CRISPR/Cas genome editing in plants: Dawn of Agrobacterium transformation for recalcitrant and transgene-free plants for future crop breeding.

Genome editing tools based on CRISPR/Cas system have been posed to solve many issues in agriculture and improve food production. Genetic engineering by Agrobacterium-mediated transformation has helped to impart specific traits straightaway in many crops. Many GM crops have also reached the field for commercial cultivation. Genetic engineering requires mostly a transformation protocol often mediated by Agrobacterium to insert a specific gene at a random locus. Genome editing with CRISPR/Cas system is a more precise technique for the targeted modification of genes/bases in the host plant genome. Unlike the conventional transformation system, wherein elimination of marker/foreign gene was possible only post-transformation, CRISPR/Cas system could generate transgene-free plants by delivering CRISPR/Cas reagents such as the Cas protein and guide RNAs gRNA(s) preassembled to form ribonucleoproteins (RNPs) into plant cells. CRISPR reagent delivery might be helpful to overcome issues with plants that are recalcitrant to Agrobacterium transformation and the legal hurdles due to the presence of the foreign gene. More recently, the grafting of wild-type shoots to transgenic donor rootstocks developed by the CRISPR/Cas system has reported transgene-free genome editing. CRISPR/Cas system also requires only a small piece of gRNA besides Cas9 or other effectors to target a specific region in the genome. So this system has been projected to be a key contributor to future crop breeding. In this article, we recap the main events of plant transformation, compare the difference between genetic transformation and CRISPR/Cas-mediated genome editing, and draw insights into the future application of the CRISPR/Cas system.

Hepatoprotective Constituents of Macrocybe gigantea (Agaricomycetes) from India.

Non-alcoholic fatty liver disease (NAFLD) is one of the most frequent, chronic liver diseases worldwide and currently has no specific therapy. Our previous study indicated the anti-NAFLD effect of Macrocybe gigantea (Massee) Pegler & Lodge in high-fat diet-fed animals. This study aimed to isolate and identify the active hepatoprotective constituents from M. gigantea using fatty acid induced steatotic HepG2 cells as in vitro model. The effect of the test materials on the viability of HepG2 cells was analyzed using MTT assay. The HepG2 cells were treated with a mixture of palmitate-oleate to induce steatosis; after 24 h of treatment with the test materials, the intracellular lipid content was estimated using Oil Red O staining. The levels of transaminases were also estimated in the spent media. Bioassay-guided isolation of hepatoprotective constituents from M. gigantea yielded two compounds viz., ergosterol and linoleic acid; their structures were confirmed using spectroscopic data. Among these two compounds, ergosterol significantly lowered the levels of intracellular triglyceride content of fatty acid induced HepG2 cells; it also lowered the leakage of transaminases. The reductions caused by linoleic acid were not statistically significant at the tested concentrations. Detailed investigations on efficacy and safety of these compounds and M. gigantea might yield some useful leads for the management of NAFLD.

In vitro growth inhibitory effect of selected 18ß-glycyrrhetinic acid esters on Theileriaannulata.

Glycyrrhetinic acid (GA) is one of the important Pentacyclic Triterpenoids (PT) found in the roots of licorice. This study aimed to evaluate the in vitro growth inhibitory effect of 18ß-GA (18ß-Glycyrrhetinic acid) and C-30 esters against Theileria annulata, the causative agent of Tropical Bovine Theileriosis. C-30 esters of 18ß-GA were synthesized and their structures were elucidated using spectroscopy. The pharmacodynamic properties of 18ß-GA and its C-30 esters were predicted using DataWarrior and Swiss ADME tools. Cattle isolates of T. annulata schizont-infected bovine lymphoblastoid cells were cultured using standard conditions and the growth inhibitory effect of GA and its esters were evaluated using MTT assay. The isopropyl ester of 18ß-GA (GI50- 1.638 µM; R2- 0.818) showed improved anti-theileriosis efficacy than other 18ß-GA derivatives. The propyl (GI50 - 5.549 µM), ethyl (GI50 - 5.638 µM), and benzyl (GI50 - 7.431 µM) esters also showed considerable inhibitory effect. The GI50 value for 18ß-GA was recorded as 6.829 µM. This study throws light on the usefulness of 18ß-GA and its esters for the treatment of Tropical Bovine Theileriosis.

Improving abiotic stress tolerance in sorghum: focus on the nutrient transporters and marker-assisted breeding.

MAIN CONCLUSION: Identification of molecular markers and characterization of nutrient transporters could help to improve the tolerance under abiotic and low nutrient stresses in sorghum ensuring higher yield to conserve food security Sorghum is an important cereal crop delivering food and energy security in the semi-arid tropics of the world. Adverse climatic conditions induced by global warming and low input agriculture system in developing countries demand for the improvement of sorghum to tolerate various abiotic stresses. In this review, we discuss the application of marker-assisted breeding and nutrient transporter characterization studies targeted towards improving the tolerance of sorghum under drought, salinity, cold, low phosphate and nitrogen stresses. Family members of some nutrient transporters such as nitrate transporter (NRT), phosphate transporter (PHT) and sulphate transporter (SULTR) were identified and characterized for improving the low nutrient stress tolerance in sorghum. Several quantitative trait loci (QTL) were identified for drought, salinity and cold stresses with an intention to enhance the tolerance of sorghum under these stresses. A very few QTL and nutrient transporters have been identified and validated under low nitrogen and phosphorus stresses compared to those under drought, salinity and cold stresses. Marker-assisted breeding and nutrient transporter characterization have not yet been attempted in sorghum under other macro- and micro-nutrient stresses. We hope this review will raise awareness among plant breeders, scientists and biotechnologists about the importance of sorghum and need to conduct the studies on marker-assisted breeding and nutrient transporter under low nutrient stresses to improve the sorghum production.

Bluemomycin, a new naphthoquinone derivative from Streptomyces sp. with antimicrobial and cytotoxic properties.

Streptomyces is one of the most prolific producers of economically important bioactive compounds used against several illnesses; it has also been found to produce industrially useful enzymes. In this study, Streptomyces sp. (ERINLG-201) was isolated from the soil sample of Kodanad forest (Southern Western Ghats), The Nilgiris, Tamil Nadu, India. ERINLG-201 isolate showed promising antibacterial activity against tested Gram-positive and Gram-negative bacteria which was confirmed by perpendicular 'T' streak method. Secondary metabolites of ERINLG-201 isolate exhibited promising antibacterial activity against tested Gram-positive and Gram-negative bacteria which was confirmed by disc diffusion method using the ethylacetate extract. Further, the ethylacetate extract of ERINLG-201 (15 g) was packed in column chromatography over silica gel and eluted; it resulted in isolation of a new naphthoquinone derivative named bluemomycin from the active fraction. Bluemomycin showed promising antibacterial activity against Gram-negative bacteria and clinical isolates at least concentration (6.25 µg/mL). Cytotoxic studies of bluemomycin showed promising activity against A549, Skvo-3 and HepG2 cell lines with IC50 values of 5.9, 24.2 and 11 µM, respectively.

Quantitative analysis of medicinal plants used to treat musculoskeletal ailments by non-institutionally trained siddha practitioners of Virudhunagar district, Tamil Nadu, India.

BACKGROUND: Siddha is one of the traditional medical systems of India; previous ethnobotanical survey in Virudhunagar district indicated a high consensus for treating musculoskeletal ailments. OBJECTIVES: This study was aimed to quantitatively document the medicinal plants used to treat musculoskeletal ailments by non-institutionally trained siddha practitioners of Virudhunagar district, Tamil Nadu, India. MATERIALS AND METHODS: This work was the outcome of free-list interviews conducted among 45 informants between April 2016 and January 2017. Sampling sufficiency was assessed by plotting Shannon-Wiener's index and cumulative number of UR. Informant consensus was assessed using Informant Consensus Factor (Fic) and Informant Agreement Ratio (IAR). RESULTS: This study recorded the data regarding 116 plant species which were used to prepare 129 formulations; analysis of the data yielded 490 UR. Among these, 65.3% of UR dealt with internal applications and 34.6% dealt with external applications. In the case of internally used formulations, pain and general musculoskeletal ailments had high Fic values. In the case of externally used formulations, headache and pain had high Fic values. CONCLUSION: Important species prescribed by the informants to treat various musculoskeletal ailments were: Azima tetracantha, Ricinus communis, Sesamum indicum, Moringa oleifera, Cardiospermum halicacabum (internal application), Calophyllum inophyllum, Justicia adhatoda, Curcuma longa, Calotropis gigantea, Zingiber officinale, Withania somnifera, Strychnos nux-vomica, Dodonaea viscosa (external application), Azadirachta indica, Clerodendrum phlomidis,Delonix elata, Pergularia daemia and Vitex negundo (internal and external applications). Robust studies on these local claims will help to improve the community healthcare and will yield some novel agents to treat musculoskeletal ailments.

Expression of PHT1 family transporter genes contributes for low phosphate stress tolerance in foxtail millet (Setaria italica) genotypes.

MAIN CONCLUSION: This is a first comprehensive study to analyze the 12 PHT1 family phosphate transporter genes in 20 foxtail millet genotypes for the improvement of millets and other crops for phosphate use efficiency. Phosphorus (P), absorbed from soil solutions as inorganic phosphate (Pi), is a limiting nutrient for plant growth and yield. Twenty genotypes of foxtail millet (Setaria italica) with contrasting degree of growth and Pi uptake responses under low Pi (LP) and high Pi (HP) supply were chosen based on a previous study. To gain molecular insights, expression dynamics of 12 PHosphate Transporter 1 (PHT1) family (SiPHT1;1 to 1;12) genes were analyzed in these 20 genotypes and compared with their Pi and total P (TP) contents. SiPHT1;1, 1;2, 1;3 and 1;8 genes were expressed in shoot tissues of three (ISe 1209, ISe 1305 and Co-6) of the LP best performing genotypes (LPBG); however, they were expressed in only one of the LP worst performing genotype (LPWG) (ISe 748). More importantly, this is correlating with higher shoot Pi and TP contents of the LPBG compared to LPWG. Apart from this condition, expression of SiPHT1 genes and their Pi and TP contents do not correlate directly for many genotypes in other conditions; genotypes with low Pi and TP contents induced more SiPHT1 genes and vice versa. Promoter analysis revealed that genotype ISe 1888 with a high level of SiPHT1;8 expression possesses two additional root box motifs compared to other genotypes. The PHT1 family genes seem to play a key role for LP stress tolerance in foxtail millet and further studies will help to improve the P-use efficiency in foxtail millet and other cereals.

Phenotypic responses of foxtail millet (Setaria italica) genotypes to phosphate supply under greenhouse and natural field conditions.

Food insecurity is a looming threat for the burgeoning world population. Phosphorus (P), which is absorbed from soil as inorganic phosphate (Pi), is an essential macronutrient for the growth of all agricultural crops. This study reports phenotype analysis for P responses in natural field and greenhouse conditions, using 54 genotypes of foxtail millet (Setaria italica) representing wide geographic origins. The genotype responses were assessed in natural field conditions in two different seasons (monsoon and summer) under Pi-fertilized (P+) and unfertilized (P-) soil for eight above-ground traits. Enormous variations were seen among the genotypes in phenotypic responses for all the measured parameters under low P stress conditions. Variations were significant for plant height, leaf number and length, tillering ability and seed yield traits. Genotypes ISe 1234 and ISe 1541 were P+ responders, and the genotypes ISe 1181, ISe 1655, ISe 783 and ISe 1892 showed tolerance to low P for total seed yield. Genotypes that performed well under P- conditions were almost as productive as genotypes that performed well under P+ conditions suggesting some genotypes are well adapted to nutrient-poor soils. In the greenhouse, most of the genotypes produced changes in root architecture that are characteristic of P- stress, but to differing degrees. Significant variation was seen in root hair density and root hair number and in fresh and dry weight of shoot and root under P- stress. However, there was not much difference in the shoot and root total P and Pi levels of five selected high and low responding genotypes. We noticed contrasting responses in the greenhouse and natural field experiments for most of these genotypes. The leads from the study form the basis for breeding and improvement of foxtail millet for better Pi-use efficiency.

The down regulation of PTP1B expression and attenuation of disturbed glucose and lipid metabolism using Borassus flabellifer (L) fruit methanol extract in high fat diet and streptozotocin induced diabetic rats.

Borassus flabellifer L. is a tall palm traditionally used for its stimulating, diuretic and anti-inflammatory activities; it is rich in fibers and various pharmacologically important secondary metabolites. This study was undertaken to evaluate the antidiabetic effects of Borassus flabellifer fruit methanol extract (BF-M) on diabetic rats induced with High Fat Diet (HFD)/streptozotocin (STZ). When BF-M (100 or 200 mg/kg) was administered for 21 days orally it led to a sharp decline in triglycerides, total cholesterol, free unsaturated fat, glucose-6-phosphate, fasting blood glucose and fructose 1,6 bisphosphatase in contrast to diabetic control. BF-M also downregulated Protein Tyrosine Phosphatase 1B. In vitro study showed the IC50 value to be 23.98 µg/mL. BF-M significantly increased serum insulin, glycogen content, and body weight. Western blot analysis exhibited significant inhibition of PTP1B in pancreatic tissue which was confirmed by histology and immunohistological studies. GC-MS analysis revelaled that the presence of major compounds such as 5-hydroxymethylfurfural (47.56%), Guanosine (21.01%) and n-hecxadeconoic acid (25.14%) in BF-M. In short, BF-M exerted antidiabetic property by down regulating PTP1B expression, and eventually enhancing glucose stimulated insulin release; it also exhibited favorable effects in diabetes and its secondary complications.

Antihyperlipidemic effect of iridoid glycoside deacetylasperulosidic acid isolated from the seeds of Spermacoce hispida L. - A traditional antiobesity herb.

ETHNOBOTANICAL RELEVANCE: The interest on herbal health supplements for obesity is increasing globally. Our previous ethnobotanical survey in Tiruvallur district, Tamil Nadu, India indicated the use of Spermacoce hispida L. seeds for the treatment of obesity. AIM OF THE STUDY: This study was aimed to validate the traditional claim and to identify the antihyperlipidemic principle in the seeds of Spermacoce hispida using bioassay guided fractionation method. METHODS: Bioassay monitored fractionation of the aqueous extract from Spermacoce hispida seeds was carried out using triton WR 1339 induced hyperlipidemic animals. It yielded deacetylasperulosidic acid (DAA) as the active ingredient. Pharmacokinetic properties of DAA were predicted using DataWarrior and SwissADME tools. In vitro antiobesity and antihyperlipidemic effects of DAA were evaluated in 3T3L1 preadipocytes and HepG2 cells, respectively. The chronic antihyperlipidemic efficacy of DAA was evaluated in high fat diet fed rats. RESULTS: DAA did not show any mutagenic and tumorigenic properties. It bound with PPARα with comparable ligand efficiency as fenofibrate. The treatment with DAA significantly lowered the proliferation of matured adipocytes, but not preadipocytes. The treatment of steatotic HepG2 cells with DAA significantly decreased the LDH leakage by 43.03% (P < 0.05) at 50 µM concentration. In triton WR 1339 induced hyperlipidemic animals, the treatment with 50 mg/kg dose significantly lowered the TC, TG and LDL-c levels by 40.27, 46.00 and 63.65% respectively. In HFD fed animals, the treatment at 10 mg/kg decreased BMI and AC/TC ratio without altering SRBG. It also improved serum lipid, transaminases and phosphatases levels of HFD fed animals. The treatment lowered adipocyte hypertrophy and steatosis of hepatocytes. CONCLUSION: This preliminary report supported the traditional use of Spermacoce hispida for the treatment of obesity. Further detailed investigations on the long term safety, efficacy and molecular mode of action of Spermacoce hispida and DAA will throw more light on their usefulness for the management of obesity.

Hepatoprotective effect of bisbenzylisoquinoline alkaloid tiliamosine from Tiliacora racemosa in high-fat diet/diethylnitrosamine-induced non-alcoholic steatohepatitis.

Non-alcoholic steatohepatitis (NASH) is one of the aggressive forms of non-alcoholic fatty liver disease (NAFLD) and is a potential risk factor of HCC. This study reports the curative effect of tiliamosine on NASH. Tiliamosine was isolated from Tiliacora racemosa Colebr. (Menispermaceae) and its structure was confirmed by studying the physical and spectroscopic data. The effects of tiliamsoine on lipid accumulation and lipotoxicity were evaluated using palmitate-oleate induced steatosis in HepG2 cells. The in vivo efficacy of tiliamosine was evaluated using HFD fed, DEN induced non-alcoholic steatohepatitis Wistar rats. In HepG2 cells, tiliamosine did not affect the cell viability up to 100 µM concentration and showed GI25 value of 264.28 µM. The treatment with tiliamsoine significantly lowered the ORO concentration by 44.17% and triglyceride accumulation by 69.32% at 50 µM concentration (P < 0.005). It also reduced the leakage of LDH and transaminases in PO-BSA induced HepG2 cells. The treatment with tiliamsoine significantly decreased the plasma levels of transaminases, phosphatase and LDH (P < 0.05) in HFD-DEN induced steatohepatitis. The histology and the immunohistochemistry of the hepatic sections were in accordance with the biochemical findings. Preliminary molecular analysis indicated that the hepatic FXR expression was upregulated and TNFα expression was downregulated by the treatment with tiliamsoine. This study provided preliminary evidence on the use of tiliamosine for the treatment of NASH.

Curative effect of arjunolic acid from Terminalia arjuna in non-alcoholic fatty liver disease models.

The prevalence of Non Alcoholic Fatty Liver Disease (NAFLD) is increasing globally. Terminalia arjuna W. & Arn. (Combretaceae) is an endemic tree found in India and Sri Lanka and used traditionally for its cardioprotective and hepatoprotective effects. Arjunolic acid (AA) is an oleanane triterpenoid found mainly in the heartwood of T. arjuna. This study was aimed to evaluate the hepatoprotective effect of AA using cellular and rodent models of NAFLD. AA was isolated from the ethyl acetate extract of the heartwood of T. arjuna. The structure of AA was confirmed by physical and spectroscopic data. Steatosis was induced in HepG2 cells using palmitate-oleate mixture and the effects of AA on triglyceride accumulation and lipotoxicity were assessed. In vivo effect of AA on NAFLD was assessed using HFD fed rats. The treatment with AA did not affect the cell viability upto 100 µM and showed GI25 value of 379.9 µM in HepG2 cells. The treatment with AA significantly lowered the ORO concentration by 35.98% and triglyceride accumulation by 66.36% at 50 µM concentration (P < 0.005) compared to the vehicle treated group. The treatment with AA also reduced the leakage of ALT and AST by 61.11 and 48.29% in a significant manner (P < 0.005). The in vivo findings clearly demonstrated that the animals treated with AA at 25 and 50 mg/kg concentrations showed a significant decrease in the levels of transaminases, phosphatase and GGT (P < 0.005). In the liver, the expression of PPARα and FXRα expressions were upregulated, while PPARγ expression was downregulated by the treatment with AA. The liver histology of the animals showed reduction in steatosis and MNC infiltration. These preliminary evidences suggested that AA might be a promising lead to treat NAFLD. Future robust scientific studies on AA will lead to tailoring it for the treatment of NAFLD.

A study on food-medicine continuum among the non-institutionally trained siddha practitioners of Tiruvallur district, Tamil Nadu, India.

BACKGROUND: Medicinal properties of the food species are one of the poorly documented and important areas of ethnopharmacology. The present survey quantitatively documented the medicinal foods prescribed by the non-institutionally trained siddha practitioners of Tiruvallur district of Tamil Nadu. METHODS: Field work was carried out between December 2014 and April 2017 using a questionnaire. The illnesses mentioned by the informants were grouped as illness categories on the basis of emic perceptions. Sufficiency of sampling of this survey was assessed by plotting the cumulative number of UR and Shannon-Wiener's index. The indices such as informant consensus factor (FIC), Index of Agreement on Remedies (IAR), and Cultural Food Significance Index (CFSI) were calculated. RESULTS: This study documented 165 medicinal foods used by 82 non-institutionally trained siddha practitioners of Tiruvallur district, and 73.93% of these preparations were plant based. Among the animal taxa, 82.05% were represented by fish taxa. The illness category gastrointestinal ailments is the majorly cited illness category treated with plant-based formulations. The illness categories viz., gastrointestinal ailments, hemorrhoids, and neural ailments had high consensus under the group of plant-based medicinal foods. In animal-based medicinal foods, kapha ailments had gained 23.07% of UR. The illness categories such as bone fractures, male reproductive ailments, blood ailments, and anabolic had high FIC values. CONCLUSIONS: Deeper studies on different dietary cultures of India may help to derive better interpretations on food-medicine continuum. This study identified some important claims such as the use of citron, pomegranate and Solanum americanum (gastrointestinal ailments), Abutilon indicum, onions and elephant foot yam (hemorrhoids), Boerhavia diffusa (urinary ailments), Moringa oleifera (anemia), Aloe vera (gynecological ailments), Eclipta prostrata (liver ailments), ivy gourd (diabetes), citron (hypertension), Centella asiatica (psychological ailments), spade nose shark (lactogogue), reticulate whipray (wheezing and bronchitis), Katelysia opima (impotence), Indian squid (anemia), and Indian oil sardine (anabolic). More studies on these claims will help identify novel functional foods to add to the field of medical nutrition therapy, with traditional brand identity. Robust studies on the documentation of the traditional knowledge on marine resources will yield a good database for various stakeholders and policy makers.

Bio-prospecting of soil Streptomyces and its bioassay-guided isolation of microbial derived auxin with antifungal properties.

The present study was aimed to isolate bioactive actinomycetes with antifungal properties. Twenty-seven distinct soil derived actinomycetes were investigated for their antifungal activities. Among these, one isolate exhibited significant antifungal activity. Phenotypic and 16s rRNA gene sequence analysis strongly suggested that the active isolate BG4 belonged to the genus Streptomyces. Further, the chemical investigation of the active extract resulted in the isolation of a major compound and it was structurally elucidated as phenyl acetic acid (PAA). PAA exhibited promising antifungal activity with 100% inhibition, ranging from 31.25 to 25µg/mL. It is to be noted that PAA is naturally occurring and biologically active auxin. In addition, it has also been hypothesized that phytohormone endorsing the source of soil-symbionts has similar pathways for synthesizing compounds and its congeners of host due to horizontal gene transfer. These findings demonstrate that microbially derived phytohormone can be used to treat fungal infections.

Hepatoprotective effect of lawsone on rifampicin-isoniazid induced hepatotoxicity in in vitro and in vivo models.

The Drug-induced liver injury is one of the common unfavourable impacts, which seriously affects any drug therapy. This study documented the hepatoprotective efficacy of lawsone, the major bioactive naphthoquinone present in Lawsonia inermis L. (Lythraceae) using in vitro and in vivo models. Lawsone was isolated from the leaves of L. inermis and its structure was confirmed using spectroscopic data. In-vitro antioxidant effect of lawsone was evaluated using ABTS assay. Hepatoprotective effect of lawsone was determined with RIF-INH treated HepG2 cells and Wistar rats. Administration of RIF-INH reduced the viability of the HepG2 cells and the treatment with lawsone significantly restored the viability of the cells even at lower concentration (7.5 µM). The other parameters such as the leakage of transaminases and MDA levels were also significantly reduced by the treatment with lawsone. Oral administration of lawsone to the animals did not show any toxicity up to 2 g/kg b.w. concentration. Treatment with lawsone to the RIF-INH administered animals significantly lowered the serum transaminases levels. The ratio of albumin to globulin was improved and the level of bilirubin was lowered. This study indicated the hepatoprotective effect of lawsone; detailed investigations will give deeper understanding of the application of lawsone for hepatoprotection.

Traditionally practiced medicinal plant extracts inhibit the ergosterol biosynthesis of clinically isolated dermatophytic pathogens.

OBJECTIVE: Dermatophytes are resistant to some available antibiotics. Development of new plant drugs to control drug resistant microbes is urgently needed. This study evaluates the antidermatophytic potential of 18 selected medicinal plants used by traditional healers in Theni and Virudhunagar Districts of Tamil Nadu, India. MATERIALS AND METHODS: Selected plant parts were collected, shade dried and powdered. Plant powders were extracted with ethanol and their antifungal potency was investigated against and clinical dermatophytes. The antioxidant effect of the extracts was screened using DPPH assay. Minimal Inhibitory Concentration (MIC) and Minimal Fungicidal Concentration (MFC) were estimated for the extracts. Ten plant extracts showed maximum MFC and they were selected to study their efficacy in interfering with ergosterol biosynthesis. Fluconazole-35µg/mL known fungicide was used as control. The most active extracts were taken for biocompatibility studies using 3T3-L1 fibroblast cell lines. RESULTS: The ethanol extract of Phyllanthus reticulates leaves showed good antifungal activity compared to other plant extracts. The MIC and MFC for Phyllanthus reticulatus were 62.5 and 250µg/mL against M. pachydermatitis and T. rubrum respectively. The ethanol extracts of Phyllanthus reticulatus, Coldenia procumbens, Thespesia populnea and Senna alata significantly lowered the release of ergosterol by 16.37, 19.53, 24.79, and 21.44%, respectively. The ethanol extract of Phyllanthus reticulatus leaves was more biocompatible to host cells than other active extracts. CONCLUSION: Our study indicated that the ethanol extract of Phyllanthus reticulates leaves showed promising activity against dermatophytes. It could be a potential material for future development of antidermatophytic agents.

Functional characterization of the PHT1 family transporters of foxtail millet with development of a novel Agrobacterium-mediated transformation procedure.

Phosphate is an essential nutrient for plant growth and is acquired from the environment and distributed within the plant in part through the action of phosphate transporters of the PHT1 family. Foxtail millet (Setaria italica) is an orphan crop essential to the food security of many small farmers in Asia and Africa and is a model system for other millets. A novel Agrobacterium-mediated transformation and direct plant regeneration procedure was developed from shoot apex explants and used to downregulate expression of 3 members of the PHT1 phosphate transporter family SiPHT1;2 SiPHT1;3 and SiPHT1;4. Transformants were recovered with close to 10% efficiency. The downregulation of individual transporters was confirmed by RT-PCR. Downregulation of individual transporters significantly reduced the total and inorganic P contents in shoot and root tissues and increased the number of lateral roots and root hairs showing they have non-redundant roles. Downregulation of SiPHT1;2 had the strongest effect on total and inorganic P in shoot and root tissues. Complementation experiments in S. cerevisiae provide evidence for the ability of SiPHT1;1, 1;2, 1;3, 1;7 and 1;8 to function as high affinity Pi transporters. This work will aid development of improved millet varieties for global food security.

Swertiamarin, a natural steroid, prevent bone erosion by modulating RANKL/RANK/OPG signaling.

Bone erosion is a central feature of rheumatoid arthritis (RA) that is characterized by the infiltration of the synovial lining by osteoclasts and lymphocytes. In the present study, swertiamarin a major secoiridoid glycoside was evaluated for anti-osteoclastogenic property to prevent bone erosion in Freund's complete adjuvant (FCA) induced in-vivo model, in-vitro osteoblast and osteoclasts as well as in co-culture system and in-silico molecular docking analysis. The swertiamarin treatment decreased the expression of TRAP, RANKL, and RANK levels and increased the levels of OPG levels significantly in both in vitro and in vivo models. In in vitro, the compound treatment significantly increased the cell proliferation and ALP levels in osteoblast cells; the high proliferation (153.8600±5.23%) and ALP release (165.6033±4.13%) were observed at 50µg/ml concentration of swertiamarin treatment. At the same time the treatment decreased the TRAP positive cells in osteoclast cells; the high reductions of TRAP positive cells (39.32±3.19%) were observed at 50µg/ml of swertiamarin treatment. The treatment modulated the levels of pro-inflammatory cytokines, MMPs and NF-κB levels in osteoblast and osteoclast co-culture system. In in silico analysis swertiamarin had affinity towards the proteins RANK, RANKL and OPG residues with low binding energy -4.5, -3.92 and -5.77kcal/mol respectively. Thus, the results of this study revealed the anti-osteoclastogenic activity of swertiamarin on the prevention of bone destruction.

Effect of two andrographolide derivatives on cellular and rodent models of non-alcoholic fatty liver disease.

The prevalence of Non-Alcoholic Fatty Liver Disease (NAFLD) is increasing and there is an increasing interest in natural products to treat NAFLD. This study aimed to evaluate the hepatoprotective effect of andrographolide and two of its derivatives; in one the OH group at C-14 was removed and in the other OH groups at C-3 and C-19 were protected. Andrographolide (AN) was isolated from the aerial parts of Andrographis paniculata Wall. Isoandrographolide (IAN) and 3,19-acetonylidene andrographolide (ANA) were derivatized from AN. Drug likeness of the compounds was studied using DataWarrior. The effect of the compounds in ameliorating hepatic steatosis and lipotoxicity was assessed using palmitate-oleate induced steatotic HepG2 cell lines. In vivo efficacy of the compounds was assessed by using HFD fed rats. IAN showed comparatively high drug score and low irritability than AN. MTT assay indicated that the treatment with IAN had comparatively less toxicity than AN and ANA to HepG2 cells. The treatment with IAN significantly reduced the lipid accumulation and the leakage of LDH and transaminases, while the treatments with AN and ANA did not prohibit the leakage. In the in vivo experiment, the treatment with IAN showed comparatively better hepatoprotection by reducing the serum lipid, transaminases and ALP levels than with AN and ANA. Our results showed that IAN could be a promising lead to treat NAFLD with comparatively low toxicity and improved efficacy.

Identification of putative QTLs for seedling stage phosphorus starvation response in finger millet (Eleusine coracana L. Gaertn.) by association mapping and cross species synteny analysis.

A germplasm assembly of 128 finger millet genotypes from 18 countries was evaluated for seedling-stage phosphorus (P) responses by growing them in P sufficient (Psuf) and P deficient (Pdef) treatments. Majority of the genotypes showed adaptive responses to low P condition. Based on phenotype behaviour using the best linear unbiased predictors for each trait, genotypes were classified into, P responsive, low P tolerant and P non-responsive types. Based on the overall phenotype performance under Pdef, 10 genotypes were identified as low P tolerants. The low P tolerant genotypes were characterised by increased shoot and root length and increased root hair induction with longer root hairs under Pdef, than under Psuf. Association mapping of P response traits using mixed linear models revealed four quantitative trait loci (QTLs). Two QTLs (qLRDW.1 and qLRDW.2) for low P response affecting root dry weight explained over 10% phenotypic variation. In silico synteny analysis across grass genomes for these QTLs identified putative candidate genes such as Ser-Thr kinase and transcription factors such as WRKY and basic helix-loop-helix (bHLH). The QTLs for response under Psuf were mapped for traits such as shoot dry weight (qHSDW.1) and root length (qHRL.1). Putative associations of these QTLs over the syntenous regions on the grass genomes revealed proximity to cytochrome P450, phosphate transporter and pectin methylesterase inhibitor (PMEI) genes. This is the first report of the extent of phenotypic variability for P response in finger millet genotypes during seedling-stage, along with the QTLs and putative candidate genes associated with P starvation tolerance.