Short-term Desiccation Research Articles

Hygrophilous old-growth forest lichens are highly capable of instantaneous photosynthesis activation after short-term desiccation stress

Hygrophilous old-growth forest lichens are highly capable of instantaneous photosynthesis activation after short-term desiccation stress

Sea Slug Mucus Production Is Supported by Photosynthesis of Stolen Chloroplasts.

Simple SummaryKleptoplasty is the capacity of a non-photosynthetic organism to acquire and maintain structurally intact chloroplasts from its algal food source(s), thereafter termed kleptoplasts. In animals, the capacity for long-term (several weeks to months) maintenance of photosynthetic active kleptoplasts is a unique characteristic in a handful of sea slugs, mostly within the genus Elysia. In this study, we investigated the role of kleptoplast photosynthesis on mucus production by the tropical sea slug Elysia crispata. Mucus secretion is paramount for mollusks, playing a role in locomotion, feeding, reproduction and protection. Limiting photosynthesis by rearing animals under reduced light led to lower mucus production and lower carbohydrate concentrations in the secreted mucus. This study indicates that production of mucus by kleptoplast-bearing sea slugs is supported by photosynthesis, confirming the biological relevance of kleptoplasty to the fitness of these peculiar mollusks.A handful of sea slugs of the order Sacoglossa are able to steal chloroplasts—kleptoplasts—from their algal food sources and maintain them functionally for periods ranging from several weeks to a few months. In this study, we investigated the role of kleptoplast photosynthesis on mucus production by the tropical sea slug Elysia crispata. Animals reared for 5 weeks in quasi dark (5 μmol photons m−2 s−1) showed similar growth to those under regular light (60–90 μmol photons m−2 s−1), showing that kleptoplast photosynthesis was not relevant for growth when sea slugs were fed ad libitum. However, when subjected to short-term desiccation stress, animals reared under regular light produced significantly more mucus. Furthermore, the carbohydrate content of secreted mucus was significantly lower in slugs limited in the photosynthetic activity of their kleptoplasts by quasi-dark conditions. This study indicates that photosynthesis supports the synthesis of protective mucus in kleptoplast-bearing sea slugs.

The specific glycerolipid composition is responsible for maintaining the membrane stability of Physcomitrella patens under dehydration stress

Land colonization is a major event in plant evolution. Little is known about the evolutionary characteristics of lipids during this process. Here, we proved that Physcomitrella patens, a bryophyte that appeared in the early evolution of terrestrial plants, has short-term desiccation resistance. The maintenance of membrane integrity is related to its specific glycerolipid composition and key genes for lipid metabolism. We analyzed 414 types of lipid molecules, and found that phospholipids accounted for 61.7%, mainly PC and PI; glycolipids accounted for only 26.5%, with a special MGDG molecular map. The most abundant MDGD, that is, MGDG34:6, contained rare 15- and 19-carbon acyl chains; the level of neutral lipids was higher. This was consistent with the results observed by TEM, with fewer lamellae and obvious lipid droplets. Slight dehydration accumulated a large number of TAG molecules, and severe dehydration degraded phospholipids and caused membrane leakage, but PA and MGDG fluctuated less. The key genes of lipid metabolism, DGAT and PAP, were actively transcribed, suggesting that PA was one of the main DAG sources for TAG synthesis. This work proves that Physcomitrella patens adopts high-constitutive PC and PI similar to plant seeds, abundant TAG, and its own specific MGDG to resist extreme dehydration. This result provides a new insight into the lipid evolution of early terrestrial plants against unfavorable terrestrial environments.

Effect of Short-Term Desiccation, Recovery Time, and CAPA-PVK Neuropeptide on the Immune System of the Burying Beetle Nicrophorus vespilloides.

Environmental conditions, especially related to winter, are crucial for shaping activity of insect immune system. However, our previous research clearly indicates differences in the immune system functioning when the cold stress was induced in the laboratory conditions and when the beetles were collected from natural environment during winter. This is probably related to the multiplication of observed effects by simultaneous presence of different stress factors characteristic of winter, including desiccation. For these reasons, our next step was analysis of the effects of short-term desiccation and recovery time on the functioning of immune system of burying beetle Nicrophorus vespilloides. Also, the effect of Tenmo–PVK-2 (tenebrionid periviscerokinin), member of the CAPA–PVK neuropeptide family, was investigated to better understand observed changes. Short-term desiccation decreases the phagocytic activity of burying beetle haemocytes, which is correlated with a reduction in their adhesive ability. On the other hand, there was a significant increase in phenoloxidase (PO) activity and the level of proPO expression, which may suggest sealing the cuticula by melanin deposition and prevention of water loss. Additionally, the elevated level of defensin expression may be associated with the cross-talk between mechanisms, which participate in insect response to environmental stress, including pathogen infection. After 1 h of recovery time, the activity of tested cellular and humoral mechanisms was mostly back to the control level. However, inhibition of the activity of PO and down-regulation of proPO were noted. These results also indicate importance of melanin deposition during water loss. Moreover, it suggests that some changes in immune system functioning during stress conditions do not have an immune function. Interestingly, part of the effects characteristic of recovery time were also observed after the application of Tenmo–PVK-2, mainly related to haemocyte morphology. These results indicate that CAPA–PVK neuropeptides may also influence on activity of burying beetle immune system. It should be also highlighted that, because of the study of the effects of CAPA–PVK neuropeptides, homologs of vertebrate neuromedin U, the results may be interesting for search evolutionary similarities in the functioning of the neuroendocrine system of insects and vertebrates.

Distribution Coefficient and Metal Pollution Index in Water and Sediments: Proposal of a New Index for Ecological Risk Assessment of Metals

The Lake of Texcoco is a closed basin with soils that confer salinity, conductivity, and alkalinity to it. It has undergone a reduction in size, incorporation of wastewater, and short-term desiccation, and includes temporary wetlands interconnected during the rainy season, some of which receive treated wastewater. Sediments contain metals, thus affecting water quality. Five artificial lakes were studied, and 12 metals (As, Ba, Cd, Cu, Cr, Fe, Mg, Mn, Hg, Ni, Pb, and Zn) were monitored bimonthly in water and sediments from June 2015 to March 2018. The Metal Pollution Index (MPI) and the Distribution Coefficient (Kd) were computed. Fe and Cd were the most and least stable metals in sediments, respectively (mean Log(Kd) = 4.24 and 2.079). Based on Log(Kd), metals were ranked as Fe > Mn > Zn > Cu > Mg > Cr > Ni > Ba > Pb > Hg > As > Cd. Log(Kd) values < 3 and > 5 indicate that metals occur mainly in water and sediments, respectively. The Mean Distribution Coefficient Log(Kd MPI) is a novel index proposed to assess ecological risk from metals in a water body. This index allows determining the phase (liquid or solid) where metals predominate, negatively affecting either free-swimming or benthic organisms. Log(Kd MPI) values indicated that metals occur primarily in the liquid phase in all lakes studied.

Tolerance of Ruppia sinensis Seeds to Desiccation, Low Temperature, and High Salinity With Special Reference to Long-Term Seed Storage.

Seeds are important materials for the restoration of globally-threatened marine angiosperm (seagrass) populations. In this study, we investigated the differences between different Ruppia sinensis seed types and developed two feasible long-term R. sinensis seed storage methods. The ability of R. sinensis seeds to tolerate the short-term desiccation and extreme cold had been investigated. The tolerance of R. sinensis seeds to long-term exposure of high salinity, cold temperature, and desiccation had been considered as potential methods for long-term seed storage. Also, three morphological and nine physiological indices were measured and compared between two types of seeds: Shape L and Shape S. We found that: (1) wet storage at a salinity of 30–40 psu and 0°C were the optimal long-term storage conditions, and the proportion of viable seeds reached over 90% after a storage period of 11 months since the seeds were collected from the reproductive shoots; (2) dry condition was not the optimal choice for long-term storage of R. sinensis seeds; however, storing seeds in a dry condition at 5°C and 33 ± 10% relative humidity for 9 months had a relatively high percentage (74.44 ± 2.22%) of viable seeds, consequently desiccation exposure could also be an acceptable seed storage method; (3) R. sinensis seeds would lose vigor in the interaction of extreme cold (-27°C) and desiccation; (4) there were significant differences in seed weight, seed curvature, and endocarp thickness between the two types of seeds. These findings provided fundamental physiological information for R. sinensis seeds and supported the long-term storage of its seeds. Our results may also serve as useful reference for seed storage of other threatened seagrass species and facilitate their ex situ conservation and habitat restoration.

Desiccation tolerance and growth-temperature requirements of Coccomyxa (Trebouxiophyceae, Chlorophyta) strains from Antarctic biological soil crusts

The genus Coccomyxa is frequently present in biological soil crusts from temperate regions but has also been found in Antarctic soils. Strain NIES 2166 (Coccomyxa subellipsoidea) from continental South Victoria Land is described as a model organism for cold adaptation (Blanc et al. 2012) although it has been cultivated for more than 50 years under artificial conditions. In order to test whether this isolate still kept physiological traits for Antarctic conditions or exhibited acclimation/adaptation to culturing, it was ecophysiologically compared with three recently isolated Coccomyxa strains from soil samples collected at the Antarctic Peninsula. Additionally, the investigated strains were tested for potential endemism. Molecular analysis based on ITS rDNA identified the three newly isolated strains also as C. subellipsoidea. The effective quantum yield of photosynthesis was examined during controlled short-term desiccation (at relative air humidity c. 10%) and after rehydration. Growth rates were determined over a temperature gradient from 4.6 to 34.8 °C. The results indicate cold tolerance and drought tolerance among all tested strains which are slightly more developed in the continental strain NIES 2166 although 50 years passed since isolation. Therefore it is assumed that strain NIES 2166 can still be recommended to test traits of cold adaptation but psychrophilic behavior, and therefore endemism, can be excluded among all str

Entransia and Hormidiella, sister lineages of Klebsormidium (Streptophyta), respond differently to light, temperature, and desiccation stress.

The green-algal class Klebsormidiophyceae (Streptophyta), which occurs worldwide, comprises the genera Klebsormidium, Interfilum, Entransia, and Hormidiella. Ecophysiological research has so far focused on the first two genera because they are abundant in biological soil crust communities. The present study investigated the photosynthetic performances of Hormidiella attenuata and two strains of Entransia fimbriata under light, temperature, and desiccation stress. Their ultrastructure was compared using transmission electron microscopy. The two Entransia strains showed similar physiological responses. They used light more efficiently than Hormidiella, as indicated by higher oxygen production and relative electron transport rate under low light conditions, lower light saturation and compensation points, and higher maximum oxygen production during light saturation. Their requirement for low light levels explains the restriction of Entransia to dim limnetic habitats. In contrast, Hormidiella, which prefers drier soil habitats, responded to light gradients similarly to other aero-terrestrial green algae. Compared to Entransia, Hormidiella was less affected by short-term desiccation, and rehydration allowed full recovery of the photosynthetic performance. Nevertheless, both strains of Entransia coped with low water availability better than other freshwater algae. Photosynthetic oxygen production in relation to respiratory consumption was higher in low temperatures (Entransia: 5 °C, Hormidiella: 10 °C) and the ratio decreased with increasing temperatures. Hormidiella exhibited conspicuous triangular spaces in the cell wall corners, which were filled either with undulating cell wall material or with various inclusions. These structures are commonly seen in various members of Klebsormidiophyceae. The data revealed significant differences between Hormidiella and Entransia, but appropriate adaptations to their respective habitats.Electronic supplementary materialThe online version of this article (doi:10.1007/s00709-015-0889-z) contains supplementary material, which is available to authorized users.

Increasing intracellular trehalose is sufficient to confer desiccation tolerance to Saccharomyces cerevisiae

Diverse organisms capable of surviving desiccation, termed anhydrobiotes, include species from bacteria, yeast, plants, and invertebrates. However, most organisms are sensitive to desiccation, likely due to an assortment of different stresses such as protein misfolding and aggregation, hyperosmotic stress, membrane fracturing, and changes in cell volume and shape leading to an overcrowded cytoplasm and metabolic arrest. The exact stress(es) that cause lethality in desiccation-sensitive organisms and how the lethal stresses are mitigated in desiccation-tolerant organisms remain poorly understood. The presence of trehalose in anhydrobiotes has been strongly correlated with desiccation tolerance. In the yeast Saccharomyces cerevisiae, trehalose is essential for survival after long-term desiccation. Here, we establish that the elevation of intracellular trehalose in dividing yeast by its import from the media converts yeast from extreme desiccation sensitivity to a high level of desiccation tolerance. This trehalose-induced tolerance is independent of utilization of trehalose as an energy source, de novo synthesis of other stress effectors, or the metabolic effects of trehalose biosynthetic intermediates, indicating that a chemical property of trehalose is directly responsible for desiccation tolerance. Finally, we demonstrate that elevated intracellular maltose can also make dividing yeast tolerant to short-term desiccation, indicating that other disaccharides have stress effector activity. However, trehalose is much more effective than maltose at conferring tolerance to long-term desiccation. The effectiveness and sufficiency of trehalose as an antagonizer of desiccation-induced damage in yeast emphasizes its potential to confer desiccation tolerance to otherwise sensitive organisms.

The influence of short term water level fluctuations and desiccation stress on periphyton development at a riparian zone of a large regulated river

The influence of short term water level fluctuations and desiccation stress on periphyton development at a riparian zone of a large regulated river

Pathogens as potential hitchhikers on intercontinental dust

Desert dust seeds distant lands and waters with minerals as well as micro-organisms raising the question of whether this ancient phenomenon also spreads pathogens across the globe. Severe dust storms require strong winds blowing over land-masses that are largely devoid of vegetation, effectively limiting the scope for winds to raise pathogens into the air. Nevertheless, changing patterns of land-use, often driven by belligerency, result in refugees spreading to areas that were previously deemed barely habitable. With the help of the International Committee of the Red Cross, a number of sand/dust samples were collected from the Republic of Chad, some near refugee camps, others further removed from human influence. In parallel studies, we documented the micro-organisms present in these samples and used a number of the isolates here to test the effect of environmental constraints on their ability to survive intercontinental flight. We also added traditional pathogens to the palette of microbes and tested the effects of UV irradiation, desiccation and temperature on survival of both bacteria and fungi. A clear trend was obvious—those microbes that are coloured or able to form conidia or spores (in other words, those that are native to deserts) were well able to resist the imposed stresses. On the other hand, most pathogens were more sensitive to stresses than the environmental isolates. Toxin production in two species of Aspergillus was also investigated. Short-term desiccation (simulating environmental conditions during intercontinental travel) of sand amended with fungal spores containing sterigmatocystin leads to increased mycotoxin contents, but significant mycotoxin production was only possible under growth-permissive conditions, e.g. at higher humidity. It thus seems likely that an ever-decreasing fraction of the initial pathogen load survives as the dust recedes from its desert source and that those organisms that land on other continents are highly enriched in desert dwellers.

Light, Temperature, and Desiccation Effects on Photosynthetic Activity, and Drought-Induced Ultrastructural Changes in the Green Alga Klebsormidium dissectum (Streptophyta) from a High Alpine Soil Crust

Members of the cosmopolitan green algal genus Klebsormidium (Klebsormidiales, Streptophyta) are typical components of terrestrial microbiotic communities such as biological soil crusts, which have many important ecological functions. In the present study, Klebsormidium dissectum (Gay) Ettl & Gärtner was isolated from a high alpine soil crust in the Tyrolean Alps, Austria. Physiological performance in terms of growth and photosynthesis was investigated under different controlled abiotic conditions and compared with ultrastructural changes under the treatments applied. K. dissectum showed very low light requirements as reflected in growth patterns and photosynthetic efficiency. Increasing temperatures from 5°C to 40°C led to different effects on respiratory oxygen consumption and photosynthetic oxygen evolution. While at low temperatures (5-10°C), respiration was not detectable or on a very low level, photosynthesis was relatively high, Reversely, at the highest temperature, respiration was unaffected, and photosynthesis strongly inhibited pointing to strong differences in temperature sensitivity between both physiological processes. Although photosynthetic performance of K. dissectum was strongly affected under short-term desiccation and recovered only partly after rehydration, this species was capable to survive even 3weeks at 5% relative air humidity. K. dissectum cells have a cell width of 5.6 ± 0.3μm and a cell length of 8.4 ± 2.0μm. Desiccated cells showed a strongly reduced cell width (46% of control) and cell length (65% of control). In addition, in desiccated cells, fewer mitochondria were stained by DIOC(6), and damaged plasma membranes were detected by FM 1-43 staining. High-pressure freeze fixation as well as chemical fixation allowed visualizing ultrastructural changes caused by desiccation. In such cells, the nucleus and chloroplast were still visibly intact, but the extremely thin cell walls (75-180nm) were substantially deformed. The cytoplasm appeared electron dense and mitochondria were altered. Although K. dissectum can be characterized as euryoecious species, all ecophysiological and ultrastructural data indicate susceptibility to desiccation. However, the steadily occurring fragmentation of filaments into smaller units leads to improved self protection and thus may represent a life strategy to better survive longer periods of drought in exposed alpine soil crusts.

AbstractsAstrobiology Graduate Conference (AbGradCon) 2011HOSTED BYMontana State UniversityBozeman, Montana • June 5–8, 2011

AbstractsAstrobiology Graduate Conference (AbGradCon) 2011HOSTED BYMontana State UniversityBozeman, Montana • June 5–8, 2011

Effects of s-ABA on the Physiology and Marketability of Various Container-Grown Taxa During Short-Term Desiccation

Abstract Reduced post harvest care of woody plant material in mass retail settings can decrease the number of days plants remain marketable. If plants are sold on consignment for growers, reduced sales can lead to poor profitability. This study investigated the effect of spray applications of s-abscisic acid (s-ABA) (ConTego™ Pro SL, Valent Biosciences Corp.) to increase the number of days of marketability for various woody taxa in a simulated retail setting. In the first stage of the study, various well-watered container-grown taxa were treated with a spray application of either 0, 1000 or 2000 mg·liter−1 of s-ABA and water was withheld. Daily, desiccation symptoms were recorded to determine if plants had reached the critical wilting point (CWP) and thus became unmarketable. Marketability was increased approximately 1–7 days for plants treated with 2000 mg·liter−1 of s-ABA compared to nontreated plants. In the second stage, marketability and physiology of Ligustrum japonicum ‘Recurvifolium’ (wavy leaf privet) were monitored after plants were treated with spray applications of 0, 500, 1000, 1500 or 2000 mg·liter−1 of s-ABA and water withheld. Stomatal conductance (gs) declined for all plants depending on the concentration applied. Plants treated with s-ABA had lower gs rates and remained marketable longer than nontreated plants. All plants fully recovered to pretreatment gs rates provided they were rewatered immediately upon reaching the CWP. Spray applications of s-ABA can increase shelf life of select woody ornamentals.

Relationship of larval desiccation to Anopheles gambiae Giles and An. arabiensis Patton survival

Relationship of larval desiccation to Anopheles gambiae Giles and An. arabiensis Patton survival

Seasonal changes in humidity level in the tropics impact body color polymorphism and desiccation resistance in Drosophila jambulina—Evidence for melanism-desiccation hypothesis

Drosophila jambulina exhibits color dimorphism controlled by a single locus but its ecological significance is not clear. Dark and light morphs differ significantly in body melanisation, desiccation resistance, rate of water loss, mating activity and fecundity. Interestingly, this species lacks clinal variation for body size, desiccation resistance and life history traits. For body melanisation, lack of geographical variation as well as plastic effects is not in agreement with a thermal melanism hypothesis. However, based on field data, there are seasonal changes in phenotypic frequencies of dark and light body color morphs which correlate significantly with variation in humidity levels. Under short-term (8 h) desiccation stress, we observed higher number of assortative matings, longer copulation period and increased fecundity for dark strains as compared with light strains. By contrast, both the morphs when exposed to high humid conditions exhibited higher assortative matings and fecundity for light strains as compared with dark strains. In tropical populations of D. jambulina, body color polymorphism seems to be maintained through humidity changes as opposed to thermal melanism. Thus, seasonal changes in the frequency of body color morphs in this tropical species supports melanism-desiccation hypothesis.

Root desiccation and drought stress responses of bareroot Quercus rubra seedlings treated with a hydrophilic polymer root dip

Root hydrogel, a hydrophilic polymer, has been used to improve transplanting success of bareroot conifer seedlings through effects on water holding capacity. We examined mechanisms by which Terra-sorb® Fine Hydrogel reduces damage that occurs when roots of 1-year old, dormant northern red oak (Quercus rubra L.) were subjected to short-term (1, 3, and 5 h) pre-transplanting desiccation and long-term (45 days) drought stress following transplanting in a controlled environment chamber or greenhouse conditions. Hydrogel-treated seedlings had 80% greater root moisture content than non-root dipped control seedlings following the pre-transplanting desiccation period. Hydrogel reduced root membrane leakiness by 31% 5 h after the desiccation exposure. Hydrogel-treated seedlings did not show greater differences in shoot length, plant dry mass, root volume, net photosynthesis, and stomatal conductance compared with control seedlings following the 45-day drought stress exposure. A reduction in mean number of days to bud break in hydrogel-treated seedlings, combined with delayed tissue moisture loss (linked to higher stem water potential), suggests that hydrogel may have provided stress protection to aid survival under short-term desiccation, which may be beneficial toward alleviating initial transplanting stress.

Glutathione status correlates with different degrees of desiccation tolerance in three lichens.

• Glutathione-related mechanisms of coping with desiccation were studied in three lichens (Pseudevernia furfuracea, Peltigera polydactyla and Lobaria pulmonaria) with different degrees of tolerance. Reduced (GSH) and oxidised glutathione (GSSG), glutathione reductase (GR) and glucose-6-phosphate dehydrogenase (G6PDH) were measured during short- and long-term desiccation, and subsequent rehydration. • Glutathione was analysed by HPLC, and enzymes by spectrophotometry. Lichens were desiccated over silica gel, and rehydrated either in liquid water or water vapour. • Desiccation caused oxidation of almost all of the GSH. After short-term desiccation, rehydration caused rapid reduction of GSSG in all three species. Following long-term desiccation, P. furfuracea (the most desiccation-tolerant lichen studied) rapidly increased GSH when rehydrated in liquid water or water vapour. However, in P. polydactyla, neither method of rehydration re-established the initial GSH pool. After long-term desiccation, L. pulmonaria regenerated initial concentrations of GSH only when rehydrated in liquid water. • Reduction of GSSG was correlated with re-establishment of the predesiccation activity of G6PDH, a key enzyme of the oxidative pentose shunt. This enzyme probably provides NADPH during the first stages of rehydration, when photosynthesis is not yet possible.

Aspects of carbon monoxide production and oxidation by marine macroalgae

Rates of macroalgal carbon monoxide (CO) production were compared among 5 taxa representing 3 major phylogenetic groups (Phaeophyta, Chlorophyta, Rhodophyta). CO production varied substantially from a minimum of about 20 ng CO gdw -1 h -1 for Fucus vesiculosus to >4000 ng CO gdw -1 h -1 for Laminaria saccharina. None of the macroalgae examined contained significantly elevated CO concentrations within their pneumatocysts (float bladders), so the variability among taxa reflects other intrinsic properties. An in vitro evaluation of Ascophyllum nodosum indicated that CO production varied as a function of temperature, desiccation and illumination. CO production increased strongly for live fronds over an ecologically relevant range (5 to 23°C), but decreased at 45°C. For non-living desiccated wrack, CO production increased consistently from 5 to 47°C. Short-term desiccation of living algae decreased CO production substantially, but long-term changes in water content appeared not to markedly alter CO production relative to fresh material. Illumination strongly increased CO production relative to dark incubations, with similar responses for living and non-living material. CO oxidation (presumably bacterial) was observed for most living algae during incubations with exogenous CO at concentrations of 100 ppm, suggesting that a microbe-alga association might limit in part CO fluxes. Extrapolation of CO production rates indicates that macroalgae likely contribute only a minor fraction (<1 %) of global marine CO emissions to the atmosphere (about 10 Tg yr -1 ).

Water loss and viability inZizania(Poaceae) seeds during short‐term desiccation

How Texas wild rice, Zizania texana, became isolated in the San Marcos River of Central Texas, hundreds of kilometres from other wild rice populations is not known. Zizania seeds are intolerant of short-term desiccation. Seeds desiccated at 14% relative humidity (RH) and 75% RH do not survive after only 5-6 d and 2-3 wk of drying. Water loss is rapid and reaches a maximum at the time of seed death due to drying. And although all Zizania seeds germinate well following a long, cold dormancy period, Z. texana seeds readily germinate in the isothermic water (22°C) of the San Marcos River and Springs without an obligate, cold dormant period. Within 30-60 d of collection, Z. texana seeds germinate in substantial numbers, unlike seeds of Z. palustris, which require a long, cold dormant period. The Texas population of Z. texana may represent a relict population of a once more widely dispersed wild rice population, since the San Marcos springs probably have never gone dry.