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Drought, salinity and extreme temperatures are the most common abiotic stresses, adversely affecting plant growth and productivity. Exposure of plants to stress activates stress signalling pathways that induce biochemical and physiological changes essential for stress acclimation. Stress tolerance is governed by multiple traits, and importance of a few traits in imparting tolerance has been demonstrated. Under drought, traits linked to water mining and water conservation, water use efficiency and cellular tolerance (CT) to desiccation are considered to be relevant. In this study, an attempt has been made to improve CT in drought hardy crop, peanut (Arachis hypogaea L., cv. TMV2) by co-expressing stress-responsive transcription factors (TFs), AtDREB2A, AtHB7 and AtABF3, associated with downstream gene expression. Transgenic plants simultaneously expressing these TFs showed increased tolerance to drought, salinity and oxidative stresses compared to wild type, with an increase in total plant biomass. The transgenic plants exhibited improved membrane and chlorophyll stability due to enhanced reactive oxygen species scavenging and osmotic adjustment by proline synthesis under stress. The improvement in stress tolerance in transgenic lines were associated with induced expression of various CT related genes like AhGlutaredoxin, AhAldehyde reductase, AhSerine threonine kinase like protein, AhRbx1, AhProline amino peptidase, AhHSP70, AhDIP and AhLea4. Taken together the results indicate that co-expression of stress responsive TFs can activate multiple CT pathways, and this strategy can be employed to improve abiotic stress tolerance in crop plants.

Basic helix-loop-helix (bHLH) transcription factors constitute one of the largest families in plants and are known to be involved in various developmental processes and stress tolerance. We report the characterization of a stress responsive bHLH transcription factor from stress adapted species finger millet which is homologous to OsbHLH57 and designated as EcbHLH57. The full length sequence of EcbHLH57 consisted of 256 amino acids with a conserved bHLH domain followed by leucine repeats. In finger millet, EcbHLH57 transcripts were induced by ABA, NaCl, PEG, methyl viologen (MV) treatments and drought stress. Overexpression of EcbHLH57 in tobacco significantly increased the tolerance to salinity and drought stress with improved root growth. Transgenic plants showed higher photosynthetic rate and stomatal conductance under drought stress that resulted in higher biomass. Under long-term salinity stress, the transgenic plants accumulated higher seed weight/pod and pod number. The transgenic plants were also tolerant to oxidative stress and showed less accumulation of H202 and MDA levels. The overexpression of EcbHLH57 enhanced the expression of stress responsive genes such as LEA14, rd29A, rd29B, SOD, APX, ADH1, HSP70 and also PP2C and hence improved tolerance to diverse stresses.

Endophytes, both of bacterial and fungal origin, are ubiquitously present in all plants. While their origin and evolution are enigmatic, there is burgeoning literature on their role in promoting growth and stress responses in their hosts. We demonstrate that a salt-tolerant endophyte isolated from salt-adapted Pokkali rice, a Fusarium sp ., colonizes the salt-sensitive rice variety IR-64, promotes its growth under salt stress and confers salinity stress tolerance to its host. Physiological parameters, such as assimilation rate and chlorophyll stability index were higher in the colonized plants. Comparative transcriptome analysis revealed 1348 up-regulated and 1078 down-regulated genes in plants colonized by the endophyte. Analysis of the regulated genes by MapMan and interaction network programs showed that they are involved in both abiotic and biotic stress tolerance, and code for proteins involved in signal perception (leucine-rich repeat proteins, receptor-like kinases) and transduction (Ca 2+ and calmodulin-binding proteins), transcription factors, secondary metabolism and oxidative stress scavenging. For nine genes, the data were validated by qPCR analysis in both roots and shoots. Taken together, these results show that salt-adapted Pokkali rice varieties are powerful sources for the identification of novel endophytes, which can be used to confer salinity tolerance to agriculturally important, but salt-sensitive rice varieties.

Improving mulberry leaf production with enhanced leaf quality holds the key to sustain the ever increasing demand for silk. Adoption of modern genomic approaches for crop improvement is severely constrained by the lack of sufficient molecular markers in mulberry. Here, we report development and validation of 206 EST derived SSR markers using transcriptome data generated from leaf tissue of a drought tolerant mulberry genotype, Dudia white. Analysis of transcriptome data containing 10169 EST sequences, revealed 1469 sequences with microsatellite repeat motifs. We designed a total of 264 primers to the most appropriate repeat regions, of which 206 were locus specific. These markers were validated with 25 diverse mulberry accessions and their transferability to closely related species belonging to family Moraceae was examined. Of these markers, 189 revealed polymorphism with up to 8 allelic forms across mulberry species, genotypes and varieties with a mean of 3.5 alleles per locus. The markers also revealed higher polymorphic information content of 0.824 among the accessions. These markers effectively segregated the species and genotypes and hence, can be used for both diversity analysis and in breeding applications. Around 40% of these markers were transferable to other closely related species. Along with the other genic and genomic markers, we report a set of over 750 co-dominant markers. Using these markers we constructed the first genetic linkage map of mulberry exclusively with co-dominant markers.

Pre-sowing seed treatment with systemic fungicides is a firmly entrenched practice for most agricultural crops worldwide. The treatment is intended to protect the crop against seed- and soil-borne diseases. In recent years, there is increasing evidence that fungicidal applications to manage diseases might inadvertently also affect non-target organisms, such as endophytes. Endophytes are ubiquitously present in plants and contribute to plant growth and development besides offering resistance to biotic and abiotic stresses. In seeds, endophytes may play a role in seed development, seed germination, seedling establishment and crop performance. In this paper, we review the recent literature on non-target effects of fungicidal applications on endophytic fungal community and discuss the possible consequences of indiscriminate seed treatment with systemic fungicide on seed endophytes. It is now well recognized that endophytes are ubiquitously present in all parts of the plant, including the seeds. They may be transmitted vertically from seed to seed as in many grasses and/or acquired horizontally from the soil and the environment. Though the origins and evolution of these organisms in plants are a matter of conjecture, numerous studies have shown that they symbiotically aid in plant growth and development, in nutrient acquisition as well in protecting the plants from abiotic and biotic stresses. Against this background, it is reasonable to assume that the use of systemic fungicides in seed treatment may not only affect the seed endophytes but also their attendant benefits to seedling growth and establishment. While there is evidence to indicate that fungicidal applications to manage plant diseases also affect foliar endophytes, there are only few studies that have documented the effect of seed treatment on seed-borne endophytes. Some of the convincing examples of the latter come from studies on the effect of fungicide application on rye grass seed endophyte AR37. More recently, experiments have shown that removal of seed endophytes by treatment with systemic fungicides leads to significant loss of seedling vigour and that such losses could be partially restored by enriching the seedlings with the lost endophytes. Put together, these studies reinforce the importance of seed endophytes to seedling growth and establishment and draw attention on how to trade the balance between the benefits of seed treatments and the direct and indirect costs incurred due to loss of endophytes. Among several approaches, use of reduced-risk fungicides and identifying fungicide-resistant endophytes are suggested to sustain the endophyte contribution to early seedling growth.

Endophytes are non-disease causing microbes (bacteria and fungi) surviving in living tissues of plants. Their intimate association and possible coevolution with their plant partners have resulted in them contributing to an array of plant growth benefits ranging from enhanced growth and biomass accumulation, tolerance to abiotic and biotic stresses and in nutrient acquisition. The last couple of decades have witnessed a burgeoning literature on the role of endophytes (Class 3 type) in regulating plant growth and development and their adaptation to abiotic and biotic stresses. Though the underlying mechanisms of plant-endophyte interactions are far from clear, several studies have raised the hope of their potential application in agriculture, especially in mitigating abiotic and biotic stresses. The use of endophytes is envisaged as a route to reduce the production cost and burden on the environment by lessening the dependence on breeding for crop improvement and agrochemicals. Unfortunately, save a few well documented examples of their use, a little of these insights has been translated into actual agricultural applications. Here, we reflect on this paucity and elaborate on some of the important bottlenecks that might stand in way of fully realizing the potential that endophytes hold for crop improvement. We stress the need to study various facets of the endophyte-plant association for their gainful application in agriculture.

Plants host numerous endophytic microbes which promote plant performance, in particular under stress. A new endophytic fungus was isolated from the leaves of a deciduous wood tree Leucas aspera. Morphological inspection and multilocus phylogeny identified the fungus as a new Trichoderma strain. If applied to Arabidopsis thaliana and Nicotiana attenuata , it mainly colonizes their roots and strongly promotes initial growth of the plants on soil. The fungus grows on high NaCl or mannitol concentrations, and shows predatory capability on the pathogenic fungus Alternaria brassicicola . Colonized Arabidopsis plants tolerate higher salt stress and show lower A. brassicicola spread in roots and shoots, while arbuscular mycorrhiza formation in N. attenuata is not affected by the Trichoderma strain. These beneficial features of the novel Trichoderma strain are important prerequisites for agricultural applications.

Drought transcriptome analysis of finger millet ( Eleusine coracana ) by cDNA subtraction identified drought responsive genes that have a potential role in drought tolerance. Through virus-induced gene silencing (VIGS) in a related crop species, maize ( Zea mays ), several genes, including a G-BOX BINDING FACTOR 3 ( GBF3 ) were identified as candidate drought stress response genes and the role of GBF3 in drought tolerance was studied in Arabidopsis thaliana . Overexpression of both EcGBF3 and AtGBF3 in A. thaliana resulted in improved tolerance to osmotic stress, salinity and drought stress in addition to conferring insensitivity to ABA. Conversely, loss of function of this gene increased the sensitivity of A. thaliana plants to drought stress. EcGBF3 transgenic A. thaliana results also suggest that drought tolerance of sensitive plants can be improved by transferring genes from far related crops like finger millet. Our results demonstrate the role of GBF3 in imparting drought tolerance in A. thaliana and indicate the conserved role of this gene in drought and other abiotic stress tolerance in several plant species.

NAC (NAM, ATAF1-2, and CUC2) proteins constitute one of the largest families of plant-specific transcription factors and have been shown to be involved in diverse plant processes including plant growth, development, and stress-tolerance. In this study, a stress-responsive NAC gene, EcNAC1, was isolated from the subtracted stress cDNA library generated from a drought adapted crop, finger millet, and characterized for its role in stress-tolerance. The expression analysis showed that EcNAC1 was highly induced during water-deficit and salt stress. EcNAC1 shares high amino acid similarity with rice genes that have been phylogenetically classified into stress-related NAC genes. Our results demonstrated that tobacco transgenic plants expressing EcNAC1 exhibit tolerance to various abiotic stresses like simulated osmotic stress, by polyethylene glycol (PEG) and mannitol, and salinity stress. The transgenic plants also showed enhanced tolerance to methyl-viologen (MV) induced oxidative stress. Reduced levels of reactive oxygen species (ROS) and ROS-induced damage were noticed in pot grown transgenic lines under water-deficit and natural high light conditions. Root growth under stress and recovery growth after stress alleviation was more in transgenic plants. Many stress-responsive genes were found to be up-regulated in transgenic lines expressing EcNAC1. Our results suggest that EcNAC1 overexpression confers tolerance against abiotic stress in susceptible species, tobacco.

Freshwater is a limited and dwindling global resource; therefore, efficient water use is required for food crops that have high water demands, such as rice, or for the production of sustainable energy biomass. We show here that expression of the Arabidopsis HARDY (HRD) gene in rice improves water use efficiency, the ratio of biomass produced to the water used, by enhancing photosynthetic assimilation and reducing transpiration. These drought-tolerant, low-water-consuming rice plants exhibit increased shoot biomass under well irrigated conditions and an adaptive increase in root biomass under drought stress. The HRD gene, an AP2/ERF-like transcription factor, identified by a gain-of-function Arabidopsis mutant hrd-D having roots with enhanced strength, branching, and cortical cells, exhibits drought resistance and salt tolerance, accompanied by an enhancement in the expression of abiotic stress associated genes. HRD overexpression in Arabidopsis produces thicker leaves with more chloroplast-bearing mesophyll cells, and in rice, there is an increase in leaf biomass and bundle sheath cells that probably contributes to the enhanced photosynthesis assimilation and efficiency. The results exemplify application of a gene identified from the model plant Arabidopsis for the improvement of water use efficiency coincident with drought resistance in the crop plant rice.