Manoj Singh

Nutraceutical Value of Finger Millet [Eleusine coracana (L.) Gaertn.], and Their Improvement Using Omics Approaches

The science of nutritional biology has progressed extensively over the last decade to develop food-based nutraceuticals as a form of highly personalized medicine or therapeutic agent. Finger millet [Eleusine coracana (L.) Gaertn.] is a crop with potentially tremendous but under-explored source of nutraceutical properties as compared to other regularly consumed cereals. In the era of growing divide and drawback of nutritional security, these characteristics must be harnessed to develop finger millet as a novel functional food. In addition, introgression of these traits into other staple crops can improve the well-being of the general population on a global scale. The objective of this review is to emphasize the importance of biofortification of finger millet in context of universal health and nutritional crisis. We have specifically highlighted the role that recent biotechnological advancements have to offer for enrichment of its nutritional value and how these developments can commission to the field of nutritional biology by opening new avenues for future research.

Identification and characterization of calcium transporter gene family in finger millet in relation to grain calcium content

Calcium (Ca) is an essential mineral for proper growth and development of plants as well as animals. In plants including cereals, calcium is deposited in seed during its development which is mediated by specialized Ca transporters. Common cereal seeds contain very low amounts of Ca while the finger millet (Eleusine coracana) contains exceptionally high amounts of Ca in seed. In order to understand the role of Ca transporters in grain Ca accumulation, developing seed transcriptome of two finger millet genotypes (GP-1, low Ca and GP-45 high Ca) differing in seed Ca content was sequenced using Illumina paired-end sequencing technology and members of Ca transporter gene family were identified. Out of 109,218 and 120,130 contigs, 86 and 81 contigs encoding Ca transporters were identified in GP-1 and GP-45, respectively. After removal of redundant sequences, a total of 19 sequences were confirmed as Ca transporter genes, which includes 11 Ca(2+) ATPases, 07 Ca(2+)/cation exchangers and 01 Ca(2+) channel. The differential expressions of all genes were analyzed from transcriptome data and it was observed that 9 and 3 genes were highly expressed in GP-45 and GP-1 genotypes respectively. Validation of transcriptome expression data of selected Ca transporter genes was performed on different stages of developing spikes of both genotypes grown under different concentrations of exogenous Ca. In both genotypes, significant correlation was observed between the expression of these genes, especially EcCaX3, and on the amount of Ca accumulated in seed. The positive correlation of seed mass with the amount of Ca concentration was also observed. The efficient Ca transport property and responsiveness of EcCAX3 towards exogenous Ca could be utilized in future biofortification program.

Effect of Japanese mint (Mentha arvensis) oil as fumigant on nutritional quality of stored sorghum

Japanese mint (Mentha arvensis) oil (JMO) can be used effectively as fumigant againstSitophilus oryzae in stored sorghum. The effect of JMO at a dose of 166 µl/l of space on nutrient composition and protein quality was studied in infested and uninfested sorghum grains stored for 3 months. The results revealed non significant effect of JMO on gran moisture, total ash, crude fibre, crude fat, crude protein and fat acidity in infested and uninfested grains at the end of 3 months storage. The JMO treatment had small but significant effect on reducing and nonreducing sugars. The values of Protein Efficiency Ratio (PER) for uninfested JMO treated grains, infested JMO treated grains and for untreated control stored for 3 months were 1.11, 1.07 and 1.09, respectively against control casein diet for which it was 2.15.

Involvement of hsr203J like gene homologue, protease and protease inhibitors in triggering differential defense response against Alternaria blight in Brassica

Transcript profiling of hsr203J, a known marker gene for Hypersensitive response (HR), was performed to delineate its role in differential defense against Alternaria brassicae in tolerant and susceptible genotypes of Brassica juncea. Reverse transcriptase (RT) PCR approach was utilized to investigate the correlation between expression of hsr203J like gene(s) and pathogenesis in stage dependent manner. It was revealed that the expression of hsr203J like gene increased as disease progressed from initial too late stage of infection in tolerant genotype. However, in susceptible genotype, its expression increased up to middle stage of infection with no expression in late stage of infection. In both genotypes, no expression of hsr203J like gene was observed in healthy leaves. It was observed that whereas three homologues of hsr203J like gene express at the late stage of infection in tolerant genotype, only single homologue of same expresses in susceptible genotype throughout all stages of infection. This indicates the role of hsr203J homologues in determining the differential defense response against Alternaria blight in Brassica. Determination of specific activity and in-gel assay revealed differential accumulation of protease and protease inhibitor in tolerant and susceptible genotypes at different stages of infection. Induction of differential protease and protease inhibitor activity appears to modulate the cell death during HR response to pathogen. Dissection of pathway leading to HR related cell death will enable us to know the molecular basis of disease resistance which will, in turn, help in engineering Brassica for resistance to Alternaria blight.

Draft genome sequence of Karnal bunt pathogen (Tilletia indica) of wheat provides insights into the pathogenic mechanisms of quarantined fungus.

Karnal bunt disease in wheat is caused by hemibiotrophic fungus, Tilletia indica that has been placed as quarantine pest in more than 70 countries. Despite its economic importance, little knowledge about the molecular components of fungal pathogenesis is known. In this study, first time the genome sequence of T. indica has been deciphered for unraveling the effectors’ functions of molecular pathogenesis of Karnal bunt disease. The T. indica genome was sequenced employing hybrid approach of PacBio Single Molecule Real Time (SMRT) and Illumina HiSEQ 2000 sequencing platforms. The genome was assembled into 10,957 contigs (N50 contig length 3 kb) with total size of 26.7 Mb and GC content of 53.99%. The number of predicted putative genes were 11,535, which were annotated with Gene Ontology databases. Functional annotation of Karnal bunt pathogen genome and classification of identified effectors into protein families revealed interesting functions related to pathogenesis. Search for effectors’ genes using pathogen host interaction database identified 135 genes. The T. indica genome sequence and putative genes involved in molecular pathogenesis would further help in devising novel and effective disease management strategies including development of resistant wheat genotypes, novel biomarkers for pathogen detection and new targets for fungicide development.

Multiplex Dipstick Technologies for Rapid and Simultaneous Screening of Analytes of Importance in Agri-Food-Nutrition and Health Care: A Review.

Dipstick test kits are being widely used for the rapid screening of a range of antigens or toxins in food, agriculture, and health care. They provide specific results on-site within 10 min with suitable accuracy and are, therefore, cost-effective. Multiplex dipsticks also provide the opportunity for simultaneous detection of multiple antigens in the target sample without using expensive instrumentation, minimizing the cost of analysis as well as the duration of assay. Because of these benefits, dipstick kits are widely being used in the simultaneous detection of several antigens/toxins in large number of samples and in high-throughput manner. This review focuses on the current status of developed multiplex strips and its working principles and future direction of the technology in the agriculture, food, nutrition, and health care sectors.

Dextrocardia and asplenia in situs inversus totalis in a baby: a case report

IntroductionSitus inversus with dextrocardia is the complete inversion of position of the thoracic and abdominal viscera. It may be isolated or associated with malformations, especially cardiac and/or alimentary. It may be discovered in infancy because of associated anomalies but often remains asymptomatic and discovered incidentally in adult life. Only a small number of cases have been reported from India.Case presentationWe report the case of a 7-month-old Indo-Aryan baby girl found to have dextrocardia with situs inversus totalis who presented with fever, cough and respiratory distress. A chest X-ray showed her heart in the right hemithorax with the cardiac apex pointing towards the right. The findings of an electrocardiogram and echocardiography confirmed the location of her heart in the right hemithorax and an abdominal sonogram showed her liver and gall bladder in midline of her abdomen whereas her stomach was located more towards the right side, her spleen was absent.ConclusionsSitus inversus totalis, although a rare condition, should be sought for when clinical and radiologic findings indicate dextrocardia, especially as it may be an incidental finding. Doctors should encourage routine medical examination for their patients which could help identify this anomaly, thereby preventing wrong diagnosis and possibly death due to delay in management.

Generation of diagnostic anti-teliospore antibodies for development of specific immunodiagnostic format for detection of Karnal bunt (Tilletia indica) of wheat

Rabbit antisera were generated against four preparations of teliospores of Tilletia indica the fungus responsible for Karnal bunt in wheat having titre ranging from 1:2500 to 1:10,000. The suitability of four solubilising agents (I–IV) used for extraction of teliospores/spore proteins was assessed, highest protein extractability was observed with solubilising agent I (0.5% sodium dodecyl sulphate). The maximum immunoreactivity in enzyme linked immunosorbent assay (ELISA) was observed with the teliosporic protein(s) extracted using solubilising agent I followed by agent II, IV and III. Further, the results of western blotting with proteins from different fungal pathogens using anti-intact teliospores antibody showed one unique immunoreactive band of 28 kDa with teliosporic protein of T. indica only. The potential diagnostic antibody was generated against the characterised protein (28 kDa) of teliospore walls antigen of T. indica. The developed diagnostic antibody and solubilising agent I could therefore be employed for development of specific and rapid immunodiagnostic format for detection of Karnal bunt.

Identification and localisation of glycosyl moieties of surface glycoproteins of teliospore wall of Karnal bunt (Tilletia indica) of wheat

Abstract This article aims to characterise and localise the glycosyl moieties of teliospore wall of Tilletia indica a quarantined fungal pathogens by biochemical and immunological approaches. Chemical enzyme modifier studies, followed by determination of structural configuration using phase contrast and SEM after periodate treatment, showed antigenic entities are glycoprotein in nature. Further characterisation using sodium dodecyl sulphate-polyacrylamide gel electrophroesis (SDS-PAGE) glycoprotein staining and western blotting using anti-teliospore antibodies showed two common proteins of molecular weight 28 and 40 kDa, which is also suggestive of glycoprotein nature of antigenic entities of teliospore wall. To study the binding patterns and localisation of glycosyl moieties on the teliospore walls, fluorescein isothiocyanate (FITC) labelled lectins [Wheat Germ Agglutinin (WGA) and Concanavilin A (Con A)] and anti-teliospore antibodies were used. The patterns of WGA and anti-teliospore antibodies binding with teliospore wall are almost similar and hence it is quite reasonable to suggest that immunodominant glycosyl entities of teliospore wall are acetylglucosamine in nature.

Determination of teliospore concentration as surface contaminants in Karnal bunt-infected seed samples of wheat using indirect ELISA

Abstract The study describes the indirect ELISA method for the determination of teliospore concentration in infected seed lots employing anti-teliospore antiserum generated against intact teliospores of Tilletia indica – a quarantined pathogen. The detection limit is as small as 5–1000 teliospores. The teliospores were extracted by washing seed either one or three times using detergent to ensure complete removal of teliospores present as surface contaminants in wheat samples with varying per cent of Karnal bunt infection. To ensure the concentration of teliospores within the sensitivity range, optimal volume of different extracted samples was determined. The ELISA reactivity was used to extrapolate the number of teliospores present in wheat samples. Forty-nine wheat samples received for certification based on per cent of infection were also subjected to indirect ELISA to determine the sensitivity of the protocol. The samples having 0.05–26% infection were having teliospore concentration in the range of 7×102 to 6.6×104/1000 seeds.