An assessment on the role of endophytic microbes in the therapeutic potential of Fagonia indica.

BACKGROUND: Natural products of animals, plants and microbes are potential source of important chemical compounds, with diverse applications including therapeutics. Endophytic bacteria that are especially associated with medicinal plants presents a reservoir of therapeutic compounds. Fagonia indica has been recently investigated by numerous researchers because of its striking therapeutic potential especially in cancer. It is also reported that endophytes play a vital role in the biosynthesis of various metabolites; therefore we believe that endophytes associated with F. indica are of crucial importance in this regard. The present study aims successful isolation, molecular identification of endophytic bacteria and their screening for bioactive metabolites quantification and in vitro pharmacological activities. METHODS: 16S rRNA gene sequencing was used for the identification of isolated endophytic bacteria. Methanolic extracts were evaluated for total phenolic contents (TPC), total flavonoids contents (TFC), DPPH free radical scavenging activity, reducing power and total anti-oxidant assays were performed. And also screened for antibacterial and antifungal activities by disc diffusion method and their MIC were calculated by broth dilution method using microplate reader. Further, standard protocols were followed for antileishmanial activity and protein kinase inhibition. Analysis and statistics were performed using SPSS, Table curve and Origin 8.5 for graphs. RESULTS: Bacterial strains belonging to various genera (Bacillus, Enterobacter, Pantoea, Erwinia and Stenotrophomonas) were isolated and identified. Total phenolic contents and total flavonoids contents varies among all the bacterial extracts respectively in which Bacillus subtilis showed high phenolic contents 243 µg/mg of gallic acid equivalents (GAE) and Stenotrophomonas maltophilia showed high flavonoids contents 15.9 µg/mg quercitin equivalents (QA), total antioxidant capacity (TAC) 37.6 µg/mg of extract, reducing power (RP) 206 µg/mg of extract and 2, 2-diphenyl-1-picrylhydrazyl (DPPH) free radical scavenging activity with 98.7 µg/mL IC50 value. Although all the extracts tested were active to inhibit growth of selected pathogenic microbes (bacteria and fungi), but significant antibacterial activity was observed against Klebsiella pneumonia and B. subtilis. An Enterobacter cloaca was active against Leishmania tropica with IC50 value of 1.4 µg/mg extracts. B. subtilis and Bacillus tequilensis correspondingly exhibit significant protein kinase inhibition of 47 ± 0.72 and 42 ± 1.21 mm bald zones, indicating anti-infective and antitumor potential. CONCLUSIONS: Our findings revealed that crude extracts of selected endophytic bacteria from F. indica possess excellent biological activities indicating their potential as an important source of antibiotics (antifungal, antibacterial) compounds.

Leishmanicidal active withanolides from a pakistani medicinal plant, Withania coagulans.

In the course of screening for leishmanicidal constituents from Asian and South American medicinal plants, a Pakistani medicinal plant, Withania coagulans, showed activity. We therefore studied the active components of the methanol extract of aerial parts of W. coagulans. From the ethyl acetate soluble fraction of the extract, four new withanolides (1-4) were isolated along with seven known withanolides (5-11). The new compounds were elucidated to be (14R,15R,17S,20S,22R)-14,15,17,20-tetrahydroxy-1-oxowitha-2,5,24-trienolide (1), (14R,15R,17S,20S,22R)-14,15,17,20-tetrahydroxy-1-oxowitha-3,5,24-trienolide (2), (14S,17R,20S,22R)-14,17,20-trihydroxy-1-oxowitha-2,5,24-trienolide (3), and (14S,17R,20S,22R)-14,17,20-trihydroxy-1-oxowitha-3,5,24-trienolide (4), from 1H-NMR, 13C-NMR, 2D-NMR and high resolution (HR)-MS data. Some of these compounds having the partial structure 1-oxo-2,5-diene showed strong leishmanicidal activity against Leishmania major.

The phytochrome-interacting factor PIF7 negatively regulates DREB1 expression under circadian control in Arabidopsis.

Transcription factors of the DRE-Binding1 (DREB1)/C-repeat binding factor family specifically interact with a cis-acting dehydration-responsive element/C-repeat involved in low-temperature stress-responsive gene expression in Arabidopsis (Arabidopsis thaliana). Expression of DREB1s is induced by low temperatures and is regulated by the circadian clock under unstressed conditions. Promoter sequences of DREB1s contain six conserved motifs, boxes I to VI. We analyzed the promoter region of DREB1C using transgenic plants and found that box V with the G-box sequence negatively regulates DREB1C expression under circadian control. The region around box VI contains positive regulatory elements for low-temperature-induced expression of DREB1C. Using yeast one-hybrid screens, we isolated cDNA encoding the transcriptional factor Phytochrome-Interacting Factor7 (PIF7), which specifically binds to the G-box of the DREB1C promoter. The PIF7 gene was expressed in rosette leaves, and the PIF7 protein was localized in the nuclei of the cells. Transactivation experiments using Arabidopsis protoplasts indicated that PIF7 functions as a transcriptional repressor for DREB1C expression and that its activity is regulated by PIF7-interacting factors TIMING OF CAB EXPRESSION1 and Phytochrome B, which are components of the circadian oscillator and the red light photoreceptor, respectively. Moreover, in the pif7 mutant, expression of DREB1B and DREB1C was not repressed under light conditions, indicating that PIF7 functions as a transcriptional repressor for the expression of DREB1B and DREB1C under circadian control. This negative regulation of DREB1 expression may be important for avoiding plant growth retardation by the accumulation of DREB1 proteins under unstressed conditions.

Promising antiviral, antimicrobial and therapeutic properties of green nanoceria.

Aim: To demonstrate synthesis of cerium oxide nanoparticles (CeO2 NPs) by a green method using Hyphaene thebaica, and investigate their therapeutic applications. Materials & methods: Structural, vibrational and luminescent properties were established using x-ray diffraction, Fourier transformed infrared spectroscopy, Raman spectroscopy, ultraviolet absorption spectroscopy, selected area electron diffraction, electron microscopy and photolumincence spectroscopy. Therapeutic properties were established using different in vitro assays. Results: CeO2 NPs were determined to be crystalline in nature with a grain size of approximately 14 nm. They had characteristic Ce-O vibration at 481 cm-1. Photoluminescence spectra revealed broad bands at 463 and 600 nm. ζ potential was recorded as -17.2 mV. Potent antimicrobial and antiviral properties with hemocompatibility were reported. Conclusion: Biosynthesized CeO2 NPs revealed multifunctional therapeutic properties.

Greener synthesis of ZnO and Ag-ZnO nanoparticles using Silybum marianum for diverse biomedical applications.

AIM: To investigate the physical and biological properties of Silybum marianum inspired ZnO nanoparticles (NPs), Ag-ZnO heterostructures. Experiment: Nanoparticles were characterized using ultraviolet-visible and infrared spectroscopy, x-ray diffraction, high resolution electron microscopy, ζ potential and thermo-gravimetric analysis etc. Results: Ag-ZnO-NPs indicated slightly higher antimicrobial potential then ZnO-NPs. Good antileishmanial (IC50 = 246 µg/ml for Ag-ZnO; 341 µg/ml for ZnO) and antioxidant potential while moderate enzyme inhibition is reported. 2, 2-Diphenyl 1-picrylhydrazyl radical scavenging of Ag-ZnO was higher relative to ZnO-NPs. Nanocosmaceutical formulation of nanoparticles indicated stable antimicrobial performance. CONCLUSION: Biosynthesized nanoparticles indicated interesting biological properties and should be subjected to further research to establish their pharmacological relevance.

Alterations in the infrared spectral signature of avian feathers reflect potential chemical exposure: a pilot study comparing two sites in Pakistan.

Chemical contamination of ecosystems is a global issue with evidence that pollutants impact on living organisms in a harmful fashion. Developing sensor approaches that would allow the derivation of biomarkers or signatures of effect in target sentinel organisms and monitor environmental chemical contamination in a high throughput manner is of utmost importance. As biomolecules absorb infrared (IR), signature vibrational spectra related to structure and function can be derived. In light of this, we tested the notion that IR spectra of bird feathers might reflect environmental chemical contaminant exposure patterns. Feathers were collected from monospecific heronries of cattle egret based in two independent locations (Trimu vs. Mailsi) in the Punjab province of Pakistan; these sites were found to differ in their chemical contamination patterns. Feather samples were chemically analyzed for polychlorinated biphenyls, polybrominated diphenyl ethers, organochlorines and heavy metals. Attenuated total reflection Fourier-transform IR (ATR-FTIR) spectroscopy was employed to derive a spectral signature of individual feathers. Resultant IR spectra were then subjected to canonical correspondence analysis (CAA) to determine whether feather spectral signatures correlate to chemical exposure. Additionally, we explored if principal component analysis (PCA) and linear discriminant analysis (LDA) could be applied to distinguish site-specific differences; linear discriminant function (LDF) was also applied to classify sites. The sampled feathers varied in their chemical exposure patterns depending on whether they were sourced from one site associated with heavy metal exposure or the other which suggested high organic pollutant exposures. CCA of chemical and spectral data showed a correlation between spectral signatures and chemical exposure. PCA-LDA readily distinguished feathers from the two different sites. Discriminating alterations were identified and these were associated with protein and lipid regions in IR spectra. Additionally, LDF showed that the classification rate of spectral categories correlated well with the two chemical exposure patterns (93.6% for Trimu feathers and 91.77% for Mailsi feathers). This pilot study suggests that IR spectra derived from feathers reflect background chemical exposure and points to a novel monitoring tool for contamination.

Interaction between two cis-acting elements, ABRE and DRE, in ABA-dependent expression of Arabidopsis rd29A gene in response to dehydration and high-salinity stresses.

Many abiotic stress-inducible genes contain two cis-acting elements, namely a dehydration-responsive element (DRE; TACCGACAT) and an ABA-responsive element (ABRE; ACGTGG/TC), in their promoter regions. We precisely analyzed the 120 bp promoter region (-174 to -55) of the Arabidopsis rd29A gene whose expression is induced by dehydration, high-salinity, low-temperature, and abscisic acid (ABA) treatments and whose 120 bp promoter region contains the DRE, DRE/CRT-core motif (A/GCCGAC), and ABRE sequences. Deletion and base substitution analyses of this region showed that the DRE-core motif functions as DRE and that the DRE/DRE-core motif could be a coupling element of ABRE. Gel mobility shift assays revealed that DRE-binding proteins (DREB1s/CBFs and DREB2s) bind to both DRE and the DRE-core motif and that ABRE-binding proteins (AREBs/ABFs) bind to ABRE in the 120 bp promoter region. In addition, transactivation experiments using Arabidopsis leaf protoplasts showed that DREBs and AREBs cumulatively transactivate the expression of a GUS reporter gene fused to the 120 bp promoter region of rd29A. These results indicate that DRE and ABRE are interdependent in the ABA-responsive expression of the rd29A gene in response to ABA in Arabidopsis.