Application of NMR-based metabolomics to the investigation of salt stress in maize (Zea mays)

Introduction – High salinity, caused by either natural (e.g. climatic changes) or anthropic factors (e.g. agriculture), is a widespread environmental stressor that can affect development and growth of salt‐sensitive plants, leading to water deficit, the inhibition of intake of essential ions and metabolic disorders. Objective – The application of an NMR‐based metabolic profiling approach to the investigation of saline‐induced stress in Maize plants is presented. Methodology – Zea Maize seedlings were grown in either 0, 50 or 150 mM saline solution. Plants were harvested after 2, 4 and 6 days (n = 5 per class and time point) and ¹H NMR spectroscopy was performed separately on shoot and root extracts. Spectral data were analysed and interpreted using multivariate statistical analyses. Results – A distinct effect of time/growth was observed for the control group with relatively higher concentrations of acetoacetate at day 2 and increased levels of alanine at days 4 and 6 in root extracts, whereas concentration of alanine was positively correlated with the shoot extracts harvested at day 2 and trans‐aconitic acid increased at days 4 and 6. A clear dose‐dependent effect, superimposed on the growth effect, was observed for saline treated shoot and root extracts. This was correlated with increased levels of alanine, glutamate, asparagine, glycine‐betaine and sucrose and decreased levels of malic acid, trans‐aconitic acid and glucose in shoots. Correlation with salt‐load shown in roots included elevated levels of alanine, γ‐amino‐N‐butyric acid, malic acid, succinate and sucrose and depleted levels of acetoacetate and glucose. Conclusions – The metabolic effect of high salinity was predominantly consistent with osmotic stress as reported for other plant species and was found to be stronger in the shoots than the roots. Using multivariate data analysis it is possible to investigate the effects of more than one environmental stressor simultaneously. Copyright © 2010 John Wiley & Sons, Ltd.     

Physiological and biochemical adaptations of Cynodon dactylon (L.) Pers. from the Salt Range (Pakistan) to salinity stress

Naturally adapted salt tolerant populations provide a valuable material for exploring the adaptive components of salt tolerance. Under this aspect, two populations of Cynodon dactylon (L.) Pers. were subjected to salt stress in hydroponics. One was collected from a heavily salt-affected soil in the vicinity of a natural salt lake, Uchhali Lake, in the Salt Range of the Punjab province of Pakistan, and the other from a normal non-saline habitat from the Faisalabad region. The NaCl treatments in Hoagland's nutrient solution were: Control (no salt), 50, 100, 150 and 200 mM of NaCl. After 8 weeks of growth in hydroponics produced biomass, ion relations, and photosynthetic capacity were measured in the differently adapted ecotypes. In the ecotype of C. dactylon from the Salt Range, shoot dry weight was only slightly affected by varying levels of salt. However, in contrast, its root weight was markedly increased. On the other hand, the ecotype from Faisalabad (non-saline habitat) showed a marked decrease in shoot and root dry weights under saline regimes. The ecotype from the Salt Range accumulated relatively less amount of Na⁺ in the shoot than did that from Faisalabad, particularly at higher salt levels. Shoot or root K⁺ and Ca²⁺ contents varied inconsistently in both ecotypes under salt stress. All the photosynthetic parameters, leaf water potential and osmotic potential, and chlorophyll content in both ecotypes were adversely affected by salt stress, but all these physiological attributes except turgor potential and soluble sugars were less affected at high salinities in the salt tolerant ecotype from Salt Range. This ecotype accumulated significantly higher organic osmotica (total free amino acids, proline, total soluble proteins, and total soluble sugars) under saline conditions than its intolerant counterpart. Overall, the salt tolerant ecotype of C. dactylon from the Salt Range showed high salt tolerance due to its restricted uptake of Na⁺ accompanied by an increased uptake of K⁺ and Ca²⁺ in the roots as well as shoot due to its higher photosynthetic capacity and accumulation of organic osmotica such as free amino acids and proline under saline conditions.     

NaCl胁迫对坪山柚、福橘实生苗矿质营养吸收特性的影响

通过沙培和水培实验,采用原子吸收分光光度计法和离子吸收动力学方法,对NaCl胁迫下坪山柚(Citrus grandis Osbeck 'Pingshanyou')和福橘(C. reticulata Blanco 'Fuju')实生苗矿质营养吸收特性进行研究. 结果表明, 随NaCl浓度增加, 坪山柚和福橘幼苗地上部及根部Na 、 Cl-含量明显增加, K 、 Ca2 含量降低. 相同浓度NaCl胁迫下, 坪山柚地上部及根部Na 、 Cl-含量分别低于福橘, 而根部K 、 Ca2 含量及地上部K 含量、 K /Na 值、 Ca2 /Na 值均高于福橘.NaCl胁迫下,坪山柚、福橘幼苗Na 吸收量高于Cl-吸收量,Na 及 Cl-的吸收速率均随NaCl浓度的增大而增加.相同浓度NaCl胁迫下,坪山柚对Na 、Cl-的吸收速率低于福橘,这与坪山柚对Na 、Cl-的吸收具有较大的Km值及较小的Vmax值一致,表明坪山柚幼苗比福橘幼苗耐盐性强.     

Protein phosphatase type 2B (calcineurin)-mediated, FK506-sensitive regulation of intracellular ions in yeast is an important determinant for adaptation to high salt stress conditions.

To assess the physiological function of Ca(2+)-dependent protein phosphatase (PP2B) in the yeast Saccharomyces cerevisiae, the phenotypes of PP2B-deficient mutants were investigated. Although PP2B was dispensable for growth under normal conditions, the mutations did, however, cause growth inhibition under certain stress circumstances. The growth of the mutants was inhibited by NaCl and LiCl, but not by KCl, CaCl2, MgCl2 or nonspecific osmotic stresses. Upon shift to high NaCl medium, intracellular Na+ levels of both wild type yeast and the mutants initially increased at a comparable rate. However, internal Na+ in wild type cells started to decline more rapidly than the mutant cells during cultivation in high NaCl medium, indicating that PP2B is important in maintaining a gradient across the membrane. The protection against salt stress was achieved, at least in part, by the stimulation of Na+ export. The maintenance of a high level of internal K+ in high NaCl medium was also PP2B-dependent. In the presence of the immunosuppressant FK506, the growth behaviour and intracellular Na+ and K+ of wild type cells in high NaCl medium became very similar to those of the PP2B-deficient mutant in a manner dependent on the presence of the FK506 binding protein.     

The status of Eurostus dussumierii and Hypsirhina chinensis (Reptilia,Squamata, Serpentes): with comments on the origin of salt tolerance in homalopsid snakes

The Kerala mud snake, Enhydris dussumierii Duméril, Bibron & Duméril, has long been known only from syntypes collected in the nineteenth century, but more recent specimens have provided the opportunity for molecular work. Using Bayesian phylogenetic analysis of 2200 base pairs of cytb, 16S, and c-mos, we recovered the Kerala mud snake as the sister species to the Chinese mud snake, Enhydris chinensis (Chinese–Vietnamese endemic). The DNA results establish the position of dussumierii and chinensis as separate from the Enhydris clade, and together they form the sister group to most of the other fanged homalopsids ((Bitia+Cantoria) + (Erpeton+Gerarda+Fordonia) + (Myron+Pseudoferania) + (Enhydris punctata) + (Homalopsis+Enhydris bocourti) + (Cerberus)). Here we resurrect Ferania Gray for Enhydris sieboldii Gray, establish a new genus for the E. chinensis clade (bennettii+chinensis), and apply the generic replacement name Dieurostus Berg to E. dussumierii Duméril, Bibron & Duméril. A biogeographic scenario is proposed based on the ecology, distribution and salt tolerance of the species composing a hypothesized (not supported with molecular evidence) Asian coastal lineage that is distributed from Pakistan's Indus River delta to temperate coastal China.     

Physiological adaptation in relation to hyperosmotic stress in the epidermis of a fresh-water teleost Barbus sophor (Cypriniformes, Cyprinidae): a histochemical study.

The changes in distribution of mucopolysaccharides, glycogen and protein bound sulphydryl groups of cysteine in the various cellular components of the epidermis of Barbus sophor along with its structural alterations as a result of hyperosmotic stress, have been described using histochemical techniques. The hyperosmotic saline induces a cyclic secretion of acid mucopolysaccharides by the mucous cells. Simultaneously the polygonal cells also show a marked disturbance in the processes of mucogenesis and keratinization, indicating an inverse relationship between the degree of keratinization and the amount of mucus secreted by epidermis. The role of glycogen in the polygonal cells has been discussed in relation to disturbed mucogenesis. The appearance of intercellular spaces in basal layer and middle layer has been correlated with the passage for movement of increased amounts of nutrients through the skin.     

Responses to Mg/Ca balance in an Iranian serpentine endemic plant, Cleome heratensis (Capparaceae) and a related non-serpentine species, C. foliolosa

A soil Ca/Mg quotient greater than unity is generally considered necessary for normal plant growth but some serpentine plants are adapted to much lower Ca/Mg quotients, resulting from a major cation imbalance in their substrata. In order to investigate the growth and tolerance responses of serpentine and non-serpentine species to varied Ca/Mg quotients, controlled nutrient solution experiments were performed using an a newly reported Iranian endemic serpentine plant, Cleome heratensis Bunge et Bien. Ex Boiss. and a related non-serpentine species Cleome foliolosa DC. and a Eurasian Ni-hyperaccumulating species Alyssum murale Waldst. and Kit. Seedlings were grown in modified Hoagland’s solutions with varying Ca and Mg concentrations (0.2–2.5 and 0.5–10 mM, respectively) in a fully factorial randomised block design. The yields of the two serpentine plants increased significantly as Mg concentrations in the nutrient solution were increased from 0.5 to 4 mM but decreased in the 10 mM Mg treatment. For C. foliolosa yields decreased significantly from 0.5 to 10 mM Mg, indicating the sensitivity of this non-serpentine plant, and the relative tolerance of the serpentine plants to extremely high levels of Mg. Shoot and root Mg and Ca concentrations in C. heratensis and A. murale were higher than those in C. foliolosa in the low and moderate Mg treatments, supporting the view that many serpentine plants have a relatively high requirement for Mg. Maximum Mg concentrations were found in the roots of C. heratensis. Yields of C. heratensis and A. murale did not change significantly as Ca levels in nutrient solution increased from 0.2 to 2.5 mM Ca, However the yield of C. foliolosa increased significantly from 0.2 to 1.5 mM Ca, indicating sensitivity in this non-serpentine plant and tolerance of the two serpentine plants to low levels of Ca correlated with tissue Ca concentrations, probably because of a greater ability for Ca uptake at low-Ca availability. Calcium deficiency in the low-Ca treatments could be a reason for reduced yield in the non-serpentine plants.     

Expression of dehydration-stress-related genes in the crowns of wheatgrass species [Lophopyrum elongatum (Host) A. Love and Agropyron desertorum (Fisch. ex Link.) Schult.] having contrasting acclimation to salt, cold and drought

Agropyron desertorum and Lophopyrum elongatum were grown in a control environment or acclimated in high‐salt (daily exposure to 75 or 150 m M NaCl for 6 d), cold (6/2 °C for 14 d) or drought environments (watering withheld for 6 d). Lophopyrum elongatum was constitutively tolerant to salt and also acclimated more to salt than did A. desertorum whereas A. desertorum acclimated more to cold and drought. Dehydrin and non‐specific lipid transfer protein (nsLTP) mRNA sequences and polypeptides increased more, during acclimation to cold and drought, in A. desertorum than in L. elongatum crowns. Expression of immunologically identified dehydrin polypeptides was much higher in drought‐acclimated A. desertorum than in any other species/treatment combination. The most strongly expressed were 42 and 20 kDa. No change in dehydrin or nsLTP polypeptides were detected in the crowns during acclimation to salt. Overall, there was stronger acclimation to dehydrative stresses, at the molecular level, in A. desertorum than in L. elongatum crowns. Differences in dehydrin and nsLTP mRNA and polypeptide expression during acclimation to different stresses indicated that plants sense the differences between different primary potential causes of cellular dehydration.     

Expression profiling of the 14-3-3 gene family in response to salt stress and potassium and iron deficiencies in young tomato (Solanum lycopersicum) roots: analysis by real-time RT-PCR

BACKGROUND AND AIMS Mineral nutrient deficiencies and salinity constitute major limitations for crop plant growth on agricultural soils. 14-3-3 proteins are phosphoserine-binding proteins that regulate the activities of a wide array of targets via direct protein-protein interactions and may play an important role in responses to mineral nutrients deficiencies and salt stress. In the present study, the expression profiling of the 14-3-3 gene family in response to salt stress and potassium and iron deficiencies in young tomato (Solanum lycopersicum) roots was investigated in order to analyse the 14-3-3 roles of the proteins in these abiotic stresses. METHODS: Sequence identities and phylogenetic tree creation were performed using DNAMAN version 4.0 (Lynnon Biosoft Company). Real-time RT-PCR was used to examine the expression of each 14-3-3 gene in response to salt stress and potassium and iron deficiencies in young tomato roots. KEY RESULTS: The phylogenetic tree shows that the 14-3-3 gene family falls into two major groups in tomato plants. By using real-time RT-PCR, it was found that (a) under normal growth conditions, there were significant differences in the mRNA levels of 14-3-3 gene family members in young tomato roots and (b) 14-3-3 proteins exhibited diverse patterns of gene expression in response to salt stress and potassium and iron deficiencies in tomato roots. CONCLUSIONS: The results suggest that (a) 14-3-3 proteins may be involved in the salt stress and potassium and iron deficiency signalling pathways in young tomato roots, (b) the expression pattern of 14-3-3 gene family members in tomato roots is not strictly related to the position of the corresponding proteins within a phylogenetic tree, (c) gene-specific expression patterns indicate that isoform-specificity may exist in the 14-3-3 gene family of tomato roots, and (d) 14-3-3 proteins (TFT7) might mediate cross-talk between the salt stress and potassium and iron-deficiency signalling pathways in tomato roots.     

A comparison of the effect of long-term starvation on responses to low-temperature stress by juvenile rainbow (Oncorhynchus mykiss) and brown (Salmo trutta) trout reveal different responses in the two species

Biometric parameters and oxidative stress indicators were measured in liver and muscle samples from rainbow trout and brown trout juveniles exposed to a 45-day starvation period at low water temperature. As a general tendency, hepatic antioxidant enzyme activities in both species increased with fasting to eliminate the harmful effects of reactive oxygen species (ROS). However, the metabolic response to food deprivation in the muscle of each species was different. Lipid peroxidation levels in both species increased with starvation. We concluded that (1) low water temperature enhances ROS production in salmonids because of increased polyunsaturated fatty acid content in the cell membrane; (2) starvation significantly impaired the growth parameters of brown trout, yet the reverse was found for rainbow trout; and (3) despite this negative interaction, brown trout juveniles can physiologically tolerate oxidative stress caused by starvation and can therefore be cultivated under stressful conditions even in their early life stages.     

Vector competence of South African Culicoides species for bluetongue virus serotype 1 (BTV-1) with special reference to the effect of temperature on the rate of virus replication in C. imicola and C. bolitinos

Abstract. The oral susceptibility of 22 South African livestock associated Culicoides species to infection with bluetongue virus serotype 1 (BTV‐1) and its replication rate in C. imicola Kieffer and C. bolitinos Meiswinkel (Diptera: Ceratopogonidae) over a range of different incubation periods and temperatures are reported. Field‐collected Culicoides were fed on sheep blood containing 7.5 log₁₀TCID₅₀/mL of BTV‐1, and then held at constant different temperatures. Virus replication was measured over time by assaying individual flies in BHK‐21 cells using a microtitration procedure. Regardless of the incubation temperatures (10, 15, 18, 23.5 and 30°C) the mean virus titre/midge, infection rates (IR) and the proportion of infected females with transmission potential (TP = virus titre/midge ≥ 3 log₁₀ TCID₅₀) were found to be significantly higher in C. bolitinos than in C. imicola. Results from days 4–10 post‐infection (dpi), at 15–30°C, shows that the mean IR and TP values in C. bolitinos ranged from 36.7 to 87.8%, and from 8.4 to 87.7%, respectively; in C. imicola the respective values were 11.0–13.7% and 0–46.8%. In both species the highest IR was recorded at 25°C and the highest TP at 30°C. The time required for the development of TP in C. bolitinos ranged from 2 dpi at 25°C to 8 dpi at 15°C. In C. imicola it ranged from 4 dpi at 30°C to 10 dpi at 23.5°C; no individuals with TP were detected at 15°C. There was no evidence of virus replication in flies held at 10°C. When, at various points of incubation, individual flies were transferred from 10°C to 23.5°C and then assayed 4–10 days later, virus was recovered from both species. The mean virus titres/midge, and proportion of individuals with TP and IR, were again significantly higher in C. bolitinos than in C. imicola. Also the infection prevalence in C. magnus Colaço was higher than in C. imicola. Low infection prevalences were found in C. bedfordi Ingram & Macfie, C. leucostictus Kieffer, C. pycnostictus Ingram & Macfie, C. gulbenkiani Caeiro and C. milnei Austen. BTV‐1 was not detected in 14 other Culicoides species tested; however, some of these were tested in limited numbers. The present study indicates a multivector potential for BTV transmission in South Africa. In C. imicola and C. bolitinos the replication rates are distinct and are significantly influenced by temperature. These findings are discussed in relation to the epidemiology of bluetongue in South Africa.     

Histological studies on the host response to Myxobolus artus Akhmerov, 1960 (Myxozoa: Myxobolidae) infection in the skeletal muscle of carp, Cyprinus carpio L.

Myxobolus artus infection of carp is reported for the first time from Japan. In most cases, it took the form of interfibrillar pseudocysts in the skeletal muscle, some showing abnormal growth, imperfect encapsulation of the parasite by the host connective tissue and irregular development of the ectoplasm. Such atypical pseudocysts probably ruptured before the completion of sporogenesis, causing lysis of adjacent muscle fibres. Intrafibrillar pseudocysts were also present. After spore formation was complete or atypical pseudocytes were ruptured host reaction was initiated. Spores were phagocytosed and transported to the melanomacrophage centres in the kidney, liver, spleen etc. Phagocytosed spores were also observed in the capillaries and epithelial tissues of various organs, suggesting that the spores were transported there from the skeletal muscle through the host vascular system. They are thought to be discharged from the host through the skin and intestine.     

The influence of the ectomycorrhizal fungus Rhizopogon subareolatus on growth and nutrient element localisation in two varieties of Douglas fir (Pseudotsuga menziesii var. menziesii and var. glauca) in response to manganese stress

Acidification of forest ecosystems leads to increased plant availability of the micronutrient manganese (Mn), which is toxic when taken up in excess. To investigate whether ectomycorrhizas protect against excessive Mn by improving plant growth and nutrition or by retention of excess Mn in the hyphal mantle, seedlings of two populations of Douglas fir (Pseudotsuga menziesii), two varieties, one being menziesii (DFM) and the other being glauca (DFG), were inoculated with the ectomycorrhizal fungus Rhizopogon subareolatus in sand cultures. Five months after inoculation, half of the inoculated and non-inoculated seedlings were exposed to excess Mn in the nutrient solution for further 5 months. At the end of this period, plant productivity, nutrient concentrations, Mn uptake and subcellular compartmentalisation were evaluated. Non-inoculated, non-stressed DFM plants produced about 2.5 times more biomass than similarly treated DFG. Excess Mn in the nutrient solution led to high accumulation of Mn in needles and roots but only to marginal loss in biomass. Colonisation with R. subareolatus slightly suppressed DFM growth but strongly reduced that of DFG (-50%) despite positive effects of mycorrhizas on plant phosphorus nutrition. Growth reductions of inoculated Douglas fir seedlings were unexpected since the degree of mycorrhization was not high, i.e. ca. 30% in DFM and 8% in DFG. Accumulation of high Mn was not prevented in inoculated seedlings. The hyphal mantle of mycorrhizal root tips accumulated divalent cations such as Ca, but not Mn, thus not providing a barrier against excessive Mn uptake into the plants associated with R. subareolatus.     

Notes on the ant genus Cataglyphis Foerster, 1850 (Hymenoptera,Formicidae) in the Arabian Peninsula with description of a new species and a key to species of the C. pallida-group

Cataglyphis fisheri sp. n. is described and illustrated from the United Arab Emirates, Oman and Kingdom of Saudi Arabia based on the worker caste. It belongs to the Cataglyphis pallida-group which is recorded for the first time from the Arabian Peninsula. Cataglyphis fisheri sp. n. is similar to Cataglyphis pallida Mayr, 1877 from Kazakhstan. Differential diagnosis between these two species is given and a key to the species of the Cataglyphis pallida-group is presented. Cataglyphis laylae Collingwood, 2011 is treated as a junior synonym of Cataglyphis saharae Santschi, 1929. Cataglyphis flavobrunnea Collingwood & Agosti, 1996 is redescribed and a lectotype for this species is designated.