Shotgun proteomic analysis of soybean embryonic axes during germination under salt stress

Seed imbibition and radicle emergence are generally less affected by salinity in soybean than in other crop plants. In order to unveil the mechanisms underlying this remarkable salt tolerance of soybean at seed germination, a comparative label‐free shotgun proteomic analysis of embryonic axes exposed to salinity during germination sensu stricto (GSS) was conducted. The results revealed that the application of 100 and 200 mmol/L NaCl stress was accompanied by significant changes (>2‐fold, P<0.05) of 97 and 75 proteins, respectively. Most of these salt‐responsive proteins (70%) were classified into three major functional categories: disease/defense response, protein destination and storage and primary metabolism. The involvement of these proteins in salt tolerance of soybean was discussed, and some of them were suggested to be potential salt‐tolerant proteins. Furthermore, our results suggest that the cross‐protection against aldehydes, oxidative as well as osmotic stress, is the major adaptive response to salinity in soybean.     

Assessing the role of endophytic bacteria in the halophyte Arthrocnemum macrostachyum salt tolerance

• There is an increasing interest to use halophytes for revegetation of salt affected ecosystems, as well as in understanding their mechanisms of salt tolerance. We hypothesized that bacteria from the phyllosphere of these plants might play a key role in its high tolerance to excessive salinity.
• Eight endophytic bacteria belonging to Bacillus and closely related genera were isolated from phyllosphere of the halophyte Arthrocnemum macrostachyum growing in salty agricultural soils. The presence of plant‐growth promoting (PGP) properties, enzymatic activities and tolerance towards NaCl was determined. Effects of inoculation on seeds germination and adult plant growth under experimental NaCl treatments (0, 510 and 1030 mM NaCl) were studied.
• Inoculation with a consortium including the best performing bacteria improved considerably the kinetics of germination and the final germination percentage of A. macrostachyum seeds. At high NaCl concentrations (1030 mM), inoculation of plants mitigated the effects of high salinity on plant growth and physiological performance and, in addition, this consortium appears to have increased the potential of A. macrostachyum to accumulate Na⁺ in its shoots, thus improving sodium phytoextraction capacity.
• Bacteria isolated from A. macrostachyum phyllosphere seem to play an important role in plant salt tolerance under stressing salt concentrations. The combined use of A. macrostachyum and its microbiome can be an adequate tool to enhance plant adaptation and sodium phytoextraction during restoration of salt degraded soils.

     

Vegetative salt tolerance of barnyard grass mutants selected for salt tolerant germination

Improving salt tolerance of economically important plants is imperative to cope with the increasing soil salinity in many parts of the world. Mutation breeding has been widely used to improve plant performance under salinity stress. In this study, we have mutagenized Echinochloa crusgalli L. with sodium azide and three selected mutants (designated fows A) with salt tolerant germination. Their vegetative growth was compared to that of the wild type after short-term and long-term salt stress. The germination of the three fows A mutants in the presence of inhibitory concentrations of NaCl, KCL, and mannitol was better than that of the wild type. Early growth of the mutants in the presence of 200 mM NaCl was also better than that of the wild type perhaps due to improved K⁺ uptake and enhanced accumulation of sugars particularly sucrose at least in two mutants. But the three mutants and the wild type responded similarly to long-term salt stress. The tolerance mechanisms during short-term and long-term salt stress are discussed.     

Germination ecology of coastal plants in relation to salt environment

Responses of seed germination to salinity were examined using 37 species collected from salt marshes, cliffs, and fore (unstable) and hind (stable) sand dunes along Japanese coasts. For comparison, seed germination of nine inland species was also examined. The soil salinities in salt marshes ranged from 150 to 300 mmol/L NaCl, whereas those in fore and hind dunes ranged from 0 to 150 mmol/L NaCl, with a few exceptions. Cliff soils showed relatively high salinities up to 300 mmol/L NaCl. Ciff and foredune soils that encountered a typhoon and storm showed high salinities >300 mmol/L NaCl. Salt tolerance in seed germination of coastal plants was ordered by comparing the responses of percentage and rate of germination to salinity conditions up to 200 mmol/L NaCl, being in the order of salt marsh>cliff>foredune≅hind dune≅inland. Thse results indicate that salt tolerance in seed germination of coastal plants is closely related to the salinity conditions of their habitats. Germination experiments under favorable conditions showed that a high percentage of the seeds of salt marsh species germinate rapidly, those of diff species germinate slowly and those of foredune species exhibit a low percentage and low rate of germination. It seems that these germination characteristics contribute to the success of germination at the ‘safe site’ and the subsequent survivorship of emerged plants in their natural habitats.     

Osmotic sensitivity in relation to salt sensitivity in germinating barley seeds

Abstract Cultivars of barley (Hordeum vulgare L.) were tested for germination sensitivity to progressively higher concentrations of salt, mannitol, and betaine. The three solutes were equally inhibitory at equal osmotic potential, but there was a consistent difference in osmotic sensitivity between two cultivars, CM-67 and Briggs (Briggs was the most sensitive). There was no difference between the two cultivars in salt or water uptake from salt solutions during imbibition. Brief presoaking in water did not improve salt resistance, indicating that a hydration-dependent decrease in membrane permeability is not involved in salt tolerance. The calcium content of Briggs was higher than CM-67. These results suggest that salt inhibits barley germination primarily by osmotic effects, and that salt influx during imbibition does not play a role in this inhibition. A hypothesis regarding salt effects on germination is discussed.     

Photoperiod buffers responses to salt and temperature during germination of two coastal salt marsh colonizers Juncus acutus and Juncus maritimus

Juncus acutus and J. maritimus are two colonizers of coastal marsh rangeland with worldwide distribution. We tested the effect of salt, temperature, and photoperiod on the germination capacity of seeds of the two Juncus from the Rhône delta (south of France). We measured the first day of germination, the mean time, speed, and rate of germination on seeds subjected to five salinity levels, three Dark–Light temperatures and two D-L photoperiods (12–12 and 10–14). The 10–14 D-L photoperiod, although it corresponds to one of the two main germination periods of J. acutus and J. maritimus in the northern Mediterranean, has never been previously used to study their germination capacity. Analyses showed significant effects of salinity and temperature on the germination parameters tested, and for these two factors we found results comparable to those of previous studies. Salinity slowed down and reduced the germination process while spring temperatures had a positive effect. A surprising result was obtained by changing the D-L photoperiod from 12–12 to 10–14 which then largely buffered the negative effect of salt and high temperatures on germination. This capacity of J. acutus and J. maritimus to withstand salty conditions during the germination phase due to the spring photoperiod could be decisive in the ability of both species to colonize saline environments.     

Effects of maternal salinity on salt tolerance during germination of Suaeda aegyptiaca,a facultative halophyte in the Arab Gulf desert

Suaeda aegyptiaca is a facultative halophyte found in saline and non‐saline habitats of the Arab Gulf desert, which produces small‐sized undispersible seeds. The interactive effects of maternal salinity and other environmental conditions, such as salinity, light and temperatures, that are prevailing during seed germination have received little attention for a facultative halophyte. This study tested the effects of maternal salinity on salt tolerance during seed germination of S. aegyptiaca under different light and temperature regimes. Seeds collected from both saline and non‐saline habitats of the United Arab Emirates (UAE) were germinated in 0, 50, 100, 200 and 400 mM NaCl, and incubated at 15/25°C, 20/30°C and 25/35°C in both 12‐h light/12‐h dark regimes and continuous darkness. Generally, seeds of the non‐saline habitat were 56% heavier and attained greater germination at the lower temperatures than seeds of the saline habitat. Seeds of the saline habitat germinated better in saline solutions at higher temperatures and in light. Germination was faster for seeds of the saline habitat than for seeds of non‐saline habitats. Germination recovery after transfer to distilled water was significantly greater for seeds from the non‐saline habitat, compared with seeds from saline habitats. Recovery was greater at lower and/or moderate temperatures, compared with at higher temperatures. Germination was significantly faster during recovery, compared with in the saline solutions. The study indicates that the maternal effect of salinity was confounded with the seed‐size effect and it cannot be conclusively confirmed.     

Helophyte germination in a Mediterranean salt marsh: Gut‐passage by ducks changes seed response to salinity

Question: In seeds which are regularly consumed by waterbirds in the field, how does gut‐passage modify their response to salinity gradients? Location: Doñana National Park salt marsh, south‐west of Spain. Methods: Seeds of Scirpus litoralis and Scirpus maritimus were collected and force fed to mallards (Anas platyrhynchos). Both the ingested seeds (passage) and non‐ingested seeds (controls) were exposed, in germination chambers, to a salinity range similar to that observed in the field (0–32 dS/m). After 30 days, the total percentage germination, the duration of the dormancy period and the germination speed were computed. The response of the different germination parameters to ingestion and salinity was analyzed using generalized lineal models. Recovery tests on seeds that did not germinate in the various treatments and tests of the effect of ingestion on the intrinsic variability in seed response were also performed. Results: An increase in salinity reduced germinability and increased the length of dormancy, while gut pas sage increased the intrinsic variability of the temporal seed response in both species. In S. litoralis there was a significant interaction between the effects of salinity and passage on germination rate. Passage increased germination rate at low salinities (≤2 dS/m) but decreased it at high salinities (≥4 dS/m). Conclusion: Gut‐passage by ducks significantly changes seed response to salinity. The outcome of plant‐animal interactions can be influenced by environmental gradients. Studies of germination in response to gut passage that do not take such gradients into account may produce misleading results.     

High salinity impacts germination of the halophyte Cakile maritima but primes seeds for rapid germination upon stress release

Seed germination recovery aptitude is an adaptive trait of overriding significance for the successful establishment and dispersal of extremophile plants in their native ecosystems. Cakile maritima is an annual halophyte frequent on Mediterranean coasts, which produces transiently dormant seeds under high salinity, that germinate fast when soil salinity is lowered by rainfall. Here, we report ecophysiological and proteomic data about (1) the effect of high salt (200 mM NaCl) on the early developmental stages (germination and seedling) and (2) the seed germination recovery capacity of this species. Upon salt exposure, seed germination was severely inhibited and delayed and seedling length was restricted. Interestingly, non‐germinated seeds remained viable, showing high germination percentage and faster germination than the control seeds after their transfer onto distilled water. The plant phenotypic plasticity during germination was better highlighted by the proteomic data. Salt exposure triggered (1) a marked slower degradation of seed storage reserves and (2) a significant lower abundance of proteins involved in several biological processes (primary metabolism, energy, stress‐response, folding and stability). Yet, these proteins showed strong increased abundance early after stress release, thereby sustaining the faster seed storage proteins mobilization under recovery conditions compared to the control. Overall, as part of the plant survival strategy, C. maritima seems to avoid germination and establishment under high salinity. However, this harsh condition may have a priming‐like effect, boosting seed germination and vigor under post‐stress conditions, sustained by active metabolic machinery.     

The effect of salinity on different developmental stages of an endemic annual plant, Aster laurentianus (Asteraceae)

Salinity reduces substrate water potential, thereby restricting water and nutrient uptake by plants; salinity may also cause ionic imbalance and toxicity. Because substrate salinity fluctuates through the growing season, a plant may be exposed to different salinity levels, at various stages of development, with potentially significant consequences on population dynamics. Here, we present the results of a study of the effect of substrate salinity on seed germination, seedling emergence, and growth of Aster laurentianus, an annual marsh plant, endemic to the Gulf of St. Lawrence and potentially threatened. Seed germination was reduced in low salt concentration (10 g sea salt/L) and completely inhibited by salinity levels >/=20 g sea salt/L. However, this inhibiting effect was reversible: seeds from the salt treatments germinated readily after being washed in distilled water. Though seedling emergence was diminished at low salinity levels, postemergence survival was little affected. Plant growth was reduced, but net carbon assimilation rate was not affected by high salinity levels. Increased root respiration and respiratory costs associated with salt tolerance might have contributed to lower C accumulation at higher salinity levels. All developmental processes considered are thus negatively affected by substrate salinity, with potentially significant consequences on population abundance and distribution in salt marshes. Yet, the tolerance of this species to high salinity levels after seedling emergence is remarkable. Seed germination represents a major bottleneck in the species life cycle, potentially controlling local distribution and abundance in the natural habitat.     

The role of Rhytidoponera metallica (Hymenoptera,Formicidae) in facilitating post‐fire seed germination of three ant‐dispersed legume species

The fire avoidance hypothesis proposes that a benefit of seed dispersal by ants (myrmecochory) is to protect seeds from being killed during fire and to facilitate post‐fire germination of seeds that require heat shock to break their physical dormancy. The aim of this study was to quantify the effect of fire and seed burial by a predominant seed‐dispersing ant, Rhytidoponera metallica (subfamily: Ectatomminae) on germination levels of three ant‐dispersed legume species (Pultenaea daphnoides, Acacia myrtifolia and Acacia pycnantha). Experimental burial of seeds within aluminium cans at a site prior to being burnt and at an adjacent unburnt site showed that fire increased germination levels, particularly for seeds buried at 1‐ and 2‐cm deep and that overall, germination levels differed among the three plant species. To quantify seed burial depths and post‐fire germination levels facilitated by R. metallica ants, seeds were fed to colonies prior to fire at the burnt and unburnt sites. Of the seeds buried within nests that were recovered, between 45% and 75% occurred within the upper 6 cm of the soil profile, although unexpectedly, greater percentages of seeds were recovered from the upper 0–2 cm of nests in the unburnt site compared with nests in the burnt site. Germination levels of buried seeds associated with R. metallica nests ranged from 21.2% to 29.5% in the burnt site compared with 3.1–14.8% in the unburnt site. While increased seed germination levels were associated with R. metallica nests following fire, most seeds were buried at depths below those where optimal temperatures for breaking seed dormancy occurred during the fire. We suggest that R. metallica ants may provide fire avoidance benefits to myrmecochorous seeds by burying them at a range of depths within a potential germination zone defined by intra‐ and inter‐fire variation in levels of soil heating.     

Light,temperature, dry after‐ripening and salt stress effects on seed germination of Phleum sardoum (Hackel) Hackel

Phleum sardoum is an endemic psammophilous species of Sardinia, growing exclusively on coastal sandy dunes. The effect of glumes on seed germination, germination requirements at constant (5–25°C) and alternating (25/10°C) temperatures, both in the light (12/12 h) and in the dark were evaluated, as well as the effect of a dry after‐ripening period (90 days at 25°C), the salt stress effect (0–600 mmol NaCl) and its recovery on seed germination. The presence of glumes reduced final germination percentages. For fresh naked seeds, high germination percentages were observed at 10°C. Dry after‐ripening increased germination rate at low temperatures, but did not affect final germination percentages. NaCl determined a secondary salt‐induced dormancy which recovery interrupted only partially. Our results highlighted that this species has its optimum of germination during autumn–winter when, under a Mediterranean climate, water availability is highest and soil salinity levels are minimal.     

Seasonal changes in the germination responses of Hordeum maritimum and H. murinum seeds in relation to salinity,temperature and after‐ripening time

Abstract

The effects of temperature and salinity (NaCl) on germination of Hordeum maritimum With, (halophyte) and H. murinum L. (glycophyte) seeds were investigated. Dehulled caryopses were used for monthly germination trials, starting from November (120 days of after‐ripening in darkness at 20±1°C). Trials were continued for one year. Differences in germination response between the two species were observed, confirming that H. Maritimum is better adapted to high salinity levels and to variations in external temperature than H. murinum. H. maritimum showed a germination control mechanism related to after‐ripening time and based on seed dormancy break/resumption. At higher temperature (30°C), thermodormancy was also recorded. No germination strategies were observed in H. murinum that is relatively insensitive to the combined effects of temperature and salinity. Thus, in virtually all treatments, H. murinum exhibited a higher germination rate compared with H. maritimum, as early as 72 h after imbibition, suggesting that dormancy, both in the presence or absence of salt, is totally abolished by early fall or at the latest in winter.     

Comparative salt tolerance of two wild Hordeum species (H. maritimum with. and H. murinum L.) from the coast of Tuscany (Italy)

ABSTRACT

The differences in salt tolerance between Hordeum maritimum and H. murinum were studied. Seeds were collected at horn maturity from wild populations growing respectively near the Orbetello Lagoon and S. Piero a Grado (Tuscany, Italy) and were used in germination and growth tests at increasing salinity (NaCl) levels. H. maritimum was confirmed to be a true halophyte as compared to H. murinum, which exhibited germination behavior typical of many wild glycophytes. The higher salt sensitivity of H. murinum compared to H. maritimum was also shown by its shoot length values, which decreased only in H. murinum, albeit in 100 mM NaCl treatment. The higher degree of salt tolerance of H. maritimum is further demonstrated by the Na : K ratio. H. maritimum can accumulate a greater amount of sodium than potassium in both roots and shoots, even in the absence of salt treatment. However, in NaCl solutions H. maritimum showed a higher Na : K ratio for shoots — an index of better uptake and translocation of sodium to leaves — as has been demonstrated for many halophyte includers. These results thus help to enhance knowledge on wild relatives of barley, whose potential contribution to genetic improvement in salinity tolerance has previously not been thoroughly explored.     

Ecological role of distinct fruit‐wing perianth color in synchronization of seed germination in Haloxylon salicornicum

Haloxylon salicornicum is a perennial shrub that grows in sandy and stony desert areas. This species produces two types of dispersal unit with yellow and pink winged perianths that are wind dispersed. This species has been considered as an excellent sand binder and can tolerate extreme temperature, drought and salinity. We compared the germination characteristics of both colored dispersal units (with and without a wing) under various light, temperature and salinity treatments. It was found that wing color, presence of wings, temperature and light have a significant effect on germination percentage and germination rate. The presence of a winged perianth significantly reduced the germination percentage and germination rate in both yellow and pink dispersal units. De‐winged yellow dispersal units were able to tolerate higher salinity and, therefore, their germination was higher in all the tested concentrations of NaCl compared with pink de‐winged dispersal units. However, salinity tolerance in both pink and yellow dispersal units decreased with increasing salinity concentration. The germination‐recovery percentage was also higher for yellow compared with pink dispersal units. The present study has demonstrated that both morphological and physiological differences exist in different colored dispersal units, which might represent the combination of different complementary adaptive strategies and have ecological significance for Haloxylon salicornicum's successful survival in harsh desert environmental conditions.     

Involvement of nitrate reductase-dependent nitric oxide production in magnetopriming-induced salt tolerance in soybean

In the present study, experiments were performed to investigate the role of nitric oxide (NO) in magnetopriming-induced seed germination and early growth characteristics of soybean (Glycine max) seedlings under salt stress. The NO donor (sodium nitroprusside, SNP), NO scavenger (2-[4-carboxyphenyl]-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide, CPTIO), inhibitors of nitrate reductase (sodium tungstate, ST) or NO synthase (N-nitro-L-Arg-methyl ester, LNAME) and NADPH oxidase inhibitor (diphenylene iodonium, DPI) have been used to measure the role of NO in the alleviation of salinity stress by static magnetic field (SMF of 200 mT, 1 h). Salt stress (50 mM NaCl) significantly reduced germination and early growth of seedlings emerged from non-primed seeds. Pre-treatment of seeds with SMF positively stimulated the germination and consequently promoted the seedling growth. ST, LNAME, CPTIO and DPI significantly decreased the growth of seedling, activities of α-amylase, protease and nitrate reductase (NR), hydrogen peroxide (H₂O₂), superoxide (O₂^•−) and NO content in roots of seedlings emerged from non-primed and SMF-primed seeds. However, the extent of reduction was higher with ST in seedlings of SMF-primed seeds under both conditions, whereas SNP promoted all the studied parameters. Moreover, the generation of NO was also confirmed microscopically using a membrane permanent fluorochrome (4-5-diaminofluorescein diacetate [DAF-2 DA]). Further, analysis showed that SMF enhanced the NR activity and triggered the NO production and NR was maximally decreased by ST as compared to LNAME, CPTIO and DPI. Thus, in addition to ROS, NO might be one of the important signaling molecules in magnetopriming-induced salt tolerance in soybean and NR may be responsible for SMF-triggered NO generation in roots of soybean.     

Effects of salinity and temperature on ex situ germination of the threatened Gulf of St. Lawrence Aster,Symphyotrichum laurentianum Fernald (Nesom)

The threatened Gulf of St. Lawrence Aster, Symphyotrichum laurentianum Fernald (Nesom), is an annual coastal halophyte of the southern Gulf of St. Lawrence, Canada. We examined the effects of salinity (0–20 g/L) and temperature (16–30°C) on germination of S. laurentianum seeds over 32 days. The time‐course of germination was significantly affected by both salinity and temperature. At lower temperatures (16°C and 23°C), germination was inhibited by salt water at days 16 and 32. However, at 30°C germination rates after 16 days were highest at an intermediate salinity, whereas after 32 days germination was uniformly high in all salinity treatments. Overall, the effect of temperature on germination was much stronger than the effect of salinity. Delays in germination resulting from exposure to salinity or from low soil temperatures could set up strong size asymmetries between seedlings of S. laurentianum and the surrounding vegetation, leading to suppression of growing seedlings via shading. Because germination has the potential to be a significant population bottleneck for this seed‐dependent annual, conservation efforts should consider microsite suitability for germination in the management of natural populations and in the selection of sites for explants.     

Effects of salinity levels and seed mass on germination in Australian species of Frankenia L. (Frankeniaceae)

Halophyte species demonstrate differing levels of salt tolerance. Understanding interspecific variation to salinity levels is of value from both the scientific perspective, which includes the identification of traits associated with salinity tolerance, as well as from an applied perspective, which includes identifying plant species for specific salinity restoration and remediation projects. This paper investigates the effects of salinity on germination of 12 Australian species of the plant genus Frankenia L. (Frankeniaceae). We use saline solutions that corresponded to the average soil–water salinity concentrations in the arid zones of inland Australia. These solutions consisted of 10 mM calcium chloride, 30 mM magnesium sulphate, and 450 mM sodium chloride. The aims of our study were: (1) to investigate the germination (germination rates, germination success) of Frankenia seeds to four salinity levels (0%, 10%, 20%, 30%), (2) to test for possible interaction effects between seed mass, germination, and salinity, and (3) to examine the effect of salinity levels on the inhibition of germination and/or seed damage. Species varied in their salt tolerance for germination rates and success. Species with larger seeds had higher germination rates and germination success for high salinity levels. Several species did not germinate well at any salinity level. Finally, no seeds were adversely affected by exposure to high salinity levels pre-germination. There is potential for including some Frankenia species in remediation and revegetation projects in areas affected by salinity, and also as garden plants in saline regions.     

Imbibition phases and germination response of Mimosa bimucronata (Fabaceae: Mimosoideae) to water submersion

Mimosa bimucronata is a pioneering tree that occurs predominantly in moist lowlands, floodplains and on margins of rivers and lakes in Latin America. The effect of submergence on seed germination in M. bimucronata was firstly studied. Patterns of water absorption by M. bimucronata seeds were investigated thereafter to assess the imbibition phases of scarified and unscarified seeds. The germination percentage was significantly higher in scarified than in unscarified seeds, and the velocity of seed germination also increased considerably in scarified seeds. Submergence duration did not significantly affect germination percentages of scarified and unscarified seeds. Therefore, seed viability after submersion suggests that M. bimucronata may display hydrochorous dispersal and also that seeds are able to germinate successfully in areas with frequent seasonal flooding. With respect to imbibition phases, phase II was very short or even absent for scarified and unscarified seeds; therefore, a plateau, where water absorption by seeds is established, was not observed. Finally, we verified that the passage from phase I to III was very tenuous and took a long time in seeds without scarification.