Sequence similarity based identification of abiotic stress responsive genes in chickpea

Chickpea (Cicer arietinum L.) is an important food legume crop, particularly for the arid regions including Indian subcontinent. Considering the detrimental effect of drought, temperature and salt stress on crop yield, efforts have been initiated in the direction of developing improved varieties and designing alternate strategies to sustain chickpea production in adverse environmental conditions. Identification of genes that confer abiotic stress tolerance in plants remains a challenge in contemporary plant breeding. The present study focused on the identification of abiotic stress responsive genes in chickpea based on sequence similarity approach exploiting known abiotic stress responsive genes from model crops or other plant species. Ten abiotic stress responsive genes identified in other plants were partially amplified from eight chickpea genotypes and their presence in chickpea was confirmed after sequencing the PCR products. These genes have been functionally validated and reported to play significant role in stress response in model plants like Arabidopsis, rice and other legume crops. Chickpea EST sequences available at NCBI EST database were used for the identification of abiotic stress responsive genes. A total of 8,536 unique coding long sequences were used for identification of chickpea homologues of these abiotic stress responsive genes by sequence similarity search (BLASTN and BLASTX). These genes can be further explored towards achieving the goal of developing superior chickpea varieties providing improved yields under stress conditions using modern molecular breeding approaches

Studies on Diagnosis and Pathogenesis of Equine Herpesviruses-1 and -4 by Polymerase Chain Reaction

The work presented in this thesis focuses on the development of polymerase chain reaction (PCR) assays for diagnosis and differentiation of equid herpes viruses-1 and -4 (EHV-1 and EHV-4) infections in suspected field cases and contact animals and in experimentally infected specific-pathogen-free (SPF) foals. The technique has been applied to study the pathogenesis of EHV-1 and EHV-4 infections in these foals. Further, the effect of homologous (EHV-1:EHV-1) challenge on nasal virus excretion and duration of viremia has been studied. Studies on heterologous (EHV-4:EHV-1) challenge have also been conducted with one objective being to determine whether challenge virus could reactivate virus inoculated to produce primary infection (reactivation of EHV-4 by EHV-1 in this instance). Following secondary challenge (homologous or heterologous), dexamethasone administration was performed in order to reactivate latent virus(es). The chapterwise details of the thesis are as follows: Chapter 1 describes the general features the family Herpesviridae structure of herpesvirion, genomes of herpesviruses, classification of herpesviruses, their replicative cycles and latency. A brief account of various equid herpesviruses (equid herpesviruses-1 to -8) is given and the literature on various aspects of EHV-1 and EHV-4 has been reviewed comprehensively. The general materials and methods used throughout the course of study are described in Chapter 2. Chapter 3 gives an account of polymerase chain reaction (PCR) and its application in various fields of science. The EHV-1/EHV-4 PCR assay is detailed. Type common primers derived from the conserved regions of homologues of HSV genes encoding glycoproteins gH and gC and a nested set comprising a primer derived from thymidine kinase (tk) gene and three from the gH gene have been used to amplify specific regions of EHV-1 and EHV-4. The targets for amplification in the PCR assays were; tissue culture supernatants from EHV-1 and EHV-4 infected cultures and recombinant plasmic DNAs containing EHV-1 and EHV-4 target. The amplified products could be distinguished by type-specific probes selected from the divergent regions of the genes and internal to the primers. Reaction conditions were optimised and assay sensitivity assessed. Chapter 4 describes the use of PCR techniques to the detection of EHV-1 or EHV-4- specific DNA in nasopharyngeal swab samples from suspected field cases and their in-contacts. The primers used in the PCR assay were: inner gH, nested set and gC. The assays were conducted blind and later decoded and compared with virus isolation data. Of 98 samples, 91 of which were EHV-1 and EHV-4-negative and 7 positive, all 7 positives were confirmed by PCR and 5 virus-negative were PCR-positive. The results indicated that PCR is a sensitive and rapid technique for the diagnosis of EHV-1 and EHV-4 infections. The studies on the experimental infections of specific-pathogen-free (SPF) foals are described in Chapter 5. The experiment was conducted in three phases: primary infection, secondary infection/challenge and reactivation. Two groups of two SPF foals were infected by inoculating 107 pfu of EHV-1 strain Ab4 or EHV-4 strain MD intranasally. Nasal secretions and peripheral blood leucocytes (PBMCs) were collected at days -2, 1, 3, 5, 8, 11 and 18 post-infection (p.i.) and analysed by PCR. Results were compared with the virus isolation and co-cultivation data. The latter work was carried out by research group at Cambridge led by Dr Hugh Field. Rectal temperatures as high as 106F were recorded in EHV-1 infected foals and classical signs of EHV-1 induced disease were observed. The EHV-4 infected foals exhibited a mild disease. EHV-1 specific DNA was detected in the nasal secretions of the EHV-1 infected foals til day 18 post-infection, the day of last sampling. In the PBMCs of EHV-1 infected foals, EHV-1 specific DNA was detectable till day 11 p.i. At the peak of infection at day 5 in one foal and at day 8 in another foal. 5 x 10+ and 5 x 104 PBMCs respectively gave a positive signal in PCR assay. Nasal virus excretion by virus isolation was demonstrable till day 11 p.i. in both the EHV-1 infected foals. EHV-1 viraemia was observed by infectious centre assay at days 3, 5 and 11 in one foal and at day 11 post infection in another. The EHV-4-specific DNA in EHV-4 infected group was detectable till day 18 and 15 respectively in the nasal secretions of the two foals. EHV-4 specific DNA was detectable till day 15 p.i.; the infection was mild throughout as 5 x 105 cells gave a positive signal in PCR assay. In another foal, EHV-4 specific DNA was detectable till day 11 p.i. in the PBMCs, the peak of infection observed at day 11 when 5 x 104 cells gave a positive signal. No infectious centre was demonstrated in co-cultivation studies of leucocytes of EHV-4 infected foals indicating thereby that the assay is less sensitive as compared to PCR. Our studies provide definitive evidence of EHV-1 and EHV-4 specific DNA sequences in PBMCs of foals infected under specific pathogen-free conditions, following primary infection with EHV-1 and EHV-4. (Abstract shortened by ProQuest.)

Behavioral problems in children with steroid-sensitive nephrotic syndrome: An ignored aspect!

Background: Nephrotic syndrome (NS) is a common chronic renal disorder in children. Oral glucocorticoids are required in high doses for prolonged periods adding to various comorbidities including psychosocial issues and behavioral problems. Objectives: This study was done to analyze the behavioral profile of children with steroid-sensitive NS (SSNS), also its association with other factors such as mothers’ education, socioeconomic status, and age distribution. Materials and Methods: This was a questionnaire (child behavior checklist)-based cross-sectional study conducted at a tertiary care center. Children from 6 to 15 years with diagnosis of SSNS for the duration of 1 year or more were included in the study. Cases were divided into two groups: Infrequent and frequent relapsing NS (IFRNS)/steroid-dependent NS (SDNS). Equal number of children in 6–15 years age group attending outpatient clinic was taken as controls. Results: Children with SSNS (n=161) had behavioral problems in significant percentage (39.13%) compared to the control group (8.75%). It was observed more in boys (73.02%) who had more hyperactive, aggressive behavior, and externalizing problems. Internalizing problems (anxious/depressed, withdrawn/depressed, and somatic complaints) were significantly more in the study group (17.39%) as compared to none in the control group. Children with FRNS/SDNS course had more significant problems compared to relapse IFRNS group. Age distribution, socioeconomic status, and mother’s education had no significant effect on its occurrence. Conclusion: Significant behavioral problems are observed in children with SSNS and are related to the type of SSNS

MicroRNAs: potential targets for developing stress-tolerant crops

Crop yield is challenged every year worldwide by changing climatic conditions. The forecasted climatic scenario urgently demands stress-tolerant crop varieties to feed the ever-increasing global population. Molecular breeding and genetic engineering approaches have been frequently exploited for developing crops with desired agronomic traits. Recently, microRNAs (miRNAs) have emerged as powerful molecules, which potentially serve as expression markers during stress conditions. The miRNAs are small non-coding endogenous RNAs, usually 20–24 nucleotides long, which mediate post-transcriptional gene silencing and fine-tune the regulation of many abiotic- and biotic-stress responsive genes in plants. The miRNAs usually function by specifically pairing with the target mRNAs, inducing their cleavage or repressing their translation. This review focuses on the exploration of the functional role of miRNAs in regulating plant responses to abiotic and biotic stresses. Moreover, a methodology is also discussed to mine stress-responsive miRNAs from the enormous amount of transcriptome data available in the public domain generated using next-generation sequencing (NGS). Considering the functional role of miRNAs in mediating stress responses, these molecules may be explored as novel targets for engineering stress-tolerant crop varieties

Expressed sequence tags (ESTs)?based computational identification of novel and conserved microRNAs in turmeric (Curcuma longa L.)

MicroRNAs (miRNAs) are endogenous, small, single stranded non‒coding 17‒24 nucleotide long RNAs that regulates gene expression in plants and animals either by direct degradation of mRNA or inhibition of translation. Conventional approaches employed for the detection of miRNA are costly and time consuming. On the other hand, comparative genomics assisted with modern tools of bioinformatics provide an efficient and cost‒effective identification of novel and conserved miRNAs via homology search within known miRNAs. The present study reports computational identification of miRNAs present in the Expressed Sequence Tags (ESTs) of turmeric (Curcuma longa L.), a plant known for its great medicinal and culinary value. A total of 12678 ESTs were assembled into 2710 contigs/unigenes and homology sequence search was performed against 4647 non‒redundant miRNAs of Viridiplantae. In total, 102 potential mature miRNAs showed homology with 51 contigs/unigenes. Subsequently, four novel miRNAs were identified in turmeric, which satisfied the potential miRNA criteria. These four miRNAs were further characterized for target gene prediction identifying 10 target genes with their putative functions. Our study provides a valuable resource on miRNAs in turmeric, and also suggests a methodology useful for the identification of novel and conserved miRNA in non‒model but important organisms, whose EST and genome sequence data are available in the public domain. Keywords: turmeric;, Curcuma longa, microRNA, expressed sequence tag

Unusual premonsoon eddy and Kelvin wave activities in the Bay of Bengal during Indian Summer monsoon deficit in June 2009 and 2012

An investigation of the eddy and coastal Kelvin wave activities in the Bay of Bengal (BoB) is carried out during premonsoon season in two years of Indian summer monsoon deficit in June (2009 and 2012), occurred in the recent warming hiatus period. Using altimeter observations, our study reveals that over the northern BoB cyclonic eddy kinetic energy is reduced by 35% and 50% from the climatology during premonsoon seasons in 2009 and 2012, respectively, while the cyclonic eddy area is reduced by 18% and 24%, respectively. A concurrent reduction is observed in the first upwelling Kelvin wave (uKW) activities in the eastern equatorial Indian Ocean as well as in the coastal BoB for these years. The reduction in the generation of the first uKW in the eastern equatorial Indian Ocean is attributed to the westerly wind anomalies in January-March of these years. Additionally, meridional wind stress anomalies during March-April in these years are found to be southerly, causing anomalous coastal downwelling in the eastern rim of BoB. This coastal downwelling blocks the propagation of the first uKW. The decrease in the first uKW activities in the coastal waveguide of the BoB reduces the radiation of upwelling Rossby waves, thereby decreasing the cyclonic eddy activities in the northern BoB. The results from this letter could be helpful for further understanding of upper ocean mixing processes in the BoB during monsoon deficit years

Unusual Premonsoon Eddy and Kelvin Wave Activities in the Bay of Bengal During Indian Summer Monsoon Deficit in June 2009 and 2012

An investigation of the eddy and coastal Kelvin wave activities in the Bay of Bengal (BoB) is carried out during premonsoon season in two years of Indian summer monsoon deficit in June (2009 and 2012), occurred in the recent warming hiatus period. Using altimeter observations, our study reveals that over the northern BoB cyclonic eddy kinetic energy is reduced by 35% and 50% from the climatology during premonsoon seasons in 2009 and 2012, respectively, while the cyclonic eddy area is reduced by 18% and 24%, respectively. A concurrent reduction is observed in the first upwelling Kelvin wave (uKW) activities in the eastern equatorial Indian Ocean as well as in the coastal BoB for these years. The reduction in the generation of the first uKW in the eastern equatorial Indian Ocean is attributed to the westerly wind anomalies in January-March of these years. Additionally, meridional wind stress anomalies during March-April in these years are found to be southerly, causing anomalous coastal downwelling in the eastern rim of BoB. This coastal downwelling blocks the propagation of the first uKW. The decrease in the first uKW activities in the coastal waveguide of the BoB reduces the radiation of upwelling Rossby waves, thereby decreasing the cyclonic eddy activities in the northern BoB. The results from this letter could be helpful for further understanding of upper ocean mixing processes in the BoB during monsoon deficit years

The dominant Anopheles vectors of human malaria in the Asia-Pacific region: occurrence data, distribution maps and bionomic précis

<p>Abstract</p> <p>Background</p> <p>The final article in a series of three publications examining the global distribution of 41 dominant vector species (DVS) of malaria is presented here. The first publication examined the DVS from the Americas, with the second covering those species present in Africa, Europe and the Middle East. Here we discuss the 19 DVS of the Asian-Pacific region. This region experiences a high diversity of vector species, many occurring sympatrically, which, combined with the occurrence of a high number of species complexes and suspected species complexes, and behavioural plasticity of many of these major vectors, adds a level of entomological complexity not comparable elsewhere globally. To try and untangle the intricacy of the vectors of this region and to increase the effectiveness of vector control interventions, an understanding of the contemporary distribution of each species, combined with a synthesis of the current knowledge of their behaviour and ecology is needed.</p> <p>Results</p> <p>Expert opinion (EO) range maps, created with the most up-to-date expert knowledge of each DVS distribution, were combined with a contemporary database of occurrence data and a suite of open access, environmental and climatic variables. Using the Boosted Regression Tree (BRT) modelling method, distribution maps of each DVS were produced. The occurrence data were abstracted from the formal, published literature, plus other relevant sources, resulting in the collation of DVS occurrence at 10116 locations across 31 countries, of which 8853 were successfully geo-referenced and 7430 were resolved to spatial areas that could be included in the BRT model. A detailed summary of the information on the bionomics of each species and species complex is also presented.</p> <p>Conclusions</p> <p>This article concludes a project aimed to establish the contemporary global distribution of the DVS of malaria. The three articles produced are intended as a detailed reference for scientists continuing research into the aspects of taxonomy, biology and ecology relevant to species-specific vector control. This research is particularly relevant to help unravel the complicated taxonomic status, ecology and epidemiology of the vectors of the Asia-Pacific region. All the occurrence data, predictive maps and EO-shape files generated during the production of these publications will be made available in the public domain. We hope that this will encourage data sharing to improve future iterations of the distribution maps.</p