Browsing by Author "Sushma Rathaur"

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Antifilarial efficacy of andrographolide: Ex vivo studies on bovine filarial parasite Setaria cervi
(Elsevier Inc., 2022) Smita Yadav; Faiyaz Ahmad; Sushma Rathaur
Lymphatic filariasis caused by filarial nematode is an important disease leading to considerable morbidity throughout tropical countries. Even after specific elimination programs, the disease continue to spread in endemic countries. Thus newer therapeutic interventions are urgently needed to control the spread. In the present study, we have seen the effect of andrographolide (andro), a diterpenoid lactone from the leaves of Andrographis paniculata on filarial parasite Setaria cervi. There was time and concentration dependent decrease in motility and viability leading to death of parasite after 6 h of the exposure of andro. Andro showed potential antifilarial activity with an IC50 value of 24.80 μM assessed through MTT assay. There was concentration dependent decrease in the antioxidant enzymes activity and increase in proapoptotic markers after 5 h exposure of andro. Further, molecular docking analysis revealed that andro binds with filarial glutathione-S-transferase at glutathione (GSH) binding site and inhibiting enzyme activity competitively. Andro induced oxidative stress mediated apoptosis in parasites as evidenced by increase in the intracellular reactive oxygen species (ROS) and apoptotic markers.Therefore this study suggested that andro could be further explored as a new antifilarial drug. © 2022 Elsevier Inc.
Antifilarial efficacy of green silver nanoparticles synthesized using Andrographis paniculata
(Editions de Sante, 2020) Smita Yadav; Shweta Sharma; Faiyaz Ahmad; Sushma Rathaur
The currently available antifilarial drugs are mostly effective against microfilariae. Antifilarial drugs have disadvantages such as toxicity and the development of resistance due to the continuous use. Therefore alternative drugs are required for the control of disease. At present, nanoparticles are used for developing anti-parasitic therapy for their special properties such as smallest in size, bio-availability, bio-compatibility and penetration capacity into a cell. In the present study green nanoparticles were biosynthesized by using leaf extract of Andrographis paniculata to evaluate its antifilarial efficacy. Biosynthesized nanoparticles were characterized by UV–Vis spectroscopy, FTIR, XRD, SEM, EDAX, TEM, HRTEM, AFM and found to be homogenous with average size of 11 nm. The antifilarial efficacy of green nanoparticles was based on adult filarial parasite motility and viability assay. These green nanoparticles (NPs) were found to have antifilarial activity with LC50 of 11.6 μg/ml against adult female filarial parasites. The green nanoparticles induced oxidative stress as evidenced by elevated ROS production and decline of parasitic GST, GR, TRxR and GSH levels in the parasite. The activation of ced-3 gene, a homolog of mammalian caspase 3, reduced expression of ced-9 and decreased activity of cytochrome c oxidase suggested induction of mitochondrial mediated apoptosis in parasite. These green NPs are more effective than the plant extract. This is the first report where green nanoparticles synthesized from A. paniculata showed antifilarial efficacy against adult filarial parasites. © 2020
Arsenic mediated modifications in Bacillus aryabhattai and their biotechnological applications for arsenic bioremediation
(Elsevier Ltd, 2016) Namrata Singh; Sunil Gupta; Naina Marwa; Vivek Pandey; Praveen C. Verma; Sushma Rathaur; Nandita Singh
The present study reports the arsenic (As) tolerance mechanism of bacteria Bacillus aryabhattai (NBRI014). The data explores the intracellular accumulation and volatilization of As from the culture medium after 48 h of exposure to 25,000 mg l−1 arsenate As(V). The study also provides the evidence of presence of ars operon in bacteria, which may have played an important role in reducing As toxicity. Additionally, we found 7 differentially expressed proteins to be up-regulated in bacterial cells upon As exposure which may have role in reducing As toxicity inside bacterial cells. Furthermore, Fourier transform infrared (FTIR) spectroscopic techniques were useful to describe the structural and compositional alterations in bacterial cells after As treatment. It showed the changes in peak positions of the spectrum pattern when NBRI014 was grown in medium containing As, indicating that these functional groups viz. (amino, alkyl halides and hydroxyl) present on bacterial surface, which may be involved in As binding. The above results signify that biotechnological application of the isolate NBRI014 could be helpful in removal of As from polluted sites. © 2016 Elsevier Ltd
Assessing the bioremediation potential of arsenic tolerant bacterial strains in rice rhizosphere interface
(Chinese Academy of Sciences, 2016) Namrata Singh; Shubhi Srivastava; Sushma Rathaur; Nandita Singh
The arsenic tolerant bacterial strains Staphylococcus arlettae (NBRIEAG-6), Staphylococcus sp. (NBRIEAG-8) and Brevibacillus sp. (NBRIEAG-9) were tested for their roles in enhancing plant growth and induction of stress-related enzymes in rice (Oryza sativa L. cv. NDR-359) plants at two different concentrations, 30 and 15 mg/kg of As(V) and As(III), respectively. An experiment was conducted to test the effect of these strains on plant growth promotion and arsenic uptake. We found 30%–40% reduction in total As uptake in bacteria-inoculated plants, with increased plant growth parameters compared to non-inoculated plants. Moreover, the bacteria-inoculated plants showed reduced activity of total glutathione (GSH) and glutathione reductase (GR) compared to their respective controls, which suggests the bacteria-mediated reduction of oxidative stress in plants. Thus, these strains were found to be beneficial in terms of the biochemical and physiological status of the plants under arsenic stress conditions. Furthermore, one-way ANOVA and principal component analysis (PCA) on enzymatic and non-enzymatic assays also revealed clear variations. The results support the distinction between control and treatments in both shoots and roots. Therefore, this study demonstrates the potential of rhizobacteria in alleviating arsenic stress in rice plants. © 2016
Brevundimonas diminuta mediated alleviation of arsenic toxicity and plant growth promotion in Oryza sativa L.
(Academic Press, 2016) Namrata Singh; Naina Marwa; Shashank k. Mishra; Jyoti Mishra; Praveen C. Verma; Sushma Rathaur; Nandita Singh
Arsenic (As), a toxic metalloid adversely affects plant growth in polluted areas. In the present study, we investigated the possibility of improving phytostablization of arsenic through application of new isolated strain Brevundimonas diminuta (NBRI012) in rice plant [Oryza sativa (L.) Var. Sarju 52] at two different concentrations [10 ppm (low toxic) and 50 ppm (high toxic)] of As. The plant growth promoting traits of bacterial strains revealed the inherent ability of siderophores, phosphate solubilisation, indole acetic acid (IAA), 1-aminocyclopropane-1-carboxylic acid (ACC) deaminase production which may be associated with increased biomass, chlorophyll and MDA content of rice and thereby promoting plant growth. The study also revealed the As accumulation property of NBRI012 strain which could play an important role in As removal from contaminated soil. Furthermore, NBRI012 inoculation significantly restored the hampered root epidermal and cortical cell growth of rice plant and root hair elimination. Altogether our study highlights the multifarious role of B. diminuta in mediating stress tolerance and modulating translocation of As in edible part of rice plant. © 2015 Elsevier Inc.
Brugia malayi and Wuchereria bancrofti: Gene comparison and recombinant expression of π-class related glutathione S-transferases
(Academic Press Inc., 2003) Sushma Rathaur; Peter Fischer; Marzena Domagalski; Rolf D. Walter; Eva Liebau
The nucleotide sequence for Brugia malayi and Wuchereria bancrofti GST have been submitted to EMBL, GenBank, and DDBJ Nucleotide Sequence Databases under Accession Nos. Y12788 and AY195867. © 2003 Elsevier Science (USA). All rights reserved.
Characterization of filarial phosphoglycerate kinase
(Elsevier B.V., 2019) Ranjeet Kumar; Faiyaz Ahmad; Sushma Rathaur
Phosphoglycerate kinase (PGK) is a key enzyme of glycolysis which also acts as a mediator of DNA replication and repair in the nucleus. We have cloned and expressed PGK in Brugia malayi. The rBmPGK was found to be 415 amino acid residues long having 45 kDa subunit molecular weight. This enzyme was also identified in different life stages of bovine filarial parasite Setaria cervi. The enzyme activity was highest in microfilarial stage followed by adult female and male as also shown by real time PCR in the present study. Further using BmPGK primers the cDNA prepared from S. cervi was amplified and sequenced which showed 100% homology with Brugia malayi PGK. B. malayi and S. cervi, PGK consists of conserved calmodulin binding domain (CaMBD) having 21 amino acids. In the present study we have shown the CaMBD binds to calcium-calmodulin and regulates its activity. The binding of calmodulin (CaM) with CaMBD was confirmed using calmodulin agarose binding pull down assay, which showed that the rBmPGK binds to CaM agarose-calcium dependent manner. The effect of CaM-Ca2+on the activity of rBmPGK was studied at different concentration of CaM (0.01–5.0 μM) and calcium chloride (0.01–100 μM). The rBmPGK was activated up to 85% in the presence of CaM at 1 μM and 10 μM concentration of CaCl2. Interestingly this activation was abrogated by metal chelator EDTA. Similar results were shown in case of Setaria cervi PGK. A significant increase (90 ± 10) % in ScPGK activity was observed in the presence of CaM and CaCl2 at 1.0 μM and 1.0 mM respectively, further increase in the conc. of CaCl2, the activity of ScPGK was found to be decreased like rBmPGK. Bioinformatics studies have also confirmed the interaction between CaMBD and CaM which showed CaM interacted to Phe 206, Gln 220, Arg 223 and Asn 224 of rBmPGK CaM binding domain. On the basis of these findings, it has been suggested that the activity of filarial PGK could be regulated in cells by Ca2+-CaM depending upon the concentration of calcium. To the best of our knowledge this is first report in filarial parasite. © 2019 Elsevier B.V. and Société Française de Biochimie et Biologie Moléculaire (SFBBM)
Characterization of secretory acetylcholinesterase from Setaria cervi microfilariae: A potential antigen for diagnosis of human filariasis
(1999) Sudha Sharma; Sushma Rathaur
Acetylcholinesterase (AChE) is released to the external medium when microfilariae (mf) of Setaria cervi, a bovine filarial parasite, are maintained in vitro. Intense enzyme staining at amphids, excretory pores, anal vesicle and phasmids suggest an active secretion of AChE from mf. Excretory-secretory products of mf displayed two electromorphic variants of AChE when resolved by 6% nondenaturing PAGE. The two isoforms of AChE (A and B) were separated on the basis of charge by DEAE sepharose CL 6B column following gel filtration. The two isoforms showed differing kinetic properties with respect to substrate specificity and inhibitor sensitivity. Anti-Nippostrongylus brasiliensis AChE antibodies cross-reacted with the affinity purified secretory AChE in ELISA. Immunoblotting of purified AChEs with cross-reacting anti-AChE antibodies revealed the presence of an approximately 75 kD protein in the isoenzyme A and an approximately 45 kD protein in B, whereas both proteins were present in the enzyme purified via affinity chromatography on edrophonium sepharose column.
Cloning, overexpression, purification and characterization of Plasmodium knowlesi lactate dehydrogenase
(2012) Vandana Singh; Deep C. Kaushal; Sushma Rathaur; Niraj Kumar; Nuzhat A. Kaushal
Plasmodial lactate dehydrogenase, key enzyme of anaerobic glycolysis, has been shown to be a potential immunodiagnostic marker as well as a novel target for chemotherapy. We have cloned, overexpressed and immunochemically characterized the recombinant lactate dehydrogenase of Plasmodium knowlesi, the fifth human malaria parasite. The P. knowlesi lactate dehydrogenase (PkLDH) gene was PCR amplified and 0.9 kb PCR product was cloned into pGEM-T Easy vector. Sequencing and BLAST analysis revealed open reading frame of 316 amino acids of PkLDH showing 96.8% homology with Plasmodium vivax LDH and around 90% with Plasmodium falciparum, Plasmodium malariae and Plasmodium ovale LDHs. The PkLDH gene was subcloned into pGEX-6P1 expression vector and the SDS-PAGE analysis revealed that about 70% of fusion protein was present in the soluble fraction. The fusion protein was cleaved with PreScission protease and recombinant PkLDH (34 kDa) was affinity purified to homogeneity. The purified PkLDH exhibited high reactivity with polyclonal and monoclonal antibodies against plasmodial LDH. The polyclonal antibody produced against purified recombinant PkLDH in rabbits showed high ELISA reactivity with both native and recombinant PkLDH and could detect parasite LDH in malaria infected blood samples by sandwich ELISA. The purified recombinant PkLDH can be used to produce P. knowlesi specific monoclonal antibodies for specific diagnosis of P. knowlesi infection in humans. © 2012 Published by Elsevier Inc.
Combination of DEC plus aspirin induced mitochondrial mediated apoptosis in filarial parasite Setaria cervi
(2010) Alka Singh; Sushma Rathaur
Diethylcarbamazine (DEC) is the main drug used against lymphatic filariasis but it is only microfilaricidal. Hence there is an urgent need for adulticidal drug. Aspirin is known nonsteroidal anti-inflammatory drugs which can inhibit prostaglandin H synthase and also induces apoptosis. Studies presented in this paper demonstrated that exposure of worms to the combination of DEC plus aspirin (DEC + A) at 100 μM concentration irreversibly paralyzed adult worms as well as microfilariae within 2 h. Some of the apoptosis markers viz; DNA fragmentation with accompanying ladder formation, upregulation of Bax expression and decrease in Bcl-2 have suggested that the parasite may be killed due to mitochondrial mediated apoptosis. The levels of several apoptosis regulating proteins and enzymes have also shown to be altered. DEC + A treated worms showed significant decrease in prostaglandin H synthase activity (PGHS) and increase in the level of nitric oxide (NO) and cysteine proteases while glutathione (GSH) and peroxidase level was found to be decreased. NO is known inducer of mitochondrial mediated apoptosis and acts by increasing the permeability of mitochondrial membrane through Bax and allowing cytochrome c to release in cytosol, inducing caspases leading to apoptosis. The DEC + A concentration used in this study is much lower than recommended dose so its intake is safe. Here we report for the first time that combination of DEC and aspirin is more effective and could be used as an adulticidal for control of human filarial infections. © 2010 Elsevier Masson SAS.
Designing, synthesis of selective and high-affinity chalcone-benzothiazole hybrids as Brugia malayi thymidylate kinase inhibitors: In vitro validation and docking studies
(Elsevier Masson SAS, 2015) Koneni V. Sashidhara; Srinivasa Rao Avula; Pawan Kumar Doharey; L. Ravithej Singh; Vishal M. Balaramnavar; Jyoti Gupta; Shailja Misra-Bhattacharya; Sushma Rathaur; Anil K. Saxena; Jitendra Kumar Saxena
In our continuing search for safe and efficacious antifilarials, a series of novel chalcone-benzothiazole hybrids have been synthesized and evaluated for their Brugia malayi thymidylate kinase (BmTMK) enzyme inhibition activity. Their selectivity towards BmTMK was studied and compared to the human TMK (HsTMK) by an in silico method. Out of seventeen derivatives, compounds 34 and 42 showed higher interactions with the BmTMK active site. MolDock docking model revealed the interactions of these two derivatives and the results corroborated well with their in vitro antifilarial activities. Our studies suggest that these hybrids are selective towards the BmTMK enzyme and may serve as potential therapeutic agents against filariasis. © 2015 Elsevier Masson SAS. All rights reserved.
Effect of CDNB on filarial thioredoxin reductase : A proteomic and biochemical approach
(Elsevier, 2015) Savitri Tiwari; Mohit Wadhawan; Neetu Singh; Sushma Rathaur
Thioredoxin reductase plays a crucial role in the maintenance of cellular redox homeostasis. In this study, we have targeted TrxR in Setaria cervi, a bovine filarial parasite using its inhibitor CDNB. It caused significant decrease in the motility and viability of these parasites leading to their death. Inhibition of TrxR leads to the downregulation of the antioxidant system followed by generation of oxidative stress in these parasites. The increased ROS level induced lipid peroxidation and protein carbonyl formation which might alter the mitochondrial membrane permeability leading to release of cytochrome c. CDNB significantly downregulated the level of ced-9 and activity of tyrosine phosphatases, cytochrome c oxidase. It also upregulated ced-3, homolog of mammalian caspase 3 suggesting initiation of intrinsic pathway of apoptosis. The proteomic profile of CDNB treated parasites showed marked alteration in abundance of different protein spots with 20% downregulated and 13% unregulated spots in comparison to control parasites. We observed a downregulation in the glycolytic enzymes such as enolase, PGK, and GAPDH thereby blocking the ATP formation in the parasite. This study suggests that TrxR inhibition disrupts the cellular homeostasis thereby generating oxidative stress followed by mitochondrial mediated apoptosis in filarial parasites leading to the death of the parasites. Biological significance: Lymphatic filariasis is one of the most prevalent tropical diseases caused by tissue dwelling parasitic nematodes viz., Wuchereria bancrofti, Brugia malayi and Brugia timori. Currently available antifilarial drugs effectively eliminate larval stages of the parasite but are ineffective against the adult worms. Therefore, there is an urgent need for finding proteins/enzymes which play a crucial role in the persistence of these parasites. Our study for the first time reports the important role played by S. cervi TrxR in its survival. Thus, suggesting filarial TrxR as a potent chemotherapeutic target against lymphatic filariasis. This would help in screening of new compounds having macrofilaricidal activity. © 2014 Elsevier B.V.
Effect of diethylcarbamazine, butylated hydroxy anisole and methyl substituted chalcone on filarial parasite Setaria cervi: Proteomic and biochemical approaches
(2011) Sushma Rathaur; Marshleen Yadav; Neetu Singh; Alka Singh
For survival, parasite exerts several lines of defense of which drug neutralization is one of the major phenomena. Lack of phase I cytochrome P450 in some of the nematode render them depend on the phase II detoxification system involving GST as a major detoxifying enzymes. In present study, the antifilarial DEC, phenolic compound BHA and methyl chalcone have been evaluated for proteomic and biochemical studies in Setaria cervi. BHA and methyl chalcone showed cytotoxic effect leading to irreversible inhibition in motility and viability of parasites. These drugs showed marked alteration in proteomic profile of S. cervi at 100 μM concentration with 10.82, 8.52 and 6.75% downregulated (< 0.5) and 7.64, 31.78 and 24.32% upregulated (> 1.5) in DEC, BHA and methyl chalcone treatment respectively. Significant depletion in GSH level with increase in NO production was observed. Amongst these compounds, methyl chalcone demonstrated significant inhibitory effect (p < 0.05) on GST, PGHS and PTP activity leading to loss of metabolic homeostasis and parasite death. The cytotoxic response and altered expression profile of major enzymes under drug exposure suggested the oxidative stress induced apoptosis as a major cause of parasite killing which was further supported by DNA fragmentation in BHA and methyl chalcone. © 2011 Elsevier B.V.
Effects of explant age, germination medium, pre-culture parameters, inoculation medium, pH, washing medium, and selection regime on Agrobacterium-mediated transformation of tomato
(2012) Govind Kumar Rai; Neha Prakash Rai; Sanjeev Kumar; Akhilesh Yadav; Sushma Rathaur; Major Singh
An efficient protocol was developed for Agrobacterium tumefaciens-mediated transformation of tomato (Solanum lycopersicum) cultivars using cotyledon explants. The transformation frequency was assessed in response to several different factors, including seed germination medium, seedling age, pre-culture duration, pre-culture and co-cultivation media, inoculation medium, medium pH, washing medium, and kanamycin concentration in initial selection medium. Cotyledons excised from 6-d-old seedlings germinated on half-strength Murashige and Skoog's (MS) basal medium containing 8.9 μM benzyladenine (BA) produced the most suitable explant material. Six days of explant pre-culture and 5 min inoculation with Agrobacterium culture in MS medium, containing 8.9 μM BA, 9.3 μM kinetin, and 0.4 mg l -1 thiamine at pH 5.0, significantly improved the transformation frequency. The addition of a tobacco feeder cell layer, however, did not lead to any significant improvement in the transformation rate. Kanamycin at 20 mg l -1 in the selection medium for the initial 10 d resulted in the highest transformation frequency. Combining the best conditions for each parameter resulted in an overall transformation efficiency of 44.3 %. Gene transfer was confirmed through PCR and Southern blot analyses. Mendelian inheritance ratios were found in 71.5 % of the independent transgenic lines from self-fertilized T 1 progeny. The optimized transformation procedure showed high transformation frequencies for all three tomato cultivars tested, namely, Kashi Vishesh (H-86), Hisar Anmol (H-24), and Kashi Amrit (DVRT-1), and is also expected to give reproducible results with other tomato cultivars. © 2012 The Society for In Vitro Biology.
Evidence for the presence of prostaglandin H synthase like enzyme in female Setaria cervi and its inhibition by diethylcarbamazine
(2009) Sushma Rathaur; Alka Singh; Marshleen Yadav; Reeta Rai
Experimental evidence has shown that Setaria cervi a bovine filarial parasite contains significant amount of prostaglandin H synthase like activity in the somatic extract of its different life stages. A protein with characteristics of prostaglandin H synthase was purified to homogeneity from female somatic extract using a combination of affinity and gel filtration chromatography. Molecular weight of purified enzyme was 70 kDa as determined by SDS-PAGE. Purified enzyme showed high activity with arachidonic acid and TMPD substrates suggests the presence of both cyclooxygenase and peroxidase activity in enzyme. Fluorescence spectroscopy and hemin-associated peroxidase activity confirmed presence of heme in purified enzyme. The Km and Vmax values using arachidonic acid were determined to be 79 ± 1.5 μM and 0.165 ± 0.2 U/ml, respectively. Further, indomethacin and aspirin, specific inhibitors for PGHS, significantly inhibited the enzyme activity. Diethylcarbamazine, an antifilarial drug inhibited the microfilarial PGHS like activity as well as their motility. Here we are reporting for the first time PGHS like activity in filarial parasite and its inhibition with DEC which provide that this enzyme could be used as a drug target. © 2009 Elsevier B.V. All rights reserved.
Expression of rd29A:: AtDREB1A/CBF3 in tomato alleviates drought-induced oxidative stress by regulating key enzymatic and non-enzymatic antioxidants
(2013) Govind Kumar Rai; Neha Prakash Rai; Sushma Rathaur; Sanjeev Kumar; Major Singh
Transgenic tomato lines (cv. Kashi Vishesh) over-expressing AtDREB1A/. CBF3 driven by stress-inducible rd29A promoter showed significantly higher activities of key antioxidant enzymes when exposed to water-deficit for 7, 14, and 21 days. Transgenic tomato plants exposed to water-deficit recorded lower levels of hydrogen peroxide and superoxide anion formation compared to the non-transgenic plants, suggesting alleviation of reactive oxygen species (ROS). A significant increase in activities of enzymes superoxide dismutase (SOD), catalase (CAT), ascorbate peroxidase (APX), glutathione reductase (GR), dehydroascorbate reductase (DHAR), and monodehydroascorbate reductase (MDHAR) was observed in response to the different durations of water-deficit conditions. In contrast, enzyme guaiacol peroxidase (POD) activity was lower in the transgenic lines and showed a negative correlation with ROS, ascorbic acid (AsA), and glutathione levels. The concentrations of AsA, glutathione and their reduced forms were higher in the transgenic plants and increased with ROS levels. These results indicate that AtDREB1A transgenic tomato lines are better adapted to water-deficit as they showed lower drought-induced oxidative stress due to activation of the antioxidant response. © 2013 Elsevier Masson SAS.
Filarial glutathione S-transferase: Its induction by xenobiotics and potential as drug target
(2005) Sarika Gupta; Sushma Rathaur
Glutathione-S-transferase (GST) a Phase-II drug detoxification enzyme, was detected in Setaria cervi, a bovine filarial parasite. In vitro effect of diethylcarbamazine, butylated hydroxyanisole and phenobarbitone on the GST of adult female S. cervi was assayed by the addition of these compounds in the maintenance medium. The specific activity of GST towards 1-chloro-2,4- dinitrobenzene was increased progressively 1.2-1.97, 1.3-2.4 and 1.2-2.7 times at 10-100 μM of diethylcarbamazine, butylated hydroxyanisole and phenobarbitone, respectively, after 5 h at 37°C. Substrate specificity studies showed a higher increase in specific activity with ethacrynic acid and no change with cumene hydroperoxide. Although the intensity of GST activity band was more in extract from diethylcarbamazine or butylated hydroxyanisole treated worms extract, an extra band of activity appeared in those worm extracts compared to control worm extract. SDS/PAGE showed increased thickness of the band corresponding to purified GST in extracts from diethylcarbamazine/butylated hydroxyanisole/phenobarbitone treated worms. Purification and quantification of GST from diethylcarbamazine and butylated hydroxyanisole treated worms indicated an increase in enzyme specific activity. The increase in GST protein by these agents was blocked by prior treatment with actinomycin D, indicative of a transcription dependent response. The role of this enzyme in motility and viability of microfilariae and adult female was tested in vitro using a range of known GST inhibitors. Of those tested, ethacrynic acid, ellagic acid, 1-chloro-2,4-dinitrobenzene, cibacron blue and butylated hydroxyanisole reduced the viability and motility of microfilariae and adult female worms at micromolar concentrations. These results suggest that S. cervi GST is inducible in response to the antifilarial drug diethylcarbamazine and may play an important role in parasite's survival, thus could be a potential drug target.
Filarial glutathione-S-transferase: A potential vaccine candidate against lymphatic filariasis
(2008) Sushma Rathaur; Marshleen Yadav; Sarika Gupta; V. Anandharaman; Maryada Venkatarami Reddy
Present report enumerates the vaccine potential of a glutathione-S-transferase purified from Setaria cervi against lymphatic filariasis. In jirds (Meriones unguiculatus) vaccination trial, a very significant 82.75% (p < 0.005) reduction in adult parasite burden was observed in ScGST immunized group after 90 days post Brugia malayi L3 challenge. An inverse correlation between the antibody level and worm burden was found in ScGST immunized group (Person's correlation r = 0.943, p < 0.05). No recoveries of worms were obtained in heart and lungs of vaccinated group. The Antibodies reactive to ScGST appeared within four weeks of first dose and were able to neutralize the GST activity up to 86%. In an earlier study we have shown vaccine potential of ScGST against B. malayi by ADCC. Evaluation of cytokine profile in T-cells isolated from BALB/c mice immunized with ScGST were also showed predominance of Th2 response which further maintained the humoral immunity generated by ScGST administration in mice. The overall observations prompted us to envisage ScGST as a potential vaccine candidate against lymphatic filariasis. © 2008 Elsevier Ltd. All rights reserved.
Filarial selenium glutathione peroxidase: A probable immunodiagnostic marker for lymphatic filariasis
(2010) Anchal Singh; Shaukat Kamal; Sushma Rathaur
Lymphatic filariasis (LF) caused by Wuchereria bancrofti is widely prevalent in tropical and subtropical countries. Night blood film examination is most commonly used for diagnosis of filariasis but is cumbersome and labour intensive. In order to develop an indirect ELISA-based immunodiagnostic test, the importance of antifilarial IgG subclasses was evaluated in bancroftian filariasis patients. Blood samples from healthy individuals and different categories of LF patients were used to estimate the diagnostic potential of selenium glutathione peroxidase antigen purified from the bovine filarial parasite Setaria cervi. This antigen reacted with both IgG1 and IgG4; however, the IgG1 response was greater in microfilaraemic patients and the IgG4 response was higher in chronic filarial patients. The diagnostic sensitivity of IgG1 and IgG4 was 97% and 96% whereas specificity was determined to be 95% and 98% respectively. Our observations suggest that SeGSHPx could be an alternative diagnostic marker for the detection of bancroftian filariasis in an endemic area. © 2010 Royal Society of Tropical Medicine and Hygiene.
Filling the gap of intracellular dephosphorylation in the Plasmodium falciparum vitamin B1 biosynthesis
(2008) Julia Knöckel; Bärbel Bergmann; Ingrid B. Müller; Sushma Rathaur; Rolf D. Walter; Carsten Wrenger
Thiamine pyrophosphate (TPP), the active form of vitamin B1, is an essential cofactor for several enzymes. Humans depend exclusively on the uptake of vitamin B1, whereas bacteria, plants, fungi and the malaria parasite Plasmodium falciparum are able to synthesise thiamine monophosphate (TMP) de novo. TMP has to be dephosphorylated prior to pyrophosphorylation in order to obtain TPP. In P. falciparum the phosphatase capable to catalyse this reaction has been identified by analysis of the substrate specificity. The recombinant enzyme accepts beside vitamin B1 also nucleotides, phosphorylated sugars and the B6 vitamer pyridoxal 5′-phosphate. Vitamin B1 biosynthesis is known to occur in the cytosol. The cytosolic localisation of this phosphatase was verified by transfection of a GFP chimera construct. Stage specific Northern blot analysis of the phosphatase clearly identified an expression profile throughout the entire erythrocytic life cycle of P. falciparum and thereby emphasises the importance of dephosphorylation reactions within the malaria parasite. © 2007 Elsevier B.V. All rights reserved.