56)
46´58 6 6´72 (30±60)
10´47 6 0´97 (8±12´5)
The fruits/¯ower ratio was recorded in individuals grown in an insectfree glasshouse, the other parameters being measured on plants cultured
in an open glasshouse. n = sample size. Seed weight refers to batches of
50 seeds.
TA B L E 2. Accumulated germination percentage (and allied parameters) of L. minoricensis seeds incubated in darkness at
various constant temperatures (T5°C±T30°C), after sea water (SW1d±SW10d) and dry-heat (DH10 m±DH90 m) pre-treatments 30 d
after sowing
Temperature
T 5°C
T 10°C
T 15°C
T 20°C
T 25°C
T 30°C
Sea water pre-treatment
SW 1 d
SW 3 d
SW 10 d
Dry-heat pre-treatment
DH 10 m
DH 60 m
DH 90 m
G
T
T50
T¢50
T100
GR
IGR
0a
9a
98b
100b
99b
98b
±
28a
12b
7c,d
5c,d
7d
±
±
14a
9b,c
9b,c
11a,b
±
29a
14b
9c,d
9c,d
11b,c
±
30a
18c,d
15d,e
15d,e
22b,c
0
4´50
16´33
12´50
9´90
6´53
0
0´32
6´79
10´85
11´21
8´75
75b
97b
93b
5c
4c
4c
6b,c
5c
7c
6c,d
5d
7d
17b,c,d
25d,e
15e
6´25
4´61
8´45
12´59
18´19
12´35
98b
98b
92b
5c
4c
4c
8c
6c
6b,c
8d
6d
6c,d
19c,d
14b
10d,e
7´00
9´80
15´33
13´40
17´28
15´20
G, Percentage of germinating seeds; T, time to ®rst observed germinant; T50, number of days until 50 % of the initial seeds had germinated; T¢50,
number of days for 50 % of the actual number of seeds germinated; T100, time to maximum germination rate; GR, maximum number of germinants/
(days to maximum germination ± days to initial germination); IGR, (100/n) S gi /ti, where n is the number of seeds incubated, gi is the number of
seeds germinated at time ti. For each parameter, values with the same superscripts are not signi®cantly different at the 1 % level of probability as
determined by Tukey's test.
562
Rossello and Mayol РSeed Germination in Lysimachia
germination experiments were conducted under saturated
water conditions, soil moisture content could be a key factor
in regulating germination responses in L. minoricensis
under natural conditions.
Laboratory germination studies may not adequately
predict the ecology of germination in the ®eld (Baskin and
Baskin, 1998). However, although the levels of germination
in L. minoricensis obtained in these experiments might not
be attainable in the ®eld, they are of interest. Absolute and
comparative results with other taxa of Lysimachia suggest
that fertility and seed viability were not the major causes of
extinction of this species. Low ®tness values at these stages
have been suggested to operate in other rare and endangered
insular species, such as Lactoris fernandeziana from the
Juan FernaÂndez archipelago (Bernardello et al., 1999) and
Schiedea membranacea from the Hawaiian Islands (Culley
et al., 1999).
It seems that the low genetic variability present in L.
minoricensis has not led to maladaptation causing failure to
survive, and there do not appear to be any highly speci®c
requirements for germination. Extrinsic factors acting
through the reproductive processes in L. minoricensis
(from ¯owering to germination), which have not been
tested in this study, could be associated with successful
plant establishment and may have been involved in the
overall failure to obtain self-sustaining re-introduced populations in the wild. Field studies investigating ¯ower and
fruit predation, allelopathic effects on seed germination and
seedling survival, seedling predation, seedling survivorship
and establishment in the presence and absence of interspeci®c competition could be very rewarding. Such studies
could help elucidate why a self-compatible plant with a high
seed set, unreliant on pollinators and without highly speci®c
germination requirements went extinct in the wild.
A C K N O WL ED G EM EN T S
We are indebted to O. IbaÂnÄez for technical assistance. G.
Nieto and J. Thompson made useful criticisms that
improved the paper.
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