High Osmotic Stress Research Articles

Investigation of the instability and low water kefir grain growth during an industrial water kefir fermentation process.

A poorly performing industrial water kefir production process consisting of a first fermentation process, a rest period at low temperature, and a second fermentation process was characterized to elucidate the causes of its low water kefir grain growth and instability. The frozen-stored water kefir grain inoculum was thawed and reactivated during three consecutive prefermentations before the water kefir production process was started. Freezing and thawing damaged the water kefir grains irreversibly, as their structure did not restore during the prefermentations nor the production process. The viable counts of the lactic acid bacteria and yeasts on the water kefir grains and in the liquors were as expected, whereas those of the acetic acid bacteria were high, due to the aerobic fermentation conditions. Nevertheless, the fermentations progressed slowly, which was caused by excessive substrate concentrations resulting in a high osmotic stress. Lactobacillus nagelii, Lactobacillus paracasei, Lactobacillus hilgardii, Leuconostoc mesenteroides, Bifidobacterium aquikefiri, Gluconobacter roseus/oxydans, Gluconobacter cerinus, Saccharomyces cerevisiae, and Zygotorulaspora florentina were the most prevalent microorganisms. Lb. hilgardii, the microorganism thought to be responsible for water kefir grain growth, was not found culture-dependently, which could explain the low water kefir grain growth of this industrial process.

Adaptive Diversity of Echinochloa Species to Osmotic Stress

Three Echinochloa species inhabit various crop fields with different soil moisture conditions. Therefore, a growth pouch test was conducted to investigate adaptive diversity of six Echinochloa species, three from Korea and three from USA, to osmotic stress by assessing shoot and root growths. Echinochloa crus-galli var. praticola showed the greatest tolerance to osmotic stress in both root (GR 50 =1316.3 g PEG L −1 ) and shoot (GR 50 =212.2 g PEG L −1 ) growths, while Korean E. oryzicola was most sensitive to osmotic stress in both root (GR 50 =116 g PEG L −1 ) and shoot (GR 50 =126.2 g PEG L −1 ) growths. Root to shoot (R/S) ratio of Echinochloa crus-galli var. praticola increased with increasing osmotic stress, while that of Korean E. oryzicola decreased, suggesting that R/S ratio is closely related to osmotic stress tolerance in Echinochloa species. Our results clearly demonstrate that E. crus-galli var. praticola maintains high R/S ratio even under high osmotic stress, which enables this species to well adapt to dry upland condition. In contrast, while E. oryzicola fails to maintain sufficiently high R/S ratio, resulting in poor adaptability to dry upland condition.

Salinity increases total body prolactin and gill and skin prolactin receptor expression in the Chinese edible frog,Hoplobatrachus rugulosus, tadpole

Water salinity has effects on growth and metamorphosis of anuran species, including Hoplobatrachus rugulosus. Previously, we reported that cultured H. rugulosus tadpoles at low salinities (2–4‰) were of bigger size and grew faster than those in fresh water (FW). However, at a higher salinity level of 6‰, their sizes were reduced and the metamorphosis was delayed. It was therefore hypothesized that high salinity-induced osmotic stress affected secretion of prolactin (PRL), which acts as osmoregulatory hormone and a regulator of metamorphosis in amphibians. In this study, transferring tadpoles into 4‰ and 6‰ brackish water increased the PRL levels by ~1.2- and ~twofold, respectively, as compared to FW group. These osmotic challenges also increased the total body fluid osmolality and levels of Na+, Cl− and Ca2+. The contents of triiodothyronine (T3) were significantly reduced in 4‰ and 6‰ groups, but not 2‰ group. Three sizes (49, 29 and 23 kDa) of H. rugulosus PRL receptors (PRLRs) were detected, and their protein expression was found in the skin, gill, tail fin, brain, intestine, heart, liver and kidney. The expression of PRLR-49k protein was significantly higher in both skin and gills in 2–4‰ groups, whereas that of PRLR-29k and PRLR-23k were higher in gills and lower in skin of the 6‰ group than FW group. In conclusion, salinity challenge, particularly 4‰ and 6‰, increased the levels of PRL, while decreasing the T3 levels, which could explain why salinity markedly modulated growth, metamorphosis and survival of tadpoles.

MAP kinase gene STK1 is required for hyphal, conidial, and appressorial development, toxin biosynthesis, pathogenicity, and hypertonic stress response in the plant pathogenic fungus Setosphaeria turcica

The mitogen-activated protein kinase (MAPK), a key signal transduction component in the MAPK cascade pathway, regulates a variety of physiological activities in eukaryotes. However, little is known of the role MAPK plays in phytopathogenic fungi. In this research, we cloned the MAPK gene STK1 from the northern corn leaf blight pathogen Setosphaeria turcica and found that the gene shared high homology with the high osmolality glycerol (HOG) MAPK gene HOG1 of Saccharomyces cerevisiae. In addition, gene knockout technology was employed to investigate the function of STK1. Gene knockout mutants (KOs) were found to have altered hyphae morphology and no conidiogenesis, though they did show similar radial growth rate compared to the wild-type strain (WT). Furthermore, microscope observations indicated that STK1 KOs did not form normal appressoria at 48 h post-inoculation on a hydrophobic surface. STK1 KOs had reduced virulence, a significantly altered Helminthosporium turcicum (HT)-toxin composition, and diminished pathogenicity on the leaves of susceptible inbred corn OH43. Mycelium morphology appeared to be significantly swollen and the radial growth rates of STK1 KOs declined in comparison with WT under high osmotic stress. These results suggested that STK1 affects the hyphae development, conidiogenesis, and pathogenicity of S. turcica by regulating appressorium development and HT-toxin biosynthesis. Moreover, the gene appears to be involved in the hypertonic stress response in S. turcica.

PiHOG1, a stress regulator MAP kinase from the root endophyte fungus Piriformospora indica, confers salinity stress tolerance in rice plants.

In this study, yeast HOG1 homologue from the root endophyte Piriformospora indica (PiHOG1) was isolated and functionally characterized. Functional expression of PiHOG1 in S. cerevisiae ∆hog1 mutant restored osmotolerance under high osmotic stress. Knockdown (KD) transformants of PiHOG1 generated by RNA interference in P. indica showed that genes for the HOG pathway, osmoresponse and salinity tolerance were less stimulated in KD-PiHOG1 compared to the wild-type under salinity stress. Furthermore, KD lines are impaired in the colonization of rice roots under salinity stress of 200 mM NaCl, and the biomass of the host plants, their shoot and root lengths, root number, photosynthetic pigment and proline contents were reduced as compared to rice plants colonized by WT P. indica. Therefore, PiHOG1 is critical for root colonisation, salinity tolerance and the performance of the host plant under salinity stress. Moreover, downregulation of PiHOG1 resulted not only in reduced and delayed phosphorylation of the remaining PiHOG1 protein in colonized salinity-stressed rice roots, but also in the downregulation of the upstream MAP kinase genes PiPBS2 and PiSSK2 involved in salinity tolerance signalling in the fungus. Our data demonstrate that PiHOG1 is not only involved in the salinity response of P. indica, but also helping host plant to overcome salinity stress.

Strain improvement of Pichia kudriavzevii TY13 for raised phytase production and reduced phosphate repression.

SummaryIn this work, we present the development and characterization of a strain of Pichia kudriavzevii (TY1322), with highly improved phytate‐degrading capacity. The mutant strain TY1322 shows a biomass‐specific phytate degradation of 1.26 mmol g−1 h−1 after 8 h of cultivation in a high‐phosphate medium, which is about 8 times higher compared with the wild‐type strain. Strain TY1322 was able to grow at low pH (pH 2), at high temperature (46°C) and in the presence of ox bile (2% w/v), indicating this strain's ability to survive passage through the gastrointestinal tract. The purified phytase showed two pH optima, at pH 3.5 and 5.5, and one temperature optimum at 55°C. The lower pH optimum of 3.5 matches the reported pH of the pig stomach, meaning that TY1322 and/or its phytase is highly suitable for use in feed production. Furthermore, P. kudriavzevii TY1322 tolerates ethanol up to 6% (v/v) and shows high osmotic stress tolerance. Owing to the phenotypic characteristics and non‐genetically modified organisms nature of TY1322, this strain show great potential for future uses in (i) cereal fermentations for increased mineral bioavailability, and (ii) feed production to increase the phosphate bioavailability for monogastric animals to reduce the need for artificial phosphate fortification.

Overexpression of a Barley Aquaporin Gene, HvPIP2;5 Confers Salt and Osmotic Stress Tolerance in Yeast and Plants.

We characterized an aquaporin gene HvPIP2;5 from Hordeum vulgare and investigated its physiological roles in heterologous expression systems, yeast and Arabidopsis, under high salt and high osmotic stress conditions. In yeast, the expression of HvPIP2;5 enhanced abiotic stress tolerance under high salt and high osmotic conditions. Arabidopsis plants overexpressing HvPIP2;5 also showed better stress tolerance in germination and root growth under high salt and high osmotic stresses than the wild type (WT). HvPIP2;5 overexpressing plants were able to survive and recover after a 3-week drought period unlike the control plants which wilted and died during stress treatment. Indeed, overexpression of HvPIP2;5 caused higher retention of chlorophylls and water under salt and osmotic stresses than did control. We also observed lower accumulation of reactive oxygen species (ROS) and malondialdehyde (MDA), an end-product of lipid peroxidation in HvPIP2;5 overexpressing plants than in WT. These results suggest that HvPIP2;5 overexpression brought about stress tolerance, at least in part, by reducing the secondary oxidative stress caused by salt and osmotic stresses. Consistent with these stress tolerant phenotypes, HvPIP2;5 overexpressing Arabidopsis lines showed higher expression and activities of ROS scavenging enzymes such as catalase (CAT), superoxide dismutase (SOD), glutathione reductase (GR), and ascorbate peroxidase (APX) under salt and osmotic stresses than did WT. In addition, the proline biosynthesis genes, Δ1-Pyrroline-5-Carboxylate Synthase 1 and 2 (P5CS1 and P5CS2) were up-regulated in HvPIP2;5 overexpressing plants under salt and osmotic stresses, which coincided with increased levels of the osmoprotectant proline. Together, these results suggested that HvPIP2;5 overexpression enhanced stress tolerance to high salt and high osmotic stresses by increasing activities and/or expression of ROS scavenging enzymes and osmoprotectant biosynthetic genes.

Characterization and functional analysis of a MADS-box transcription factor gene (GbMADS9) from Ginkgo biloba

The MADS-box transcription factors play diverse roles in plant development, but little is known about the role of MADS-box genes in Ginkgo biloba. Here a MADS-box transcription factor gene, designated as GbMADS9, was cloned and functionally characterized from G. biloba. The open reading frame of GbMADS9 comprised 684bp encoding a polypeptide of 227 amino acids. Phylogenetic analysis showed that GbMADS9 belonged to the GGM13 clade of Bsister-class MADS-box proteins. Real-time PCR results revealed that GbMADS9 was preferentially expressed in strobili and ovules. In addition, its expression was up-regulated with the increase in the size of ginkgo pulp, indicating that GbMADS9 had a positive function in ginkgo ovule development. The expression of GbMADS9 in ginkgo was up-regulated in response to salinity, drought, and cold stress, as well as to phytohormones gibberellic acid and abscisic acid. GbMADS9 transgenic Arabidopsis plants presented early flowering phenotypes compared with wild-type plants. The expression of the FLOWERING LOCUS T (FT), APETALA1 (AP1), LEAFY (LFY), and SUPPRESSOR OF OVEREXPRESSION OF CO 1 (SOC1) genes involved in flowering was up-regulated, whereas the expression of AGAMOUS-LIKE24 (AGL24) and SHORT VEGETATIVE PHASE (SVP) was down-regulated in the GbMADS9 transgenic lines. The GbMADS9 transgenic plants exhibited better growth than wild-type plants under high osmotic stress. Enhanced tolerance of transgenic Arabidopsis plants to osmotic stress was confirmed by changes in the chlorophyll, proline, and malondialdehyde contents. Moreover, increased tolerance to osmotic stress in GbMADS9 transgenic lines was associated with improved antioxidant enzymatic activities. These findings suggested that GbMADS9 might be a promising gene to manipulate flowering time and improve abiotic stress tolerance in G. biloba.

The occurrence of spoilage yeasts in cream-filled bakery products

Filling creams can provide an adequate substrate for spoilage yeasts because some yeasts can tolerate the high osmotic stress in these products. To discover the source of spoilage of a cream-filled baked product, end products, raw materials, indoor air and work surfaces were subjected to microbiological and molecular analyses. The efficacy of disinfectants against spoilage yeasts was also assessed. The analyses on end products revealed the presence of the closest relatives to Zygosaccharomyces bailii with counts ranging from 1.40 to 4.72 log cfu g-1 . No spoilage yeasts were found in the indoor air and work surfaces. Polymerase chain reaction-denaturing gradient gel electrophoresis analysis, carried out directly on filling creams collected from unopened cans, showed the presence of bands ascribed to the closest relatives to Z. bailii sensu lato, although with counts < 1 log cfu g-1 . Susceptibility testing of yeast isolates to disinfectants showed a significantly lower effect of 10% alkyl dimethyl benzyl ammonium chloride. Different responses of isolates to the tested disinfectants were seen. To guarantee the quality of end products, reliable and sensitive methods must be used. Moreover, hygiene and the application of good manufacturing practices represent the most efficient way for the prevention and minimization of cross-contamination. © 2016 Society of Chemical Industry.

Identification and expression analysis of cytokinin response regulators in Fragaria vesca

Cytokinin response regulators (RRs) are important components of the two component signal systems, which are involved in the regulation of plant growth and development, and in the response to abiotic stress. In this study, 18 cytokinin RR genes were identified in Fragaria vesca through the genome-wide search. They were further classified into three types: type-A (FvRR1–7), type-B (FvRR8–14) and type-C (FvRR15–18) according to the domain architecture and the phylogeny. Phylogenetic analysis demonstrated that most cytokinin response regulators of F. vesca and Arabidopsis formed clear orthologous pairs. Expression patterns of the cytokinin FvRR genes in various tissues and organs at reproductive stages were detected in this study. Additionally, gene expression response patterns to ABA and abiotic stresses including high temperature and osmotic stress were investigated. The results showed that different types of cytokinin FvRRs have different expression patterns, suggesting the functional differentiation of cytokinin FvRRs during the evolution. This systematic study provides insights into possible functions of the cytokinin FvRR genes and a basis for further functional analysis.

Salinity and osmotic stress trigger different antioxidant responses related to cytosolic ascorbate peroxidase knockdown in rice roots

Salinity and osmotic stress trigger distinct signals in roots, which might induce differences in antioxidant responses. To clarify these relationships, transgenic rice plants silenced in both cytosolic ascorbate peroxidases (apx1/2) and non-transformed (NT) were exposed to iso-osmotic concentrations of NaCl and mannitol. Under both stress conditions, apx1/2 roots did not suffer oxidative stress, revealing that cytosolic APXs were not crucial to oxidative protection. Silenced and non-transformed roots triggered different responses to high salinity and osmotic stress and these stressful factors induced also distinct antioxidant changes. High salinity up-regulated expression of important OsAPX isoforms and these changes were related to increased APX activity, especially in NT roots. Intriguingly, salt stress triggered up-regulation of OsCAT isoforms but CAT activity did not change in both genotypes. In contrast, mannitol trigged very low increment in expression of OsAPX isoforms but induced substantial up-regulation in APX activity in NT roots. Mannitol also remarkably up-regulated OsCATB expression in parallel to CAT activity, in both apx1/2 and NT roots. POD and GPX (glutathione peroxidases) activities were strongly increased by high salinity but did not change in response to mannitol, in both genotypes. The two stress types as well as apx1/2 and NT roots displayed different response in terms of modulation in the H2O2 levels but lipid peroxidation did not change. Membrane integrity was drastically affected by both stressful factors and similarly in both genotypes, whereas root fresh matter was affected only by salt stress. Altogether, the obtained data reveal that high salinity and osmotic stress trigger different antioxidant responses and these strategies were genotype-dependent. The different antioxidant molecular-biochemical mechanisms employed by cytosolic APX knockdown and non-transformed roots allowed reaching similar physiological performance.

Comparative genomic and transcriptomic analyses of the Fuzhuan brick tea-fermentation fungus Aspergillus cristatus.

BackgroundAspergillus cristatus is the dominant fungus involved in the fermentation of Chinese Fuzhuan brick tea. Aspergillus cristatus is a homothallic fungus that undergoes a sexual stage without asexual conidiation when cultured in hypotonic medium. The asexual stage is induced by a high salt concentration, which completely inhibits sexual development. The taxon is therefore appropriate for investigating the mechanisms of asexual and sexual reproduction in fungi. In this study, de novo genome sequencing and analysis of transcriptomes during culture under high- and low-osmolarity conditions were performed. These analyses facilitated investigation of the evolution of mating-type genes, which determine the mode of sexual reproduction, in A. cristatus, the response of the high-osmolarity glycerol (HOG) pathway to osmotic stimulation, and the detection of mycotoxins and evaluation of the relationship with the location of the encoding genes.ResultsThe A. cristatus genome comprised 27.9 Mb and included 68 scaffolds, from which 10,136 protein-coding gene models were predicted. A phylogenetic analysis suggested a considerable phylogenetic distance between A. cristatus and A. nidulans. Comparison of the mating-type gene loci among Aspergillus species indicated that the mode in A. cristatus differs from those in other Aspergillus species. The components of the HOG pathway were conserved in the genome of A. cristatus. Differential gene expression analysis in A. cristatus using RNA-Seq demonstrated that the expression of most genes in the HOG pathway was unaffected by osmotic pressure. No gene clusters associated with the production of carcinogens were detected.ConclusionsA model of the mating-type locus in A. cristatus is reported for the first time. Aspergillus cristatus has evolved various mechanisms to cope with high osmotic stress. As a fungus associated with Fuzhuan tea, it is considered to be safe under low- and high-osmolarity conditions.Electronic supplementary materialThe online version of this article (doi:10.1186/s12864-016-2637-y) contains supplementary material, which is available to authorized users.

Oleanolic Acid Inhibits High Salt-Induced Exaggeration of Warburg-like Metabolism in Breast Cancer Cells.

Cancer cells have a proliferative advantage by utilizing intermediates of aerobic glycolysis (Warburg effect) for their macromolecule synthesis. Although the exact causes of this Warburg effect are unclear, high osmotic stress in solid tumor microenvironment is considered one of the important factors. Oleanolic acid (OA) is known to exert anti-inflammatory and anti-cancer effect. In our current studies, using breast cancer cell lines, we determined the protective role of OA in high salt-mediated osmotic stress-induced cancer growth. Hypertonic (0.16M NaCl) culture conditions enhanced the cancer cell growth (26%, p<0.05) and aerobic glycolysis as marked by increased glucose consumption (34%, p<0.05) and lactate production (25%, p<0.05) over untreated cells. This effect was associated with increased expression and activity of key rate-limiting enzymes of aerobic glycolysis, namely hexokinase, pyruvate kinase type M2, and lactate dehydrogenase A. Interestingly, this high salt-mediated enhanced expression of aerobic glycolytic enzymes was efficiently reversed by OA along with the decreased cancer cell proliferation. In cancer cells, enhanced aerobic glycolysis is associated with the decreased mitochondrial activity and mitochondrial-associated caspase activity. As expected, high salt further inhibited the mitochondrial related cytochrome oxidase and caspase-3 activity. However, OA efficiently reversed the high salt-mediated inhibition of cytochrome oxidase, caspase activity, and pro-apoptotic Bax expression, thus suggesting that OA induced mitochondrial activity and enhanced apoptosis. Taken together, our data indicate that OA efficiently reverses the enhanced Warburg-like metabolism induced by high salt-mediated osmotic stress along with potential application of OA in anti-cancer therapy.

Stress promotes Arabidopsis - Piriformospora indica interaction

ABSTRACTThe endophytic fungus Piriformospora indica colonizes Arabidopsis thaliana roots and promotes plant performance, growth and resistance/tolerance against abiotic and biotic stress. Here we demonstrate that the benefits for the plant increase when the two partners are co-cultivated under stress (limited access to nutrient, exposure to heavy metals and salt, light and osmotic stress, pathogen infection). Moreover, physical contact between P. indica and Arabidopsis roots is necessary for optimal growth promotion, and chemical communication cannot replace the physical contact. Lower nutrient availability down-regulates and higher nutrient availability up-regulates the plant defense system including the expression of pathogenesis-related genes in roots. High light, osmotic and salt stresses support the beneficial interaction between the plant and the fungus. P. indica reduces stomata closure and H2O2 production after Alternaria brassicae infection in leaves and suppresses the defense-related accumulation of the phytohormone jasmonic acid. Thus, shifting the growth conditions toward a stress promotes the mutualistic interaction, while optimal supply with nutrients or low stress diminishes the benefits for the plant in the symbiosis.

Effect of Heat Shock Treatment on Endothelial Cell Integrity after Histamine Challenge

Endothelial cells and endothelial cell integrity play a central role in circulatory homeostasis. Our previous study indicated that heat shock treatment reduces protein leakage and attenuates the hypotension caused by anaphylactic shock. Also, heat shock treatment maintains cerebral vascular permeability, while the blood brain barrier is opened by high osmotic stress. In this study we investigated the potential mechanism by which heat shock protein 72 (Hsp72), the major product after heat shock, maintains cellular integrity of cultured histamine-induced human umbilical vein endothelial cells (HUVECs) upon challenge by histamine. Tight junction proteins (ZO-1 and occludin) and a cytoskeleton component (F-actin) were evaluated by an immunocytochemical study. The immunochemical density of ZO-1 and occludin was clearly decreased in the non-heated cells after histamine challenge, and associated with the proportional decline of F-actin. The adverse phenomenon was prevented in the heated cells in which Hsp72 was significantly over-expressed. It was further demonstrated that Hsp72 could be exclusively coimmunoprecipitated with ZO-1 or occludin in heated cells. In conclusion, histamine-induced conformational alteration of HUVECs is attenuated by previous heat shock treatment, and the Hsp72-mediated preservation of tight junction proteins, as well as cytoskeletal structure, could be the possible mechanism of action.

Transgenic NfFeSOD Sedum alfredii plants exhibited profound growth impairments and better relative tolerance to long-term abiotic stresses

Transgenic research was preformed by transferring a cyanobacterial (Nostoc flagelliforme) iron superoxide dismutase gene (NfFeSOD) into heavy metal hyperaccumulator Sedum alfredii via Agrobacterium-mediated method. Beyond expectation, NfFeSOD-overexpressing S. alfredii plants exhibited profound impairments, including plant growth retardation, abnormal root architecture, and reduced leaf greenness, photosynthetic efficiency and metal accumulation efficiency. Although transgenic plants appeared physiologically sensitive to high temperature, a higher relative biomass growth was still observed under long-term high temperature and osmotic stresses. Further investigation found that reactive oxygen species (ROS) homeostasis of transgenic plants was significantly affected, being ~50 % reduction of H2O2 level relative to wild-type plants. Gene transcription including ROS responsive genes was overall attenuated in transgenic plants, being more significant at normal temperature than at high temperature. In addition, ascorbate peroxidase (APX) activity was increased nearly twofolds in transgenic plants as compared to wild-type control. It may be inferred that ectopic NfFeSOD overexpression gives rise to a substantial increase of APX activity and leads to a sharp reduction of H2O2 level, thus impairing basal ROS signaling and plant growth. Specific genetic background of S. alfredii may be responsible for this sharp reduction of H2O2 level induced by NfFeSOD overexpression. S. alfredii plant has acclimated to elevated levels of ROS induced by heavy metals in native habitats and should require high ROS levels for basal signaling. We thus suppose that a sustained disturbance of high basal ROS signaling in metal hyperaccumulators may instead incur very sensitive response and thus result in profound growth impairments.

ZxSKOR is important for salinity and drought tolerance of Zygophyllum xanthoxylum by maintaining K+ homeostasis

As an important nutrient element, K+ plays a crucial role in plant stress resistance. It was reported that the stelar K+ outward rectifying channel (SKOR) is involved in loading K+ into xylem for its transport from roots to shoots. Zygophyllum xanthoxylum, a succulent woody xerophyte, could maintain stable K+ concentration in leaves to adapt to salt and arid environments. Here we characterized ZxSKOR from Z. xanthoxylum, ZxSKOR expression patterns and Na+ and K+ accumulation in Z. xanthoxylum treated with various concentrations of KCl and NaCl and −0.5 MPa osmotic stress were investigated in order to assess the contribution of ZxSKOR to K+ homeostasis. The results showed that ZxSKOR was predominantly expressed in roots and stems rather than in leaves. Its expression levels in roots and stems increased significantly accompanied by an increase in K+ concentration in leaves when plants were exposed to 5–10 mM KCl. Moreover, a positive correlation was identified not only between ZxSKOR expression in roots and K+ accumulation in shoots, but also between ZxSKOR expression in stems and K+ accumulation in leaves. Transcription levels of ZxSKOR in roots and stems under high salinity (100–150 mM NaCl) and osmotic stress (−0.5 MPa) were 2.0–2.8 times those in plants grown in the absence of NaCl or osmotic stress. Concomitantly, the expression level of ZxSKOR in roots under osmotic stress plus salt (−0.5 MPa +50 mM NaCl) was significantly higher than that under osmotic stress (−0.5 MPa) alone during 12–48 h of treatment. We propose that ZxSKOR in roots and stems is well-coordinated to mediate long-distance K+ transport and perhaps plays an important role in K+ accumulation and homeostasis in Z. xanthoxylum under salt as well as drought stress.

Development of a HOG-based real-time PCR method to detect stress response changes in mycotoxigenic moulds

There is a need to understand the mechanism of adaptation of toxigenic fungal species which are able to colonise highly specialised foods such as cured meats where there is a high osmotic stress due to the presence up to 20–22% NaCl during the ripening process. A new tool able to detect changes in stress related genes would be useful to understand the ecological reasons for the ability of these species to grow in specialised niches. In this work a real-time PCR (qPCR) using SYBR Green was developed. Primers were designed from the Hog1 gene involved in osmo-adaptation in fungi. For this, conserved regions resulting from the alignment of 26 published partial sequences of such gene were used. Specificity of primers HogF2/R2 was demonstrated when amplified, producing a unique 131-bp PCR product with a Tm value of 84 °C. The qPCR method showed an efficiency of 98%, R2 value > 0.99 and a detection limit of 0.7 log Hog1 gene copies. The qPCR method to measure changes in the Hog1 gene expression in relation to growth in ionic and non-ionic stressed environments (using 10–40% NaCl and sorbitol concentrations) was found to be suitable for two mycotoxigenic species (Penicillium nordicum, P. expansum). This assay will be a valuable tool for generating relevant Hog1 expression data from different mould species in relation to different stresses in food habitats. It will also be a good tool for a better understanding of the ability of xerophilic and xerotolerant species to colonise extreme environments.

The Microbial Opsin Homolog Sop1 is involved in Sclerotinia sclerotiorum Development and Environmental Stress Response.

Microbial opsins play a crucial role in responses to various environmental signals. Here, we report that the microbial opsin homolog gene sop1 from the necrotrophic phytopathogenic fungus Sclerotinia sclerotiorum was dramatically up-regulated during infection and sclerotial development compared with the vegetative growth stage. Further, study showed that sop1 was essential for growth, sclerotial development and full virulence of S. sclerotiorum. Sop1-silenced transformants were more sensitive to high salt stress, fungicides and high osmotic stress. However, they were more tolerant to oxidative stress compared with the wild-type strain, suggesting that sop1 is involved in different stress responses and fungicide resistance, which plays a role in the environmental adaptability of S. sclerotiorum. Furthermore, a Delta blast search showed that microbial opsins are absent from the genomes of animals and most higher plants, indicating that sop1 is a potential drug target for disease control of S. sclerotiorum.

Cloning and Functional Analysis of wzy2-1 Gene in Wheat

Dehydrins (DHNs) are identified as the group II of LEA proteins and involved in plant abiotic stress tolerance. In this study, we isolated a novel Kn-type dehydrin gene from wheat cultivar Zhengyin 1, which was designated wzy2-1. The full length of wzy2-1 is 1740 bp, encoding 579 amino acids and containing nine conserved K-fragments. Sequence alignment indicated that wzy2-1 had high homology to Dhn5 gene in Hordeum vulgare. The WZY2-1 protein was predicted to be a highly-hydrophilic and disordered protein. The WZY2-1 protein was successfully expressed in E. coli strain BL21 (DE3). We found that WZY2-1 protein improved the tolerance to low or high temperature, salt and osmotic stresses in E. coli. The qRT-PCR assay indicated that the ex- pression of wzy2-1 gene was induced by low temperature, PEG, and salt stresses rather than ABA. Thus, we conclude that wzy2-1 is an ABA-independent gene.