Search results for: Jitendra Panwar
Commenced in January 2007
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Edition: International
Paper Count: 41

Search results for: Jitendra Panwar

11 In vitro Regeneration of Neural Cells Using Human Umbilical Cord Derived Mesenchymal Stem Cells

Authors: Urvi Panwar, Kanchan Mishra, Kanjaksha Ghosh, ShankerLal Kothari

Abstract:

Background: Day-by-day the increasing prevalence of neurodegenerative diseases have become a global issue to manage them by medical sciences. The adult neural stem cells are rare and require an invasive and painful procedure to obtain it from central nervous system. Mesenchymal stem cell (MSCs) therapies have shown remarkable application in treatment of various cell injuries and cell loss. MSCs can be derived from various sources like adult tissues, human bone marrow, umbilical cord blood and cord tissue. MSCs have similar proliferation and differentiation capability, but the human umbilical cord-derived mesenchymal stem cells (hUCMSCs) are proved to be more beneficial with respect to cell procurement, differentiation to other cells, preservation, and transplantation. Material and method: Human umbilical cord is easily obtainable and non-controversial comparative to bone marrow and other adult tissues. The umbilical cord can be collected after delivery of baby, and its tissue can be cultured using explant culture method. Cell culture medium such as DMEMF12+10% FBS and DMEMF12+Neural growth factors (bFGF, human noggin, B27) with antibiotics (Streptomycin/Gentamycin) were used to culture and differentiate mesenchymal stem cells into neural cells, respectively. The characterisations of MSCs were done with Flow Cytometer for surface markers CD90, CD73 and CD105 and colony forming unit assay. The differentiated various neural cells will be characterised by fluorescence markers for neurons, astrocytes, and oligodendrocytes; quantitative PCR for genes Nestin and NeuroD1 and Western blotting technique for gap43 protein. Result and discussion: The high quality and number of MSCs were isolated from human umbilical cord via explant culture method. The obtained MSCs were differentiated into neural cells like neurons, astrocytes and oligodendrocytes. The differentiated neural cells can be used to treat neural injuries and neural cell loss by delivering cells by non-invasive administration via cerebrospinal fluid (CSF) or blood. Moreover, the MSCs can also be directly delivered to different injured sites where they differentiate into neural cells. Therefore, human umbilical cord is demonstrated to be an inexpensive and easily available source for MSCs. Moreover, the hUCMSCs can be a potential source for neural cell therapies and neural cell regeneration for neural cell injuries and neural cell loss. This new way of research will be helpful to treat and manage neural cell damages and neurodegenerative diseases like Alzheimer and Parkinson. Still the study has a long way to go but it is a promising approach for many neural disorders for which at present no satisfactory management is available.

Keywords: bone marrow, cell therapy, explant culture method, flow cytometer, human umbilical cord, mesenchymal stem cells, neurodegenerative diseases, neuroprotective, regeneration

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10 Fructose-Aided Cross-Linked Enzyme Aggregates of Laccase: An Insight on Its Chemical and Physical Properties

Authors: Bipasa Dey, Varsha Panwar, Tanmay Dutta

Abstract:

Laccase, a multicopper oxidase (EC 1.10.3.2) have been at the forefront as a superior industrial biocatalyst. They are versatile in terms of bestowing sustainable and ecological catalytic reactions such as polymerisation, xenobiotic degradation and bioremediation of phenolic and non-phenolic compounds. Regardless of the wide biotechnological applications, the critical limiting factors viz. reusability, retrieval, and storage stability still prevail. This can cause an impediment in their applicability. Crosslinked enzyme aggregates (CLEAs) have emerged as a promising technique that rehabilitates these essential facets, albeit at the expense of their enzymatic activity. The carrier free crosslinking method prevails over the carrier-bound immobilisation in conferring high productivity, low production cost owing to the absence of additional carrier and circumvent any non-catalytic ballast which could dilute the volumetric activity. To the best of our knowledge, the ε-amino group of lysyl residue is speculated as the best choice for forming Schiff’s base with glutaraldehyde. Despite being most preferrable, excess glutaraldehyde can bring about disproportionate and undesirable crosslinking within the catalytic site and hence could deliver undesirable catalytic losses. Moreover, the surface distribution of lysine residues in Trametes versicolor laccase is significantly less. Thus, to mitigate the adverse effect of glutaraldehyde in conjunction with scaling down the degradation or catalytic loss of the enzyme, crosslinking with inert substances like gelatine, collagen, Bovine serum albumin (BSA) or excess lysine is practiced. Analogous to these molecules, sugars have been well known as a protein stabiliser. It helps to retain the structural integrity, specifically secondary structure of the protein during aggregation by changing the solvent properties. They are comprehended to avert protein denaturation or enzyme deactivation during precipitation. We prepared crosslinked enzyme aggregates (CLEAs) of laccase from T. versicolor with the aid of sugars. The sugar CLEAs were compared with the classic BSA and glutaraldehyde laccase CLEAs concerning physico-chemical properties. The activity recovery for the fructose CLEAs were found to be ~20% higher than the non-sugar CLEA. Moreover, the 𝐾𝑐𝑎𝑡𝐾𝑚⁄ values of the CLEAs were two and three-fold higher than BSA-CLEA and GACLEA, respectively. The half-life (t1/2) deciphered by sugar-CLEA was higher than the t1/2 of GA-CLEAs and free enzyme, portraying more thermal stability. Besides, it demonstrated extraordinarily high pH stability, which was analogous to BSA-CLEA. The promising attributes of increased storage stability and recyclability (>80%) gives more edge to the sugar-CLEAs over conventional CLEAs of their corresponding free enzyme. Thus, sugar-CLEA prevails in furnishing the rudimentary properties required for a biocatalyst and holds many prospects.

Keywords: cross-linked enzyme aggregates, laccase immobilization, enzyme reusability, enzyme stability

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9 The Relationship between Risk and Capital: Evidence from Indian Commercial Banks

Authors: Seba Mohanty, Jitendra Mahakud

Abstract:

Capital ratio is one of the major indicators of the stability of the commercial banks. Pertinent to its pervasive importance, over the years the regulators, policy makers focus on the maintenance of the particular level of capital ratio to minimize the solvency and liquidation risk. In this context, it is very much important to identify the relationship between capital and risk and find out the factors which determine the capital ratios of commercial banks. The study examines the relationship between capital and risk of the commercial banks operating in India. Other bank specific variables like bank size, deposit, profitability, non-performing assets, bank liquidity, net interest margin, loan loss reserves, deposits variability and regulatory pressure are also considered for the analysis. The period of study is 1997-2015 i.e. the period of post liberalization. To identify the impact of financial crisis and implementation of Basel II on capital ratio, we have divided the whole period into two sub-periods i.e. 1997-2008 and 2008-2015. This study considers all the three types of commercial banks, i.e. public sector, the private sector and foreign banks, which have continuous data for the whole period. The main sources of data are Prowess data base maintained by centre for monitoring Indian economy (CMIE) and Reserve Bank of India publications. We use simultaneous equation model and more specifically Two Stage Least Square method to find out the relationship between capital and risk. From the econometric analysis, we find that capital and risk affect each other simultaneously, and this is consistent across the time period and across the type of banks. Moreover, regulation has a positive significant impact on the ratio of capital to risk-weighted assets, but no significant impact on the banks risk taking behaviour. Our empirical findings also suggest that size has a negative impact on capital and risk, indicating that larger banks increase their capital less than the other banks supported by the too-big-to-fail hypothesis. This study contributes to the existing body of literature by predicting a strong relationship between capital and risk in an emerging economy, where banking sector plays a majority role for financial development. Further this study may be considered as a primary study to find out the macro economic factors which affecting risk and capital in India.

Keywords: capital, commercial bank, risk, simultaneous equation model

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8 Comparison of Iodine Density Quantification through Three Material Decomposition between Philips iQon Dual Layer Spectral CT Scanner and Siemens Somatom Force Dual Source Dual Energy CT Scanner: An in vitro Study

Authors: Jitendra Pratap, Jonathan Sivyer

Abstract:

Introduction: Dual energy/Spectral CT scanning permits simultaneous acquisition of two x-ray spectra datasets and can complement radiological diagnosis by allowing tissue characterisation (e.g., uric acid vs. non-uric acid renal stones), enhancing structures (e.g. boost iodine signal to improve contrast resolution), and quantifying substances (e.g. iodine density). However, the latter showed inconsistent results between the 2 main modes of dual energy scanning (i.e. dual source vs. dual layer). Therefore, the present study aimed to determine which technology is more accurate in quantifying iodine density. Methods: Twenty vials with known concentrations of iodine solutions were made using Optiray 350 contrast media diluted in sterile water. The concentration of iodine utilised ranged from 0.1 mg/ml to 1.0mg/ml in 0.1mg/ml increments, 1.5 mg/ml to 4.5 mg/ml in 0.5mg/ml increments followed by further concentrations at 5.0 mg/ml, 7mg/ml, 10 mg/ml and 15mg/ml. The vials were scanned using Dual Energy scan mode on a Siemens Somatom Force at 80kV/Sn150kV and 100kV/Sn150kV kilovoltage pairing. The same vials were scanned using Spectral scan mode on a Philips iQon at 120kVp and 140kVp. The images were reconstructed at 5mm thickness and 5mm increment using Br40 kernel on the Siemens Force and B Filter on Philips iQon. Post-processing of the Dual Energy data was performed on vendor-specific Siemens Syngo VIA (VB40) and Philips Intellispace Portal (Ver. 12) for the Spectral data. For each vial and scan mode, the iodine concentration was measured by placing an ROI in the coronal plane. Intraclass correlation analysis was performed on both datasets. Results: The iodine concentrations were reproduced with a high degree of accuracy for Dual Layer CT scanner. Although the Dual Source images showed a greater degree of deviation in measured iodine density for all vials, the dataset acquired at 80kV/Sn150kV had a higher accuracy. Conclusion: Spectral CT scanning by the dual layer technique has higher accuracy for quantitative measurements of iodine density compared to the dual source technique.

Keywords: CT, iodine density, spectral, dual-energy

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7 Internal Financing Constraints and Corporate Investment: Evidence from Indian Manufacturing Firms

Authors: Gaurav Gupta, Jitendra Mahakud

Abstract:

This study focuses on the significance of internal financing constraints on the determination of corporate fixed investments in the case of Indian manufacturing companies. Financing constraints companies which have less internal fund or retained earnings face more transaction and borrowing costs due to imperfections in the capital market. The period of study is 1999-2000 to 2013-2014 and we consider 618 manufacturing companies for which the continuous data is available throughout the study period. The data is collected from PROWESS data base maintained by Centre for Monitoring Indian Economy Pvt. Ltd. Panel data methods like fixed effect and random effect methods are used for the analysis. The Likelihood Ratio test, Lagrange Multiplier test, and Hausman test results conclude the suitability of the fixed effect model for the estimation. The cash flow and liquidity of the company have been used as the proxies for the internal financial constraints. In accordance with various theories of corporate investments, we consider other firm specific variable like firm age, firm size, profitability, sales and leverage as the control variables in the model. From the econometric analysis, we find internal cash flow and liquidity have the significant and positive impact on the corporate investments. The variables like cost of capital, sales growth and growth opportunities are found to be significantly determining the corporate investments in India, which is consistent with the neoclassical, accelerator and Tobin’s q theory of corporate investment. To check the robustness of results, we divided the sample on the basis of cash flow and liquidity. Firms having cash flow greater than zero are put under one group, and firms with cash flow less than zero are put under another group. Also, the firms are divided on the basis of liquidity following the same approach. We find that the results are robust to both types of companies having positive and negative cash flow and liquidity. The results for other variables are also in the same line as we find for the whole sample. These findings confirm that internal financing constraints play a significant role for determination of corporate investment in India. The findings of this study have the implications for the corporate managers to focus on the projects having higher expected cash inflows to avoid the financing constraints. Apart from that, they should also maintain adequate liquidity to minimize the external financing costs.

Keywords: cash flow, corporate investment, financing constraints, panel data method

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6 The Impact of Monetary Policy on Aggregate Market Liquidity: Evidence from Indian Stock Market

Authors: Byomakesh Debata, Jitendra Mahakud

Abstract:

The recent financial crisis has been characterized by massive monetary policy interventions by the Central bank, and it has amplified the importance of liquidity for the stability of the stock market. This paper empirically elucidates the actual impact of monetary policy interventions on stock market liquidity covering all National Stock Exchange (NSE) Stocks, which have been traded continuously from 2002 to 2015. The present study employs a multivariate VAR model along with VAR-granger causality test, impulse response functions, block exogeneity test, and variance decomposition to analyze the direction as well as the magnitude of the relationship between monetary policy and market liquidity. Our analysis posits a unidirectional relationship between monetary policy (call money rate, base money growth rate) and aggregate market liquidity (traded value, turnover ratio, Amihud illiquidity ratio, turnover price impact, high-low spread). The impulse response function analysis clearly depicts the influence of monetary policy on stock liquidity for every unit innovation in monetary policy variables. Our results suggest that an expansionary monetary policy increases aggregate stock market liquidity and the reverse is documented during the tightening of monetary policy. To ascertain whether our findings are consistent across all periods, we divided the period of study as pre-crisis (2002 to 2007) and post-crisis period (2007-2015) and ran the same set of models. Interestingly, all liquidity variables are highly significant in the post-crisis period. However, the pre-crisis period has witnessed a moderate predictability of monetary policy. To check the robustness of our results we ran the same set of VAR models with different monetary policy variables and found the similar results. Unlike previous studies, we found most of the liquidity variables are significant throughout the sample period. This reveals the predictability of monetary policy on aggregate market liquidity. This study contributes to the existing body of literature by documenting a strong predictability of monetary policy on stock liquidity in an emerging economy with an order driven market making system like India. Most of the previous studies have been carried out in developing economies with quote driven or hybrid market making system and their results are ambiguous across different periods. From an eclectic sense, this study may be considered as a baseline study to further find out the macroeconomic determinants of liquidity of stocks at individual as well as aggregate level.

Keywords: market liquidity, monetary policy, order driven market, VAR, vector autoregressive model

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5 Isolation and Probiotic Characterization of Lactobacillus plantarum and Lactococcus lactis from Gut Microbiome of Rohu (Labeo rohita)

Authors: Prem Kumar, Anuj Tyagi, Harsh Panwar, Vaneet Inder Kaur

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Though aquaculture started as an occupation for poor and weak farmers for livelihood, it has now acquired the shape of one of the biggest industry to grow live protein in the form of aquatic organisms. Industrialization of the aquaculture sector has led to intensification resulting in stress on aquatic organisms and frequent disease outbreaks leading to huge economic impacts. Indiscriminate use of antibiotics as growth promoter and prophylactic agent in aquaculture has resulted in rapid emergence and spread of antibiotic resistance in bacterial pathogens. Over the past few years, use of probiotics (as an alternative of antibiotics) in aquaculture has gained attention due to their immunostimulant and growth promoting properties. It has now well known that after administration, a probiotic bacterium has to compete and establish itself against native microbiota to show its eventual beneficial properties. Due to their non-fish origin, commercial probiotics sometimes may display poor probiotic functionalities and antagonistic effects. Thus, isolation and characterization of probiotic bacteria from same fish host is very much necessary. In this study, attempts were made to isolate potent probiotic lactic acid bacteria (LAB) from intestinal microflora of rohu fish. Twenty-five experimental rohu fishes (mean weight 400 ± 20gm, mean standard length 20 ± 3cm) were used in the study to collect fish gut after dissection in a sterile condition. A total of 150 tentative LAB isolates from selective agar media (de Man-Rogosa-Sharpe (MRS)) were screened for their antimicrobial activity against Aeromonas hydrophila and Microccocus leuteus. A total of 17 isolates, identified as Lactobacillus plantarum and Lactococcus lactis, identified by biochemical tests and PCR amplification and sequencing of 16S rRNA gene fragment, displayed promising antimicrobial activity against both the pathogens. Two isolates from each species (FLB1, FLB2 from L. plantarum; and FLC1, FLC2 from L. lactis) were subjected to downstream probiotic potential characterization. These isolates were compared in vitro for their hemolytic activity, acid and bile tolerance for growth kinetics, auto-aggregation, cell-surface hydrophobicity against xylene, and chloroform, tolerance to phenol, cell adhesion, and safety parameters (by intraperitoneal and intramuscular injections). None of the tested isolates showed any hemolytic activity indicating their potential safety. Moreover, these isolates were tolerant to 0.3% bile (75-82% survival), phenol stress (96-99% survival) with 100% viability at pH 3 over a period of 3 h. Antibiotic sensitivity test revealed that all the tested LAB isolates were resistant to vancomycin, gentamicin, streptomycin, and erythromycin and sensitive to Erythromycin, Chloramphenicol, Ampicillin, Trimethoprim, and Nitrofurantoin. Tetracycline resistance was found in L. plantarum (FLB1 and FLB2 isolates), whereas L. lactis were susceptible to it. Intramuscular and intraperitoneal challenges to fingerlings of rohu fish (5 ± 1gm weight) with FLB1 showed no pathogenicity and occurrence of disease symptoms in fishes over an observation period of 7 days. The results revealed FLB1 as a potential probiotic candidate for aquaculture application among other isolates.

Keywords: aquaculture, Lactobacillus plantarum, Lactococcus lactis, probiotics

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4 Impact of Financial Factors on Total Factor Productivity: Evidence from Indian Manufacturing Sector

Authors: Lopamudra D. Satpathy, Bani Chatterjee, Jitendra Mahakud

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The rapid economic growth in terms of output and investment necessitates a substantial growth of Total Factor Productivity (TFP) of firms which is an indicator of an economy’s technological change. The strong empirical relationship between financial sector development and economic growth clearly indicates that firms financing decisions do affect their levels of output via their investment decisions. Hence it establishes a linkage between the financial factors and productivity growth of the firms. To achieve the smooth and continuous economic growth over time, it is imperative to understand the financial channel that serves as one of the vital channels. The theoretical or logical argument behind this linkage is that when the internal financial capital is not sufficient enough for the investment, the firms always rely upon the external sources of finance. But due to the frictions and existence of information asymmetric behavior, it is always costlier for the firms to raise the external capital from the market, which in turn affect their investment sentiment and productivity. This kind of financial position of the firms puts heavy pressure on their productive activities. Keeping in view this theoretical background, the present study has tried to analyze the role of both external and internal financial factors (leverage, cash flow and liquidity) on the determination of total factor productivity of the firms of manufacturing industry and its sub-industries, maintaining a set of firm specific variables as control variables (size, age and disembodied technological intensity). An estimate of total factor productivity of the Indian manufacturing industry and sub-industries is computed using a semi-parametric approach, i.e., Levinsohn- Petrin method. It establishes the relationship between financial factors and productivity growth of 652 firms using a dynamic panel GMM method covering the time period between 1997-98 and 2012-13. From the econometric analyses, it has been found that the internal cash flow has a positive and significant impact on the productivity of overall manufacturing sector. The other financial factors like leverage and liquidity also play the significant role in the determination of total factor productivity of the Indian manufacturing sector. The significant role of internal cash flow on determination of firm-level productivity suggests that access to external finance is not available to Indian companies easily. Further, the negative impact of leverage on productivity could be due to the less developed bond market in India. These findings have certain implications for the policy makers to take various policy reforms to develop the external bond market and easily workout through which the financially constrained companies will be able to raise the financial capital in a cost-effective manner and would be able to influence their investments in the highly productive activities, which would help for the acceleration of economic growth.

Keywords: dynamic panel, financial factors, manufacturing sector, total factor productivity

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3 Fischer Tropsch Synthesis in Compressed Carbon Dioxide with Integrated Recycle

Authors: Kanchan Mondal, Adam Sims, Madhav Soti, Jitendra Gautam, David Carron

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Fischer-Tropsch (FT) synthesis is a complex series of heterogeneous reactions between CO and H2 molecules (present in the syngas) on the surface of an active catalyst (Co, Fe, Ru, Ni, etc.) to produce gaseous, liquid, and waxy hydrocarbons. This product is composed of paraffins, olefins, and oxygenated compounds. The key challenge in applying the Fischer-Tropsch process to produce transportation fuels is to make the capital and production costs economically feasible relative to the comparative cost of existing petroleum resources. To meet this challenge, it is imperative to enhance the CO conversion while maximizing carbon selectivity towards the desired liquid hydrocarbon ranges (i.e. reduction in CH4 and CO2 selectivities) at high throughputs. At the same time, it is equally essential to increase the catalyst robustness and longevity without sacrificing catalyst activity. This paper focuses on process development to achieve the above. The paper describes the influence of operating parameters on Fischer Tropsch synthesis (FTS) from coal derived syngas in supercritical carbon dioxide (ScCO2). In addition, the unreacted gas and solvent recycle was incorporated and the effect of unreacted feed recycle was evaluated. It was expected that with the recycle, the feed rate can be increased. The increase in conversion and liquid selectivity accompanied by the production of narrower carbon number distribution in the product suggest that higher flow rates can and should be used when incorporating exit gas recycle. It was observed that this process was capable of enhancing the hydrocarbon selectivity (nearly 98 % CO conversion), reducing improving the carbon efficiency from 17 % to 51 % in a once through process and further converting 16 % CO2 to liquid with integrated recycle of the product gas stream and increasing the life of the catalyst. Catalyst robustness enhancement has been attributed to the absorption of heat of reaction by the compressed CO2 which reduced the formation of hotspots and the dissolution of waxes by the CO2 solvent which reduced the blinding of active sites. In addition, the recycling the product gas stream reduced the reactor footprint to one-fourth of the once through size and product fractionation utilizing the solvent effects of supercritical CO2 were realized. In addition to the negative CO2 selectivities, methane production was also inhibited and was limited to less than 1.5%. The effect of the process conditions on the life of the catalysts will also be presented. Fe based catalysts are known to have a high proclivity for producing CO2 during FTS. The data of the product spectrum and selectivity on Co and Fe-Co based catalysts as well as those obtained from commercial sources will also be presented. The measurable decision criteria were the increase in CO conversion at H2:CO ratio of 1:1 (as commonly found in coal gasification product stream) in supercritical phase as compared to gas phase reaction, decrease in CO2 and CH4 selectivity, overall liquid product distribution, and finally an increase in the life of the catalysts.

Keywords: carbon efficiency, Fischer Tropsch synthesis, low GHG, pressure tunable fractionation

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2 DNA Barcoding for Identification of Dengue Vectors from Assam and Arunachal Pradesh: North-Eastern States in India

Authors: Monika Soni, Shovonlal Bhowmick, Chandra Bhattacharya, Jitendra Sharma, Prafulla Dutta, Jagadish Mahanta

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Aedes aegypti and Aedes albopictus are considered as two major vectors to transmit dengue virus. In North-east India, two states viz. Assam and Arunachal Pradesh are known to be high endemic zone for dengue and Chikungunya viral infection. The taxonomical classification of medically important vectors are important for mapping of actual evolutionary trends and epidemiological studies. However, misidentification of mosquito species in field-collected mosquito specimens could have a negative impact which may affect vector-borne disease control policy. DNA barcoding is a prominent method to record available species, differentiate from new addition and change of population structure. In this study, a combined approach of a morphological and molecular technique of DNA barcoding was adopted to explore sequence variation in mitochondrial cytochrome c oxidase subunit I (COI) gene within dengue vectors. The study has revealed the map distribution of the dengue vector from two states i.e. Assam and Arunachal Pradesh, India. Approximate five hundred mosquito specimens were collected from different parts of two states, and their morphological features were compared with the taxonomic keys. The analysis of detailed taxonomic study revealed identification of two species Aedes aegypti and Aedes albopictus. The species aegypti comprised of 66.6% of the specimen and represented as dominant dengue vector species. The sequences obtained through standard DNA barcoding protocol were compared with public databases, viz. GenBank and BOLD. The sequences of all Aedes albopictus have shown 100% similarity whereas sequence of Aedes aegypti has shown 99.77 - 100% similarity of COI gene with that of different geographically located same species based on BOLD database search. From dengue prevalent different geographical regions fifty-nine sequences were retrieved from NCBI and BOLD databases of the same and related taxa to determine the evolutionary distance model based on the phylogenetic analysis. Neighbor-Joining (NJ) and Maximum Likelihood (ML) phylogenetic tree was constructed in MEGA6.06 software with 1000 bootstrap replicates using Kimura-2-Parameter model. Data were analyzed for sequence divergence and found that intraspecific divergence ranged from 0.0 to 2.0% and interspecific divergence ranged from 11.0 to 12.0%. The transitional and transversional substitutions were tested individually. The sequences were deposited in NCBI: GenBank database. This observation claimed the first DNA barcoding analysis of Aedes mosquitoes from North-eastern states in India and also confirmed the range expansion of two important mosquito species. Overall, this study insight into the molecular ecology of the dengue vectors from North-eastern India which will enhance the understanding to improve the existing entomological surveillance and vector incrimination program.

Keywords: COI, dengue vectors, DNA barcoding, molecular identification, North-east India, phylogenetics

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1 Effect of Radioprotectors on DNA Repair Enzyme and Survival of Gamma-Irradiated Cell Division Cycle Mutants of Saccharomyces pombe

Authors: Purva Nemavarkar, Badri Narain Pandey, Jitendra Kumar

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Introduction: The objective was to understand the effect of various radioprotectors on DNA damage repair enzyme and survival in gamma-irradiated wild and cdc mutants of S. pombe (fission yeast) cultured under permissive and restrictive conditions. DNA repair process, as influenced by radioprotectors, was measured by activity of DNA polymerase in the cells. The use of single cell gel electrophoresis assay (SCGE) or Comet Assay to follow gamma-irradiation induced DNA damage and effect of radioprotectors was employed. In addition, studying the effect of caffeine at different concentrations on S-phase of cell cycle was also delineated. Materials and Methods: S. pombe cells grown at permissive temperature (250C) and/or restrictive temperature (360C) were followed by gamma-radiation. Percentage survival and activity of DNA Polymerase (yPol II) were determined after post-irradiation incubation (5 h) with radioprotectors such as Caffeine, Curcumin, Disulphiram, and Ellagic acid (the dose depending on individual D 37 values). The gamma-irradiated yeast cells (with and without the radioprotectors) were spheroplasted by enzyme glusulase and subjected to electrophoresis. Radio-resistant cells were obtained by arresting cells in S-phase using transient treatment of hydroxyurea (HU) and studying the effect of caffeine at different concentrations on S-phase of cell cycle. Results: The mutants of S. pombe showed insignificant difference in survival when grown under permissive conditions. However, growth of these cells under restrictive temperature leads to arrest in specific phases of cell cycle in different cdc mutants (cdc10: G1 arrest, cdc22: early S arrest, cdc17: late S arrest, cdc25: G2 arrest). All the cdc mutants showed decrease in survival after gamma radiation when grown at permissive and restrictive temperatures. Inclusion of the radioprotectors at respective concentrations during post irradiation incubation showed increase in survival of cells. Activity of DNA polymerase enzyme (yPol II) was increased significantly in cdc mutant cells exposed to gamma-radiation. Following SCGE, a linear relationship was observed between doses of irradiation and the tail moments of comets. The radioprotection of the fission yeast by radioprotectors can be seen by the reduced tail moments of the yeast comets. Caffeine also exhibited its radio-protective ability in radio-resistant S-phase cells obtained after HU treatment. Conclusions: The radioprotectors offered notable radioprotection in cdc mutants when added during irradiation. The present study showed activation of DNA damage repair enzyme (yPol II) and an increase in survival after treatment of radioprotectors in gamma irradiated wild type and cdc mutants of S. pombe cells. Results presented here showed feasibility of applying SCGE in fission yeast to follow DNA damage and radioprotection at high doses, which are not feasible with other eukaryotes. Inclusion of caffeine at 1mM concentration to S phase cells offered protection and did not decrease the cell viability. It can be proved that at minimal concentration, caffeine offered marked radioprotection.

Keywords: radiation protection, cell cycle, fission yeast, comet assay, s-phase, DNA repair, radioprotectors, caffeine, curcumin, SCGE

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