Seed dormancy pattern of the annuals Argemone ochroleuca and A. mexicana (Papaveraceae)
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Flora (2003) 198, 329–339
http://www.urbanfischer.de/journals/flora
Seed dormancy pattern of the annuals Argemone ochroleuca and
A. mexicana (Papaveraceae)
Laila M. Karlsson1*, T. Tamado2 and Per Milberg1
1
Department of Biology-IFM, Linköping University, SE-581 83 Linköping, Sweden
2
Department of Plant Science, Alemaya University, PO Box 138, Dire Dawa, Ethiopia
Submitted: Nov 12, 2002, in revised form: Mar 11, 2003 · Accepted: Mar 12, 2003
Summary
Seed dormancy of the tropical annual weeds Argemone ochroleuca Sweet and Argemone mexicana L. was studied in a series of
experiments. Factors investigated included: light, darkness, five temperature regimes and combinations of periods of light,
darkness and the different temperature regimes for various time periods (up to one year). Furthermore, germination in solutions
of gibberellic acid, after leaching, after mechanical treatments, and after periods of dry storage, was studied. Since the seeds have
underdeveloped embryos, their growth was also studied. There was no, or very little, germination (< 5%) or embryo growth in
most experiments, irrespective of light and temperature conditions. Argemone ochroleuca after-ripened slightly and germinated
to 10–20% after one year of dry storage. Germination to 100% occurred only after artificial treatments, either after removal of
part of the seed coat and endosperm or after incubation in gibberellic acid solution. The significance of the strong dormancy was
confirmed in the field for A. ochroleuca when only 8% of seeds sown produced seedlings in the first season after shedding. We
conclude that the majority of seeds normally do not germinate during their first season after shedding, but instead enter into the
seed bank, thus producing seedlings, even in a well-maintained field, probably for several years.
Key words: Argemone mexicana, Argemone ochroleuca, germination, Mexican poppy, prickly poppy, seed dormancy, tropical
weed
1. Introduction 1992; Wilson et al. 1995), India (Ramakrishnan &
Gupta 1972), and in several parts of Africa (Schwer-
The genus Argemone (Papaveraceae) contains almost zel & Mabasa 1986; Parsons & Cuthbertson 1992;
30 species, all with prickly stems, leaves and capsules Tamado & Milberg 2000). Argemone mexicana occurs
(Schwarzbach & Kadereit 1999). The annual herbs as a serious weed in Australia (Wilson et al. 1995) and
studied here, A. ochroleuca Sweet and A. mexicana L., in several countries of Latin America, Africa and Asia
are closely related (Schwarzbach & Kadereit 1999). (Häfliger & Wolf 1988). Besides being competitive
They show a similar leaf development, and all leaf seg- weeds in agriculture, Argemone species are toxic: seeds
ments terminate in a prickle (Gleisberg 1998). Their confused with those of mustard, and consumed, can lead
seeds are spherical, diameter ca. 1.5 – 2.5 mm, and dark to illness and even death (Das & Khanna 1997).
brown. Both species originated from the region of Cen- Not much is known about the ecology of Argemone.
tral America and south-western North America (Par- The two species studied in the present paper seem to
sons & Cuthbertson 1992; Schwarzbach & Kade- thrive in tropical and subtropical climates. In Puebla and
reit 1999). Nowadays, they are widespread around the Tlaxcala, Mexico, A. ochroleuca was found mostly in
warmer parts of the world. Argemone ochroleuca is a cultivations (Vibrans 1997), occurring in 47 of 378
serious weed in Australia (Parsons & Cuthbertson maize fields investigated (Vibrans 1998). A survey of
* Corresponding author: Laila M. Karlsson, Department of Biology-IFM, Linköping University, SE-581 83 Linköping,
Sweden, e-mail: laika@ifm.liu.se
0367-2530/03/198/04-329 $ 15.00/0 FLORA (2003) 198 329weeds in 240 crop fields in eastern Ethiopia indicated (Vleeshouwers et al. 1995; Baskin & Baskin 1998).
that occurrence of A. ochroleuca is not restricted to any Knowledge about seed dormancy and germination helps
of 23 environmental and crop management variables to explain the occurrence of weeds, and can be im-
recorded, e.g. altitude, soil type, rainfall, or crop species portant when developing, or foreseeing the effect of,
(Tamado & Milberg 2000). In Cuba, A. mexicana various weed-control methods (Forcella et al. 1993;
occurs in a wide range of cropping situations (sugarca- Mortimer 1997; Benech-Arnold et al. 2000).
ne, vegetable, coffee, cacao, citrus fruit and tobacco cul- Argemone ochroleuca and A. mexicana have linear
tivations, and in pastures) but was lacking in rice culti- embryos (Martin 1946; Gunn 1980). Thus, the two
vations (Acuña 1974). Hussein et al. (1983) found A. species have a type of underdeveloped embryo and
mexicana in all of twenty 5 × 5m quadrates investigated therefore some kind of morphological dormancy ac-
in a desert area in Saudi Arabia. Argemone ochroleuca cording to Nikolaeva (1977) and Baskin & Baskin
and A. mexicana are better suited to grow in deficiency (1998), i.e. the embryo needs to grow inside the seed
of phosphorus and nitrogen, respectively, but the two before germination. The purpose of this study was to
species co-occurred as weeds in eight of thirty sites further investigate this type of dormancy, its initial level
examined at Chandigarh, India (Ramakrishnan & and how this changes when seeds are subjected to envi-
Gupta 1972). Hence, the overall impression from the ronmental alternations.
scattered reports above is that of two ubiquitous tropical
weed species with no obvious restriction to particular
agronomic or environmental situations.
Many species have seeds showing some type of dor-
mancy, i.e. they do not germinate immediately when
2. Materials and methods
placed in their appropriate germination conditions (e.g.
Baskin & Baskin 1998). Dormancy breaking factor 2.1. Seed collection
could be, for example, high or low temperature periods,
environmental factors that change physics or chemistry We collected the seeds of Argemone ochroleuca on 3 October
of the seed, or also a combination of different factors 2000, 3km west of Harar (9°N, 42°E) in eastern Ethiopia, at
(Nikolaeva 1977; Baskin & Baskin 1998). So far, 16 an altitude of about 1800 m asl. Seeds of A. mexicana were
collected for us around 7 February 2001 at La Compañia
different dormancy types have been distinguished
(11°54N, 86°09W) in the pacific region of Nicaragua, at an
(Frost-Christensen 1974; Nikolaeva 1977; Baskin altitude of about 450m asl. The seeds of A. ochroleuca were
& Baskin 1990). The dormancy level of seeds can immediately transferred to Sweden (where the laboratory
change on a presumably continuous scale. Seeds of experiments were conducted), while the seeds of A. mexicana
many species can react on the environment and adjust were allowed to dry about 4 weeks at room temperature (ca.
the level of dormancy by deepening or weakening it 25°C) before being sent to Sweden.
Table 1. Outline of the different germination experiments. Species were either Argemone ochroleuca (o) or A. mexicana (m).
Storage times refer to dry storage at c. 20°C before the start of the experiments. See text for further explanations.
Experiment Species Storage Experiment Number Number of Light regime
time time of seeds seeds in the during
(weeks) (weeks) per dish experiment* incubation
Light and temperature o 3 52 50 45600 L, D
m 1 33 30 20690 L, D
Gibberellic acid (GA) o 21 23 50 4800 L, D
m 1 22 30 2880 L, D
Incubation + GA o 3 52 + 9 50 1800 L
Mechanical treatment o 23 21 30 1680 L
m 2 21 30 1440 L
Dry-storage o 3 37 50 11700 L
o 21 23 50 600 L
o 40 16 20 300 L
o 52 16 50 1200 L
Leaching & mechanical o 52 16 40 1920 L
* Including seeds used as controls
L = light for 12h per day, D = continuous darkness.
330 FLORA (2003) 1982.2. General treatments 2.4. Germination
Once the seeds had arrived in Sweden they were dry-stored 2.4.1. In the field
(Table 1) at room temperature (ca. 20°C) until the experiments
commenced. An overview of the experiments is given in Fresh seeds of A. ochroleuca were sown in eastern Ethiopia
Table 1, including the length of each experiment, number of (Alemaya University campus, 9°26’N, 42°03’E) immediately
seeds used and light/dark regimes. after collection. Seeds were buried (in order to allow imbibi-
Each germination experiment involved three Petri dishes tion and to protect the seeds from birds) ca. 2cm in local soil
(diam. 5cm) per treatment. The dishes were sealed with para- (well drained loam). Fifty seeds were sown in each of five
film. In those cases where seeds were subjected to darkness, plastic pots (diam. 25cm, 23.5cm deep) and placed unprotect-
the dishes were individually wrapped in aluminium foil. The ed outdoors, exposed to natural temperature, daylight and rain-
seeds were placed on moist (2.2mL deionized water) filter fall. The pots were checked for seedlings at least monthly.
paper (two pieces of Munkell 1003), the only exceptions being Daily precipitation and temperature were measured. The expe-
the seeds of A. mexicana in the experiment (described below) riment time included one short (ca. 1 month) and one long (ca.
with germination in different light and temperature regimes 6 months) rainy period.
and in the embryo growth experiment; these seeds were placed
on three pieces of Munkell 3, moistened with 2.4mL deionized
water.
Five temperature regimes were used in the study: 2°C con- 2.4.2. After dry storage
tinuously in a refrigerator (used only in some experiments with
A. ochroleuca), and 15/5°C, 20/10°C, 25/15°C and 30/20°C Seeds of A. ochroleuca were dry-stored at room temperature,
as maximum/minimum daily temperatures in germination ca. 20°C, for 3, 21, 40 and 52 weeks before being tested for
incubators equipped with light (Rubarth Apparatebau, Laat- germination at the four temperature regimes in the incubators.
zen, Germany). Light tubes gave 22–42 µmol m – 2 s–1 (photo-
meter: SKP 200, sensor SKP 215, Skye Instruments Ltd,
Wales, measure precision 1 µmol m – 2 s–1) with a 660/730nm
photon fluence ratio of 9.4 to 9.5 (photometer: SKR 100, sen- 2.4.3. At different temperatures and light regimes
sor SKR 110, Skye Instruments Ltd, Wales). The incubators
had a diurnal rhythm, with 10h of maximum and minimum The purpose of this experiment was to investigate germination
temperatures and 4h of continuous transition between the two. after a change of temperature and/or light regime. During a
The “light” treatments had a dark period coinciding with the total of up to 52 weeks (A. ochroleuca) or 33 weeks (A. mexi-
low temperature. cana) seeds were subjected to combinations of two incubation
All dishes in light treatments were checked for seedlings, conditions. These incubation periods are hereafter referred to
moisture conditions and fungi, at least biweekly. Seedlings as “first” and “second” incubation. In total, 320 and 96 com-
and obviously dead seeds were then counted and removed, and binations were used for A. ochroleuca and A. mexicana, re-
deionized water was added if necessary. A few times there was spectively.
mould overgrowing the filter paper (ca. 5 Petri dishes out of
1600) and in these cases the seeds were transferred to new
filter paper. The aluminium foil, enclosing the Petri dishes Table 2. Incubation periods for seeds of Argemone ochroleu-
with seeds kept in darkness, was not removed until an experi- ca and A. mexicana. Seeds were incubated in combinations of
ment was terminated. Germination was defined as the time two different temperature and/or light regimes during investi-
when the embryo had grown enough to split the endosperm, gation of germination and embryo growth. To both first and
which was clearly visible in a dissection microscope second incubation period 15/5°C, 20/10°C, 25/15°C and
(40× magnification). 30/20°C day/night temperature regimes were used, and seeds
were subjected to either light during 12 h/day or to continuous
darkness. At the end of every first incubation period seeds
were transferred to every other temperature regime. Argemo-
2.3. Imbibition ne ochroleuca was also subjected to changes of light regime
and to 2°C during first incubation. See text for further descrip-
To check if the seed coat was permeable to water, a batch of
tions of the experiment and of temperature and light regimes.
50 dry seeds of each of A. ochroleuca and A. mexicana were
weighed (measure precision 1.0mg) and placed on moist filter Argemone ochroleuca Argemone mexicana
paper in light at 15/5°C. The seed batches were then weighed
First Second First Second
after 5, 10, 20, 45 and 90min, and then 3, 6, 12, 24, 48 and 96h.
incubation incubation incubation incubation
Before being placed on the scale, the seeds were placed on dry
(weeks) (weeks) (weeks) (weeks)
filter paper to remove any excess water. The seeds were placed
back on the moist filter paper between weighings. The initial
3 3 3 15
weight of 50 dry seeds was 0.117g and 0.088g for A. ochro-
6 5 6 12
leuca and A. mexicana, respectively. The seeds of A. ochro-
11 26 12 6
leuca and A. mexicana placed on moist filter paper imbibed and
37 15 18 15
stabilized at 29.1% and 22.7% weight increase, respectively.
FLORA (2003) 198 331For both species, each combination of the four incubator 0.56–0.86mm or (iii) 0.98–1.17 mm, measured with a micro-
temperatures (15/5°C, 20/10°C, 25/15°C and 30/20°C) was meter scale in a dissection microscope (measure precision
used in the different incubation periods (Table 2). Seeds were 2.5 · 10 – 2 mm). In treatment (i) the endosperm was not affect-
subjected to either light during 12h/day or to continuous dar- ed, while in treatments (ii) and (iii) a piece of endosperm was
kness in each such combination. For A. ochroleuca the first removed. A total of 480 seeds were used for each treatment
incubation also included 2°C in darkness combined with the level, while 480 intact seeds were used as controls. Batches of
different incubator temperatures during second incubation. 40 seeds of A. ochroleuca were placed in nylon mesh (0.5mm
For A. ochroleuca germination after a change of light mesh size) bags (ca. 5 × 10cm), which were then placed on
regime, or combined change of light regime and temperature moist filter paper (one piece of Munkell 1003) in Petri dishes.
regime, was also investigated. After every first incubation The dishes were placed at 30/20°C in light for two days to
period (Table 2) light treated seeds were placed in darkness imbibe. Then the bags were put in individual beakers in
without temperature change, and vice versa. Seeds were also 125mL deionized water once a day for ten days at 30/20°C in
subjected to change of light regime at the same time as they light. Three bags were left in the water for 0, 0.1, 1 or 10 hours
were transferred between the temperatures regimes used. This a day. The rest of the time the bags were on the filter paper in
was done for all combinations of incubation conditions men- the Petri dishes. After the ten days, seeds were removed from
tioned above. the bags and placed on moist filter paper (two pieces of Mun-
Seeds of both species were also incubated continuously at kell 1003) in Petri dishes at 30/20°C in light.
one temperature and light regime. These dishes were checked
for seedlings, and then discarded, after 3, 6, 11, 37 and 52
weeks for A. ochroleuca and after 3, 6, 12, 18 and 33 weeks 2.5. Embryo growth
for A. mexicana, i.e. at end of the first incubation periods, and
at end of the experiment. To measure embryo length, the seed was bisected with a scal-
pel and the embryo was taken out and measured with a micro-
meter scale in a dissection microscope (measure precision
2.4.4. In gibberellic acid solution 2.5 ·10–2 mm). In most cases the embryos were slightly bent
or folded, yet could be easily straightened out under the dis-
Four concentrations of gibberellic acid, 0, 10, 100 and section microscope. In those cases where the embryos had
1000 mg L–1 GA3 (C19H21O6K, Sigma Chemical Co, USA) grown considerably, they had first become folded and then
were used. Seeds were incubated at the different incubator twisted and were therefore impossible to unfold. These em-
temperatures and in both light and darkness. The experiment bryos were therefore cut into parts and the pieces measured
was terminated when germination in light had ceased. individually.
A similar experiment was done with A. ochroleuca after
52 weeks of incubation in light at 20/10°C, 25/15°C and
30/20°C. These seeds were transferred to new filter papers 2.5.1. At different temperatures and light regimes
moistened with GA3 solution in concentrations as above. The
seeds were then incubated, at the same temperature as their The experiment was started together with, and ran parallel to,
first incubation period, for 9 weeks. the germination experiment involving different temperature
and light regimes (described in 2.4.3.).
The initial embryo length of 25 seeds of A. ochroleuca and 15
seeds of A. mexicana was measured after imbibition. Six and
2.4.5. After mechanical treatments four petri dishes, for A. ochroleuca and A. mexicana respect-
Seeds were abraded on the surface by being tumbled with ively, with ca. 30 seeds were put at each temperature and light re-
sand. For each species, two batches of 360 seeds were mixed gime used. The embryo length of 20–25 seeds of A. ochroleuca
with 500 mL mineral (90% gneiss and/or granite, 10% other was measured after one and two weeks in each treatment. The
minerals) sand (0.2–0.6 mm grain size). The mixture was embryo length of 10–20 seeds of each species was measured
placed in a test sieve shaker (Pascall’s Inclyno Test Sieve at the end of each first incubation period (Table 2).
Shaker, Model 1, Pascall Engineering, England) between two After the second incubation period, if conducted in dark-
sieves with 0.02mm mesh size. For each species, the shaker ness, embryo lengths of 10–20 nongerminated seeds from
was run for 1min with one batch of 360 seeds mixed with sand each treatment were measured. Embryo lengths of A. ochro-
and for 10min with the other. Careful sieving (0.6mm mesh leuca with a first incubation period of three or six weeks and
size) then separated the seeds from the sand. The seeds were the second period in light were also measured. Embryo lengths
then incubated in light at the four incubator temperatures. were measured in all treatments after a total incubation time of
52 and 33 weeks for A. ochroleuca and A. mexicana, respec-
tively.
2.4.6. After leaching combined
with mechanical treatment 2.5.2. After partial seed coat removal
After 52 weeks of dry storage, the seed coats of A. ochroleuca After 36 weeks of dry storage, the seed coat of 400 A. ochro-
seeds were cut away to three different extents, making holes in leuca seeds was cut away at the end opposite to the embryo,
the seed coat with diameters of: (i) 0.19–0.37mm, (ii) making a hole with a diameter of ca. 0.75mm, and simul-
332 FLORA (2003) 198taneously removing a small piece of endosperm. The embryo rainy period began after five months, and continued for
itself remained unaffected. The seeds were placed in Petri six months. Precipitation varied during the period, giv-
dishes, 20 seeds in each dish, and placed at the four incubator ing in total 720mm. Most of the time it rained frequently
temperatures in light. The embryo length in nongerminated but, about once a month, there was a period of five to
seeds was measured after 3, 6 and 9 days at 30/20°C and
seven days without rain. The soil in pots should not have
25/15°C, after 3, 6 and 11 days at 20/10°C, and after 6, 11 and
13 days for seeds at 15/5°C. On the first two occasions, for been able to dry out to the depth where seeds were
each incubator, the nongerminated seeds in one dish were buried, during this rainy period. The 1st month after
measured. On the third occasion, the remaining nongerminat- sowing, the mean daily air temperature was ca. 15°C,
ed seeds from the three remaining dishes in each incubator the 2nd–6th month ca. 10–15°C and the 6th–12th ca.
were used. Critical germination length (defined as the embryo 15–20°C. There was no relationship between rainfall or
length just before the endosperm split), at the different tempe- temperature and germination (data not shown).
ratures, was estimated from 2–5 germinating seeds in each
incubator.
3.1.2. After dry storage
2.5.3. In excised embryos Argemone ochroleuca germinated to 15–20% at
30/20°C after 40 and 52 weeks of dry storage. Other-
Embryos of A. ochroleuca were carefully excised from seeds wise, there was no increased germination with storage
and put on filter paper in Petri dishes. Twenty embryos on fil- times (Table 3).
ter paper moistened with deionized water were put in each
incubator, and 20 seeds on filter paper moistened with gibbe-
rellic acid (1000 mg L–1) were put at 25/15°C. Embryo lengths 3.1.3. At different temperatures and light regimes
were measured immediately, and then once a week for three
weeks. Germination was low in all treatments, and there was no
apparent difference between different combinations of
treatments (Table 4). Argemone ochroleuca did not
2.6. Statistical analyses exceed 1% germination in any treatment. The majority
of these plants germinated during the first six weeks, but
Germination was determined as the fraction of seeds that ger- germination did occasionally occur after up to 52 weeks
minated in the different treatments, excluding obviously dead of incubation. Argemone mexicana did not exceed 5%
seeds. The germination results are presented with 95% confi-
dence intervals for binomial parameters (Zar 1999).
Due to very limited germination, data from experiment Table 3. Germination (%) of Argemone ochroleuca after
2.4.3. were summed according to temperature or light/dark various periods of dry storage at ca. 20°C. Seeds were incu-
regimes, to examine the general response to; (i) time, (ii) light bated in light for 12 hour a day and at different temperature
or dark treatment, (iii) incubation temperatures and (iv) trans- regimes. The confidence intervals are based on statistics for
ference between different incubation temperatures. binomial distributions.
Embryo lengths were calculated as the arithmetic mean
with standard error (SE), with exception of the ones in the Temperature Storage time Germination
experiment with embryo growth in cut-off seeds, which are (°C) (weeks) (95% C.I.)
presented with 95% confidence intervals. The results of
embryo growth at all the separate temperatures and light regi- 30/20 3 0.1 (0.02–0.18)
mes were analysed with Dunett’s test for comparison of the 21 2.0 (0.41–5.79)
mean of a control group to the mean of each other group (Zar 40 20.0 (10.77–32.40)
1999) using 95% confidence limits. 52 14.8 (9.49–22.03)
25/15 3 0.1 (0.06–0.14)
21 2.0 (0.41–5.79)
3. Results 40 0.0 (0.00–5.96)
52 4.0 (1.48–8.58)
3.1. Germination 20/10 3 0.0 (0.00–0.10)
21 0.0 (0.00–2.43)
40 0.0 (0.00–5.96)
3.1.1. In the field 52 0.7 (0.02–3.69)
Circa 50 weeks after sowing, 8.0% (95% C.I. = 15/5 3 0.2 (0.09–0.34)
4.87–12.3) of the seeds sown in Ethiopia had produced 21 0.0 (0.00–2.43)
40 0.0 (0.00–5.96)
seedlings. A short rainy period occurred one month after
52 0.0 (0.00–2.43)
sowing, giving 100mm in a few days. The longer annual
FLORA (2003) 198 333Table 4. Germination (%) of Argemone ochroleuca and A. mexicana after various time periods of incubation, and in various
combinations of incubation in light or in darkness and at different temperatures. The confidence intervals are based on statistics
for binomial distributions.
Investigated Treatment Argemone ochroleuca Argemone mexicana
factor Germination (95% C.I.) Germination (95% C.I.)
Time 1–6 weeks 0.1 (0.06–0.16) 1.5 (1.16–1.88)
7–52 weeks 0.0 (0.00–0.01) –
7–33 weeks – 3.9 (2.90–5.06)
Continuous Light 0.1 (0.06–0.18) 4.0 (3.29–4.86)
light regime Darkness 0.1 (0.08–0.20) 3.0 (2.42–3.59)
Continuous 30/20°C 0.1 (0.02–0.18) 0.7 (0.32–1.44)
temperature 25/15°C 0.1 (0.06–0.27) 1.3 (0.71–2.18)
regime 20/10°C 0.0 (0.00–0.10) 2.7 (1.78–3.90)
15/5°C 0.2 (0.09–0.33) 4.4 (3.08–5.93)
Transference 30/20°C → 25/15°C 0.0 (0.00–0.27) 1.9 (1.01–3.12)
to cooler 30/20°C → 20/10°C 0.0 (0.00–0.27) 2.0 (1.08–3.31)
temperature 30/20°C → 15/5°C 0.0 (0.00–0.27) 1.4 (0.67–2.45)
regime 25/15°C → 20/10°C 0.0 (0.00–0.27) 0.2 (0.03–0.89)
25/15°C → 15/5°C 0.0 (0.00–0.27) 0.4 (0.08–1.10)
20/10°C → 15/5°C 0.0 (0.00–0.27) 0.6 (0.20–1.45)
Transference 15/5°C → 20/10°C 0.0 (0.00–0.27) 0.0 (0.09–0.46)
to warmer 15/5°C → 25/15°C 0.0 (0.00–0.27) 0.0 (0.09–0.46)
temperature 15/5°C → 30/20°C 0.0 (0.00–0.27) 0.0 (0.09–0.46)
regime 20/10°C → 25/15°C 0.0 (0.00–0.27) 0.1 (0.00–0.69)
20/10°C → 30/20°C 0.1 (0.07–0.41) 0.0 (0.09–0.46)
25/15°C → 30/20°C 0.0 (0.00–0.27) 0.0 (0.09–0.46)
germination in any treatment. This germination occur- were abraded on the surface, but there were no seeds
red mainly at the cooler temperatures and after transfer with the seed coat punctured or removed. Generally, the
from 30/20 °C to a cooler temperature regime (Table 4). treatment did not stimulate germination (Table 5); in
Seed mortality was low, a total of 0.30% of the seeds only one species and temperature there was a non-over-
were scored dead when terminating the germination lapping confidence interval between non-abraded seeds
experiments. and abraded ones (A. ochroleuca at 25/15°C).
3.1.6. After leaching combined
3.1.4. In gibberellic acid solution
with mechanical treatment
Germination in light ceased 22 weeks after the start of
the experiment (data not shown). Both species respond- There was a clear germination response to mechanical
ed to gibberellic acid solutions, both in light and in dark- treatment (Table 6), with full germination after the inter-
ness. Maximum germination occurred in 1000mg L–1 mediate (ii) treatment, but no effect of leaching
and there was detectable germination in 10mg L–1 (Table 6).
(Fig. 1). The germination of A. ochroleuca in gibberel-
lic acid following 52 weeks of incubation on filter
papers moistened with water was similar; full germina- 3.2. Embryo growth
tion was achieved in 1000mg L–1 after 9 weeks (data not
shown). 3.2.1. At different temperatures
and light regimes
3.1.5. After mechanical treatments Mean initial embryo length was 1.00mm (SE = 0.015)
and 0.82 mm (SE = 0.025) for A. ochroleuca and
After the treatment in the test sieve shaker, the seeds A. mexicana, respectively. There was no embryo growth
334 FLORA (2003) 198Fig. 1. Germination of Argemone ochro-
leuca and A. mexicana in various con-
centrations of gibberellic acid and under
various light and temperature conditions.
Germination with 95% confidence inter-
val, based on statistics for binomial dis-
tribution, is shown.
Table 5. Germination (%) of Argemone ochroleuca and A. mexicana after being abraded on the surface when tumbled with sand.
The confidence intervals are based on statistics for binomial distributions.
Incubation Abraded Argemone ochroleuca Argemone mexicana
temperature (°C) on surface Germination (95% C.I.) Germination (95% C.I.)
30/20 not at all 2 (0.4–5.7) 3 (0.7– 9.5)
slight 1 (0.0–6.1) 3 (0.7–9.5)
heavy 8 (3.2–15.2) 4 (1.2–11.0)
25/15 not at all 2 (0.4–5.7) 2 (0.3–9.5)
slight 12 (6.2–20.9) 1 (0.0–6.1)
heavy 9 (3.2–15.2) 2 (0.3–7.8)
20/10 not at all 0 (0.0–2.4) 1 (0.0–6.1)
slight 0 (0.0–4.0) 3 (0.7–9.5)
heavy 1 (0.0–6.1) 6 (1.8–12.5)
15/5 not at all 0 (0.0–2.4) 5 (1.3–11.3)
slight 1 (0.0–6.1) 4 (1.2–11.0)
heavy 0 (0.0–4.0) 6 (1.8–12.5)
in response to temperature, light or darkness, or after ferent treatments varied from 0.92–1.07mm (all SE
changes between the various incubation conditions < 0.05) for A. ochroleuca and 0.77–0.95 mm (all SE
(data not shown). Embryo lengths were measured 279 < 0.04) for A. mexicana. The mean embryo lengths from
and 100 times for A. ochroleuca and A. mexicana, re- the various incubation treatments and time periods were
spectively (Fig. 2). The mean embryo length in the dif- not significantly (all p > 0.05) greater than the mean
FLORA (2003) 198 335embryo length in imbibed fresh seeds.
3.2.2. After partial seed coat removal
Embryos in cut-off A. ochroleuca grew faster at higher
temperatures. In nine days the mean initial length had
tripled at 30/20°C and 25/15°C. At 20/10°C and 15/5°C,
lengths had doubled after 11 and 13 days, respectively
(Fig. 3). The critical germination length increased from
2.8mm at 15/5°C to 4.6mm at 30/20°C (Fig. 3).
3.2.3. In excised embryos
There was no growth in excised embryos, neither when
Fig. 2. Embryo lengths of Argemone ochroleuca and A. mexi- the embryos where put on deionized water nor when
cana after incubation in different light and temperature they where put in gibberellic acid solution (data not
regimes over various time periods. Each point represents the shown). On the contrary, the embryos shrunk in the
mean of one combination of light regime, temperature, and treatment with gibberellic acid.
time period.
4. Discussion
4.1. Germination
Both species had seeds that were highly dormant, with
very little initial germination and no substantial (up to
20% germination) after-ripening effect (Table 3), nor
any response to various combinations of incubation
environments (Table 4). This would suggest that very
few seeds would germinate in the season following seed
dispersal, which was also confirmed when seeds of one
of the species produced less than 10% seedlings.
Substantial germination occurred only in response to
gibberellic acid (Fig. 1) and for A. ochroleuca after some
mechanical treatment (Table 6; A. mexicana was not
included in the latter experiment).
Fig. 3. Embryo lengths of Argemone ochroleuca after various
times of incubation in light at different temperatures. The seed
coats were cut off at the opposite end to the embryo, to the
extent that a small piece of endosperm was removed, before 4.2. Embryo growth
the start of the experiment. Mean embryo length (open sym-
bols) with 95% confidence interval and estimated critical ger- The lack of growth or elongation in the excised embryos
mination length (filled symbols) are shown.
Table 6. Germination of Argemone ochroleuca after different degrees of mechanical treatment and
different times of leaching. Germination percentages with 95% confidence intervals, based on statis-
tics for binomial distributions, are shown. See text for further details.
Mechanical Time for leaching (hours)
damage 0 1 10 100
none 25 (18.0–35.0) 23 (15.6–32.4) 22 (14.4–30.4) 18 (11.5–26.9)
small (i) 22 (14.4–30.4) 38 (29.4–47.9) 45 (36.3–55.5) 33 (23.7–41.4)
intermediate (ii) 100 (97.0–100) 99 (95.4–100) 100 (97.0–100) 100 (97.0–100)
large (iii) 71 (61.6–79.0) 37 (27.7–46.2) 38 (29.4–47.9) 58 (48.1–66.6)
336 FLORA (2003) 198indicated that the initiation of their growth was depen- exhibit deep simple double morphophysiological dor-
dent on the endosperm. This is to be expected for under- mancy.
developed embryos since, up to now, excised embryos The seed coats of A. ochroleuca and A. mexicana do
that have been reported to grow (and produce a normal not inhibit imbibition and therefore the species do not
or abnormal seedling) belong to developed embryos in have physical dormancy. There are no visible structures
seeds with physiological dormancy (Baskin & Baskin in the seeds of A. ochroleuca and A. mexicana that could
1998). restrict embryo growth and/or germination. Further,
Nothing in the results indicated that there was a slow seeds of both species did germinate, without any mecha-
embryo growth, or a delay before the seeds with full- nical treatment of the seeds, in response to gibberellic
grown embryos germinated (Fig. 2). On the contrary, it acid (Fig. 1). Therefore, the two species do not have
seems that, in every single seed, growth of the embryo mechanical dormancy. The germination in response to
and its germination occurs as a continuum that takes 1 removal of part of the seed coat (Table 6) might be
to 2 weeks (Fig. 3), depending on temperature. explained by a hypothetical germination-inhibiting sub-
stance being present, and that this substance is more
easily leached out of the seeds after the treatment.
However, this seems less likely since there was no res-
4.3. Dormancy classification ponse to the different times with water treatment in the
leaching experiment (Table 6), and therefore at least
Because they are linear (Martin 1946; Gunn 1980) A. ochroleuca do not have chemical dormancy.
and underdeveloped at the time of dispersal (Nikolae- Shortage of seeds of A. mexicana prevented its inclu-
va 1977 ; Baskin & Baskin 1998), the embryos of sion in the leaching experiment, but we suspect that this
A. ochroleuca and A. mexicana need to grow before ger- species exhibits the same type of dormancy syndrome as
mination (Fig. 3). Consequently, the two species have A. ochroleuca. Hence, we conclude that the two species
some type of morphological dormancy according to do not easily fit into the dormancy types currently distin-
Nikolaeva (1977) and Baskin & Baskin (1998). Fresh guished.
seeds of species having (only) morphological dormancy
should begin their embryo growth when placed in
appropriate conditions for germination. None of the 4.4. Comparison with other Papaveraceae
two species germinated in response to the different species and tropical species
temperatures and light regimes tested (Table 4), despite
the fact that the experiment well covered the possible There appears to be no previous study conducted on the
temperatures occurring at the sites where seeds were germination of Argemone. As far as is known, all Papa-
collected. Nor did the species germinate after combina- veraceae have either rudimentary or linear embryos
tions of the wide range of temperatures, light regimes (Martin 1946; Gunn 1980), and thus must have some
and incubating periods covered. This excludes all ex- kind of morphological dormancy (Nikolaeva 1977;
pect one of the known combinations of morphological Baskin & Baskin 1998). Most species studied, accord-
and physiological (called morphophysiological) dor- ing to Baskin & Baskin (1998), seem to have mor-
mancy types. Physiological dormancy is broken by phophysiological dormancy, i.e. Chelidonium majus
alternations of temperature (Nikolaeva 1977; Baskin (Grime et al. 1981), Dendromecon rigida, Meconopsis
& Baskin 1998), which enables the seeds to germinate cambricum (Schroeder & Barton 1939), Glaucium
at an appropriate time of a year. However, the experi- flavum (Thanos et al. 1988), Papaver californicum
ments did not involve three, or more, temperature (Keeley & Keeley 1987), Papaver dubium, P. somni-
combinations. The only (known) dormancy type with a ferum (Grime et al. 1981), P. rhoeas (Grime et al. 1981;
demand for three different temperatures before ger- Baskin et al. 2002), and Romneya trichocalyx (Keeley
mination is the so-called “deep simple double morpho- & Keeley 1987). These studies all involve species from
physiological dormancy”. Species possessing this dor- arctic, cold temperate or warm temperate climatic
mancy type demand combinations of warm and cold zones, and morphophysiological dormancy therefore
stratification to break dormancy for the hypocotyl and seems to be the norm for Papaveraceae in such climates.
the epicotyl separately before germination (Nikolaeva Nevertheless, one cannot assume that Papaveraceae nor-
1977). This kind of dormancy is not known to be broken mally have morphophysiological dormancy in the tropi-
by gibberellic acid (Baskin & Baskin 1998) and thus, cal zone as well. In tropical areas, seasonal differences
because of the clear germination response to gibberellic in temperature are small, and maybe other environ-
acid (Fig. 1) and the continuous embryo growth and ger- mental cues than seasonal temperature changes are bet-
mination when part of the seeds coat was removed ter indicators for an appropriate time to germinate in the
(Fig. 3), A. ochroleuca and A. mexicana probably do not tropics. Germination of tropical species has not been
FLORA (2003) 198 337much studied (e.g. Baskin & Baskin 1998), and very a substantial part of the Argemone seeds that ripe every
little is known about germination ecology of tropical year will enter into the seed bank, and will probably ger-
weeds. In a field experiment conducted in Zimbabwe 13 minate in an unpredictable pattern during several con-
weedy species were sown ; for every species emergence secutive years. It is likely that this germination pattern
was less than 10% the first year and 6 species, including is the norm worldwide in tropic and subtropical regions,
Argemone subfusiformis, produced seedlings even after where the species occur as weeds. For a farmer, it is
10 years (Schwerzel & Madasa 1996). Fenner important to know that removing Argemone before
(1980) performed germination experiments in light and shedding will give a benefit of decreasing Argemone
darkness in a non-heated greenhouse with mean daily germination, though this will not be obvious in the short
temperatures between 15 and 28°C, investigating 32 time span. An already existing seed bank will continue
East African weed species. For 14 of the species no sub- to give rise to seedlings for some time, probably many
stantial germination occurred, indicating that time alone years, thus producing Argemone seedlings even in a
may be an important dormancy-breaking factor for more well-managed field.
tropical species than only Argemone.
Acknowledgement
4.5. Possible dormancy-breaking factors
We thank Freddy Aleman for supplying us with seeds of
in the field A. mexicana from Nicaragua, Carol Baskin for various com-
ments, Inga-Maj Kaller and Karsten Håkansson at the
There was no indication of a slow embryo growth Swedish Geotechnical Institute for access to the test sieve
(Fig. 2). Instead, the quick embryo growth in seeds of shaker and CF Lundströms Stiftelse for financial support.
A. ochroleuca with part of the seed coats removed
(Fig. 3) indicated that the species does not have a
demand for any second germination cue once embryo
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