Bhatt, Bhat, Phartyal and Gallacher (2020). Seed Science and Technology, 48, 2, 247-255. https://doi.org/10.15258/sst.2020.48.2.12
Research Note
Dry-storage and light exposure reduce dormancy of Arabian desert legumes more than temperature
Arvind Bhatt1,2, N.R. Bhat1, Shyam S. Phartyal3* and David J. Gallacher4
1 Kuwait Institute for Scientific Research, P.O. Box 24885 Safat, 13109, Kuwait 2 Lushan Botanical Garden, Chinese Academy of Sciences, Lushan, Jiujiang City, China 3 Nalanda University, School of Ecology and Environment Studies, Rajgir, 803116, India 4 School of Life and Environmental Sciences, The University of Sydney, Narrabri NSW 2390, Australia * Author for correspondence (E-mail: [email protected])
(Submitted January 2020; Accepted May 2020; Published online May 2020)
Abstract
Propagation and conservation of desert plants are assisted by improved understanding of seed germination ecology. The effects of dry-storage on dormancy and germination were studied in seven desert legumes. Mature seeds were collected in summer 2017 and germinated within one week of collection (fresh) and after six months (dry-storage) under two temperature and two light regimes. Seed weight of two species increased 22-55% within 24 hours of water imbibition but others increased ≤ 7%. Germination ranged from 0-32% in fresh and 2-92% in dry-stored seeds, indicating a mix of non- and physically-dormant seeds at maturity. Dry-storage at ambient room temperature was effective at relieving dormancy, though the extent was species-dependent. Germination percentage increased in response to light exposure during incubation, while the effect of temperature was species-dependent. This variable response to dormancy alleviation may assist to spread the population risk of seedling survival in the harsh and variable environment of the Arabian desert.
Keywords: arid, dry-storage, legumes, seed dormancy, seed germination, water-gap
Experimental and discussion
Desert plants are subjected to high temperature and high salinity, low soil water availability and acute nutrient deficiencies (Abu Sukar et al., 2007; Rewald et al., 2011). Each species exhibits strategies for population survival in these severe environmental conditions (Gutterman, 1993). Annual species allocate a higher proportion of resources to sexual reproduction than perennial species (Karlsson and Méndez, 2005). Populations of
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247 A. BHATT, N.R. BHAT, S.S. PHARTYAL AND D.J. GALLACHER annual species are dependent on seed bank persistence, frequency of successful seedling establishment and rapid transition from germination to seed production (Gutterman, 2000). In contrast, populations of perennial species are dependent on producing multiple seed batches, thus reducing the reliance on seed banks and successful germination events (Escós et al., 2000). Populations of annual desert species are more sensitive to environmental variation since they are more dependent on successful seedling establishment in unfavourable seasons (Li, 2010). Interspecific variation in germination timing enables different species to occupy specific niches (Rathcke and Lacey, 1985). Germination timing affects population abundance by influencing seedling survival and is thus a crucial component of a species’ life-history strategy (Handley and Davy, 2005; Shahba and Qian, 2008). Dormancy at maturation is common in desert seeds, preventing germination during the unfavourably hot and dry summer (Iglesias-Fernández et al., 2011; Baskin and Baskin, 2014). Most species that produce mature seeds during summer thus do not germinate until the following winter, when soil moisture is more likely to persist after a rain event and seeds are able to germinate and seedlings to establish. The Arabian Peninsula is home to 50 Fabaceae species from 22 genera, with diverse growth habits, from annual herbs to woody perennials (Mandaville, 1990). Legumes are adapted to nitrogen-poor soils through their symbiotic association with rhizobia (Crews, 1999) which sometimes also benefits other plant species. Nitrogen is typically low in desert soils (Wolde-Meskel et al., 2004). Legumes are a preferred feed source for many desert ungulates (Graham and Vence, 2003). Physical dormancy (PY) is prevalent in legume seeds, due to a water-impenetrable palisade cell layer in the seed and/or fruit coat (Baskin and Baskin, 2014). PY is alleviated when the ‘water-gap’ in the palisade layer is breached and water enters the seed (Baskin and Baskin, 2014). This usually occurs when conditions are suitable for seedling establishment (Jayasuriya et al., 2007). The strength of the water-gap gradually declines over time, largely due to the seasonal and diurnal changes in temperature and moisture. These factors are therefore environmental signal detectors for dormancy break (Baskin and Baskin, 2000, 2014). PY can be broken in the laboratory through mechanical or acid scarification, percussion, or exposure to heat (moist or dry) or storage (moist or dry) (Phartyal et al., 2005; Bhatt et al., 2016; Koutouan- Kontchoi et al., 2020). Although most seeds in the studied genera are known to possess PY (Kondo, 1993; Patanè and Gresta, 2006; Kim et al., 2008; Long et al., 2012; Mondoni et al., 2013), the role of dry-storage in breaking PY is not well known. We compared the germination rate of freshly collected and dry-stored seeds of seven Arabian desert legumes. All species are widely distributed across the Arabian Peninsula and produce mature seeds in summer (April-June), with new seedlings emerging in early winter (Norton et al., 2009). We hypothesised that (1) fresh seeds would exhibit PY; (2) dry-storage would alleviate PY; and (3) that the tested temperature and light regimes for germination would reflect natural winter conditions. Mature seeds of all species were collected in late April 2017 from natural populations in Kuwait (table 1). Seeds of each species were collected from 30-35 spatially separated plants to represent the genetic diversity of the population. Seeds of each species were cleaned and divided into one batch for immediate germination (fresh) and another for
248 SEED DORMANCY IN ARABIAN DESERT LEGUMES dry-storage. Fresh seeds were tested for germination within one week of collection. Dry- stored seeds were kept indoors in brown paper bags at room storage (temperature 20 ± 2°C, relative humidity 15 ± 2%) and tested for germination in the first week of November. The dry-storage conditions are thought to be optimal for most orthodox seeds (Ellis et al., 1991), and November represents the month in Kuwait when soil moisture is likely to persist after rains. The mass of fresh seeds was determined by weighing three 50- seed replicates. Water permeability was assessed by recording the mass of three 25-seed replicates before and after placement on two sheets of Whatman No. 1 filter paper for
24 hours, moistened with 10 ml distilled water. Seed mass varied seven-fold from 0.52 mg for Lotus halophilus to 3.76 mg for Astragalus spinosus. It reveals perennial species (A. spinosus) produce heavier seeds than those of the annuals, reflecting previous findings of a positive correlation between seed mass and species’ lifespan (Baker, 1972; Leishman et al., 2000). In two of the seven species, water imbibition rate varied moderately (22.5 to 52.6%) while the remaining species showed only a slight (≤ 7%) gain during the first 24 hours of imbibition (table 1). Most legume seeds, if permeable, imbibe water close to 100% or more prior to seed germination (Baskin and Baskin, 2014). The moderate percentage of water imbibition indicates that the water-gap was open in only a moderate proportion of fresh seed and germination was therefore relatively low (6-32%). Fresh and dry-stored seeds were tested for germination at two night/day temperature regimes (15/20 and 20/30°C) and two light regimes (dark/light and dark/dark). Four 25- seed replicates per treatment were placed in 90 mm-diameter Petri-dishes containing two disks of Whatman No. 1 filter paper moistened with 10 ml of distilled water. Darkness was obtained by wrapping Petri-dishes in aluminum foil. The number of germinated seeds was counted daily for 28 days for seeds in the light and at the end of 28 days for seeds in the dark. Germination was defined as the emergence of the radicle to a length > 1 mm. All non-germinated seeds were cut with a scalpel to evaluate the status of the embryo. Seeds with a turgid whitish embryo were considered viable and those with a brownish (necrotised) embryo were considered non-viable. Germination percentage was calculated based on viable seeds. The relationship between mean germination percentage (dependent variable) and the value of three predictors (dry-storage, photoperiod and temperature) was compared for each species using a generalised linear model with a Poisson probability distribution and identity-link function. The Bonferroni correction was applied to assess significance, in which α = 0.0012. The effects of three predictors and their interaction in the model were tested by Wald chi-square values. All statistical analyses were carried out using SPSS 21. Germination of dry-stored seeds was significantly greater than fresh seeds for all species (figure 1) except A. corrugatus, which exhibited dormancy even after storage (table 1). Germination varied among species from 0 to 32% and 2 to 92% in fresh and dry- stored seeds, respectively. Dry-storage and photoperiod had a greater influence over seed germination than temperature regime in most species (table 2). Of the six species exhibiting germination, all exhibited a greater percentage with dry-storage, five with light exposure, and two were influenced by temperature regime but in opposite directions. However, significant first order interactions between dry-storage and temperature indicate that temperature plays a significant role in two further species; A. annularis and A. tribuloides.
249 A. BHATT, N.R. BHAT, S.S. PHARTYAL AND D.J. GALLACHER
100 Astragalus annularis 100 Astragalus schimperi
80 (% ) (% ) 80 Fresh seed Stored seed 60 60
40 40
20 20 Germination Germination 0 0 Light Dark Light Dark Light Dark Light Dark 15/20 C 20/30 C 15/20 C 20/30 C
100 Astragalus spinosus 100 Astragalus tribuloides
80 (% ) (% ) 80
60 60
40 40
20 20 Germination Germination 0 0 Light Dark Light Dark Light Dark Light Dark 15/20 C 20/30 C 15/20 C 20/30 C
Lotus halophilus 100 100 Trigonella stellata
80 (% ) (% ) 80
60 60
40 40
20 20 Germination Germination 0 0 Light Dark Light Dark Light Dark Light Dark 15/20 C 20/30 C 15/20 C 20/30 C
Seed incubation conditions Seed incubation conditions
Figure 1. Influence of dry-storage, incubation temperatures, and photoperiod on mean seed germination percentage of desert legume species. Significance of factors is shown in table 2.
250 SEED DORMANCY IN ARABIAN DESERT LEGUMES P 0.000 0.000 0.108 0.025 0.915 0.748 0.748 2 55 92 42 12 42
42 stellata 2 X Trigonella Trigonella 2.58 5.05 0.01 0.10 0.10 94.91
Wald’s Germination (%) Germination 6 0 32 22 10 20
25 P Fresh Dry-stored 0.000 0.093 23.21 0.005 0.040 0.575 0.852 0.191 Lotus Lotus 2 halophilus 2.83 7.86 4.23 0.31 0.04 1.71 X 694.53
Wald’s P 0.000 0.526 0.000 0.000 0.016 0.004 0.526 06.27 ± 0.24 ± 06.27 22.49 ± 4.03 ± 22.49 02.15 ± 0.56 ± 02.15 0.36 ± 52.65 07.02 ± 2.17 ± 07.02 02.37 ± 0.33 ± 02.37 01.23 ± 0.16 ± 01.23 after 24 hours 24 after imbibition (% ± SD) ± (% imbibition 2 Increase in seed mass seed in Increase Astragalus tribuloides 5.80 8.50 0.40 X 0.402 35.49 19.68 127.25
Wald’s
P 0.000 0.000 0.000 0.242 0.242 0.010 0.483 2 spinosus Seed mass Seed (mg ± SD) ± (mg Astragalus Astragalus 0.70 ± 0.01 ± 0.70 0.52 ± 0.04 ± 0.52 1.33 ± 0.01 ± 1.33 0.04 ± 3.76 0.65 ± 0.01 ± 0.65 0.79 ± 0.01 ± 0.79 1.73 ± 0.02 ± 1.73 X 1.37 1.37 6.63 0.49 28.99 34.25
Wald’s P 0.000 0.072 12.33 0.000 0.678 0.213 0.019 0.213 Site 2 schimperi Astragalus Astragalus 3.23 0.17 1.55 5.53 1.55 X 62.18 318.49
Wald’s P Annual Sulabia Annual Sulabia Perennial Julaia Annual Sulabia Annual Sulabia Annual Sulabia Annual Sulabia Life form Life 0.000 0.808 0.000 0.000 0.808 0.465 0.808 2 annularis Astragalus Astragalus X 0.06 0.06 0.53 0.06 72.59
26.13 13.33
Wald’s Delile Bertol 1 1 1 1 1 1 1 Boiss Forssk ) shown in bold are significant at 0.05 level. 0.05 at significant are bold in shown ) Vahl Forssk Boiss. and Spruner and Boiss. Significance of dry-storage (D), incubation temperature (T) and photoperiod (P) on seed germination percentage of desert legume species, using a using a species, legume desert of germination percentage seed and photoperiod (P) on (T) temperature of dry-storage (D), incubation Significance D × P × D D × P × D D × T × D Factor df D × T × P × T × D Dry-storage (D) Dry-storage Temperature (T) Temperature Photoperiod (P) Photoperiod Sulabia 29.1616N, 47.6928E; Julaia 28.8915N, 48.2381E 28.8915N, Julaia 47.6928E; 29.1616N, Sulabia Trigonella stellata Trigonella Astragalus spinosus spinosus Astragalus Astragalus tribuloides Astragalus halophilus Lotus Astragalus schimperi schimperi Astragalus Astragalus corrugates corrugates Astragalus Astragalus annularis annularis Astragalus Table 2. Table function. identity-link an and distribution probability Poisson a with model linear generalised Table 1. Summary of study species, collection site and morphophysiological parameters. For specific germination parameters, see figure 1. figure see parameters, germination specific For parameters. morphophysiological and site collection species, study of Summary 1. Table Species Probability values (P values Probability
251 A. BHATT, N.R. BHAT, S.S. PHARTYAL AND D.J. GALLACHER
In both cases, germination of dry-stored seeds was higher than of fresh seeds at the lower germination temperature. Other first-order interactions were not significant after application of the Bonferroni correction. In Astragalus annularis seeds, germination percentage was significantly increased by dry-storage and exposure to light. Lower temperature resulted in higher germination in dry-stored seeds, but lower germination in fresh seeds. Seeds therefore appear to adjust temperature requirements for germination as dormancy declines toward winter months. No fresh mature seeds of A. corrugatus germinated under any treatments. Germination was low (2%) even after six months of dry-storage, indicating strong dormancy or lack of a suitable environment for dormancy-break and seed germination. In contrast, germination of A. schimperi seeds was significantly increased by dry-storage and exposure to light. Incubation temperature and the first order interaction of temperature with photoperiod had no effect. In the case of A. spinosus, seed germination percentage was significantly increased by dry-storage, exposure to light and low incubation temperature. The first order interaction of temperature with photoperiod was not significant. Similarly, in seeds of A. tribuloides germination percentage was significantly increased by dry-storage and exposure to light. Lower temperature increased germination percentage of dry-stored seed but reduced it in fresh seed. This suggests that the seeds adjust temperature requirements for germination as dormancy declines toward winter months. Other first order interactions were not significant after application of the Bonferroni correction. Lotus halophilus seed germination percentage was significantly increased by dry-storage and exposure to light. Incubation temperature and the first order interaction of temperature with dry-storage had no effect. Seeds of Trigonella stellata germinated significantly higher percentage after dry-storage and at higher incubation temperature. Photoperiod and the first order interaction of temperature with dry-storage had no effect. There was inconsistency in water imbibition and seed germination percentage. The possible explanation for this inconsistency is that seeds of different species have different rates of water imbibition, some may have imbibed water with a quick rate (A. spinosus) and others with slow rate (A. annularis). In the present study, we monitored imbibition once at 24 hours, so those germinated in moderate proportion (A. annularis) could have been imbibed enough water with slow rate prior to germinate and others that imbibed water quickly but had low germination (A. spinosus) may have dormancy other than PY. Even after dry-storage, germination of A. spinosus was not increased, which further confirms other kinds of dormancy. The remaining ungerminated seeds of most species exhibited PY similar to most legume species (Jayasuriya et al., 2013; Baskin and Baskin, 2014). Batches of PY seeds can exhibit variation in their water permeability and dormancy release during dry-storage that can span several months (Baskin and Baskin, 2014). This is particularly true for legumes in the subfamily Papilionoideae (Cavanagh, 1987) in which all the studied species are placed. Dry-storage reduced the percentage of dormant seeds in the studied species to a large extent since germination percentage increased to a moderately high proportion (42-92%) after dry-storage in most of species except A. corrugatus and A. spinosus (table 1). Dry-storage is thought to make seeds sensitive for dormancy-break through the develop ment of cracks in the seed coat (Gama-Arachchige et al., 2011; Baskin and Baskin, 2014).
252 SEED DORMANCY IN ARABIAN DESERT LEGUMES
However, almost all A. corrugatus seeds retained dormancy after six months of dry- storage at room temperature. Storage in cooler indoor temperatures may not be suitable for dormancy alleviation of these species, which naturally grow in conditions of extreme heat and dryness. However, the most effective dry-storage conditions vary among species and might not be controlled by temperature alone (Ellis et al., 2018; Jaganathan et al., 2018). For example, dry-stored PY seeds of Lotus corniculatus var. japonicas exhibited 65% germination after 12 months storage at 3°C, but < 10% when stored in warmer conditions of 10-35°C (Kondo, 1993). Similarly, dry-stored PY seeds of Ipomoea purpurea germinated to 3, 76 and 95%, respectively when stored at 15, 25 and 35°C, respectively for six months (Brechu-Franco et al., 2000). These studies also indicate that it is common for germination to be < 100% of viable seeds, even after extended dry- storage if temperature is not optimum. Seeds of all investigated species are desiccation-tolerant, disperse from the maternal plant in summer with a low moisture content (Royal Botanic Gardens Kew, 2019) and possess PY (Baskin and Baskin, 2014). Seeds with a low moisture content have a short window of opportunity to germinate, following rains either in autumn (Quinlivan, 1968) or winter. PY in most of the studied species could be broken through acid scarification, after which > 90% germinated at a temperature range of 15-25°C (Royal Botanic Gardens Kew, 2019). Most studied species exhibited positive photoblastism, indicating a preference for non- burial. Nevertheless, germination still occurred in darkness. Positive photoblastism has been reported for legumes Astragalus bibullatus and A. membranaceus (Morris et al., 2002; Zhou et al., 2012). Generally, the light requirement for germination varies among species, influencing seedling establishment (Fenner and Thompson, 2005; Baskin and Baskin, 2014). Desert annuals usually complete their life cycle in 8-10 weeks in summer if soil moisture is available (Mulroy and Rundel, 1977). However, summer germination is highly risky for annual species since failure could substantially erode the seed bank. Winter germination, starting from November, carries much lower risk. The tested temperature regimes in this study of 15/20 to 20/30°C correspond to typical field temperature conditions of Kuwait from November to March (Omar, 2007) with a higher possibility of soil moisture. To conclude, seeds of most of the tested desert legumes were physically dormant at maturity, which was moderately released by six months dry-storage at room temperature. The incomplete germination of most studied species indicates that dry-storage conditions and duration were not optimal for all seeds within a batch. Overall, dry-storage and light had a strong effect on seed germination, while temperatures had less. These results are of particular interest for the artificial propagation and in-situ conservation of these important desert species.
Acknowledgements
This work was supported by the Kuwait Institute for Scientific Research (KISR).
253 A. BHATT, N.R. BHAT, S.S. PHARTYAL AND D.J. GALLACHER
Conflict of interest No potential conflict of interest was reported by the authors.
References
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