Desiccation Sensitivity Research Articles

Desiccation tolerance of Tapirira obtusa seeds collected from different environments

This study was aimed at evaluating the desiccation sensitivity in seeds of the tree Tapirira obtusa (Benth.) J. D. Mitchell collected from three different environments and subjected to two distinct drying speeds. Seeds were collected from a rocky area, in the "Cerrado", and in a riparian forest area, in the region of municipality of Lavras, State of Minas Gerais. The seeds were subjected to drying with magnesium chloride (slow drying) or silica gel (fast drying), into closed environment, until moisture contents of 40%, 30%, 20% and 10%, considering as control, the percentage of germination at the initial moisture content in each environment, which varied from 47% to 50%. Percentages of germination and normal seedlings as well as germination speed index were assessed. For the three environments studied, there was no effect of slow drying on seed germination. Seeds from area of Cerrado, however, have shown a slight reduction on germination when subjected to fast drying. Oppositely, seeds from rocky area had germination increased when subjected to fast drying. Seeds from riparian forest area had no reduction on germination percentage, independent of drying speed. Results suggest that seeds of T. obtusa are not sensitive to desiccation.

The Re-Establishment of Desiccation Tolerance in Germinated Arabidopsis thaliana Seeds and Its Associated Transcriptome

The combination of robust physiological models with “omics” studies holds promise for the discovery of genes and pathways linked to how organisms deal with drying. Here we used a transcriptomics approach in combination with an in vivo physiological model of re-establishment of desiccation tolerance (DT) in Arabidopsis thaliana seeds. We show that the incubation of desiccation sensitive (DS) germinated Arabidopsis seeds in a polyethylene glycol (PEG) solution re-induces the mechanisms necessary for expression of DT. Based on a SNP-tile array gene expression profile, our data indicates that the re-establishment of DT, in this system, is related to a programmed reversion from a metabolic active to a quiescent state similar to prior to germination. Our findings show that transcripts of germinated seeds after the PEG-treatment are dominated by those encoding LEA, seed storage and dormancy related proteins. On the other hand, a massive repression of genes belonging to many other classes such as photosynthesis, cell wall modification and energy metabolism occurs in parallel. Furthermore, comparison with a similar system for Medicago truncatula reveals a significant overlap between the two transcriptomes. Such overlap may highlight core mechanisms and key regulators of the trait DT. Taking into account the availability of the many genetic and molecular resources for Arabidopsis, the described system may prove useful for unraveling DT in higher plants.

Trehalose transporter from African chironomid larvae improves desiccation tolerance of Chinese hamster ovary cells

Dry preservation has been explored as an energy-efficient alternative to cryopreservation, but the high sensitivity of mammalian cells to desiccation stress has been one of the major hurdles in storing cells in the desiccated state. An important strategy to reduce desiccation sensitivity involves use of the disaccharide trehalose. Trehalose is known to improve desiccation tolerance in mammalian cells when present on both sides of the cell membrane. Because trehalose is membrane impermeant the development of desiccation strategies involving this promising sugar is hindered. We explored the potential of using a high-capacity trehalose transporter (TRET1) from the African chironomid Polypedilum vanderplanki[21] to introduce trehalose into the cytoplasm of mammalian cells and thereby increase desiccation tolerance. When Chinese hamster ovary cells (CHO) were stably transfected with TRET1 (CHO–TRET1 cells) and incubated with 0.4M trehalose for 4h at 37°C, a sevenfold increase in trehalose uptake was observed compared to the wild-type CHO cells. Following trehalose loading, desiccation tolerance was investigated by evaporative drying of cells at 14% relative humidity. After desiccation to 2.60g of water per gram dry weight, a 170% increase in viability and a 400% increase in growth (after 7days) was observed for CHO–TRET1 relative to control CHO cells. Our results demonstrate the beneficial effect of intracellular trehalose for imparting tolerance to partial desiccation.

CONSERVING AUSTRALIA'S UNIQUE RAINFOREST FRUITS AND WILD RELATIVES

Australian rainforests have many unique edible fruits including wild relatives of worldwide economically important species such as citrus and macadamia. One of the key risks of projected climate change is its effect on Eastern Australian rainforests. The interaction of climate change with other threats, such as weeds and habitat fragmentation, is thus likely to impact on already vulnerable environments such as the World Heritage listed Gondwana Rainforests of S. Queensland and N. New South Wales. Seed banking is a cost effective way of conserving unique and vulnerable diversity ex situ for utilisation (e.g., plant breeding and restoration programs). However, the seeds of many rainforest species cannot survive the desiccation required in standard seed banking procedures (i.e., non-orthodox seed storage characteristics). This paper reports on preliminary findings on biology and storage characteristics of seeds of 11 significant and/or threatened Australian rainforest fruits. Of these, 4 were desiccation sensitive (DS) and living collections and technology development (e.g., cryopreservation) needs to be prioritised for ex situ conservation. Of the 7 desiccation tolerant (DT) seeds, only one is expected to be viable in long-term storage by standard seedbanking methods. The oil content and thermal analysis of oils is described for 5 of these species and indicates the need to investigate alternative storage, particularly cryopreservation, for these species.

Protein expression upon desiccation and imbibition of Magnolia ovata A. St.-Hil seeds

The effect of seed drying and imbibition was studied by differential protein expression using two dimensional gel electrophoresis. After drying to a range of water contents, seeds were germinated to assess the viability. Seeds of Magnolia ovata did not withstand the desiccation down to 0.10 g H2O · g-1 dw. The critical water content below which the desiccation sensitivity became apparent was around 0.18 g H2O · g-1 dw (-26.5 MPa). Total protein was extracted and separated by 2D electrophoresis from fresh seeds (0.28 g H2O · g-1 dw), mild dried seeds (0.25 g H2O · g-1 dw) and seeds at low water content (0.10 g H2O · g-1 dw) before and after imbibition for 10 days. The proteome profile revealed the presence of 588 spots on each silver stained gel, from which 21 showed differential expression, correlated with desiccation and germination, by increased or decreased expression. After MS/MS sequencing, three protein spots produced spectra that matched to a Magnolia salicifolia legumin precursor. Results suggested an involvement of this protein in the events taking place during the drying and subsequent imbibitions of the dried seeds.

Partial dehydration and cryopreservation of Citrus seeds

Three categories of seed storage behavior are generally recognized among plant species: orthodox, intermediate and recalcitrant. Intermediate seeds cannot be stored in liquid nitrogen (LN) without a previous partial dehydration process. The water content (WC) of the seeds at the moment of immersion in LN must be regarded as the most critical factor in cryopreservation. The purpose of this study was to investigate the basis of the optimal hydration status for cryopreservation of Citrus seeds: C. sinensis (sweet orange), C. paradisi (grapefruit), C. reticulata (mandarin) in LN. To study the tolerance to dehydration and LN exposure, seeds were desiccated by equilibration at relative humidities between 11 and 95%. Sorption isotherms were determined and modeled; lipid content of the seeds was measured. Seed desiccation sensitivity was quantified by the quantal response model. Differential scanning calorimetry (DSC) thermograms were determined on cotyledon tissue at different moisture contents to measure ice melting enthalpies and unfrozen WC. Samples of total seed lipid extract were also analyzed by DSC to identify lipid transitions in the thermograms. The limit of hydration for LN Citrus seeds treatment corresponded to the unfrozen WC in the tissue, confirming that seed survival strictly depended on avoidance of intracellular ice formation.

A sister group contrast using untargeted global metabolomic analysis delineates the biochemical regulation underlying desiccation tolerance in Sporobolus stapfianus.

Understanding how plants tolerate dehydration is a prerequisite for developing novel strategies for improving drought tolerance. The desiccation-tolerant (DT) Sporobolus stapfianus and the desiccation-sensitive (DS) Sporobolus pyramidalis formed a sister group contrast to reveal adaptive metabolic responses to dehydration using untargeted global metabolomic analysis. Young leaves from both grasses at full hydration or at 60% relative water content (RWC) and from S. stapfianus at lower RWCs were analyzed using liquid and gas chromatography linked to mass spectrometry or tandem mass spectrometry. Comparison of the two species in the fully hydrated state revealed intrinsic differences between the two metabolomes. S. stapfianus had higher concentrations of osmolytes, lower concentrations of metabolites associated with energy metabolism, and higher concentrations of nitrogen metabolites, suggesting that it is primed metabolically for dehydration stress. Further reduction of the leaf RWC to 60% instigated a metabolic shift in S. stapfianus toward the production of protective compounds, whereas S. pyramidalis responded differently. The metabolomes of S. stapfianus leaves below 40% RWC were strongly directed toward antioxidant production, nitrogen remobilization, ammonia detoxification, and soluble sugar production. Collectively, the metabolic profiles obtained uncovered a cascade of biochemical regulation strategies critical to the survival of S. stapfianus under desiccation.

Desiccation Sensitivity and Heat Tolerance ofPrunus ilicifoliaSeeds Dispersed by American Black Bears (Ursus americanus)

ABSTRACT. Carnivore consumption of fruit is a principle means by which many fleshy-fruited plant species achieve long-distance seed dispersal. We examined carnivore dispersal of hollyleaf cherry (Prunus ilicifolia) seeds, specifically assessing the survival, desiccation sensitivity, and germination of seeds found in bear scats. Studies were conducted both in the laboratory and in 2 burn areas in Los Padres National Forest, California. Bear scats containing P. ilicifolia seeds were collected in burned and unburned chaparral. We counted seeds in each scat and noted whether endocarps had tooth punctures or rattled audibly when shaken. For comparative germination trials, we also collected fruits and seeds directly from mature shrubs. In the laboratory, following a cold-moist stratification period, seeds were assessed for germinability. In the field, we compared desiccation rates and germinability of seeds from bear scats and freshly collected seeds. We compared rates of moisture loss and germination for seeds...

Desiccation Sensitivity and Germination of Recalcitrant Garcinia kola Heckel Seeds

Desiccation Sensitivity and Germination of Recalcitrant Garcinia kola Heckel Seeds

Conservation technologies for safeguarding and restoring threatened flora: case studies from Eastern Australia

This paper highlights recent advances and improved scientific understanding of conservation technologies through selected case studies on threatened plant species indigenous to Eastern Australia. This includes investigations into seed desiccation, storage responses and cryopreservation in rainforest species, particularly the socio-economically important Australian native Citrus spp., Davidsonia spp. (Davidson’s plum) and Syzygium spp. This work also (1) increases our understanding of ecological correlates of seed desiccation sensitivity for predictive use and (2) improves restoration practice through better understanding of seed storage and germination requirements. The use of in vitro conservation technologies in support of conservation actions for endangered species is outlined in case studies on Wollemia nobilis (Wollemi pine), epiphytic and terrestrial orchid species, and an endangered fern species.

In vitro conservation of Malaysian biodiversity—achievements, challenges and future directions

Malaysia is fortunate and proud to contain some of the world’s richest biodiversity. In Malaysia, there are an estimated 185,000 species of fauna and 12,500 species of flowering plants, many of which are endemic to tropical forests in this region. Indeed, such diversity is an important and invaluable national asset to safeguard both present and future generations. In vitro conservation offers possible techniques for the preservation of plant germplasm that at present is difficult to maintain or is maintained with limited success. Research at the Universiti Kebangsaan Malaysia (The National University of Malaysia) focuses on the cryopreservation of woody fruit species with seeds that cannot tolerate cryopreservation (recalcitrant or intermediate). Among the plants with recalcitrant seeds are such traditionally important edible tropical fruits as mangosteen, langsat, and rambai (Garcinia mangostana, Lansium domesticum, and Baccaurea motleyana). Citrus aurantifolia, Citrus suhuiensis, Citrus madurensis, Citrus hystrix, and Fortunella polyandra are among the Citrus and Citrus-related species studied. Cryopreservation studies include the Nepenthes species (pitcher plants) of Malaysia. Fundamental research on desiccation and low-temperature tolerance and on the physiology of desiccation are used to understand seed behavior, a prerequisite for the development of successful conservation techniques. At the same time, cryopreservation protocols for several Citrus and forestry species were developed for embryonic axes and adventitious shoots, mainly using rapid dehydration and PVS2 vitrification techniques. There are no successful standard techniques or protocols for species with highly recalcitrant seeds such as Garcinia species. Modification of existing protocols or development of new methods is required, but this can be accomplished only when a detailed understanding of the recalcitrant nature of the seeds or explants is achieved. While we have considerable knowledge concerning the basics of biochemical processes and some molecular data from work on desiccation-tolerant seeds, a great need remains for understanding the cause of the recalcitrance or desiccation sensitivity of these seeds. It may be necessary to use a systems biology approach that exploits the “omics” technologies to generate global molecular data. In combination with bioinformatics for data integration and analyses, this approach would move toward improved modeling of the biological pathways associated with the development of recalcitrant seeds.

Desiccation sensitivity and tolerance in the moss Physcomitrella patens: assessing limits and damage

The moss Physcomitrella patens is becoming the model of choice for functional genomic studies at the cellular level. Studies report that Physcomitrella survives moderate osmotic and salt stress, and that desiccation tolerance can be induced by exogenous ABA. Our goal was to quantify the extent of dehydration tolerance in wild type moss and to examine the nature of cellular damage caused by desiccation. We exposed Physcomitrella to humidities that generate water potentials from −4 (97% RH) to −273 MPa (13% RH) and monitored water loss until equilibrium. Water contents were measured on a dry matter basis to determine the extent of dehydration because fresh weights (FW) were found to be variable and, therefore, unreliable. We measured electrolyte leakage from rehydrating moss, assessed overall regrowth, and imaged cells to evaluate their response to drying and rehydration. Physcomitrella did not routinely survive water potentials <−13 MPa. Upon rehydration, moss dried to water contents >0.4 g g dm−1 maintained levels of leakage similar to those of hydrated controls. Moss dried to lower water contents leaked extensively, suggesting that plasma membranes were damaged. Moss protonemal cells were shrunken and their walls twisted, even at −13 MPa. Moss cells rehydrated after drying to −273 MPa failed to re-expand completely, again indicating membrane damage. ABA treatment elicited tolerance of desiccation to at least −273 MPa and limited membrane damage. Results of this work will form the basis for ongoing studies on the functional genomics of desiccation tolerance at the cellular level.

Identification of a novel ABC transporter required for desiccation tolerance, and biofilm formation in Rhizobium leguminosarum bv. viciae 3841.

Rhizobium leguminosarum is a soil bacterium with the ability to form nitrogen-fixing nodules on the roots of leguminous plants. Soil-dwelling, free-living R. leguminosarum often encounters desiccation stress, which impacts its survival within the soil. The mechanisms by which soil bacteria resist the effects of desiccation stress have been described. However, the role of the cell envelope in the desiccation tolerance mechanisms of rhizobia is relatively uncharacterized. Using a transposon mutagenesis approach, a mutant of R. leguminosarum bv. viciae was isolated that was highly sensitive to desiccation. The mutation is located in the ATP-binding protein of an uncharacterized ATP-binding cassette transporter operon (RL2975-RL2977). Exopolysaccharide accumulation was significantly lower in the mutant and the decrease in desiccation tolerance was attributed to the decreased accumulation of exopolysaccharide. In addition to desiccation sensitivity, the mutant was severely impaired in biofilm formation, an important adaptation used by soil bacteria for survival. This work has identified a novel transporter required for physiological traits that are important for the survival of R. leguminosarum in the rhizosphere environment.

The Response Difference of Mitochondria in Recalcitrant Antiaris toxicaria Axes and Orthodox Zea mays Embryos to Dehydration Injury

Long-term preservation of recalcitrant seeds is very difficult because the physiological basis on their desiccation sensitivity is poorly understood. Survival of Antiaris toxicaria axes rapidly decreased and that of immature maize embryos very slowly decreased with dehydration. To understand their different responses to dehydration, we examined the changes in mitochondria activity during dehydration. Although activities of cytochrome (Cyt) c oxidase and malate dehydrogenase of the A. toxicaria axis and maize embryo mitochondria decreased with dehydration, the parameters of maize embryo mitochondria were much higher than those of A. toxicaria, showing that the damage was more severe for the A. toxicaria axis mitochondria than for those of maize embryo. The state I and III respiration of the A. toxicaria axis mitochondria were higher than those of maize embryo, the former rapidly decreased, and the latter slowly decreased with dehydration. The proportion of Cyt c pathway to state III respiration for the A. toxicaria axis mitochondria was low and rapidly decreased with dehydration, and the proportion of alternative oxidase pathway was high and slightly increased with dehydration. In contrast, the proportion of Cyt c pathway for maize embryo mitochondria was high, and that of alternative oxidase pathway was low. Both pathways decreased slowly with dehydration.

Impact of body melanisation on contrasting levels of desiccation resistance in a circumtropical and a generalist Drosophila species

We investigated the role of cuticular lipids, body melanisation and body size in conferring contrasting levels of desiccation resistance in latitudinal populations of Drosophila melanogaster and Drosophila ananassae on the Indian subcontinent. Contrary to the well known role of cuticular lipids in water proofing in diverse insect taxa, there is lack of geographical variations in the amount of cuticular lipids per fly in both the species. In D. ananassae, quite low levels of body melanisation are correlated with lower desiccation resistance. By contrast, increased levels of desiccation resistance are correlated with quite high melanisation in D. melanogaster. Thus, species specific cuticular melanisation patterns are significantly correlated with varying levels of desiccation resistance within as well as between populations and across species. Role of body melanisation in desiccation resistance is further supported by the fact that assorted dark and light flies differ significantly in cuticular water loss, hemolymph and dehydration tolerance. However, similar patterns of body size variation do not account for contrasting levels of desiccation resistance in these two Drosophila species. Climatic selection is evidenced by multiple regression analysis with seasonal amplitude of thermal and humidity changes (Tcv and RHcv) along latitude on the Indian subcontinent. Finally, the contrasting levels of species specific distribution patterns are negatively correlated with RHcv of sites of origin of populations i.e. a steeper negative slope for D. ananassae corresponds with its desiccation sensitivity as compared with D. melanogaster. Thus, evolutionary changes in body melanisation impact desiccation resistance potential as well as distribution patterns of these two Drosophila species on the Indian subcontinent.

Desiccation sensitivity of excised embryonic axes of selected amaryllid species

Desiccation sensitivity of excised embryonic axes of selected amaryllid species

Classificação de sementes de espécies florestais dos gêneros Nectandra e Ocotea (Lauraceae) quanto ao comportamento no armazenamento

O conhecimento do comportamento das sementes no armazenamento permite a utilização de condições adequadas para a manutenção de sua viabilidade, o planejamento de programas de produção de mudas e a conservação ex situ. O objetivo deste estudo foi o de classificar sementes de cinco espécies de Lauraceae de ocorrência no sul de Minas Gerais, Brasil, quanto ao comportamento no armazenamento. Sementes de Nectandra grandiflora, Nectandra lanceolata, Nectandra oppositifolia, Ocotea corymbosa e Ocotea pulchella foram submetidas à avaliação da germinação e do grau de umidade antes e após a secagem artificial. As sementes apresentam comportamento recalcitrante, devido à sensibilidade à secagem.

From Avicennia to Zizania: seed recalcitrance in perspective.

Considered only in terms of tolerance of, or sensitivity to, desiccation (which is an oversimplification), orthodox seeds are those which tolerate dehydration and are storable in this condition, while highly recalcitrant seeds are damaged by loss of only a small proportion of water and are unstorable for practical purposes. Between these extremes, however, there may be a gradation of the responses to dehydration--and also to other factors--suggesting perhaps that seed behaviour might be best considered as constituting a continuum subtended by extreme orthodoxy and the highest degree of recalcitrance. As the characteristics of seeds of an increasing number of species are elucidated, non-orthodox seed behaviour is emerging as considerably more commonplace--and its basis far more complex--than previously suspected. Whatever the post-harvest responses of seeds of individual species may be, they are the outcome of the properties of pre-shedding development, and a full understanding of the subtleties of various degrees of non-orthodox behaviour must await the identification of, and interaction among, all the factors conferring extreme orthodoxy. Appreciation of the phenomenon of recalcitrance is confounded by intra- and interseasonal variability across species, as well as within individual species. However, recent evidence suggests that provenance is a pivotal factor in determining the degree of recalcitrant behaviour exhibited by seeds of individual species. Non-orthodox--and, in particular, recalcitrant--seed behaviour is not merely a matter of desiccation sensitivity: the primary basis is that the seeds are actively metabolic when they are shed, in contrast to orthodox types which are quiescent. This affects all aspects of the handling and storage of recalcitrant seeds. In the short to medium term, recalcitrant seeds should be stored in as hydrated a condition as when they are shed, and at the lowest temperature not diminishing vigour or viability. Such hydrated storage has attendant problems of fungal proliferation which, unless minimized, will inevitably and significantly affect seed quality. The life span of seeds in hydrated storage even under the best conditions is variable among species, but is curtailed (days to months), and various approaches attempting to extend non-orthodox seed longevity are discussed. Conservation of the genetic resources by means other than seed storage is then briefly considered, with detail on the potential for, and difficulties with, cryostorage highlighted. There appears to be little taxonomic relationship among species exhibiting the phenomenon of seed recalcitrance, suggesting that it is a derived trait, with tolerance having been lost a number of times. Although recalcitrant seededness is best represented in the mesic tropics, particularly among rainforest climax species, it does occur in cooler, drier and markedly seasonal habitats. The selective advantages of the trait are considered.

Duplicated P5CS genes of Arabidopsis play distinct roles in stress regulation and developmental control of proline biosynthesis

Delta-1-pyrroline-5-carboxylate synthetase enzymes, which catalyse the rate-limiting step of proline biosynthesis, are encoded by two closely related P5CS genes in Arabidopsis. Transcription of the P5CS genes is differentially regulated by drought, salinity and abscisic acid, suggesting that these genes play specific roles in the control of proline biosynthesis. Here we describe the genetic characterization of p5cs insertion mutants, which indicates that P5CS1 is required for proline accumulation under osmotic stress. Knockout mutations of P5CS1 result in the reduction of stress-induced proline synthesis, hypersensitivity to salt stress, and accumulation of reactive oxygen species. By contrast, p5cs2 mutations cause embryo abortion during late stages of seed development. The desiccation sensitivity of p5cs2 embryos does not reflect differential control of transcription, as both P5CS mRNAs are detectable throughout embryonic development. Cellular localization studies with P5CS-GFP gene fusions indicate that P5CS1 is sequestered into subcellular bodies in embryonic cells, where P5CS2 is dominantly cytoplasmic. Although proline feeding rescues the viability of mutant embryos, p5cs2 seedlings undergo aberrant development and fail to produce fertile plants even when grown on proline. In seedlings, specific expression of P5CS2-GFP is seen in leaf primordia where P5CS1-GFP levels are very low, and P5CS2-GFP also shows a distinct cell-type-specific and subcellular localization pattern compared to P5CS1-GFP in root tips, leaves and flower organs. These data demonstrate that the Arabidopsis P5CS enzymes perform non-redundant functions, and that P5CS1 is insufficient for compensation of developmental defects caused by inactivation of P5CS2.

Desiccation sensitivity in the Arecaceae: Correlates and frequency

Desiccation sensitivity in the Arecaceae: Correlates and frequency