Lived experiences of stroke survivors in India: A phenomenological study.

Background: Stroke is a major cause of long-term disability and has a potentially enormous emotional and socio-economic impact on patients, their families, and health services. Perceptions of patients with stroke have already been studied worldwide, which are unique in terms of their cultural background. However, in India, there is a lack of studies about the experience of the disease by stroke survivors and their perspectives of understanding the situation. Objective: This study aimed to explore lived experience of stroke survivors in India. Methods: A phenomenological study design was used. In-depth interviews were conducted with a purposive sample of ten stroke survivors who had experienced post-stroke deficits three months to one year after stroke. Data were analyzed using Diekelmann's hermeneutical approach to identify underlying themes. Results: Two main themes emerged: (1) emergence of stroke (actual occurrence, mental perception, and recognition of illness) and (2) therapeutic concerns (enhanced and weakened recovery). Conclusion: Recognizing how patients experience the illness is crucial in planning care for stroke survivors. Strengthening factors enhancing recovery and limiting the hindering factors through effective therapeutic management is a necessity. The findings might also contribute to refining existing interventions and designing holistic multi-component rehabilitation programs that facilitate easy recovery. The study also highlights the need for providing information to general public on recognizing warning signs of stroke.

Unraveling the mechanisms of resistance to Sclerotium rolfsii in peanut (Arachis hypogaea L.) using comparative RNA-Seq analysis of resistant and susceptible genotypes.

Stem rot, a devastating fungal disease of peanut, is caused by Sclerotium rolfsii. RNA-sequencing approaches have been used to unravel the mechanisms of resistance to stem rot in peanut over the course of fungal infection in resistant (NRCG-CS85) and susceptible (TG37A) genotypes under control conditions and during the course of infection. Out of about 290 million reads, nearly 251 million (92.22%) high-quality reads were obtained and aligned to the Arachis duranensis and Arachis ipaensis genomes with the average mapping of 78.91% and 78.61%, respectively. In total, about 48.6% of genes were commonly regulated, while approximately 21.8% and 29.6% of uniquely regulated genes from A. duranensis and A. ipaensis genomes, respectively, were identified. Several annotated transcripts, such as receptor-like kinases, jasmonic acid pathway enzymes, and transcription factors (TFs), including WRKY, Zinc finger protein, and C2-H2 zinc finger, showed higher expression in resistant genotypes upon infection. These transcripts have a known role in channelizing the downstream of pathogen perception. The higher expression of WRKY transcripts might have induced the systemic acquired resistance (SAR) by the activation of the jasmonic acid defense signaling pathway. Furthermore, a set of 30 transcripts involved in the defense mechanisms were validated with quantitative real-time PCR. This study suggested PAMP-triggered immunity as a probable mechanism of resistance, while the jasmonic acid signaling pathway was identified as a possible defense mechanism in peanut. The information generated is of immense importance in developing more effective ways to combat the stem rot disease in peanut.

Advances in the development and use of DREB for improved abiotic stress tolerance in transgenic crop plants.

Abiotic stresses negatively influence the survival, biomass production, and yield of crops. Tolerance to diverse abiotic stresses in plants is regulated by multiple genes responding differently to various stress conditions. Genetic engineering approaches have helped develop transgenic crops with improved abiotic stress tolerance including yields. The dehydration-responsive element binding protein (DREB) is a stress-responsive transcription factor that modulates the expression of downstream stress-inducible genes, which confer simultaneous tolerance to multiple stresses. This review focuses on advances in the development of DREB transgenic crops and their characterization under various abiotic stress conditions. It further discusses the mechanistic aspects of abiotic stress tolerance, yield gain, the fate of transgenic plants under controlled and field conditions and future research directions toward commercialization of DREB transgenic crops.

Regulation of antioxidant mechanisms by AtDREB1A improves soil-moisture deficit stress tolerance in transgenic peanut (Arachis hypogaea L.).

The present study evaluated the soil-moisture deficit stress tolerance of AtDREB1A transgenic peanut lines during reproductive stages using lysimetric system under controlled glasshouse conditions. The antioxidant activities of AtDREB1A transgenic lines were measured by biochemical assays. The transgenic peanut lines recorded significantly lower accumulation of malondialdehyde and hydrogen peroxide than the wild-type. Whereas, specific activity of catalase, guaiacol peroxidase, ascorbate peroxidase, glutathione reductase and ascorbic acid were found to be significantly higher in transgenic lines than in the wild-type line under drought stress. The results showed that the transgenic lines expressed lower oxidative damage than wild-type and could protect themselves from the elevated levels of reactive oxygen species under drought stress. This could be attributed to the regulation of various stress-inducible genes by AtDREB1A transcription factor. Improved photosynthetic and growth parameters were also recorded in transgenic lines over wild-type under drought stress. Improved physio-biochemical mechanisms in transgenic peanut lines might have resulted in improved growth-related traits as significant correlations were observed between physio-biochemical parameters and growth-related traits under drought stress. The potential target genes of AtDREB1A transcription factor in transgenic peanut lines during drought stress were identified, which helped in understanding the molecular mechanisms of DREB-regulated stress responses. The transgenic line D6 reported the best physio-biochemical mechanisms and growth-related parameters under drought stress over other transgenic lines and wild-type, suggesting it may be used to develop high yielding and terminal drought-tolerant peanut varieties.

Genotyping-by-sequencing based genetic mapping reveals large number of epistatic interactions for stem rot resistance in groundnut.

KEY MESSAGE: Genetic mapping identified large number of epistatic interactions indicating the complex genetic architecture for stem rot disease resistance. Groundnut (Arachis hypogaea) is an important global crop commodity and serves as a major source of cooking oil, diverse confectionery preparations and livestock feed. Stem rot disease caused by Sclerotium rolfsii is the most devastating disease of groundnut and can cause up to 100% yield loss. Genomic-assisted breeding (GAB) has potential for accelerated development of stem rot resistance varieties in short period with more precision. In this context, linkage analysis and quantitative trait locus (QTL) mapping for resistance to stem rot disease was performed in a bi-parental recombinant inbred line population developed from TG37A (susceptible) × NRCG-CS85 (resistant) comprising of 270 individuals. Genotyping-by-sequencing approach was deployed to generate single nucleotide polymorphism (SNP) genotyping data leading to development of a genetic map with 585 SNP loci spanning map distance of 2430 cM. QTL analysis using multi-season phenotyping and genotyping data could not detect any major main-effect QTL but identified 44 major epistatic QTLs with phenotypic variation explained ranging from 14.32 to 67.95%. Large number interactions indicate the complexity of genetic architecture of resistance to stem rot disease. A QTL of physical map length 5.2 Mb identified on B04 comprising 170 different genes especially leucine reach repeats, zinc finger motifs and ethyleneresponsive factors, etc., was identified. The identified genomic regions and candidate genes will further validate and facilitate marker development to deploy GAB for developing stem rot disease resistance groundnut varieties.

Transgenic Peanut (Arachis hypogaea L.) Overexpressing mtlD Gene Showed Improved Photosynthetic, Physio-Biochemical, and Yield-Parameters under Soil-Moisture Deficit Stress in Lysimeter System.

Peanut, an important oilseed crop, frequently encounters drought stress (DS) during its life cycle. In this study, four previously developed mtlD transgenic (T) peanut lines were used for detailed characterization under DS, at the reproductive stage using lysimeter system under controlled greenhouse conditions. In dry-down experiments, T lines maintained better photosynthetic machinery, such as, photosynthesis rate, stomatal conductance, transpiration rate, and SPAD (Soil-Plant Analyses Development) values, and had lower oxidative damage, including lipid membrane peroxidation and hydrogen peroxide and superoxide radical accumulation than WT, when exposed to 24 days of DS. WT plants had a more negative water potential (WP; up to -3.22 MPa) than T lines did (-2.56 to -2.71 MPa) at day 24 of DS treatment. During recovery, T lines recovered easily whereas 67% of WT plants failed to recover. In T lines, the rate of photosynthesis strongly and positively correlated with the transpiration rate (r = 0.92), RWC (r = 0.90), WP (r = 0.86), and total chlorophyll content (r = 0.75), suggesting its strong correlation with water retention-related parameters. Furthermore, yield parameters such as, pod weight and harvest index of T lines were up to 2.19 and 1.38 times more than those of WT plants, respectively. Thus, the significantly better performance of mtlD T peanut lines than of WT plants under DS could be attributed to the accumulation of mannitol, which in turn helped in maintaining the osmoregulation and ROS scavenging activity of mannitol and ultimately conferred water-economizing capacity and higher yield in T lines than in WT plants.

Insights into the Indian Peanut Genotypes for ahFAD2 Gene Polymorphism Regulating Its Oleic and Linoleic Acid Fluxes.

In peanut (Arachis hypogaea L.), the customization of fatty acid profile is an evolving area to fulfill the nutritional needs in the modern market. A total of 174 peanut genotypes, including 167 Indian cultivars, 6 advanced breeding lines and "SunOleic95R"-a double mutant line, were investigated using AS-PCRs, CAPS and gene sequencing for the ahFAD2 allele polymorphism, along with its fatty acid compositions. Of these, 80 genotypes were found having substitution (448G>A) mutation only in ahFAD2A gene, while none recorded 1-bp insertion (441_442insA) mutation in ahFAD2B gene. Moreover, 22 wild peanut accessions found lacking both the mutations. Among botanical types, the ahFAD2A mutation was more frequent in ssp. hypogaea (89%) than in ssp. fastigiata (17%). This single allele mutation, found affecting not only oleic to linoleic acid fluxes, but also the composition of other fatty acids in the genotypes studied. Repeated use of a few selected genotypes in the Indian varietal development programs were also eminently reflected in its ahFAD2 allele polymorphism. Absence of known mutations in the wild-relatives indicated the possible origin of these mutations, after the allotetraploidization of cultivated peanut. The SNP analysis of both ahFAD2A and ahFAD2B genes, revealed haplotype diversity of 1.05% and 0.95%, while Ka/Ks ratio of 0.36 and 0.39, respectively, indicating strong purifying selection pressure on these genes. Cluster analysis, using ahFAD2 gene SNPs, showed presence of both mutant and non-mutant genotypes in the same cluster, which might be due the presence of ahFAD2 gene families. This investigation provided insights into the large number of Indian peanut genotypes, covering various aspects related to O/L flux regulation and ahFAD2 gene polymorphism.

Stress Inducible Expression of AtDREB1A Transcription Factor in Transgenic Peanut (Arachis hypogaea L.) Conferred Tolerance to Soil-Moisture Deficit Stress.

Peanut, an important oilseed crop, is gaining priority for the development of drought tolerant genotypes in recent times, since the area under drought is constantly on the rise. To achieve this, one of the important strategies is to genetically engineer the ruling peanut varieties using transcription factor regulating the expression of several downstream, abiotic-stress responsive gene(s). In this study, eight independent transgenic peanut (cv. GG20) lines were developed using AtDREB1A gene, encoding for a transcription factor, through Agrobacterium-mediated genetic transformation. The transgene insertion was confirmed in (T0) using PCR and Dot-blot analysis, while copy-number(s) was ascertained using Southern-blot analysis. The inheritance of AtDREB1A gene in individual transgenic plants (T1 and T2) was confirmed using PCR. In homozygous transgenic plants (T2), under soil-moisture deficit stress, elevated level of AtDREB1A transgene expression was observed by RT-PCR assay. The transgenic plants at 45-d or reproductive growth stage showed tolerance to severe soil-moisture deficit stress. Physio-biochemical parameters such as proline content, osmotic potential, relative water content, electrolytic leakage, and total-chlorophyll content were found positively correlated with growth-related traits without any morphological abnormality, when compared to wild-type. qPCR analysis revealed consistent increase in expression of AtDREB1A gene under progressive soil-moisture deficit stress in two homozygous transgenic plants. The transgene expression showed significant correlation with improved physio-biochemical traits. The improvement of drought-stress tolerance in combination with improved growth-related traits is very essential criterion for a premium peanut cultivar like GG20, so that marginal farmers of India can incur the economic benefits during seasonal drought and water scarcity.

Novel and Stress Relevant EST Derived SSR Markers Developed and Validated in Peanut.

With the aim to increase the number of functional markers in resource poor crop like cultivated peanut (Arachis hypogaea), large numbers of available expressed sequence tags (ESTs) in the public databases, were employed for the development of novel EST derived simple sequence repeat (SSR) markers. From 16424 unigenes, 2784 (16.95%) SSRs containing unigenes having 3373 SSR motifs were identified. Of these, 2027 (72.81%) sequences were annotated and 4124 gene ontology terms were assigned. Among different SSR motif-classes, tri-nucleotide repeats (33.86%) were the most abundant followed by di-nucleotide repeats (27.51%) while AG/CT (20.7%) and AAG/CTT (13.25%) were the most abundant repeat-motifs. A total of 2456 EST-SSR novel primer pairs were designed, of which 366 unigenes having relevance to various stresses and other functions, were PCR validated using a set of 11 diverse peanut genotypes. Of these, 340 (92.62%) primer pairs yielded clear and scorable PCR products and 39 (10.66%) primer pairs exhibited polymorphisms. Overall, the number of alleles per marker ranged from 1-12 with an average of 3.77 and the PIC ranged from 0.028 to 0.375 with an average of 0.325. The identified EST-SSRs not only enriched the existing molecular markers kitty, but would also facilitate the targeted research in marker-trait association for various stresses, inter-specific studies and genetic diversity analysis in peanut.

Morphological and toxigenic variability in the Aspergillus flavus isolates from peanut (Arachis hypogaea L.) production system in Gujarat (India).

Morphological and toxigenic variability in 187 Aspergillus flavus isolates, collected from a major Indian peanut production system, from 10 districts of Gujarat was studied. On the basis of colony characteristics, the isolates were grouped as group A (83%), B (11%) and G (6%). Of all the isolates, 21%, 47% and 32% were found to be fast-growing, moderately-fast and slow-growing respectively, and nosclerotia and sclerotia production was recorded in 32.1% and 67% isolates respectively. Large, medium and small number of sclerotia production was observed in 55, 38 and 34 isolates respectively. Toxigenic potential based on ammonia vapour test was not found reliable, while ELISA test identified 68.5%, 18.7% and 12.8% isolates as atoxigenic, moderately-toxigenic and highly-toxigenic, respectively. On clustering, the isolates were grouped into 15 distinct clusters, 'A' group of isolates was grouped distinctly in different clusters, while 'B' and 'G' groups of isolates were clustered together. No association was observed between morphological-diversity and toxigenic potential of the isolates. From the present investigation, most virulent isolates were pooled to form a consortium for sick-plot screening of germplasm, against Aspergillus flavus. In future, atoxigenic isolates may be evaluated for their potential to be used as bio-control agent against toxigenicisolates.

Heterologous expression of the AtDREB1A gene in transgenic peanut-conferred tolerance to drought and salinity stresses.

Research on genetic transformation in various crop plants using the DREB1A transcription factor has shown better abiotic stress tolerance in transgenic crops. The AtDREB1A transgenic peanut (Arachis hypogaea L. cv. GG 20), which was previously developed, was characterized in terms of its physio-biochemical, molecular and growth parameters. The tolerance of this transgenic peanut to drought and salinity stresses was evaluated at the seedling (18 days old) and maturity stages. Transgenic peanut lines showed improved tolerance to both stresses over wild-type, as observed by delayed and less severe wilting of leaves and by improved growth parameters that were correlated with physio-biochemical parameters such as proline content, total chlorophyll content, osmotic potential, electrolytic leakage and relative water content. The expression pattern of the AtDREB1A gene evaluated using qPCR at different time points demonstrated that transgene expression was induced within two hours of stress imposition. The better performance of transgenic AtDREB1A peanut at the seedling stage and the improved growth parameters were due to the expression of the transgene, which is a transcription factor, and the possible up-regulation of various stress-inducible, downstream genes in the signal transduction pathway under abiotic stress.