This paper discusses how field and laboratory experiments, using a variety of genetic material, can be combined to investigate the genetic basis of germination under realistic ecological conditions, and it reviews some of our recent work on germination phenology of Arabidopsis thaliana in the field. Our results indicate that the genetic basis of germination depends on the environment. In particular, the conditions during seed maturation interact with post-dispersal environmental factors to determine germination phenology, and these interactions have a genetic basis. Therefore genetic studies of germination need to consider carefully the environment – both during seed maturation and after dispersal – in which the experiments are conducted in order to characterize genetic pathways involved with germination in the field. Laboratory studies that explicitly manipulate ecologically relevant environmental factors can be combined with manipulative field studies. These studies can identify the particular environmental cues to which seeds respond in the field and characterize the genetic basis of germination responses to those cues. In addition, a variety of genetic material – including mutant and transgenic lines, intact natural genotypes, recombinant genotypes, and near isogenic lines – can be used in field studies as tools to characterize genetic pathways involved in germination schedules under natural ecological conditions.

Higher plants have several strategies to perpetuate themselves under adequate ecophysiological conditions. The production of heterogeneous seeds is one such strategy. That is, to ensure the survival of the next generation, an individual plant might produce seeds that are heterogeneous with respect to the extent of dormancy, dispersion and persistence within the seed bank. Heterogeneity can affect not only certain physiological and molecular properties related to seed germination, but also such characteristics as colour, size and shape, parameters commonly used to differentiate morphs within a heterogeneous seed population. In heterogeneous seeds, the above features determine seed behaviour and alter their mechanism of germination. In this work, emphasis is placed on the existence of seed mutants having major alterations in characteristics of the testa and hormonal response. These mutants constitute a valuable tool for elucidating the mechanism of dormancy, germination and perpetuation of seeds. Finally, ontogeny and heterogeneity are reviewed, providing the first data related to the possible hormonal control of heterogeneity in seeds. These results raise the hypothesis that one of the factors triggering differences in germination among heterogeneous seeds may be an alteration in the signalling and action mechanism of ethylene and abscisic acid (ABA).

Germinated seeds of Medicago truncatula Gaertn. with a protruded radicle length of 2.7 mm did not survive drying below 0.2 g H2O g–1 dw, as indicated by vital stain assays and the absence of growth resumption after rehydration. The re-establishment of desiccation tolerance was achieved using an osmotic treatment with polyethylene glycol (PEG), combined with a cold treatment. The ability to regain desiccation tolerance after germination was restricted to a period of growth characterized by radicle lengths between 1 and 3 mm. After PEG treatment of germinated seeds with 2.7 mm long radicles at –1.7 MPa at 10°C for 3 d and subsequent drying to 0.04 g H2O g–1 dw, 90% survived and developed into normal seedlings after rehydration. Desiccation tolerance could also be re-established in excised radicles, demonstrating that cotyledons were not essential for this process. Upon PEG incubation, sucrose accumulated rapidly prior to the re-establishment of desiccation tolerance in germinated radicles, regardless of the presence of cotyledons. Induction of MtDHN (a dehydrin) gene expression was correlated with the re-establishment of desiccation tolerance. Furthermore, the PEG-induced expression of MtDHN was repressed when fluridone was added to the PEG solution.

This study examines whether post-dispersal seed predators could be an important selective force in determining the seed bank strategies of grassland plants. It tests the hypothesis that species with persistent seed banks should sustain proportionally less predation of buried seeds than species which have transient seed banks and that this should be true irrespective of seed size. Results are drawn from a field experiment examining the relative susceptibility of surface versus buried seeds for 19 herbaceous taxa exhibiting different degrees of seed bank persistence. The data were consistent with the hypothesis that seed predators (rodents) influence the seed bank characteristics of seeds. Rodents removed proportionally more large seeds than small seeds and removed a smaller proportion of seeds with persistent rather than transient seed banks, independently of seed size. On average, burial reduced seed removal by almost 50%. The decrease in rates of seed removal following burial was marked for seeds with persistent seed banks but negligible for seeds with transient seed banks. Herbaceous plants with relatively large seeds (seed mass > 1 mg) that form persistent seed banks were either completely avoided or only consumed in small quantities by rodents. In contrast, large-seeded species with transient seed banks suffer high rates of seed predation. Models of life-history evolution predict trade-offs between seed dormancy and seed mass since dormancy and seed size are correlated traits that both reduce risk in variable environments and thus will show patterns of negative covariation. This paper presents an alternative explanation for this trade-off based on experimental evidence of a negative relationship between seed bank persistence and predation risk.

Seed size is a good predictor of seed persistence in soil for British, Argentinean, Iranian and – to some extent – New Zealand species. It has been suggested that seed shape should also be linked to the ease of burial and, thus, to seed persistence, even if some studies failed to show this. The relationship between seed size and shape and persistence in soil was analysed for 259 species of the Italian flora, belonging to a wide range of habitats, from alpine pasture to limestone prairies and meadows of the Prealps, and from woodlands to Mediterranean maquis and garigues. Seed size was related to persistence in soil in the same way as in most other floras examined. Furthermore, seed shape was highly related to persistence in soil among the species analysed, when considered both altogether and divided among the different habitats. Our results suggest that not only seed size, but also seed shape, are key factors in determining seed fate and seed persistence in soil.

Ecophysiological aspects of seed germination were investigated in the widely distributed Mediterranean-endemic, aromatic plants thyme (Coridothymus capitatus), savory (Satureja thymbra) and oregano (Origanum vulgare subsp. hirtum). Thyme seed germination is light indifferent, whereas oregano seeds have an absolute light requirement; their germination can be promoted even by green safelight or far-red light. In savory, a portion of the seeds germinates in the dark and germination can be either increased or decreased by the appropriate illumination. All three species show more germination at a relatively low temperature range, a Mediterranean characteristic, with an optimum around 15–20°C. The rate of germination is considerably higher in the tiny seeds of thyme and oregano than in the larger seeds of savory. In the latter species, germination is also dependent upon the age of the seeds; old seeds germinate to a higher percentage than fresh ones, as already observed by Theophrastus, possibly as a result of the volatilization of the essential oils present on the nutlet coat. Seeds of thyme, savory and oregano are dispersed within the persistent fruiting calyces and the seeds eventually germinate within it. Essential oils in the calyx strongly inhibit germination of the enclosed seeds; germination is much suppressed in thyme and to a lesser extent in savory and oregano. This diaspore dormancy caused by essential oils apparently is overcome under natural conditions by leaching of the inhibitors with rainwater. It is suggested that this dormancy operates as an adaptation strategy that delays germination by acting as a rain gauge. In this way, seed germination and subsequent seedling establishment are prevented during the early phase of the rainy period, which is usually interrupted by drought spells in the Mediterranean climate.

The ascorbic acid (ASC) system functions dynamically in seeds, although the strategies for ASC production and utilization may vary according to seed developmental and functional stages. In orthodox seeds, ASC content and ASC peroxidase activity increase during the early stages of development, then decrease during the desiccation stage, so that, at quiescence, seeds have neither ASC nor ASC peroxidase, but retain a small amount of dehydroascorbic acid (DHA) and significant activities of ASC recycling enzymes. ASC and ASC peroxidase activity re-start after a few hours from the onset of imbibition. In contrast, the ASC system is little affected during germination of recalcitrant seeds. Although the presence of the ASC system in seeds has often been considered only within the framework of seed antioxidant defences, ASC function in seeds is also likely to be related to its action as a specific co-substrate required for the activity of dioxygenases (e.g. 1-aminocyclopropane carboxylate oxidase, gibberellic acid hydroxylases and 9-cis-epoxycarotenoid dioxygenases) involved in the synthesis of ethylene, gibberellins and abscisic acid, respectively. The possible role of ASC in coordinating the activities of these key enzymes is discussed.

A common feature of desiccation-tolerant organisms, such as orthodox seeds, is the presence of large quantities of sugars, especially di- and oligosaccharides. These sugars may be one component of the suite of adaptations that allow anhydrobiotes to survive the loss of most of their cellular water. This paper describes the physical effects of dehydration on cellular ultrastructure, with particular emphasis on membranes, and explains quantitatively how sugars and other solutes can influence these physical effects. As a result of dehydration, the surfaces of membranes are brought into close approach, which causes physical stresses that can lead to a variety of effects, including demixing of membrane components and fluid-to-gel phase transitions of membrane lipids. The presence of small solutes, such as sugars, between membranes can limit their close approach and, thereby, diminish the physical stresses that cause lipid fluid-to-gel phase transitions to occur during dehydration. Thus, in the presence of intermembrane sugars, the lipid fluid-to-gel phase transition temperature (Tm) does not increase as much as it does in the absence of sugars. Vitrification of the intermembrane sugar solution has the additional effect of adding a mechanical resistance to the lipid phase transition; therefore, when sugars vitrify between fluid phase bilayers, Tm is depressed below its fully hydrated value (To). These effects occur only for solutes small enough to remain in very narrow spaces between membranes at low hydration. Large solutes, such as polymers, may be excluded from such regions and, therefore, do not diminish the physical forces that lead to membrane changes at low hydration.

Temperature is a key factor affecting both dormancy and germination. In non-dormant seeds, when temperature is within the thermal range permissive for germination, it just regulates germination velocity, while in seeds presenting dormancy it can also be affecting dormancy level, dormancy termination and the expression of dormancy itself. This dual effect of temperature on dormancy and germination often leads to misinterpretation of obtained germination results and confounds the analysis of temperature effects in seed populations presenting some degree of dormancy. In the present paper we discuss the effect of temperature in the regulation of dormancy level and its implications in dormancy expression, as an attempt to construct a conceptual framework that allows distinguishing between the effects of temperature on dormancy and germination. Finally, we present examples of how a better understanding of these effects could help us to interpret the mixed effects of temperature on both processes during incubation of seeds presenting dormancy.

Strigolactones are apocarotenoids regulating shoot branching. They are also known to be exuded by plant roots at very low concentrations, stimulating hyphal branching of arbuscular mycorrhizal fungi and germination of root parasitic weed seeds. We show that strigolactones play a major role in host specificity of Orobanche and Phelipanche (the broomrapes) seed germination. This observation confirms that host-derived germination stimulants are an important component determining the host specificity of these parasitic plants. Weedy broomrape species were less specialized in germination requirements than the non-weedy species except for O. cumana and O. foetida var. broteri. Similar results were obtained with the root exudates. Some species, such as P. aegyptiaca and O. minor, showed a broad spectrum of host specificity in terms of seed germination, which was stimulated by exudates from the majority of species tested, whereas others, such as O. cumana, O. hederae and O. densiflora, were highly specific. Some species, such as O. minor, P. aegyptiaca and P. nana, were responsive to the three strigolactones studied, whereas others were induced by only one of them, or did not respond to them at all. The synthetic strigolactone analogue GR24, generally used as a standard for germination tests, was not effective on some Orobanche and Phelipanche species. Seeds of some species that did not respond to GR24 were induced to germinate in the presence of fabacyl acetate or strigol, confirming the role of strigolactones in host specificity.

Books and review articles in various areas of ecology and seed and plant biology continue to report that dormancy-break in seeds (and fruits) with water-impermeable coats (i.e. physical dormancy) occurs via soil-microbial action and/or abrasion by soil particles. However, there is little evidence in the scientific literature to support these assumptions, which, in fact, do not make good evolutionary sense for two related reasons. First, several types of anatomically specialized water-restriction structures have evolved as part of the seed or fruit coat of taxa with physical dormancy. These structures act as ‘signal detectors’ of physical-environmental changes that cause seeds (and fruits) to become water-permeable only at these sites, in seasons and habitats in which there is a good chance that some seedlings will become established. Second, seed (and fruit) coat breakdown by microbial action or by abrasion likely would occur in seasons and habitats in which seedlings could not survive, thus lowering the fitness (λ) of the plant taxa in question.

Germination is an energy-demanding process that requires the operation of mitochondria, which must survive desiccation in the quiescent seed and become rapidly functional after imbibition to meet the ATP demand. The relationship between germination and mitochondrial performance was addressed by analysing the properties of mitochondria isolated from control, primed and aged pea (Pisum sativum L.) seeds. Mitochondria were isolated and purified at early stages of germination (before radicle protrusion), and their oxidative properties, membrane integrity and ultrastructure were examined. Mitochondria isolated after 12 h of imbibition readily oxidized exogenous NADH and Krebs cycle substrates at high rates. However, their phosphorylation efficiency was restricted by poor membrane integrity. After 22 h from the beginning of imbibition, purified seed mitochondria had intact outer membranes and oxidized the substrates at slightly lower rates, but with higher respiratory control (improved capacity for phosphorylation). Purified seed mitochondria were always found to be deficient in endogenous NAD, although the organelles were capable of importing and retaining the cofactor. While the priming treatment appeared to slightly increase the performance of mitochondria, seed deterioration by accelerated ageing strongly affected the oxidative properties of mitochondria, which were badly impaired in ATP production. Outer and inner membrane integrity was identified as the primary target for desiccation and ageing stress. A link between mitochondrial function and seed quality was also corroborated by respiration measurements of seed fragments at the onset of imbibition.

Trials were carried out to investigate natural weed seed burial in a no-till agricultural system. With this aim, 30 important weed species were selected to explore if, and to what extent, seed vertical movement is conditioned by soil texture, cumulative rainfall or by seed characteristics. Without soil tillage, the vertical position of the seeds was only a few millimetres below the surface, but it was dependent on soil texture. Seed burial was much slower (and to lesser depths) in clay soils than in sandy soils. Seeds reached greater depths after 1 year in sandy soil (10% >6 mm) than in clay soils (2% >6 mm). Burial dynamics were also influenced by rainfall and seed weight. The amount of rain necessary to halve weed seeds observable on the soil surface was a function of the soil texture and seed weight; linear regressions between total rainfall amounts and seed weight showed that the slope of the relationship increased as the sand component of the soil decreased (21.1, 35.4 and 39.4 in sandy, loam and silty-clay soil, respectively). A polynomial regression, carried out in sandy soil, between 1000-seed weight and their burial capacity showed an inverse relation (P>0.05) between these two parameters. Seed shape and coat microsculpture also influenced movement, but only minimally.

Development of the highly desiccation-sensitive (recalcitrant) seeds of primarily one species, Avicennia marina, is reviewed and compared with the ontogeny of desiccation-tolerant (orthodox) seeds. A. marina seeds undergo no maturation drying and remain metabolically active throughout development, which grades almost imperceptibly into germination. While PGR control of histodifferentiation is essentially similar to that characterizing desiccation-tolerant seeds, the phase of growth and reserve deposition is characterized by exceedingly high cytokinin levels which, it is proposed, promote a sink for assimilate import. While some starch accumulation does occur, the predominant reserves are soluble sugars which are readily available for the immediate onset of seedling establishment upon shedding. ABA levels are negligible in the embryo tissues during seed maturation, but increase in the pericarp, which imposes a constraint upon germination until these outer coverings are sloughed or otherwise removed. The pattern of proteins synthesized remains qualitatively similar throughout seed development in A. marina, and no LEA proteins are produced. This suggests both that seedling establishment is independent of maturation proteins and that the absence of LEAs and desiccation sensitivity might be causally related. The study on A. marina reveals that for this recalcitrant seed-type, germination per se cannot be defined: rather, it is considered as the continuation of development temporarily constrained by the pericarp ABA levels. This leads to a reexamination of the role of rehydration as key event sensu stricto, in the germination processes in desiccation-tolerant (orthodox) seeds.

Dormancy in seeds of Dodonaea viscosa is due to a water-impermeable seed coat (physical dormancy, PY). Thus, mechanically scarified seeds imbibed water (c. 95% increase in mass) and germinated to high percentages over a wide range of temperature regimes in both white light and darkness, whereas non-scarified seeds did not take up water. Dry heat at 80–160°C and dipping in boiling water for 1–60 s also broke dormancy in a high percentage of the seeds, and continuous far-red light was not inhibitory to germination. However, dry storage in the laboratory for >1 year did not overcome dormancy. Seeds made water-permeable by boiling imbibed water, and thus germinated, at a much slower rate than those made water-permeable by mechanical scarification. We suggest that boiling opened the ‘water gap’ in the seed coat (not yet described in Sapindaceae but present in other taxa with PY) and that water entered the seed only through this small opening, thereby accounting for the slow rate of imbibition and subsequent germination. Physical dormancy has now been shown to occur in seeds of this polymorphic, worldwide species from Australia, Brazil, Hawaii, Mexico and New Zealand. The low level of dormancy reported for seed lots of D. viscosa in China, India and Pakistan is probably due to collection of seeds before they dried to the critical moisture content for development of water-impermeability of the seed coat. Germination of non-dormant seeds over a wide range of temperatures and in white light, far-red (leaf-canopy shade) light and darkness are part of the germination strategy of D. viscosa and of other taxa whose seeds have PY at maturity.

Computer software called WeedCast was developed to simulate weed seed dormancy, timing of seedling emergence, and seedling height growth in crop environments in real-time and using actual or forecasted weather data. Weather data include daily rainfall and minimum and maximum air temperatures. Air temperatures are converted to average daily soil temperature at 5-cm soil depth using a series of equations that are specific for soil type, tillage system and previous year's crop-residue type. Daily rainfall and soil temperature estimates are combined to determine soil water potential (in megapascals) at 5-cm depth. Daily estimated soil water potential or soil temperatures are matched to empirically-derived threshold values that induce secondary dormancy in seeds of certain species. Soil growing degree days (GDD), calculated from soil temperatures, are used to project maximum emergence rates of weed seedlings. Emergence ceases on days when soil water potential falls below threshold values specific to each species. GDD based on air temperatures are used to estimate post-emergence seedling height growth. All three types of simulation provide information that allows users to answer important weed management questions in real-time. These types of questions include but are not limited to the following: (1) Are soil-applied treatments necessary? (2) How late can pre-emergence herbicides be applied? (3) When should mechanical control be implemented? (4) When should field-scouting commence and end? (5) When should post-emergence herbicides be applied?

Anhydrobiosis, or life without water, is the remarkable ability of certain types of plants and animals to survive almost total dehydration. This phenomenon requires a coordinated series of events within the cells of anhydrobiotes that protect their cellular components, particularly proteins and lipid membranes, from damage caused by the removal of water. Much of what is now understood about preserving biological samples during drying was learned by studying naturally desiccation-tolerant organisms and extended using model systems such as phospholipid vesicles. Most anhydrobiotic organisms accumulate disaccharides in their cells and tissues during the dehydration process. These carbohydrates, usually sucrose or trehalose, satisfy two criteria that appear to be necessary for protecting membranes during desiccation and during storage in the dry state. These requirements include: (1) depression of the gel-to-liquid crystalline phase transition temperature (Tm) in the dehydrated lipid to a temperature at or near that of the hydrated lipid, a process that appears to require a direct interaction between the carbohydrates and the lipid molecules of the membrane; and (2) formation of a carbohydrate glass with a relatively high glass transition temperature, leading to inhibition of fusion between the vesicles.

Suaeda salsa is an annual herbaceous euhalophyte in the family Chenopodiaceae that produces dimorphic seeds on the same plant under natural conditions. In order to determine the effect of salinity on seed quality traits during seed formation, seeds from plants grown under control conditions and on 200 mM NaCl were used to investigate the effect of NaCl on seed production and seed germination. Results showed that size and weight of both black and brown seeds generated from 200 mM NaCl-treated plants were markedly greater than those from controls. The germination percentage of brown seeds from both control and NaCl-treated plants was higher than that of black seeds. Furthermore, the germination percentage of the black seeds generated from 200 mM NaCl-treated plants was significantly higher than that of the control at different concentrations of NaCl, although germination percentage declined with the increase NaCl concentration. Surprisingly, NaCl did not affect germination of the brown seeds. The germination index and vigour index of both black and brown seeds from the control plants were significantly lower than those of seeds from the different NaCl treatments. Seed starch, soluble sugar, protein and lipid content of both black and brown seeds generated from the 200 mM NaCl-treated plants were significantly higher than those from the control. These results suggest that a certain concentration of NaCl plays a pivotal role in seed vitality of the euhalophyte S. salsa through increasing seed weight and contents of storage compounds such as protein, starch and fatty acids.

A large fraction of tree species forming persistent soil seed-banks and with dormant seeds are expected to be found in strongly seasonal ecosystems such as Neotropical savannas, where seedling recruitment could be highly variable. In the savannas of Central Brazil, we studied seed characteristics (type of dormancy, longevity and moisture content) of 14 representative woody species differing in seed dispersal season. We also studied the dynamics of soil seed-banks and similarity patterns in woody species composition among seed rain, soil seed-bank, seedling bank and standing vegetation along shallow topographic gradients that differ in canopy cover. Woody species composition of the soil seed-bank largely differed from the standing vegetation, the seed rain and the seedling bank species composition, suggesting low recruitment of woody species from the soil seed-bank. Seeds of the 14 woody species remained viable for less than 16 months in laboratory dry-storage conditions. Of those, most seeds dispersed in the dry season were dormant and exhibited low moisture content, while most seeds dispersed in the wet season were non-dormant and exhibited high moisture content. Longevity of these seeds dispersed in the dry and the wet seasons did not differ significantly. This study shows that both timing of seed dispersal and dormancy appear to control timing of seed germination and seedling recruitment of most Neotropical savanna woody species, which did not form persistent soil seed-banks. This study contributes to the understanding of tree/grass coexistence and tree density variations along topographic gradients in tropical savannas.

Dehydrins are late embryogenesis abundant (LEA) proteins that accumulate during seed maturation and in response to abiotic stresses in vegetative tissues. They are thought to protect cellular components from dehydration stress. However, whether they play a role in survival in the dry state is not clear. In this study, an RNA interference (RNAi)-construct against the seed-expressed dehydrin of Arabidopsis thaliana, LEA14 (At2g21490), was introduced to wild-type plants, which led to a strong reduction in transcript abundance of the target gene as well as that of two other seed-expressed dehydrin homologues, XERO1 (At3g50980) and RAB18 (responsive to abscisic acid 18, At5g66400) in the transformants. Mature, dry seeds from the RNAi plants germinated to at least 95% after rehydration, indicating that seed desiccation tolerance was not affected, while they exhibited a twofold reduction in longevity. When stored at 75% relative humidity and 35°C, the seeds of two independent RNAi lines lost 50% of their viability in 10 d and 5 d, respectively, while it took 17 d for wild-type seeds to lose 50% viability. In addition, when seeds were imbibed in the presence of 100 mM NaCl, the seeds of RNAi plants exhibited reduced germination compared to wild-type seeds, suggesting that at least one of the three seed-specific dehydrins plays a role both against deterioration during storage at low moisture content and when imbibed tissues are submitted to salt stress at high moisture.