A rice glutaredoxin regulate the expression of aquaporin genes and modulate root responses to provide arsenic tolerance.

Rice is the most consumed food crop and essential determinant in global food security program. Currently, arsenic (As) accumulation in rice is a critical concern in terms of both crop productivity and grain quality; therefore, it is an urgent need to reduce As accumulation. Here, we selected a glutaredoxin (OsGrx_C7) gene that plays an essential role in AsIII tolerance in rice. To explore the mechanism, we raised OsGrx_C7 overexpression (OE) rice lines, which showed improved plant AsIII tolerance and lowered its accumulation in grains. Arsenic accumulation in husk, unpolished, and polished rice reduced by ca. 65%, 67%, and 85%, respectively, in OE lines, compared to wild-type (WT) plants. To know the rationale, expression of AsIII transporters (aquaporins) in root and shoot tissues were examined, and revealed that OsGrx_C7 regulates the expression of these genes, which ultimately reduces root to shoot AsIII translocation. Additionally, OsGrx_C7 improves root growth by regulating the expression of oxidative stress-induced root expansion related genes, promote root growth and plant health. Overall, current study suggested that AsIII induced OsGrx_C7 markedly enhanced tolerance to AsIII with reduced accumulation in grains by regulating root expansion and controlling root to shoot As transport by altered expression of AsIII aquaporins.

Response of two rice cultivars differing in their sensitivity towards arsenic, differs in their expression of glutaredoxin and glutathione S transferase genes and antioxidant usage.

Embodied study investigates the role of GRX and associated antioxidant enzymes in the detoxification mechanism between arsenic (As) sensitive (Usar-3) and tolerant cultivar (Pant Dhan 11) of Oryza sativa against As(III) and As(V), under GSH enriched, and GSH deprived conditions. The overall growth and physiological parameters in sensitive cultivar were lower than the tolerant cultivar, against various treatments of As(III) and As(V). The As accumulation in sensitive cv. against both As(III) and As(V) was lower than the corresponding treatments in tolerant cv. However, the As translocation against As(V) was lower (35% and 64%, resp.) than that of As(III), in both the cultivars. In sensitive cv. translocation of Zn and Cu was influenced by both As(V) and As(III) whereas, in tolerant cv. the translocation of Cu, Mn and Zn was influenced only by As(III). Translocation of Fe was negatively influenced by translocation of As in sensitive cv. and positively in tolerant cv. Strong correlation between H2O2, SOD, GRX, GR, GST and GSH/GSSG in sensitive cv. and between DHAR, APX, MDHAR and AsA in tolerant cv. demonstrates the underlying preference of GSH as electron donor for detoxification of H2O2 in sensitive cv. and AsA in tolerant cv. Higher expression of the four GRX and two GST genes in the sensitive cv. than tolerant cv, suggests that under As stress, GRX are synthesized more in the sensitive cv. than tolerant cv. Also, the expression of four GRX genes were higher against As(V) than As(III). The higher As accumulation in the tolerant cv. is due to lower GST expression, is attributed to the absence of thiolation and sequestration of As in roots, the translocation of As to shoots is higher.

Temperature differences between sites lead to altered phenylpropanoid metabolism in a varietal dependent manner.

Elevated temperature has already caused a significant loss of wine growing areas and resulted in inferior fruit quality, particularly in arid and semi-arid regions. The existence of broad genetic diversity in V. vinifera is key in adapting viticulture to climate change; however, a lack of understanding on the variability in berry metabolic response to climate change remains a major challenge to build ad-hoc strategies for quality fruit production. In the present study, we examined the impact of a consistent temperature difference between two vineyards on polyphenol metabolism in the berries of 20 red V. vinifera cultivars across three consecutive seasons (2017-2019). The results emphasize a varietal specific response in the content of several phenylpropanoid metabolites; the interaction factor between the variety and the vineyard location was also found significant. Higher seasonal temperatures were coupled with lower flavonol and anthocyanin contents, but such reductions were not related with the level of expression of phenylpropanoid related genes. Hierarchical clustering analyses of the metabolic data revealed varieties with a location specific response, exceptional among them was Tempranillo, suggesting a greater susceptibility to temperature of this cultivar. In conclusion, our results indicate that the extensive genetic capacity of V. vinifera bears a significant potential to withstand temperature increase associated with climate change.

Assessment of supernatural attitude toward mental health among tribal and non-tribal populations.

Background: In India, only about 10% of persons with psychiatric illnesses receive evidence-based treatments. Stigma, mistrust in allopathic medicine, allegiance toward faith healers, cultural and traditional practices, and beliefs are some of the major hindrances to the treatment of mentally ill patients. Aim: To assess supernatural attitudes toward mental health among tribal and non-tribal populations. Materials and Methods: A total of 100 subjects were included in the study after informed consent, 50 of whom were tribal and 50 non-tribal. All were required to fill a self-constructed proforma, the Supernatural Attitude Questionnaire (SAQ), and the Attitudes Toward Seeking Professional Psychological Help Scale-Short Form (ATSPPH-SF). The scales were scored, compiled, and statistically analyzed. Results: The tribal and non-tribal subjects were well-matched on socio-demographic variables. The score of SAQ was significantly higher in the tribal group as compared to the non-tribal group. On the ATSPPH-SF, the scores of the two groups were not significant. Conclusion: Even though the tribal populations of India hold a higher supernatural attitude toward mental illnesses as compared to the non-tribal populations, their treatment-seeking attitude remains unaffected.

Influence of late pruning practice on two red skin grapevine cultivars in a semi-desert climate.

Continually increasing global temperature could severely affect grape berry metabolite accumulation and ultimately wine polyphenol concentration and color intensity. To explore the effect of late shoot pruning on grape berry and wine metabolite composition, field trials were carried out on Vitis vinifera cv. Malbec and cv. Syrah grafted on 110 Richter rootstock. Fifty-one metabolites were detected and unequivocally annotated employing UPLC-MS based metabolite profiling. Integrating the data using hierarchical clustering showed a significant effect of late pruning treatments on must and wine metabolites. Syrah metabolite profiles were characterized by a general trend of higher metabolite content in the late shoot pruning treatments, while Malbec profiles did not show a consistent trend. In summary, late shoot pruning exerts a significant effect, though varietal specific, on must and wine quality-related metabolites, possibly related to enhanced photosynthetic efficiency, which should be taken into consideration when planning mitigating strategies in warm climates.

Genome-wide identification of rice class I metallothionein gene: tissue expression patterns and induction in response to heavy metal stress.

Metallothioneins (MTs) are members of a family of cysteine-rich low molecular weight polypeptides which play an important role in heavy metal detoxification and homeostasis of intracellular metal ions in plant. Though MT genes from some selected plants have been characterized with respect to their protein sequences, kinetic properties and tissue-specific localization, no detailed study has been carried out in rice. Here, we present genome-wide identification, structural and expression analyses of rice MT gene family. Our analysis suggests presence of 11 class I MT genes in rice genome (Release 7 of the MSU Rice Genome Annotation Project) which are differentially expressed during growth and development, in various tissues and during biotic and abiotic stresses. Our analyses suggest that class I MT proteins in rice differ in tissue localization as well as in heavy metal coordination chemistry. We also suggest that some MTs have a predominant role in detoxification of As (V) in arsenic-tolerant rice cultivars. Our analysis suggests that apart from transcriptional regulation, post-transcriptional alternative splicing in some members of this family takes place during growth and development, in various tissues and during biotic and abiotic stresses.

Root system architecture in rice: impacts of genes, phytohormones and root microbiota.

To feed the continuously expanding world's population, new crop varieties have been generated, which significantly contribute to the world's food security. However, the growth of these improved plant varieties relies primarily on synthetic fertilizers, which negatively affect the environment and human health; therefore, continuous improvement is needed for sustainable agriculture. Several plants, including cereal crops, have the adaptive capability to combat adverse environmental changes by altering physiological and molecular mechanisms and modifying their root system to improve nutrient uptake efficiency. These plants operate distinct pathways at various developmental stages to optimally establish their root system. These processes include changes in the expression profile of genes, changes in phytohormone level, and microbiome-induced root system architecture (RSA) modification. Several studies have been performed to understand microbial colonization and their involvement in RSA improvement through changes in phytohormone and transcriptomic levels. This review highlights the impact of genes, phytohormones, and particularly root microbiota in influencing RSA and provides new insights resulting from recent studies on rice root as a model system and summarizes the current knowledge about biochemical and central molecular mechanisms.

Evaluation of Clinical Profile and Various Treatment Modalities in Oral Sub Mucous Fibrosis in North India: Our Experience.

Oral submucous fibrosis is a chronic disease affecting oral cavity and sometimes the pharynx. Etiology seems to be local irritants such as capsaicin, tobacco, areca nut and spicy foods. The main concern in this is the management of trismus and burning sensation of the oral mucosa. The aim of this study was to compare various medical treatment protocol of OSMF. 210 patients were divided randomly in 3 groups. In Group A, patients were given biweekly intralesional Hyaluronidase/Dexamethasone for 6 weeks. Group B patients were given tablet Pentoxifylline 400 mg TDS.Group C patients were given Eprisone hydrochloride. All three groups were given Lycopene 10,000 mcg for period of 6 weeks. All patients were given topical Triamcelone for local application. The examinations for mouth opening were repeated at weekly intervals for a period of 6 weeks.The most common complaint was burning sensation in 75.98% cases, difficulty in mouth opening in77.45% and difficulty in swallowing food in 61.76% cases. Group A showed improvement in 41.17% cases presenting with burning sensation followed by decreased mouth opening 39.70%. Group B showed improvement in 45.58% burning sensation, 17.64% with decreased mouth opening. Group C showed improvement in 48.52% patients having pain with spicy food, 32.35% with decreased mouth opening and 17.64% with difficulty in swallowing. We conclude that patients which received intralesional dexamethasone and hyaluronidase along with oral Lycopene showed better clinical and symptomatic improvement, and at present appears to be best non-surgical treatment.

Fosfomycin versus Nitrofurantoin for the Treatment of Lower UTI in Outpatients.

Introduction Fosfomycin and nitrofurantoin are increasingly being prescribed in outpatients for the oral treatment of urinary tract infection (UTI). Although ample literature is available on the in vitro sensitivity pattern of fosfomycin and nitrofurantoin in UTI cases, clinical data are scant. Methodology Voided midstream urine, collected from patients ≥ 16 years of age of both genders with suspected sign and symptoms, was plated on cystine lactose electrolyte-deficient agar. Uropathogen was defined as an organism known to be associated with the signs and symptoms of UTI with > 10 5 colony forming units/mL of urine. Antimicrobial susceptibility testing was determined by Kirby-Bauer disc diffusion method. Further, for fosfomycin, agar dilution method was also performed. Results A total of 143 patients, 47 treated with fosfomycin and 96 with nitrofurantoin, were followed for clinical outcome. The most common isolated uropathogen was Escherichia coli . In vitro susceptibility rate of uropathogens against fosfomycin and nitrofurantoin was 99.3% and 81.2%, respectively. Overall, the clinical cure rate with fosfomycin and nitrofurantoin treatment groups was 80.85% and 90.06% respectively (not statistically significant). Conclusion Fosfomycin and nitrofurantoin showed good in vitro activity against uropathogens from lower UTI and can be used for empirical therapy in our area. Multiple confounding factors may have contributed to the discrepancy between in vitro susceptibility and clinical cure, which needs to be studied further.

CC-type glutaredoxin, OsGrx_C7 plays a crucial role in enhancing protection against salt stress in rice.

Soil salinity is one of the critical issue worldwide that adversely affect soil fertility. Salt stress significantly limits crop yield and grain quality; therefore, there is an urgent need to develop a strategy to improve salt stress tolerance. In present study, we reported that rice glutaredoxin (OsGrx_C7) plays a positive response in salt induced stress. Gene expression analysis, silencing, and overexpression of OsGrx_C7 gene were used to discover the role of OsGrx_C7 in response to salt stress. Gene expression analysis suggested that OsGrx_C7 expression was induced under salt stress and ubiquitously expressed in rice including root and shoot. The silencing of osgrx_c7 gene leads to increased sensitivity to salt stress, indicating its importance in salt stress tolerance. A gain-of-function approach showed that OsGrx_C7 may act as an important determinant in salt stress, compared with WT, and revealed higher biomass accumulation, improved root and plant growth under salt stress. Under salt stress condition, OsGrx_C7 overexpressing rice plants showed lower level of lipid peroxidation and Na+/K+ ratio, while proline accumulation, soluble sugar content and GSH/GSSG ratio was higher compared to WT. Furthermore, expression analysis suggested that OsGrx_C7 acted as positive regulator of salt tolerance by reinforcing the expression of transporters (OsHKT2;1, OsHKT1;5 and OsSOS1) engaged in Na+ homeostasis in overexpressing plants. Overall our study revealed that OsGrx_C7 emerged as a key mediator in response to salt stress in rice and could be used for engineering tolerance against salt stress in rice and other crops.

Hospital dependency: an existing concept.

Chronically institutionalized patients demonstrate a behavioral pattern characterized by unwillingness to get discharged from the hospital and severe anxiety during such attempts leading to continued hospitalization. These patients usually have poor self-concept and low evaluation in getting employment, which requires vocational rehabilitation. To prevent this, mental health professionals should make active efforts to develop a favorable attitude of the chronic mentally ill patients towards their home.

Family psychoeducation with caregivers of schizophrenia patients: Impact on perceived quality of life.

BACKGROUND: Schizophrenia is a devastating and chronic mental illness. Considering the nature of illness along with routine psychiatric care, various supportive therapies are recommended. Family psychoeducational approach has been developed to increase patients' as well as their caregivers' knowledge and insight into their illness. It is postulated that this increased knowledge and insight will enable people with schizophrenia and their caregivers to cope in a more effective way with the consequences of their illness, thereby improving prognosis. AIM: The aim of this study is to assess the efficacy of family psychoeducation intervention on the caregivers of schizophrenia patients with respect to their perceived quality of life. MATERIALS AND METHODS: A total of 30 caregivers of male schizophrenia patients were selected through purposive sampling technique. Patients were divided into two groups, namely, experimental and control group. Family psychoeducation intervention was given on twice-monthly basis for 6 months to the experimental group caregivers. Baseline assessment was carried out with the help of WHO Quality of Life-BREF followed by intervention and then posttherapeutic assessment was done with same tool. RESULTS: Most of the caregivers were above 40 years of age group, married, and male. Following family psychoeducation, significant improvement in overall quality of life scores was observed in experimental group caregivers compared to control group caregivers where no such intervention was provided. CONCLUSION: Family psychoeducation is feasible and useful in our clinical population.

Vancomycin, linezolid and daptomycin susceptibility pattern among clinical isolates of methicillin-resistant Staphylococcus aureus (MRSA) from Sub- Himalyan Center.

INTRODUCTION: The efficacy of vancomycin, drug of choice for methicillin-resistant Staphylococcus aureus (MRSA), has become questionable due to the emergence of MRSA isolates with reduced susceptibility. The present study was conducted to determine the vancomycin, linezolid, and daptomycin susceptibility pattern in clinical isolates of MRSA and to observe minimum inhibitory concentration (MIC) creep over 2 years if any. MATERIALS AND METHODS: MIC of vancomycin, linezolid, and daptomycin were determined by E-test in 198 MRSA isolates and their MIC 50, MIC 90, and geometric mean MIC were calculated. RESULTS: While all isolates were sensitive to vancomycin, linezolid, and daptomycin, MIC 90 of vancomycin increased from 1.5 µg/ml in 2015 to 2 µg/ml in 2016. The percentage of isolates with vancomycin MIC >2 µg/ml doubled in 2016 (12.9%) as compared to 2015 (6.1%). MIC 90 for linezolid remained steady as 3 µg/ml, but geometric mean MIC increased from 2.20 µg/ml in 2015 to 2.29 µg/ml in 2016, and more than 40% isolates showed MIC 3 µg/ml. MIC 90 and geometric mean MIC of daptomycin decreased from 0.75 µg/ml to 0.5 µg/ml and 0.50 µg/ml to 0.36 µg/ml in 2015 and 2016, respectively. CONCLUSION: MIC creep was observed with vancomycin. Although linezolid MIC was within the susceptible zone, more than 40% strains showing MIC 3 µg/ml may herald the future development of either resistant or heteroresistant. Daptomycin showed good sensitivity against MRSA isolates. Therefore, it could be considered as an alternative agent for the treatment of infections caused by MRSA. However, it should be reserved where this class has a clear therapeutic advantage over other anti-MRSA drugs.

A novel fungal arsenic methyltransferase, WaarsM reduces grain arsenic accumulation in transgenic rice (Oryza sativa L.).

Rice (Oryza sativa L.) grown on arsenic-containing soil and water become a primary dietary source of arsenic and pose a significant health risk. Gene modification is an important and practical approach to reduce arsenic accumulation in rice grains. Here, we reported a WaarsM gene of soil fungus Westerdykella aurantiaca, expressed in rice able to convert toxic inorganic arsenicals to methylated arsenic species, therefore, reduce arsenic accumulation in rice grains. In response to arsenic treatment in hydroponics, WaarsM expressing transgenic lines showed a marked increase in arsenic resistance and reduces its accumulation compared to NT. Also, WaarsM expressing transgenic Line 1 evolved ca. 157ng and ca. 43ng volatile arsenicals (mg-1 fresh weight) after 72h of exposure to 25µM AsIII and 250µM AsV, respectively. Transgenic Line 1, grown in soil irrigated with arsenic-containing water accumulates about 50% and 52% lower arsenic than NT in shoot and root, respectively; while arsenic concentration in polished seeds and husk of the transgenic line was reduced by 52% compared to NT. Thus, the present study demonstrates that the expression of WaarsM in rice induces arsenic methylation and volatilization, provides a potential strategy to reduce arsenic accumulation in rice grain.

A novel arsenic methyltransferase gene of Westerdykella aurantiaca isolated from arsenic contaminated soil: phylogenetic, physiological, and biochemical studies and its role in arsenic bioremediation.

Elevated arsenic concentration in the environment and agricultural soil is a serious concern to crop production and human health. Among different detoxification mechanisms, the methylation of arsenic is a widespread phenomenon in nature. A number of microorganisms are able to methylate arsenic, but less is known about the arsenic metabolism in fungi. We identified a novel arsenic methyltransferase (WaarsM) gene from a soil fungus, Westerdykella aurantiaca. WaarsM showed sequence homology with all known arsenic methyltransferases having three conserved SAM binding motifs. The expression of WaarsM enhanced arsenic resistance in E. coli (Δars) and S. cerevisiae (Δacr2) strains by biomethylation and required endogenous reductants, preferably GSH, for methyltransferase activity. The purified WaarsM catalyzes the production of methylated arsenicals from both AsIII and AsV, and also displays AsV reductase activity. It displayed higher methyltransferase activity and lower KM 0.1945 ± 0.021 mM and KM 0.4034 ± 0.078 mM for AsIII and AsV, respectively. S. cerevisiae (Δacr2) cells expressing WaarsM produced 2.2 ppm volatile arsenic and 0.64 ppm DMA(v) with 0.58 ppm volatile arsenicals when exposed to 20 ppm AsV and 2 ppm AsIII, respectively. Arsenic tolerance in rice after co-culture with genetically engineered yeast suggested its potential role in arsenic bioremediation. Thus, characterization of WaarsM provides a potential strategy to reduce arsenic concentration in soil with reduced arsenic accumulation in crops grown in arsenic contaminated areas, and thereby alleviating human health risks.

Overexpression of rice glutaredoxins (OsGrxs) significantly reduces arsenite accumulation by maintaining glutathione pool and modulating aquaporins in yeast.

Arsenic (As) is an acute poison and class I carcinogen, can cause a serious health risk. Staple crops like rice are the primary source of As contamination in human food. Rice grown on As contaminated areas accumulates higher As in their edible parts. Based on our previous transcriptome data, two rice glutaredoxins (OsGrx_C7 and OsGrx_C2.1) were identified that showed up-regulated expression during As stress. Here, we report OsGrx_C7 and OsGrx_C2.1 from rice involved in the regulation of intracellular arsenite (AsIII). To elucidate the mechanism of OsGrx mediated As tolerance, both OsGrxs were cloned and expressed in Escherichia coli (Δars) and Saccharomyces cerevisiae mutant strains (Δycf1, Δacr3). The expression of OsGrxs increased As tolerance in E. coli (Δars) mutant strain (up to 4 mM AsV and up to 0.6 mM AsIII). During AsIII exposure, S. cerevisiae (Δacr3) harboring OsGrx_C7 and OsGrx_C2.1 have lower intracellular AsIII accumulation (up to 30.43% and 24.90%, respectively), compared to vector control. Arsenic accumulation in As-sensitive S. cerevisiae mutant (Δycf1) also reduced significantly on exposure to inorganic As. The expression of OsGrxs in yeast maintained intracellular GSH pool and increased extracellular GSH concentration. Purified OsGrxs displays in vitro GSH-disulfide oxidoreductase, glutathione reductase and arsenate reductase activities. Also, both OsGrxs are involved in AsIII extrusion by altering the Fps1 transcripts in yeast and protect the cell by maintaining cellular GSH pool. Thus, our results strongly suggest that OsGrxs play a crucial role in the maintenance of the intracellular GSH pool and redox status of the cell during both AsV and AsIII stress and might be involved in regulating intracellular AsIII levels by modulation of aquaporin expression and functions.

Transformed yeast (Schizosaccharomyces pombe) overexpressing rice Tau class glutathione S-transferase (OsGSTU30 and OsGSTU41) shows enhanced resistance to hexavalent chromium.

Extensive use of hexavalent chromium [Cr(VI)] in leather tanning, stainless-steel production, wood preservatives and electroplating industries has resulted in widespread environmental pollution and poses a serious threat to human health. A plant's response to Cr(VI) stress results in growth inhibition and toxicity leading to changes in components of antioxidant systems. In a previous study, we observed that a large number of glutathione S-transferase (GST) genes were up-regulated under Cr(VI) stress in rice. In this study, two rice root-specific Tau class GST genes (OsGSTU30 and OsGSTU41) were introduced into yeast (Schizosaccharomyces pombe). Transformed yeast cells overexpressing OsGSTU30 and OsGSTU41 had normal growth, but had much higher levels of GST activities and showed enhanced resistance to Cr(VI) as compared to control cells (transformed with empty vector). Also, a higher accumulation of chromium was found in the transformed yeast cells as compared to the control cells. Manipulation of glutathione biosynthesis by exogenous application of buthionine sulfoximine abolishes the protective effect of OsGSTs against Cr(VI) stress. These results suggest that Tau class OsGSTs play a significant role in detoxification of Cr(VI), probably by chelating and sequestrating glutathione-Cr(VI) complexes into vacuoles.

Detection of Different ß-Lactamases and their Co-existence by Using Various Discs Combination Methods in Clinical Isolates of Enterobacteriaceae and Pseudomonas spp.

BACKGROUND: Resistance to broad spectrum beta-lactams mediated by extended spectrum ß-lactamase (ESBL), AmpC, and metallobetalactamase (MBLs) enzymes are an increasing problem worldwide. The study was aimed to detect occurrence rate and to evaluate different substrates and inhibitors by disc combination method for detecting varying degree of ß-lactamase enzymes and their co-production. MATERIALS AND METHODS: A disc panel containing imipenem (IMP), IMP/EDTA, ceftazidime (CA), ceftazidime-tazobactum (CAT), CAT/cloxacillin (CLOX), ceftazidime-clavulanic acid (CAC), CAC/CLOX, cefoxitin (CN), and CN/CLOX in a single plate was used to detect presence of ESBLs, AmpC, and MBLs and/or their co-existence in 184 consecutive, nonrepetitive, clinical isolates of Enterobacteriace (n = 96) and Pseudomonas spp. (n = 88) from pus samples of hospitalized patients, resistant to 3(rd) generation cephalosporins. RESULTS: Out of a total of 96 clinical isolates of Enterobacteriaceae, 18.7, 20.8, and 27% were pure ESBL, AmpC, and MBL producers, respectively. ESBL and AmpC were co-produced by 25% isolates. Among 88 Pseudomonas spp. 38.6, 13, and 6% were pure MBL, ESBL, and AmpC producers, respectively. ESBL/AmpC and MBL/AmpC co-production was seen in 20% and 18% isolates, respectively. Among ESBL and AmpC co-producers, CA/CAC/CLOX disc combination (DC) missed 7 of the 24 ESBL producers in Enterobacteriace and 4 of the 18 ESBL in Pseudomonas spp., which were detected by CA/CAT/CLOX DC. No mechanism was detected among 8.3% Enterobacteriaceae and 2.3% Pseudomonas isolates. CONCLUSION: Diagnostic problems posed by co-existence of different classes of ß-lactamases in a single isolate could be solved by disc combination method by using simple panel of discs containing CA, CAT, CAT/CLOX, IMP, and IMP/EDTA.

An improved Agrobacterium-mediated transformation of recalcitrant indica rice (Oryza sativa L.) cultivars.

Agrobacterium-mediated transformation of indica rice varieties has been quite difficult as these are recalcitrant to in vitro responses. In the present study, we established a high-efficiency Agrobacterium tumefaciens-mediated transformation system of rice (Oryza sativa L. ssp. indica) cv. IR-64, Lalat, and IET-4786. Agrobacterium strain EHA-101 harboring binary vector pIG121-Hm, containing a gene encoding for ß-glucuronidase (GUS) and hygromycin resistance, was used in the transformation experiments. Manipulation of different concentrations of acetosyringone, days of co-culture period, bacterial suspension of different optical densities (ODs), and the concentrations of L-cysteine in liquid followed by solid co-culture medium was done for establishing the protocol. Among the different co-culture periods, 5 days of co-culture with bacterial cells (OD600 nm = 0.5-0.8) promoted the highest frequency of transformation (83.04 %) in medium containing L-cysteine (400 mg l(-1)). Putative transformed plants were analyzed for the presence of a transgene through genomic PCR and GUS histochemical analyses. Our results also suggest that different cultural conditions and the addition of L-cysteine in the co-culture medium improve the Agrobacterium-mediated transformation frequencies from an average of 12.82 % to 33.33 % in different indica rice cultivars.