Higher cadmium and zinc accumulation in parsley (Petroselinum crispum) roots activates its antioxidants defense system.

Parsley (Petroselinum crispum) is herb with many biological and medicinal benefits for humans. However, growth on zinc (Zn) and cadmium (Cd) contaminated sites might get severely affected due to over accumulation of heavy metals (HM) in different plant tissues. Antioxidants play a crucial role in minimizing the negative effects of HM. The present study investigates the effects of Zn and Cd stress on P. crispum morphological parameters, enzymatic/non-enzymatic antioxidant profiling and metal accumulation in shoot/root. Plants were exposed to different concentrations of Zn (50, 100, 150 and 200 µM) and Cd (10, 20, 40 and 80 µM) along with control (no stress), in soil-less Hoagland's solution. The results showed that Zn and Cd substantially decrease the growth parameters with increased contents of malondialdehyde (MDA), hydrogen peroxide (H2O2), and electrolyte leakage (EL). Non-enzymatic antioxidant activities, like total phenolic contents (TPC) and ferric reducing antioxidant power (FRAP), were induced high in leaves only upon Cd stress and contrarily decreased upon Zn stress. Total flavonoid contents (TFC) were decreased under Zn and Cd stress. Enzymatic antioxidant activities like superoxide dismutase (SOD), catalase (CAT) and ascorbate peroxidase (APX) were also strongly induced upon Cd stress. At the same time, SOD and guaiacol peroxidase (GPX) activity was induced significantly upon Zn stress. Cd uptake and accumulation was notably high in roots as compared to shoots, which suggests P. crispum have a reduced ability to translocate Cd towards aboveground parts (leaves). Additionally, strong induction of antioxidants by P. crispum under Cd stress might indicate the capacity to effectively re-modulate its physiological response. However, further investigations regarding other HMs and experiments at the molecular level are still needed.

Cyclic guanosine monophosphate improves salt tolerance in Solanum lycopersicum.

The cyclic nucleotide cyclic guanosine monophosphate (cGMP) is a powerful cell signaling molecule involved in biotic and abiotic stress perception and signal transduction. In the model plant Arabidopsis thaliana, salt and osmotic stress rapidly induce increase in cGMP which plays role by modulating the activity of monovalent cation transporters, possibly by direct binding to these proteins and by altering the expression of many abiotic stress responsive genes. In a recent study, a membrane permeable analogue of cGMP (8-bromo-cGMP) was found to have a promotive effect on soluble sugar, flavonoids and lignin content, and membrane integrity in Solanum lycopersicum seedlings under salt stress. However, it remains to be elucidated how salt stress affects the endogenous cGMP level in S. lycopersicum and if Br-cGMP-induced improvement in salt tolerance in S. lycopersicum involves altered cation fluxes. The current study was conducted to answer these questions. A rapid increase (within 30 s) in endogenous cGMP level was determined in S. lycopersicum roots after treatment with 100 mM NaCl. Addition of membrane permeable Br-cGMP in growth medium remarkably ameliorated the inhibitory effects of NaCl on seedlings' growth parameters, chlorophyll content and net photosynthesis rate. In salt stressed plants, Br-cGMP significantly decreased Na+ content by reducing its influx and increasing efflux while it improved plants K+ content by reducing its efflux and enhancing influx. Furthermore, supplementation with Br-cGMP improved plant's proline content and total antioxidant capacity, resulting in markedly decreased electrolyte leakage under salt stress. Br-cGMP increased the expression of Na+/H+ antiporter genes in roots and shoots of S. lycopersicum growing under salt stress, potentially enhancing plant's ability to sequester Na+ into the vacuole. The findings of this study provide insights into the mechanism of cGMP-induced salt stress tolerance in S. lycopersicum.

Real-time expression and in silico characterization of pea genes involved in salt and water-deficit stress.

BACKGROUND: To tolerate salt and water-deficit stress, the plant adapts to the adverse environment by regulating its metabolism and expressing certain stress-induced metabolic pathways. This research analyzed the relative expression of four pea genes (P5CR, PAL1, SOD, and POX) in three pea varieties (Climax, Green grass, and Meteor) under different levels of salt and water-deficit stress. METHODS AND RESULTS: The experiments on salt stress and water-deficit stress were carried out within greenhouse settings under controlled environment. The saturation percentage was employed to create artificial salinity conditions: Control without NaCl treatment, Treatment 1: 50 mM NaCl treatment, Treatment 2: 75 mM NaCl treatment, and Treatment 3: 100 mM NaCl treatment. Field capacity (FC) was used for the development of artificial water-deficit treatments in the pots, i.e., Treatment 1 (Control; water application 100% of FC), Treatment 2 (water application 75% of FC), and Treatment 3 (water application 50% of FC). Pea genes involved in biosynthetic pathways of proline, flavonoids, and enzymatic antioxidant enzymes including P5CR, PAL1, SOD, and POX were selected based on literature. Quantitative real-time PCR using cDNA as a template was used to analyze the gene expression. Pea genes were analyzed for phylogenetic analysis in closely related crops having similarity percent identity 80% and above. In silico characterization of selected proteins including the family classification was done by the NCBI CDD and INTERPRO online servers. Results from RT-qPCR analysis showed increased expression of P5CR, PAL1, and POX genes, while SOD gene expression decreased under both stresses. Climax exhibited superior stress tolerance with elevated expression of P5CR and PAL1, while Meteor showed better tolerance through increased POX expression. Phylogenetic analysis revealed common ancestry with other species like chickpea, red clover, mung bean, and barrel clover, suggesting the cross relationship among these plant species. Conserved domain analysis of respective proteins revealed that these proteins contain PLNO 2688, PLN02457, Cu-Zn Superoxide dismutase, and secretory peroxidase conserved domains. Furthermore, protein family classification indicated that the oxidation-reduction process is the most common chemical process involved in these stresses given to pea plant which validates the relationship of these proteins. CONCLUSIONS: Salt and water-deficit stresses trigger distinct metabolic pathways, leading to the up-regulation of specific genes and the synthesis of corresponding proteins. These findings further emphasize the conservation of stress-tolerance-related genes and proteins across various plant species. This knowledge enhances our understanding of plant adaptation to stress and offers opportunities for developing strategies to improve stress resilience in crops, thereby addressing global food security challenges.

Quantification of rosmarinic acid from different plant species of lower Himalayan region and expression analysis of underlying L-Phenylalanine pathway.

This study adopts a very effective high-performance liquid chromatography (HPLC) technique for the quantitative determination of rosmarinic acid (RA) and PCR-based amplification of biosynthetic key regulators in Isodon rugosus, Daphne mucronata, and Viburnum grandiflorum from the lower Himalayan regions. Rosmarinic acid is engaged in a variety of biological processes and has significant industrial significance. In this study, it was identified from crude methanolic extract using thin-layer chromatography with a standard, and its content was quantified using HPLC without interrupting spikes using a mixture of methanol and deionized water containing acetonitrile (70:30 v/v) and acetic acid (0.1% v/v) at UV 310 nm absorption. We used RT-PCR to identify cDNAs encoding PAL, C4H, and RAS, and Image J's semi-quantitative analysis to quantify the expression levels of genes involved in RA production from chosen plant material. The highest levels of PAL, C4H, and RAS were detected, by band intensity, in the leaves and flowers of I. rugosus, which also exhibited a substantial quantity of RA. However, in V. grandiflorum and D. mucronata the transcript of the given genes was low. The concentration of RA ranged from 187.7 to 21.2 mg g-1 for I. rugosus, 17.42 to 5.42 mg g-1 for V. grandiflorum, and 15.19 mg g-1 for D. mucronata. This study demonstrated that the method for quantifying RA from a crude methanolic extract was effective, indicating that I. rugosus might be used as an indigenous alternative source of RA.

Detection of hepatitis through proteases and protease inhibitors genes expression and identification of HCV untypable genotype in Abbottabad, Pakistan.

Liver cancer is the 5th most common cancer caused mainly due to the late detection of hepatitis. Therefore, the early detection of hepatitis through genetic markers boosts effective and remedial management. In addition, to determine the occurrence of hepatitis C virus (HCV), genotyping is indispensable as majority of hepatitis cases remain undiagnosed. The current study was designed to find the gene expression of proteases and proteases inhibitors in different hepatitis patients and to determine HCV genotypes mainly focusing on untypeable genotypes of HCV in Abbottabad, Pakistan. PCR was conducted to find the expression of proteases and protease inhibitors genes in hepatitis patients and healthy individuals. HCV genotyping was done by PCR based method and untypeable genotypes were sequenced and verified using online tools. Controlled individuals showed normal expression of cystatin C and leptin, low expression of cathepsin B while high expression of other studied genes including cathepsin D and G, TPP1 and serpin B1 could be seen. Hepatitis A patients showed high expression of leptin while other genes showed low expression. Hepatitis B patients revealed considerable variations in the cathepsin and cystatin C gene expression. Therefore, low cystatin C (high cathepsin B) and/or high cystatin C (low cathepsin B) levels can be regarded as a potential marker for hepatitis B. Hepatitis C infected patients showed high gene expression of cystatin C and leptin, so they could be useful markers for the diagnostics and prediction of the severity of HCV infections. While genotyping findings showed that about 45% of total PCR positive samples (110) were found to be of 3a genotype followed by 3b in 18%, 1a in 13.6% and 1b in 10%. About 9% of infections turned out to be mixed infections, whereas only 4.5% were untraceable by our genotyping system. Sequencing of untypeable genotypes and applying online tools revealed that the described untypeable genotypes of HCV were in fact variants of 3a genotype. Furthermore, full length characterization of these variants could help to classify them into types and subtypes. Keywords: hepatitis; genotyping; genes expression; proteases and protease inhibitors; ML; NJ.

Ethnobotanical and antimicrobial study of some selected medicinal plants used in Khyber Pakhtunkhwa (KPK) as a potential source to cure infectious diseases.

BACKGROUND: Present investigation deals with antimicrobial screening of ten medicinally important plants used by the inhabitants of district Haripur, Khyber Pakhtunkhwa (KPK) for different infectious diseases. METHODS: Aqueous, n-hexane and ethanolic extracts of each plant were tested for their antimicrobial activity against both Gram positive and Gram negative strains of bacteria, as well as strain of yeast. Agar well diffusion and broth dilution methods were used to determine the antimicrobial activity of different plant extracts. RESULTS: The results indicated that all plants exhibited antimicrobial activity against one or more test pathogens. Interestingly, extracts of three plants showed strong and broad spectrum activity as compared to rest of the extracts which demonstrated the moderate activity. On the whole ethanolic extracts exhibited maximum antimicrobial effect than their corresponding aqueous and n-hexane extracts, when compared with standard antibiotics i.e., Streptomycin and Tetracycline. Among various extracts, only ethanolic extract of Azadirachta indica and aqueous and ethanolic extracts of Eucalyptus globulus and Bergenia ciliata and ethanolic extract of Punica granatum were found to have potentially promising activity against test microorganisms. CONCLUSION: Different plant extracts show promising antimicrobial activity justifying their usage in traditional medicines. This study will be continued to identify more plants with potential antimicrobial components.

Antioxidant, Cytotoxic, and Antimicrobial Potential of Silver Nanoparticles Synthesized using Tradescantia pallida Extract.

Silver nanoparticles have received much attention, due to their wide range of biological applications as an alternative therapy for disease conditions utilizing the nanobiotechnology domain for synthesis. The current study was performed to examine the antioxidant, anticancer, antibacterial, and antifungal potential of biosynthesized silver nanoparticles (TpAgNPs) using plant extract. The TpAgNPs were produced by reacting the Tradescantia pallida extract and AgNO3 solution in nine various concentration ratios subjected to bioactivities profiling. According to the current findings, plant extract comprising phenolics, flavonoids, and especially anthocyanins played a critical role in the production of TpAgNPs. UV-visible spectroscopy also validated the TpAgNP formation in the peak range of 401-441 nm. Further, the silver ion stabilization by phytochemicals, face-centered cubic structure, crystal size, and spherical morphology of TpAgNPs were analyzed by FTIR, XRD, and SEM. Among all TpAgNPs, the biosynthesized TpAgNP6 with a medium concentration ratio (5:10) and the plant extract had effective antioxidant potentials of 77.2 ± 1.0% and 45.1 ± 0.5% free radical scavenging activity, respectively. The cytotoxic activity of TpAgNP6 in comparison to plant extract for the rhabdomyosarcoma cell line was significantly the lowest with IC50 values of 81.5 ± 1.9 and 90.59 ± 1.6 µg/ml and cell viability % of 24.3 ± 1.62 and 27.4 ± 1.05, respectively. The antibacterial and antifungal results of TpAgNPs revealed significant improvement in comparison to plant extract, i.e., minimum inhibition concentration (MIC) 64 µg/ml against Gram-negative Pseudomonas aeruginosa while, in the case of antifungal assay, TpAgNP6 was active against Candida parapsilosis. These TpAgNPs play a crucial role in determining the therapeutic potential of T. pallida due to their biological efficacy.

Expression of cyanobacterial genes enhanced CO2 assimilation and biomass production in transgenic Arabidopsis thaliana.

BACKGROUND: Photosynthesis is a key process in plants that is compromised by the oxygenase activity of Rubisco, which leads to the production of toxic compound phosphoglycolate that is catabolized by photorespiratory pathway. Transformation of plants with photorespiratory bypasses have been shown to reduce photorespiration and enhance plant biomass. Interestingly, engineering of a single gene from such photorespiratory bypasses has also improved photosynthesis and plant productivity. Although single gene transformations may not completely reduce photorespiration, increases in plant biomass accumulation have still been observed indicating an alternative role in regulating different metabolic processes. Therefore, the current study was aimed at evaluating the underlying mechanism (s) associated with the effects of introducing a single cyanobacterial glycolate decarboxylation pathway gene on photosynthesis and plant performance. METHODS: Transgenic Arabidopsis thaliana plants (GD, HD, OX) expressing independently cyanobacterial decarboxylation pathway genes i.e., glycolate dehydrogenase, hydroxyacid dehydrogenase, and oxalate decarboxylase, respectively, were utilized. Photosynthetic, fluorescence related, and growth parameters were analyzed. Additionally, transcriptomic analysis of GD transgenic plants was also performed. RESULTS: The GD plants exhibited a significant increase (16%) in net photosynthesis rate while both HD and OX plants showed a non-significant (11%) increase as compared to wild type plants (WT). The stomatal conductance was significantly higher (24%) in GD and HD plants than the WT plants. The quantum efficiencies of photosystem II, carbon dioxide assimilation and the chlorophyll fluorescence-based photosynthetic electron transport rate were also higher than WT plants. The OX plants displayed significant reductions in the rate of photorespiration relative to gross photosynthesis and increase in the ratio of the photosynthetic electron flow attributable to carboxylation reactions over that attributable to oxygenation reactions. GD, HD and OX plants accumulated significantly higher biomass and seed weight. Soluble sugars were significantly increased in GD and HD plants, while the starch levels were higher in all transgenic plants. The transcriptomic analysis of GD plants revealed 650 up-regulated genes mainly related to photosynthesis, photorespiratory pathway, sucrose metabolism, chlorophyll biosynthesis and glutathione metabolism. CONCLUSION: This study revealed the potential of introduced cyanobacterial pathway genes to enhance photosynthetic and growth-related parameters. The upregulation of genes related to different pathways provided evidence of the underlying mechanisms involved particularly in GD plants. However, transcriptomic profiling of HD and OX plants can further help to identify other potential mechanisms involved in improved plant productivity.

A cyanobacterial photorespiratory bypass model to enhance photosynthesis by rerouting photorespiratory pathway in C3 plants.

Plants employ photosynthesis to produce sugars for supporting their growth. During photosynthesis, an enzyme Ribulose 1,5 bisphosphate carboxylase/oxygenase (Rubisco) combines its substrate Ribulose 1,5 bisphosphate (RuBP) with CO2 to produce phosphoglycerate (PGA). Alongside, Rubisco also takes up O2 and produce 2-phosphoglycolate (2-PG), a toxic compound broken down into PGA through photorespiration. Photorespiration is not only a resource-demanding process but also results in CO2 loss which affects photosynthetic efficiency in C3 plants. Here, we propose to circumvent photorespiration by adopting the cyanobacterial glycolate decarboxylation pathway into C3 plants. For that, we have integrated the cyanobacterial glycolate decarboxylation pathway into a kinetic model of C3 photosynthetic pathway to evaluate its impact on photosynthesis and photorespiration. Our results show that the cyanobacterial glycolate decarboxylation bypass model exhibits a 10% increase in net photosynthetic rate (A) in comparison with C3 model. Moreover, an increased supply of intercellular CO2 (Ci) from the bypass resulted in a 54.8% increase in PGA while reducing photorespiratory intermediates including glycolate (- 49%) and serine (- 32%). The bypass model, at default conditions, also elucidated a decline in phosphate-based metabolites including RuBP (- 61.3%). The C3 model at elevated level of inorganic phosphate (Pi), exhibited a significant change in RuBP (+ 355%) and PGA (- 98%) which is attributable to the low availability of Ci. Whereas, at elevated Pi, the bypass model exhibited an increase of 73.1% and 33.9% in PGA and RuBP, respectively. Therefore, we deduce a synergistic effect of elevation in CO2 and Pi pool on photosynthesis. We also evaluated the integrative action of CO2, Pi, and Rubisco carboxylation activity (Vcmax) on A and observed that their simultaneous increase raised A by 26%, in the bypass model. Taken together, the study potentiates engineering of cyanobacterial decarboxylation pathway in C3 plants to bypass photorespiration thereby increasing the overall efficiency of photosynthesis.

Characterization and synergistic antibacterial potential of green synthesized silver nanoparticles using aqueous root extracts of important medicinal plants of Pakistan.

In the past few years, biologically synthesized silver nanoparticles (AgNPs) have been standout amongst the most utilized nanoparticles both in the field of therapeutics and clinical practices. Therefore, the current study aimed to synthesize AgNPs for the first time using aqueous root extracts of important plants of Pakistan i.e. Bergenia ciliata, Bergenia stracheyi, Rumex dantatus and Rumex hastatus and characterize them. In addition, antibacterial activity of synthesized AgNPs at 30-150 µg/well was assessed using well diffusion method against Staphylococcus aureus, Staphylococcus haemolyticus, Bacillus cereus, Escherichia coli, Salmonella typhi and Pseudomonas aeruginosa bacterial strains that are considered most harmful bacteria for human beings. The characterization of synthesized AgNPs showed the absorption maxima ranged from 434 to 451 nm and XRD confirmed the crystalline nature of AgNPs as well as FTIR elucidated the involvement of biomolecules for reduction and capping of AgNPs. SEM determined the average size of AgNPs ranging from 25 to 73 nm and strong signals of silver were captured in EDX images. The result of antibacterial activity showed that only aqueous root extracts of all selected plants were inactive against all the tested bacterial strains. However, importantly, direct relationship between zone of inhibition of S. aureus, S. typhi and P. aeruginosa was found with increasing concentration of AgNPs of each selected plant. Moreover, S. haemolyticus was only inhibited by R. hastatus based AgNPs at only high concentrations and E. coli was inhibited by R. dantatus and R. hastatus based AgNPs. However, B. cereus was not inhibited by any AgNPs except R. hastatus and R. hastatus based AgNPs have greater antibacterial potential among all the synthesized AgNPs. These results suggest that synthesized AgNPs have improved antibacterial potential of root extracts of each selected plant and these synthesized AgNPs could be used in pharmaceutical and homeopathic industry for the cure of human diseases.

Correction: Bioactivity-guided isolation of rosmarinic acid as the principle bioactive compound from the butanol extract of Isodon rugosus against the pea aphid, Acyrthosiphon pisum.

[This corrects the article DOI: 10.1371/journal.pone.0215048.].

Bioactivity-guided isolation of rosmarinic acid as the principle bioactive compound from the butanol extract of Isodon rugosus against the pea aphid, Acyrthosiphon pisum.

Aphids are agricultural pest insects that transmit viruses and cause feeding damage on a global scale. Current pest control practices involving the excessive use of synthetic insecticides over many years have resulted in aphid resistance to a number of pesticides. In nature, plants produce secondary metabolites during their interaction with insects and these metabolites can act as toxicants, antifeedants, anti-oviposition agents and deterrents towards the insects. In a previous study, we demonstrated that the butanol fraction from a crude methanolic extract of an important plant species, Isodon rugosus showed strong insecticidal activity against the pea aphid, Acyrthosiphon pisum. To further explore this finding, the current study aimed to exploit a bioactivity-guided strategy to isolate and identify the active compound in the butanol fraction of I. rugosus. As such, reversed-phase flash chromatography, acidic extraction and different spectroscopic techniques were used to isolate and identify the new compound, rosmarinic acid, as the bioactive compound in I. rugosus. Insecticidal potential of rosmarinic acid against A. pisum was evaluated using standard protocols and the data obtained was analyzed using qualitative and quantitative statistical approaches. Considering that a very low concentration of this compound (LC90 = 5.4 ppm) causes significant mortality in A. pisum within 24 h, rosmarinic acid could be exploited as a potent insecticide against this important pest insect. Furthermore, I. rugosus is already used for medicinal purposes and rosmarinic acid is known to reduce genotoxic effects induced by chemicals, hence it is expected to be safer compared to the current conventional pesticides. While this study highlights the potential of I. rugosus as a possible biopesticide source against A. pisum, it also provides the basis for further exploration and development of formulations for effective field application.

Ethnobotanical appraisal and cultural values of medicinally important wild edible vegetables of Lesser Himalayas-Pakistan.

BACKGROUND: The association among food and health is momentous as consumers now demand healthy, tasty and natural functional foods. Knowledge of such food is mainly transmitted through the contribution of individuals of households. Throughout the world the traditions of using wild edible plants as food and medicine are at risk of disappearing, hence present appraisal was conducted to explore ethnomedicinal and cultural importance of wild edible vegetables used by the populace of Lesser Himalayas-Pakistan. METHODS: Data was collected through informed consent semi-structured interviews, questionnaires, market survey and focus group conversation with key respondents of the study sites including 45 female, 30 children and 25 males. Cultural significance of each species was calculated based on use report. RESULTS: A total of 45 wild edible vegetables belonging to 38 genera and 24 families were used for the treatment of various diseases and consumed. Asteraceae and Papilionoideae were found dominating families with (6 spp. each), followed by Amaranthaceae and Polygonaceae. Vegetables were cooked in water (51%) followed by diluted milk (42%) and both in water and diluted milk (7%). Leaves were among highly utilized plant parts (70%) in medicines followed by seeds (10%), roots (6%), latex (4%), bark, bulb, flowers, tubers and rhizomes (2% each). Modes of preparation fall into seven categories like paste (29%), decoction (24%), powder (14%), eaten fresh (12%), extract (10%), cooked vegetable (8%) and juice (4%). Ficus carica was found most cited species with in top ten vegetables followed by Ficus palmata, Bauhinia variegata, Solanum nigrum, Amaranthus viridis, Medicago polymorpha, Chenopodium album, Cichorium intybus, Amaranthus hybridus and Vicia faba. CONCLUSIONS: Patterns of wild edible plant usage depend mainly on socio-economic factors compare to climatic conditions or wealth of flora but during past few decades have harshly eroded due to change in the life style of the inhabitants. Use reports verified common cultural heritage and cultural worth of quoted taxa is analogous. Phytochemical analysis, antioxidant activities, pharmacological applications; skill training in farming and biotechnological techniques to improve the yield are important feature prospective regarding of wild edible vegetables.

Phytoremediation potential of Arundo donax in arsenic-contaminated synthetic wastewater.

The present study reports the potential of Arundo donax for phytoextraction of arsenic from synthetic wastewater. A. donax plants were grown under greenhouse conditions in pots containing a nutrient solution amended with increasing doses of As (0, 50, 100, 300, 600 and 1000 microg L(-1)) for 21 days in a completely randomized design. Shoot and roots dry matter production, growth parameters, arsenic and nutrient tissue concentrations were measured at the end of the experiment. Increasing As concentration in nutrient solution caused an increase in shoot and root biomass without toxicity symptoms in A. donax growing under a range of As concentration from 50 to 600 microg L(-1). Elevated oxidative stress was observed at As supplied level of 1000 microg L(-1). The As doses up to 600 microg L(-1) did not affect the growth of A. donax. It is suggested that A. donax plants may be employed to treat contaminated waters containing arsenic concentrations up to 600 microg L(-1).