Molecular modeling, simulation and docking of Rv1250 protein from Mycobacterium tuberculosis.

Computational prediction and protein structure modeling have come to the aid of various biological problems in determining the structure of proteins. These technologies have revolutionized the biological world of research, allowing scientists and researchers to gain insights into their biological questions and design experimental research much more efficiently. Pathogenic Mycobacterium spp. is known to stay alive within the macrophages of its host. Mycobacterium tuberculosis is an acid-fast bacterium that is the most common cause of tuberculosis and is considered to be the main cause of resistance of tuberculosis as a leading health issue. The genome of Mycobacterium tuberculosis contains more than 4,000 genes, of which the majority are of unknown function. An attempt has been made to computationally model and dock one of its proteins, Rv1250 (MTV006.22), which is considered as an apparent drug-transporter, integral membrane protein, and member of major facilitator superfamily (MFS). The most widely used techniques, i.e., homology modeling, molecular docking, and molecular dynamics (MD) simulation in the field of structural bioinformatics, have been used in the present work to study the behavior of Rv1250 protein from M. tuberculosis. The structure of unknown TB protein, i.e., Rv1250 was retrived using homology modeling with the help of I-TASSER server. Further, one of the sites responsible for infection was identified and docking was done by using the specific Isoniazid ligand which is an inhibitor of this protein. Finally, the stability of protein model and analysis of stable and static interaction between protein and ligand molecular dynamic simulation was performed at 100 ns The designing of novel Rv1250 enzyme inhibitors is likely achievable with the use of proposed predicted model, which could be helpful in preventing the pathogenesis caused by M. tuberculosis. Finally, the MD simulation was done to evaluate the stability of the ligand for the specific protein.

Environmentally relevant concentrations of Triclosan cause transcriptomic and biomolecular alterations in the hatchlings of Labeo rohita.

Suppression (p ≤ 0.05) of antioxidative/detoxification (except GPx and CYP3a) and cytoskeletal (except DHPR) genes but induction of metabolic (except for AST and TRY) and heat shock (except HSP60) genes of Labeo rohita hatchlings after 14 days of exposure to environmentally relevant concentrations of Triclosan (0.0063, 0.0126, 0.0252 and 0.06 mg/L) was followed by an increase (p ≤ 0.05) for most of the genes after 10 days recovery period. After recovery, LDH, ALT, CK, CHY, PA, HSP47 and DHPR declined, while SOD, CAT, GST, GR, GPx, CYP1a, CYP3a, AST, AChE, TRY, HSP60, HSP70, HSc71, HSP90 MLP-3, α-tropomyosin, desmin b and lamin b1 increased over exposure. Peak area of biomolecules (except 3290-3296, 2924-2925 and 2852-2855 cm-1) declined (p ≤ 0.01) more after recovery [except for an increase (p ≤ 0.01) at 1398-1401 cm-1]. CYP3a, CK, HSP90, MLP-3 and secondary structure of amide A are the most sensitive markers for the environmentally relevant concentrations of Triclosan.

MSMEG_3955 from Mycobacterium smegmatis is a FMN bounded homotrimeric NAD(P)H:Flavin mononucleotide (FMN) oxidoreductase.

BACKGROUND: Tuberculosis (TB) remains an important public health problem since it is the major cause of elevated morbidity and mortality globally. Previous works have shown that Mycobacterium tuberculosis (Mtb); the prime causative agent of the deadly disease has dormancy survival regulator (DosR) regulon, a two-component regulatory system which controls the transcription of more than 50 genes. However, the structure and detailed functions of these DosR regulated genes are largely undetermined. Out of many DosR regulon genes, Rv3131 gets up regulated in hypoxic conditions and was believed to encode for a nitroreductase flavoprotein. The utilization of mycobacteria-specific model systems has greatly added to our understanding of the molecular mechanisms involved in the life cycle and pathogenesis of Mtb. RESULTS: In this study the non-pathogenic mycobacterial model organism Mycobacterium smegmatis (Msmeg) was used to reveal the structure and function of MSMEG_3955; which is a homologue of Rv3131 from Mtb. Using chromatography and spectroscopy techniques it was revealed that cofactor flavin mononucleotide (FMN) was bound to flavoprotein MSMEG_3955. Consistent with the homology modelling predictions, Circular Dichroism (CD) analysis indicated that the MSMEG_3955 is composed of 39.3% α-helix and 24.9% ß-pleated sheets. In contrast to the current notions, the enzymatic assays performed in the present study revealed that MSMEG_3955 was not capable of reducing nitro substrates but showed NADPH dependent FMN oxidoreductase activity. Also, gel permeation chromatography, dynamic light scattering and native acidic gels showed that MSMEG_3955 exists as a homotrimer. Furthermore, the presence of NADPH dependent FMN oxidoreductase and homotrimeric existence could be an alternative function of the protein to help the bacteria survive in dormant state or may be involved in other biochemical pathways. CONCLUSION: MSMEG_3955 is a FMN bound flavoprotein, which exits as a trimer under in vitro conditions. There is no disulphide linkages in between the three protomers of the homotrimer MSMEG_3955. It has a NADPH dependent FMN oxidoreductase activity.

Genomic markers for the biological responses of Triclosan stressed hatchlings of Labeo rohita.

Triclosan (TCS) used commonly in pharmaceuticals and personal care products has become the most common pollutant in water. Three-day-old hatchlings of an indigenous fish, Labeo rohita, were given 96h exposure to a nonlethal (60 µg L-1) and two moderately lethal concentrations (67 and 97 µg L-1) of TCS and kept for 10 days of recovery for recording transcriptomic alterations in antioxidant/detoxification (SOD, GST, CAT, GPx, GR, CYP1a and CYP3a), metabolic (LDH, ALT and AST) and neurological (AchE) genes and DNA damage. The data were subjected to principal component analysis (PCA) for obtaining biomarkers for the toxicity of TCS. Hatchlings were highly sensitive to TCS (96h LC50 = 126 µg L-1 and risk quotient = 40.95), 96h exposure caused significant induction of CYP3a, AChE and ALT but suppression of all other genes. However, expression of all the genes increased significantly (except for a significant decline in ALT) after recovery. Concentration-dependent increase was also observed in DNA damage [Tail Length (TL), Tail Moment (TM), Olive Tail Moment (OTM) and Percent Tail DNA (TDNA)] after 96 h. The damage declined significantly over 96h values at 60 and 67 µg L-1 after recovery, but was still several times more than control. TCS elicited genomic alterations resulted in 5-11% mortality of exposed hatchlings during the recovery period. It is evident that hatchlings of L. rohita are a potential model and PCA shows that OTM, TL, TM, TDNA, SOD and GR (association with PC1 during exposure and recovery) are the biomarkers for the toxicity of TCS. Graphical abstract.

Effect of gallic acid on the larvae of Spodoptera litura and its parasitoid Bracon hebetor.

The antibiosis effect of gallic acid on Spodoptera litura F. (Lepidoptera: Noctuidae) and its parasitoid evaluated by feeding six days old larvae on artificial diet incorporated with different concentrations (5 ppm, 25 ppm, 125 ppm, 625 ppm, 3125 ppm) of the phenolic compound revealed higher concentration (LC50) of gallic acid had a negative impact on the survival and physiology of S. litura and its parasitoid Bracon hebetor (Say) (Hymenoptera:Braconidae). The mortality of S. litura larvae was increased whereas adult emergence declined with increasing concentration of gallic acid. The developmental period was delayed significantly and all the nutritional indices were reduced significantly with increase in concentration. Higher concentration (LC50) of gallic acid adversely affected egg hatching, larval mortality, adult emergence and total development period of B. hebetor. At lower concentration (LC30) the effect on B. hebetor adults and larvae was non-significant with respect to control. Gene expression for the enzymes viz., Superoxide dismutase, Glutathione peroxidase, Peroxidase, Esterases and Glutathione S transferases increased while the total hemocyte count of S. litura larvae decreased with treatment. Our findings suggest that gallic acid even at lower concentration (LC30) can impair the growth of S. litura larvae without causing any significant harm to its parasitoid B. hebetor and has immense potential to be used as biopesticides.

Exploration of insecticidal potential of Cry protein purified from Bacillus thuringiensis VIID1.

Insect pests are a threat to agriculture as they cause a loss of 15-22% to economically important crops every year. Bacillus thuringiensis produces parasporal crystal inclusions that have insecticidal 'Cry' proteins which are toxic to insect larvae of the order Lepidoptera, Coleoptera and Diptera, etc. In the present study, 40 different soil samples from Amritsar and its surrounding areas were selected for isolation of B. thuringiensis. The rod shaped, gram-positive bacterial isolates were further analyzed for characteristic crystal formation using phase contrast and scanning electron microscopy. 6 Bacillus samples containing cry genes were identified using the universal primers for cry genes, of which one isolate exhibited a protein band of ~95 kDa. This protein was purified using a Sephadex G-75 column. The insecticidal assays conducted with purified Cry protein on insect larvae of lepidopteran and dipteran orders viz. Spodoptera litura, Galleria malonella, Bactrocera cucurbitae and Culex pipens revealed considerable detrimental effects. A significant increase in larval mortality was observed for the larvae of all insects in a concentration dependent manner when treated with Cry protein purified from B. thuringenisis VIID1. The purified Cry protein did not have any significant effect on honey bee larvae.

5-aminolevulinic acid regulates Krebs cycle, antioxidative system and gene expression in Brassica juncea L. to confer tolerance against lead toxicity.

Heavy metal pollution seriously impairs crop production and poses serious concerns for human health. Exogenous application of biomolecules has been efficiently tested for enhancing plant resistance to metal toxicity. Current study evaluates the possible effect of 5-aminolevulinic acid (ALA) in Brassica juncea L. seedlings subjected to lead (Pb) stress. Our results showed that shoot length, root length and chlorophyll contents were significantly recovered in Pb stressed seedlings after ALA application, accompanied by reduction in the Pb accumulation. Significant reduction in the contents of reactive oxygen species (ROS) like superoxide anion, hydrogen peroxide and malondialdehyde were also observed in ALA treated seedlings under Pb stress. Furthermore, we also noticed enhancement in the activities of antioxidative enzymes like superoxide dismutase (SOD), catalase (CAT), guaiacol peroxidase (POD), glutathione reductase (GR), glutathione-S-transferase (GST) and dehydroascorbate reductase (DHAR). We further noticed that ALA upregulated the expression of SOD (7.30 folds), POD (6.11 folds), CAT (3.52 folds), DHAR (6.42 folds), GR (6.04 folds), and GST (5.58 folds) under the Pb stress. However, RBOH1 (gene involved in ROS generation) and CHLASE (chlorophyllase) expressions were reduced in ALA treated seedlings grown under Pb stress (RBOH1 expression decreased to 3.44 from 6.50 fold and CHLASE expression decreased to 2.97 from 5.58 fold). Phenolic contents were increased in the presence of ALA and expression of genes like CHS (chalcone synthase; 7.50 fold) and PAL (phenylalanine ammonia lyase; 4.77 fold) was also stimulated by ALA under Pb stress. Furthermore, contents of the Krebs cycle metabolites (fumarate, succinate, malate and citrate) were also enhanced accompanied by upregulated expression of genes like CS (citrate Synthase; 8.13 fold), SUCLG1 (succinyl CoA ligase 1; 7.40 fold), SDH (succinate dehydrogenase; 5.10 fold) and FH (fumarate hydratase; 5.65 fold). In conclusion, current investigation revealed that ALA attenuated Pb toxicity by modulating the transcription patterns of key enzymes involved in plant defense system.

Development of mitochondrial replacement therapy: A review.

Mitochondrial replacement therapy (MRT) is a new form of reproductive invitro fertilization (IVF) which works on the principle of replacing a women's abnormal mitochondrial DNA (mt-DNA) with the donor's healthy one. MRT include different techniques like spindles transfer (ST), pronuclear transfer (PNT) or polar body transfer (PBT). Transmission of defective mitochondrial DNA to the next generation can also be prevented by using these approaches. The development of healthy baby free from genetic disorders and to terminate the lethal mitochondrial disorders are the chief motive of this technique. In aged individuals, through in vitro fertilization, MRT provides the substitution of defective cytoplasm with cured one to enhance the expectation of pregnancy rates. However, moral, social, and cultural objections have restricted its exploration. Therefore, this review summarizes the various methods involved in MRT, its global status, its exaggerated censure over the years which depicts a strong emphasis for social acceptance and clinical application in the world of medical science.

Deterioration of digestive physiology of Bactrocera cucurbitae larvae by trypsin inhibitor purified from seeds of Mucuna pruriens.

Peptidase inhibitors (PIs) are plant proteins that are found to be effective against various digestive peptidases of insects. The present study isolated and characterized a trypsin inhibitor from mature dry seeds of Mucuna pruriens and investigated its effect against Bactrocera cucurbitae larvae, a major pest of cucurbitaceae crops, for its inhibitory activity. The purified trypsin inhibitor from M. pruriens seeds gave a molecular weight of ~11 kDa on SDS-PAGE. M. pruriens trypsin inhibitor (MPTI) exhibited inhibitory effect on growth of melon fruit fly larvae (64-72 h old) as it resulted in prolongation of larval, pupal and total development period. There was a significant increase in larval mortality with increase in concentration of MPTI. Nutritional indices decreased significantly at all the concentrations of MPTI. Quantitative RT- PCR revealed that the mRNA expression level of trypsin and chymotrypsin genes was reduced while that of GST, esterases, AP, SOD and catalase were enhanced. It can therefore be inferred that MPTI can serve as a promising agent for biocontrol that can reduce the problems caused by fruit fly and other similar catastrophic pests. This study provides the fundamental information for future successful strategies for pest management.

Antimicrobial Colloidal Complexes of Lysozyme with Bio-Based Surface Active Ionic Liquids in Aqueous Medium.

Bio-based surface-active ionic liquids (SAILs) have been synthesized and investigated for their complexation with lysozyme (LYZ) in an aqueous medium to develop antimicrobial SAIL-LYZ colloidal complexes. The synthesized SAILs, [Cho][Sar] and [Cho][Doc], are comprised of choline ([Cho]+) and lauryl sarcosinate ([Sar]-) or deoxycholate ([Doc]-). The constituent anions of the investigated SAILs are structurally dissimilar and thus resulted in contrasting complexation behavior toward LYZ, as suggested by the results obtained from different techniques. The interfacial behavior is monitored using tensiometry. Zeta-potential, turbidity, and dynamic light scattering results provide insights into the complexation phenomenon in bulk. The observations made from fluorescence and circular dichroism (CD) spectroscopy give information about the alterations in the inherent structure of LYZ. The thermodynamics of the binding of SAILs with LYZ is monitored using isothermal titration calorimetry (ITC). Computer simulations have been utilized to determine the preferential binding site of SAILs on LYZ, which supports the results obtained from different techniques. Interestingly, LYZ complexed with the investigated SAILs, which are non-antimicrobial, is found to exhibit enhanced antimicrobial activity depending upon the concentration regime of the used SAIL. In this way, we have developed new antimicrobial colloidal complexes of SAILs and LYZ. The present study provides useful insights to synthesize new bio-based SAILs to be utilized for creating colloidal formulations applicable in enzyme/protein stabilization, storage, and other biomedical applications.

Enhanced oral bioavailability and anti-diabetic activity of canagliflozin through a spray dried lipid based oral delivery: a novel paradigm.

AIM: Canagliflozin (CFZ), a novel SGLT II antagonist, exhibits erratic absorption after oral administration. The current study entails development and evaluation of spray dried lipid based formulation (solid SMEDDS) for enhancing oral bioavailability and anti-diabetic activity of CFZ. METHODS: Solid SMEDDS developed through spray drying containing Neusilin US2 as an adsorbent. The formed solid SMEDDS were characterized for physicochemical and solid state attributes. Scanning Electron Microscopy (SEM) and Transmission Electron Microscopy (TEM) were used to confirm the spherical morphology. In vitro dissolution, ex vivo permeability and in vivo pharmacokinetic studies were conducted to determine the release rate, permeation rate and absorption profile of CFZ, respectively. Pharmacodynamic studies were done as per standard protocols. RESULTS: The optimized solid SMEDDS exhibited acceptable practical yield and flow properties and is vouched with enhanced amorphization, nanoparticulate distribution and acceptable drug content. The spherical morphology of solid SMEDDS and reconstituted SMEDDS were confirmed in SEM and TEM, respectively. In vitro dissolution studies revealed multi-fold release behavior in CFZ in various dissolution media, whereas, remarkable permeability was observed in jejunum segment of rat intestine. Pharmacokinetic studies of CFZ in solid SMEDDS demonstrated 2.53 and 1.43 fold enhancement in Cmax and 2.73 and 1.98 fold in AUC 0-24h, as compared to pure API and marketed formulation, respectively. Pharmacological evaluation of solid SMEDDS revealed enhanced anti-diabetic activity of CFZ through predominant SGLT II inhibition in rats, as evident from evaluation of biochemical levels, urinary glucose excretion studies and SGLT II expression analysis. CONCLUSION: The current work describes significant improvement biopharmaceutical properties of CFZ in solid SMEDD formulation. Graphical abstract Graphical Abstract: Enhanced oral bioavailability and anti-diabetic activity of canagliflozin through a spray dried lipid based oral delivery: a novel paradigm.

Molecular characterization and antimicrobial susceptibility of bacterial isolates present in tap water of public toilets.

BACKGROUND: The present study was carried out to investigate the tap water quality of public toilets in Amritsar, Punjab, India. METHODS: Water samples from the taps of the public toilets were collected in sterile containers and physicochemical and bacteriological analysis was performed using standard methods. Also, genotypic and phenotypic characterization of the bacterial isolates was performed using different biochemical tests and 16S ribosomal RNA analysis. An antibiotic susceptibility test was performed using antibiotics based on their mode of action. A biofilm assay was performed to assess the adhesion potential of the isolates. RESULTS: A total of 25 bacterial isolates were identified from the water samples, including Acinetobacter junii, Acinetobacter pittii, Acinetobacter haemolyticus, Bacillus pumilus, Bacillus megaterium, Bacillus marisflavi, Bacillus flexus, Bacillus oceanisediminis, Pseudomonas otitidis, Pseudomonas sp. RR013, Pseudomonas sp. RR021, Pseudomonas sp. RR022, Escherichia coli and Enterobacter cloacae. The results of the antimicrobial susceptibility test revealed that the antibiotics cefodroxil, aztreonam, nitrofurantoin, cefepime, ceftazidime and amoxyclav were found to be mostly ineffective against various isolates. The biofilm assay revealed the weak, moderate and strong biofilm producers among them. CONCLUSIONS: The tap water in the public toilets was microbially contaminated and needs to be monitored carefully. The antibiotic susceptibility profile showed that of 25 bacterial isolates, 5 were multidrug resistant. Bacterial isolates exhibited strong to weak adhesion potential in the biofilm assay.

Molecular identification and phylogenetic relationship of Demodex mites based on mitochondrial 16S rDNA.

BACKGROUND AND OBJECTIVE: Demodex mites are tiny parasites that live around hair follicles of mammals. The two main species of Demodex i.e. Demodex folliculorum and Demodex brevis present in humans are found near the hair follicles of eyes. The present study was to understand the presence of Demodex mites in people suffering from blepharitis in Amritsar, Punjab. MATERIAL AND METHODS: Demodex mites samples present in blepharitis patients were isolated from the eyelashes. DNA was isolated from three mites and used for PCR amplification of mitochondrial (mt) 16S rDNA. The amplified PCR product were purified and used for molecular identification. RESULTS: The amplified mt16s rDNA product was sequenced and subjected to BLAST search in the NCBI database for molecular identification. The identified mite belongs to Demodex folliculorum species. The phylogenetic tree constructed by using mt16s rDNA sequence suggests that D. folliculorum is closer to D. canis than to D. brevis. CONCLUSION: All the three isolates belong to D. folliculorum and the mitochondrial DNA 16S rDNA partial sequence is applicable for phylogenetic relationship analysis.

Exploration of anti-insect potential of trypsin inhibitor purified from seeds of Sapindus mukorossi against Bactrocera cucurbitae.

Peptidase inhibitors (PIs) are defense proteins of plants which are active against gut peptidases of different insects. Sapindus mukorossi was identified as a source of bioactive PIs which could confer resistance against Bactrocera cucurbitae, a most devastating pest of several economically important crops. In the present study, a trypsin inhibitor was purified from mature dry seeds of S. mukorossi and characterized for its biochemical properties as well as its potential for bio control of B. cucurbitae. The purified fractions from RP- HPLC through SDS-PAGE gave an apparent molecular weight of ~29 kDa. S. mukorossi trypsin inhibitor (SMTI) was found to be a non-competitive inhibitor which was active over a broad range of temperature (10-100 °C) and pH (6-11). SMTI when incorporated in artificial diet inhibited the growth and development of B. cucurbitae larvae. Gene expression analysis of trypsin and chymotrypsin genes via qRT-PCR indicated that their mRNA expression was down-regulated while that of other genes namely, Catalase, Elastase, Superoxide Dismutase, Glutathione -S-transferase and Alkaline Phosphatase was up regulated. SMTI also showed deleterious effects against different bacterial strains. The results of this study indicated that S. mukorossi trypsin inhibitor has potential to be used as a bio control agent that can reduce the harm caused by melon fruit fly and other devastating pests.

Purification of a trypsin inhibitor from Psoralea corylifolia seeds and its influence on developmental physiology of Bactrocera cucurbitae.

A trypsin inhibitor was purified from the seeds of Psoralea corylifolia by ion-exchange and affinity chromatography. The purified fractions were subjected to RP- HPLC which resolved into a single peak. SDS-PAGE analysis gave an apparent molecular weight of 18 kDa. P. corylifolia trypsin inhibitor (PCTI) was found to be a competitive inhibitor and was active over a broad temperature (10-100 °C) and pH (6-11) range. It was shown to have a deleterious effect on growth and development of larvae of the melon fruit fly, Bactrocera cucurbitae, when incorporated in artificial diet using various concentrations. The larval, pupal, total development period and larval mortality significantly increased during the treatment. Inhibitory effects were also observed on percentage emergence which was significantly reduced. Nutritional indices namely food assimilated (FA) and mean relative growth rate (MRGR) also decreased significantly with increase in concentration of PCTI. qRT-PCR results indicated that the expression of trypsin and chymotrypsin genes were down-regulated while elastase, catalase, GST, SOD and AP were up-regulated. PCTI was also effective against certain bacterial strains. These results indicated that the peptidase inhibitor from P. corylifolia may be a potential bio-control agent which can decrease the damage caused by B. cucurbitae and other related destructive pests.

Characterization of a cold-active, detergent-stable metallopeptidase purified from Bacillus sp. S1DI 10 using Response Surface Methodology.

The colder regions of Earth are inhabited by cold-adapted microorganisms designated as psychrophiles that are known to produce cold-active enzymes, such as peptidases, chaperones, lipases, cellulases, and phosphatases. These types of enzymes are a major part of the market of industrial enzymes. Bacteria isolated from water samples collected from the Chamba region in the Himalayas were screened for peptidase production using skim milk agar plates. Among the peptidase-producing bacteria isolated, 20% of the isolates exhibited fast growth and maximum zones of clearance, and thus, were used for further studies. The 16S rDNA sequence analysis of isolate S1DI 10 identified it as a Bacillus sp. The peptidase was cloned in pET28a vector and expressed in Escherichia coli BL21(DE3) and the His-tagged recombinant protein was purified using Ni-NTA column. The purified peptidase of SIDI 10 was found to be an alkaline, cold-active peptidase with optimal enzyme activity at 10°C and pH 8. An approach of one variable at a time was used to further study the effect of various metal ions, organic solvents and detergents on the peptidase enzyme. The peptidase activity was enhanced in the presence of Fe2+ and Mn2+ (metal ions), hexane (organic solvent), SDS- sodium dodecyl sulfate (anionic detergent) and Tween 80 (nonionic detergent). Response surface methodology (RSM) was used to determine the cumulative effect of these five variables. A 25 full factorial central composite design was applied for the five independent variables to determine the optimal combinations of these constituents at the maximum peptidase activity.

24-epibrassinolide stimulates imidacloprid detoxification by modulating the gene expression of Brassica juncea L.

BACKGROUND: Pesticides cause oxidative stress to plants and their residues persist in plant parts, which are a major concern for the environment as well as human health. Brassinosteroids (BRs) are known to protect plants from abiotic stress conditions including pesticide toxicity. The present study demonstrated the effects of seed-soaking with 24-epibrassinolide (EBR) on physiological responses of 10-day old Brassica juncea seedlings grown under imidacloprid (IMI) toxicity. RESULTS: In the seedlings raised from EBR-treated seeds and grown under IMI toxicity, the contents of hydrogen peroxide (H2O2) and superoxide anion (O.2-) were decreased, accompanied by enhanced activities of superoxide dismutase (SOD), catalase (CAT), glutathione reductase (GR), glutathione-S-transferase (GST), guaiacol peroxidase (POD) and the content of glutathione (GSH). As compared to controls, the gene expressions of SOD, CAT, GR, POD, NADH (NADH-ubiquinone oxidoreductase), CXE (carboxylesterase), GSH-S (glutathione synthase), GSH-T (glutathione transporter-1), P450 (cytochrome P450 monooxygenase) and GST1-3,5-6 were enhanced in the seedlings raised from EBR-treated seeds and grown in IMI supplemented substratum. However, expression of RBO (respiratory burst oxidase, the gene responsible for H2O2 production) was decreased in seedlings raised from EBR treated seeds and grown under IMI toxicity. Further, the EBR seed treatment decreased IMI residues by more than 38% in B. juncea seedlings. CONCLUSIONS: The present study revealed that EBR seed soaking can efficiently reduce oxidative stress and IMI residues by modulating the gene expression of B. juncea under IMI stress. In conclusion, exogenous EBR application can protect plants from pesticide phytotoxicity.