Evaluation of antifungal potentials of Albizia kalkora extract as a natural fungicide: In vitro and computational studies.

The antifungal bioactivity potential of the organic extract of silk tree (Albizia kalkora) was investigated in the current study. The crude extracts of A. kalkora and methanol, n-hexane, chloroform, and ethyl acetate fractions were prepared. The antifungal activity of obtained fractions of A. kalkora was studied at different concentrations ranging from 0.39-50 µg/mL. Dimethyl sulfoxide (DMSO) was taken as a toxicity control, whereas thiophanate methyl (TM) as a positive control. All the fractions significantly reduced the FOL growth (methanolic: 9.49-94.93 %, n-hexane: 11.12-100 %, chloroform: 20.96-91.41 %, and ethyl acetate: 18.75-96.70 %). The n-hexane fraction showed 6.25 µg/mL MIC as compared to TM with 64 µg/mL MIC. The non-polar (n-hexane) fraction showed maximum antifungal bioactivity against FOL in comparison with chloroform, methanol, and ethyl acetate fractions. GC/MS analysis exhibited that the n-hexane fraction contained hexadecanoic acid, 9,12,15-octadecatrienoic acid, 9,12-octadecadienoic acid, bis(2-ethylhexyl) phthalate, methyl stearate, and [1,2,4]triazolo[1,5-a]pyrimidine-6-carboxylic acid. The results of in vitro antifungal inhibition were further reinforced by molecular docking analysis. Five virulence proteins of FOL i.e., pH-responsive PacC transcription factor (PACC), MeaB, TOR; target of rapamycin (FMK1), Signal transducing MAP kinase kinase (STE-STE7), and High Osmolarity Glycerol 1(HOG1) were docked with identified phytocompounds in the n-hexane fraction by GC/MS analysis. MEAB showed maximum binding affinities with zinnimide (-12.03 kcal/mol), HOG1 and FMK1with α-Tocospiro-B (-11.51 kcal/mol) and (-10.55 kcal/mol) respectively, STE-STE7 with docosanoic acid (-11.31 kcal/mol), and PACC with heptadecanoic acid (-9.88 kcal/mol) respectively with strong hydrophobic or hydrophilic interactions with active pocket residues. In conclusion, the n-hexane fraction of the A. kalkora can be used to manage FOL.

Comparative Effect of Seed Coating and Biopriming of Bacillus aryabhattai Z-48 on Seedling Growth, Growth Promotion, and Suppression of Fusarium Wilt Disease of Tomato Plants.

Beneficial plant microbes can enhance the growth and quality of field crops. However, the benefits of microbes using cheap and efficient inoculation methods are still uncommon. Seed coating with biocontrol agents can reduce the amount of inocula along with having the potential for large-scale application. Hence, in this research work, the comparative potential of tomato seed coating and biopriming with Bacillus aryabhattai Z-48, harboring multiple plant-beneficial traits, to suppress Fusarium wilt disease along with its beneficial effect on seedling and plant growth promotion was analyzed. Among two bacterial strains, B. aryabhattai Z-48 was able to antagonize the mycelial growth of Fusarium oxysporum f.sp. lycopersici in vitro and its application as a seed coating superiorly benefited seedling traits like the germination percentage, vigor index, and seedling growth index along with a reduced germination time. The seed coating with B. aryabhattai Z-48 resulted in significant increases in the shoot length, root length, dry biomass, and total chlorophyll contents when compared with the bioprimed seeds with the same bacterial strain and non-inoculated control plants. The seed coating with B. aryabhattai Z-48 significantly reduced the disease index (>60%) compared with the pathogen control during pot trials. Additionally, the seed coating with B. aryabhattai Z-48 resulted in a significantly higher production of total phenolics, peroxidase, polyphenol oxidase, and phenylalanine ammonia lyase enzyme in tomato plants. The GC/MS-based non-targeted metabolic profiling indicated that the seed coating with B. aryabhattai Z-48 could cause large-scale metabolite perturbations in sugars, sugar alcohols, amino acids, and organic acids to increase the fitness of tomato plants against biotic stress. Our study indicates that a tomato seed coating with B. aryabhattai Z-48 can improve tomato growth and suppress Fusarium wilt disease effectively under conventional agricultural systems.

Harnessing the Biocontrol Potential of Bradyrhizobium japonicum FCBP-SB-406 to Manage Charcoal Rot of Soybean with Increased Yield Response for the Development of Sustainable Agriculture.

Plant growth-promoting bacteria (PGPRs) have the potential to act as biofertilizers and biopesticides. This study was planned to explore indigenously isolated PGPRs as a potential candidate to control charcoal rot that affects various crops including soybean. Among the four different tested species of PGPRs, Bradyrhizobium japonicum (FCBP-SB-406) showed significant potential to enhance growth and control soil borne pathogens such as Macrophomina phaseolina. Bacillus subtilis (FCBP-SB-324) followed next. Bradyrhizobium japonicum (FCBP-SB-406) reduced disease severity up to 81.25% in comparison to the control. The strain showed a strong fertilizing effect as a highly significant increase in biomass and other agronomic parameters was recorded in plants grown in its presence. The same was supported by the Pearson's correlation and principal component analysis. A decrease in disease incidence and severity may be due to the induced resistance imparted by the bacterium. This resulted in significant increments in quantities of defense enzymes, including catalase, peroxidase (PO), polyphenol oxidase (PPO), phenylalanine ammonia lyase (PAL) and superoxide dismutase (SOD). A significant production of proteases, catalases and hydrogen cyanide by B. japonicum (FCBP-SB-406) can also be associated to mycoparasitism. The establishment of PGPRs in treated soils also showed positive effects on soil health. Total metabolite profiling of treated plants in comparison to the control showed the upregulation of many flavonoids, isoflavonoids and amino acids. Many of these compounds have been well reported with antimicrobial activities. Bradyrhizobium japonicum (FCBP-SB-406) can be employed for the production of a potential formulation to support sustainable agriculture by reducing the input of synthetic pesticides and fertilizers.

The Imperative Use of Bacillus Consortium and Quercetin Contributes to Suppress Fusarium Wilt Disease by Direct Antagonism and Induced Resistance.

Fusarium wilt diseases severely influence the growth and productivity of numerous crop plants. The consortium of antagonistic rhizospheric Bacillus strains and quercetin were evaluated imperatively as a possible remedy to effectively manage the Fusarium wilt disease of tomato plants. The selection of Bacillus strains was made based on in-vitro antagonistic bioassays against Fusarium oxysporum f.sp. lycoprsici (FOL). Quercetin was selected after screening a library of phytochemicals during in-silico molecular docking analysis using tomato LysM receptor kinases "SILKY12" based on its dual role in symbiosis and plant defense responses. After the selection of test materials, pot trials were conducted where tomato plants were provided consortium of Bacillus strains as soil drenching and quercetin as a foliar spray in different concentrations. The combined application of consortium (Bacillus velezensis strain BS6, Bacillus thuringiensis strain BS7, Bacillus fortis strain BS9) and quercetin (1.0 mM) reduced the Fusarium wilt disease index up to 69%, also resulting in increased plant growth attributes. Likewise, the imperative application of the Bacillus consortium and quercetin (1.0 mM) significantly increased total phenolic contents and activities of the enzymes of the phenylpropanoid pathway. Non-targeted metabolomics analysis was performed to investigate the perturbation in metabolites. FOL pathogen negatively affected a range of metabolites including carbohydrates, amino acids, phenylpropanoids, and organic acids. Thereinto, combined treatment of Bacillus consortium and quercetin (1.0 mM) ameliorated the production of different metabolites in tomato plants. These findings prove the imperative use of Bacillus consortium and quercetin as an effective and sustainable remedy to manage Fusarium wilt disease of tomato plants and to promote the growth of tomato plants under pathogen stress conditions.

Chitinase of Trichoderma longibrachiatum for control of Aphis gossypii in cotton plants.

Chitinase-producing fungi have now engrossed attention as one of the potential agents for the control of insect pests. Entomopathogenic fungi are used in different regions of the world to control economically important insects. However, the role of fungal chitinases are not well studied in their infection mechanism to insects. In this study, Chitinase of entomopathogenic fungi Trichoderma longibrachiatum was evaluated to control Aphis gossypii. For this purpose, fungal chitinase (Chit1) gene from the genomic DNA of T. longibrachiatum were isolated, amplified and characterised. Genomic analysis of the amplified Chit1 showed that this gene has homology to family 18 of glycosyl hydrolyses. Further, Chit1 was expressed in the cotton plant for transient expression through the Geminivirus-mediated gene silencing vector derived from Cotton Leaf Crumple Virus (CLCrV). Transformed cotton plants showed greater chitinase activity than control, and they were resistant against nymphs and adults of A. gossypii. About 38.75% and 21.67% mortality of both nymphs and adults, respectively, were observed by using Chit1 of T. longibrachiatum. It is concluded that T. longibrachiatum showed promising results in controlling aphids by producing fungal chitinase in cotton plants and could be used as an effective method in the future.

Bio-fabrication of Zinc Oxide nanoparticles to rescue Mung Bean against Cercospora leaf spot disease.

Plant disease management using nanotechnology is evolving continuously across the world. The purpose of this study was to determine the effect of different concentrations of green synthesized zinc oxide nanoparticles (ZnO NPs) using Trachyspermum ammi seed extract on Cercospora leaf spot disease in mung bean plants under in-vitro and in-planta conditions. Additionally, the effects on mung bean agronomic and physiological parameters were also assessed. The green synthesized ZnO NPs were characterized using UV-visible spectroscopy, Fourier-transform infrared spectroscopy (FTIR), X-ray diffraction (XRD) and Scanning electron microscopy (SEM). Green synthesized NPs were tested for their ability to inhibit fungal growth at five different concentrations under in-vitro experiment. After 7 days of inoculation, ZnO NPs (1200 ppm) inhibited mycelial growth substantially (89.86% ± 0.70). The in-planta experiment showed statistically significant result of disease control (30% ± 11.54) in response to 1200 ppm ZnO NPs. The same treatment showed statistically significant improvements in shoot length, root length, number of leaves, number of pods, shoot fresh weight (28.62%), shoot dry weight (85.18%), root fresh weight (38.88%), and root dry weight (38.88%) compared to the control. Our findings show that green synthesized ZnO NPs can control Cercospora canescens in mung bean, pointing to their use in plant disease control and growth enhancement.

Green synthesis and application of GO nanoparticles to augment growth parameters and yield in mungbean (Vigna radiata L.).

Plant growth promotion has long been a challenge for growers all over the world. In this work, we devised a green nanomaterial-assisted approach to boost plant growth. It has been reported that carbon nanomaterials are toxic to plants because they can inhibit the uptake of nutrients if employed in higher concentrations, however this study shows that graphene oxide (GO) can be used as a regulator tool to improve plant growth and stability. Graphene oxide in different concentrations was added to the soil of mungbean. It is proved that when a suitable amount of graphene oxide was applied, it had a good influence on plant growth by enhancing the length of roots and shoots, number of leaves, number of root nodules per plant, number of pods, and seeds per pod. We presume that the use of bio-fabricated graphene oxide as a strategy would make it possible to boost both plant growth and the significant increase in the number of seeds produced by each plant.

Magnetofection approach for the transformation of okra using green iron nanoparticles.

Climate change, pesticide resistance, and the need for developing new plant varieties have galvanized biotechnologists to find new solutions in order to produce transgenic plants. Over the last decade scientists are working on green metallic nanoparticles to develop DNA delivery systems for plants. In the current study, green Iron nanoparticles were synthesized using leaf extract of Camellia sinensis (green tea) and Iron Chloride (FeCl3), the characterization and Confirmation was done using UV-VIS Spectroscopy, FTIR, SEM, and TEM. Using these nanoparticles, a novel method of gene transformation in okra plants was developed, with a combination of different Magnetofection factors. Maximum gene transformation efficiency was observed at the DNA to Iron-nanoparticles ratio of 1:20, by rotation of mixture (Plasmid DNA, Iron-nanoparticles, and seed embryo) at 800 rpm for 5 h. Using this approach, the transformation of the GFP (green fluorescent protein) gene was successfully carried out in Abelmoschus esculentus (Okra plant). The DNA transformation was confirmed by observing the expression of transgene GFP via Laser Scanning Confocal Microscope (LSCM) and PCR. This method is highly economical, adaptable, genotype independent, eco-friendly, and time-saving as well. We infer that this approach can be a potential solution to combat the yield and immunity challenges of plants against pathogens.

Antifungal Potential of Green Synthesized Magnetite Nanoparticles Black Coffee-Magnetite Nanoparticles Against Wilt Infection by Ameliorating Enzymatic Activity and Gene Expression in Solanum lycopersicum L.

Tomato plants are prone to various biotic and abiotic stresses. Fusarium wilt is one of the most devasting diseases of tomatoes caused by Fusarium oxysporum f. sp. lycopersici, causing high yield and economic losses annually. Magnetite nanoparticles (Fe3O4 NPs) are one of the potent candidates to inhibit fungal infection by improving plant growth parameters. Spinach has been used as a starting material to synthesize green-synthesized iron oxide nanoparticles (IONPs). Various extracts, i.e., pomegranate juice, white vinegar, pomegranate peel, black coffee (BC), aloe vera peel, and aspirin, had been used as reducing/stabilizing agents to tune the properties of the Fe3O4 NPs. After utilizing spinach as a precursor and BC as a reducing agent, the X-ray diffraction (XRD) pattern showed cubic magnetite (Fe3O4) phase. Spherical-shaped nanoparticles (∼20 nm) with superparamagnetic nature indicated by scanning electron microscopy (SEM) monographs, whereas energy-dispersive X-ray gives good elemental composition in Fe3O4 NPs. A characteristic band of Fe-O at ∼ 561 cm-1 was exhibited by the Fourier transform infrared (FTIR) spectrum. X-ray photoelectron spectroscopy (XPS) results confirmed the binding energies of Fe 2p3/2 (∼710.9 eV) and Fe 2p1/2 (∼724.5 eV) while, Raman bands at ∼310 cm-1 (T2 g ), ∼550 cm-1 (T2 g ), and 670 cm-1 (A1 g ) indicated the formation of Fe3O4 NPs synthesized using BC extract. The in vitro activity of BC-Fe3O4 NPs significantly inhibited the mycelial growth of F. oxysporum both at the third and seventh day after incubation, in a dose-dependent manner. In vivo studies also exhibited a substantial reduction in disease severity and incidence by improving plant growth parameters after treatment with different concentrations of BC-Fe3O4 NPs. The increasing tendency in enzymatic activities had been measured after treatment with different concentrations of NPs both in roots and shoot of tomato plants as compared to the control. Correspondingly, the upregulation of PR-proteins and defense genes are in line with the results of the enzymatic activities. The outcome of the present findings suggests that Fe3O4 NPs has the potential to control wilt infection by enhancing plant growth. Hence, Fe3O4 NPs, being non-phytotoxic, have impending scope in the agriculture sector to attain higher yield by managing plant diseases.

Structure Elucidation and Toxicity Analysis of the Byproducts Formed after Biodegradation of Aflatoxins B1 and B2 Using Extracts of Mentha arvensis.

The aqueous extracts of leaves and shoots of Mentha arvensis were checked for their potential to biodegrade aflatoxin B1 and B2 (AFB1; 100 µg/L and AFB2; 50 µg/L) through in vitro assays. Overall, the results showed that leaf extract degrades aflatoxins more efficiently than the shoot extract. First, the pH, temperature and incubation time were optimized for maximum degradation by observing this activity at different temperatures between 25 and 60 °C, pH between 2 and 10 and incubation time from 3 to 72 h. In general, an increase in all these parameters significantly increased the percentage of biodegradation. In vitro trials on mature maize stock were performed under optimized conditions, i.e., pH 8, temperature 30 °C and an incubation period of 72 h. The leaf extract resulted in 75% and 80% biodegradation of AFB1 and AFB2, respectively. Whereas the shoot extract degraded both toxins up to 40-48%. The structural elucidation of degraded toxin products by LCMS/MS analysis showed seven degraded products of AFB1 and three of AFB2. MS/MS spectra showed that most of the products were formed by the loss of the methoxy group from the side chain of the benzene ring, the removal of the double bond in the terminal furan ring and the modification of the lactone group, indicating less toxicity compared to the parent compounds. The degraded products showed low toxicity against brine shrimps, confirming that M. arvensis leaf extract has significant potential to biodegrade aflatoxins.

Sustainable synthesis of microwave-assisted IONPs using Spinacia oleracea L. for control of fungal wilt by modulating the defense system in tomato plants.

BACKGROUND: Changing climate enhances the survival of pests and pathogens, which eventually affects crop yield and reduces its economic value. Novel approaches should be employed to ensure sustainable food security. Nano-based agri-chemicals provide a distinctive mechanism to increase productivity and manage phytopathogens, with minimal environmental distress. In vitro and in greenhouse studies were conducted to evaluate the potential of green-synthesized iron-oxide nanoparticles (IONPs) in suppressing wilt infection caused by Fusarium oxysporum f. sp. lycospersici, and improving tomato growth (Solanum lycopersicum) and fruit quality. RESULTS: Various microwave powers (100-1000 W) were used to modulate the properties of the green-synthesized IONPs, using spinach as a starting material. The IONPs stabilized with black coffee extract were substantively characterized using X-ray diffraction analysis (XRD), Fourier-transform infrared spectroscopy, dielectric and impedance spectroscopy, X-ray photoelectron spectroscopy (XPS), scanning and transmission electron microscopy (SEM and TEM, respectively), and magnetization analysis. XRD revealed a cubic magnetite (Fe3O4) phase with super-paramagnetic nature, detected at all microwave powers. The binding energies of Fe 2p3/2 (710.9 eV) and Fe 2p1/2 (724.5 eV) of Fe3O4 NPs were confirmed using XPS analysis at a microwave power of 1000 W. Uniform, spherical/cubical-shaped particles with an average diameter of 4 nm were confirmed using SEM and TEM analysis. A significant reduction in mycelial growth and spore germination was observed upon exposure to different IONP treatments. Malformed mycelium, DNA fragmentation, alternation in the cell membrane, and ROS production in F. oxysporum indicated the anti-microbial potential of the IONPs. The particles were applied both through the root (before transplantation) and by means of foliar application (after two weeks) to the infected seedlings. IONPs significantly reduced disease severity by an average of 47.8%, resulting in increased plant growth variables after exposure to 12.5 µg/mL of IONPs. Analysis of photosynthetic pigments, phenolic compounds, and anti-oxidant enzymes in the roots and shoots showed an increasing trend after exposure to various concentrations of IONPs. Correspondingly, lycopene, vitamin C, total flavonoids, and protein content were substantially improved in tomato fruits after treatment with IONPs. CONCLUSION: The findings of the current investigation suggested that the synthesized IONPs display anti-fungal and nutritional properties that can help to manage Fusarium wilt disease, resulting in enhanced plant growth and fruit quality.

First Report of Pseudopestalotiopsis theae Causing Leaf Spot of Date Palm (Phoenix dactylifera) in China.

Date palm (Phoenix dactylifera L.) is a popular landscape tree in Fujian province, in South China. In November 2018 and June 2019, a leaf spot disease was observed on date palm in Fuzhou city. A survey of date palm plants grown in four different locations revealed that the disease incidence was almost 20%. The spots were brown with a yellow margin, 1 to 20 mm in diameter, and oval to irregular. In later stages, the spots gradually expanded and coalesced, became dry and died. For isolation, small pieces (0.5 cm2) were cut from leaf spots obtained from seven trees and disinfested with 70% alcohol. Leaf pieces were then placed onto 2% potato dextrose agar (PDA) and incubated at 25±2°C for 3 to 4 days. One fungus was consistently isolated from fifteen leaves. Fungal colonies were white with undulating margins and a light cream on the reverse side. Black globose to oblate conidiomata were irregularly distributed throughout ten-day-old colonies. The conidiogenous cells were septate, colorless, smooth-walled, straight to slightly curved, ampulliform or subcylindrical, and 6.0 to 13.5 × 1.3 to 3.0 µm [(n=50); x̄ ± SD = 9.5 ± 2× 2 ± 0.5µm]. Conidia were fusiform and five-celled with constrictions at the septa, measuring 18.5 to 31.5 × 5.0 to 7.5 µm [(n=50); x̄ ± SD = 25.5 ± 2 × 6.5± 0.2µm]. The three median cells were light to dark brown and the two end cells were colorless. Apical cells had 2 to 4 appendages ranging from 10.2 to 22.5 µm long. Basal cells had one appendage ranging from 3.5 to 5.5 µm long. The internal transcribed spacer (ITS) region of the ribosomal DNA and translation elongation factor 1-alpha (TEF1-α) gene of fungus were amplified using primers ITS1/ITS4 and EF1728F/EF1986R, respectively. Amplified products (ITS: MN294700 and TEF1-α: MN970514) showed 99% sequence identity to Pestalotiopsis sp., and Pseudoestalotiopsis theae sequences in GenBank. A comparison of MRC12 sequences with the type culture sequences (ITS: JQ683727 and TEF1-a: JQ683743) also showed high similarity, where ITS sequences exhibited only a three-nucleotide difference at the start of the sequences. No differences, however, were found between the TEF1-α sequences. On the basis of morphology and molecular characteristics, the fungus was identified as Ps. theae (Sawada) Maharachch., K.D. Hyde & Crous Steyaert (Maharachchikumbura et al. 2014). To confirm pathogenicity, five disinfested leaves on three healthy five-year-old date palm plants in a nursery (average temperature 26°C), were punctured 3 to 5 times with a sterilized needle, and then 10 to 15 mL conidial suspension (105 conidia/mL in sterilized distilled water) was sprayed over punctured areas of the leaves. For the control treatment, punctured leaves were sprayed with sterilized distilled water. All inoculated leaves plus the control were covered with plastic bags. After 10 days, brown leaf spots similar in appearance to those observed in the field appeared on all wounded leaves, and Ps. theae was successfully re-isolated; the control leaves remained asymptomatic. Previously, Ps. theae was reported on oil palm (Elaeis guineensis Jacq.) from Sierra Leone and Thailand (Turner, 1971; Suwannarach et al. 2013). To our knowledge, this is the first report of Ps. theae on date palm in China. This report expands the host range Ps. theae to date palm and underscores the potential threat of an emerging leaf spot pathogen on Phoenix species. References Maharachchikumbura, K.D., et al. 2014. Stud. Mycol. 79: 121-186. Suwannarach, N., et al. 2013. J. Gen. Plant Pathol. 79: 277-279. Turner, P.D. 1971. Phytopathol. 14: 1-58.

Microwave-Assisted Green Synthesis and Characterization of Silver Nanoparticles Using Melia azedarach for the Management of Fusarium Wilt in Tomato.

These days, research in agriculture is focusing on the theme of sustainability along with protection of agriculture produce. Nanotechnology in the agriculture sector aims for the enhancement of agricultural produce and the reduction of pesticides through providing innovative agrochemical agents and their novel delivery mechanisms. The current investigation involved the green synthesis of silver nanoparticles (AgNPs) from the aqueous leaf extract of Melia azedarach by following a microwave-assisted method to control Fusarium oxysporum, the causal agent of tomato wilt. Biosynthesized Melia leaf extract (MLE)-AgNPs were characterized by UV-visible spectroscopy, Fourier-transform infrared (FTIR) spectroscopy, X-ray diffraction (XRD), energy dispersive X-ray (EDX) spectrometry, dynamic light scattering (DLS), scanning electron microscopy (SEM), transmission electron microscopy (TEM), and zeta potential analysis. The intensity of the peak at 434 nm in UV-vis spectra, attributed to the surface plasmon resonance of MLE-AgNPs, changes with reaction parameters. TEM exhibits spherical shaped nanoparticles with an average particle size range from 12 to 46 nm. Efficient inhibition of F. oxysporum, the causal agent of tomato wilt, was achieved after exposure to MLE-AgNPs both in vivo and in vitro. In vitro studies exhibited repressed fungal mycelial growth with 79-98% inhibition as compared to the control. Significant increases in growth parameters of tomato seedlings were observed after treatment with biosynthesized nanoparticles as compared to F. oxysporum-infected plants grown without them under greenhouse conditions. Furthermore, SEM imaging was done to reveal the prominent damage on the cell wall of hyphae and spores after MLE-AgNP treatment. Propidium iodide (PI) staining of mycelium indicated the extent of cell death, causing irretrievable damage and disintegration of cellular membranes by altering the membrane permeability. Also, 2',7'-dichlorofluorescin diacetate (DCFH-DA) fluorescence specifies intracellular reactive oxygen species (ROS) production in F. oxysporum after treatment with MLE-AgNPs. The current investigation suggested that biosynthesized nanoparticles can revolutionize the field of plant pathology by introducing an environment-friendly approach for disease management and playing a potential part in agriculture industry. However, to date, little work has been done to integrate nanotechnology into phytopathology so, this area of research is in need of adoption and exploration for the management of plant diseases.

Genetic diversity and biogeography of T. officinale inferred from multi locus sequence typing approach.

Taraxacum officinale (Asteraceae) is widely distributed weedy plant used as a traditional medicinal herb. The population genetics and historical biogeography of this plant have remained relatively unexplored. This study explores phylogeny, population genetics and ancestral reconstructions adopting multi locus sequence typing (MLST) approach. MLST sequences dataset was generated from genomics and chloroplast DNA sequences obtained from 31 T. officinale haplotypes located in 16 different countries. Phylogenetic analysis distributed these haplotypes in well differentiated geographic clades. The study suggested a close relationship between Europe and adjacent Asian countries. Populations of these regions predominantly formed common haplogroups, showed considerable level of gene flow and evidence for recombination events across European and Asian population. Biogeographical inferences obtained by applying statistical dispersal-vicariance analysis (S-DIVA) and Bayesian binary MCMC (BBM) analysis showed that T. officinale was putatively originated in Europe. Molecular clock analysis based on ITS dataset suggested that the divergence between Europe and East Asian populations can be dated to 1.07 Mya with subsequent dispersal and vicariance events. Among different spatial process long distance seed dispersal mediated by wind had potentially assisted the population expansion of T. officinale.

Identification of a Potential ISR Determinant from Pseudomonas aeruginosa PM12 against Fusarium Wilt in Tomato.

Biocontrol of plant diseases through induction of systemic resistance is an environmental friendly substitute to chemicals in crop protection measures. Different biotic and abiotic elicitors can trigger the plant for induced resistance. Present study was designed to explore the potential of Pseudomonas aeruginosa PM12 in inducing systemic resistance in tomato against Fusarium wilt. Initially the bioactive compound, responsible for ISR, was separated and identified from extracellular filtrate of P. aeruginosa PM12. After that purification and characterization of the bacterial crude extracts was carried out through a series of organic solvents. The fractions exhibiting ISR activity were further divided into sub-fractions through column chromatography. Sub fraction showing maximum ISR activity was subjected to Gas chromatography/mass spectrometry for the identification of compounds. Analytical result showed three compounds in the ISR active sub-fraction viz: 3-hydroxy-5-methoxy benzene methanol (HMB), eugenol and tyrosine. Subsequent bioassays proved that HMB is the potential ISR determinant that significantly ameliorated Fusarium wilt of tomato when applied as soil drench method at the rate of 10 mM. In the next step of this study, GC-MS analysis was performed to detect changes induced in primary and secondary metabolites of tomato plants by the ISR determinant. Plants were treated with HMB and Fusarium oxysporum in different combinations showing intensive re- modulations in defense related pathways. This work concludes that HMB is the potential elicitor involved in dynamic reprogramming of plant pathways which functionally contributes in defense responses. Furthermore the use of biocontrol agents as natural enemies of soil borne pathogens besides enhancing production potential of crop can provide a complementary tactic for sustainable integrated pest management.

Structural Analysis and Biological Toxicity of Aflatoxins B1 and B2 Degradation Products Following Detoxification by Ocimum basilicum and Cassia fistula Aqueous Extracts.

This study showed the comparison between Ocimum basilicum and Cassia fistula (leaves and branch) aqueous extracts for their ability to detoxify of aflatoxins B1 and B2 (AFB1; 100 µg L(-1) and AFB2; 50 µg L(-1)) by In Vitro assays and decontamination studies. Results indicated that O. basilicum leaves extract was found to be highly significant (P < 0.05) in degrading AFB1 and AFB2, i.e., 90.4 and 88.6%, respectively. However, O. basilicum branch, C. fistula leaves and branch extracts proved to be less efficient in degrading these aflatoxins, under optimized conditions, i.e., pH 8, temperature 30°C and incubation period of 72 h. Moreover the antifungal activity of these plants extracts were also tested. The findings depicted that O. basilicum leaves extract showed maximum growth inhibition of aflatoxigenic isolates, i.e., 82-87% as compared to other tested plants extracts. The structural elucidation of degraded toxin products by LCMS/MS analysis showed that nine degraded products of AFB1 and AFB2 were formed. MS/MS spectra showed that most of the products were formed by the removal of double bond in the terminal furan ring and modification of lactone group indicating less toxicity as compared to parent compounds. Brine shrimps bioassay further confirmed the low toxicity of degraded products, showing that O. basilicum leaves extract can be used as an effective tool for the detoxification of aflatoxins.

Phenylacetic Acid Is ISR Determinant Produced by Bacillus fortis IAGS162, Which Involves Extensive Re-modulation in Metabolomics of Tomato to Protect against Fusarium Wilt.

Bacillus fortis IAGS162 has been previously shown to induce systemic resistance in tomato plants against Fusarium wilt disease. In the first phase of current study, the ISR determinant was isolated from extracellular metabolites of this bacterium. ISR bioassays combined with solvent extraction, column chromatography and GC/MS analysis proved that phenylacetic acid (PAA) was the potential ISR determinant that significantly ameliorated Fusarium wilt disease of tomato at concentrations of 0.1 and 1 mM. In the second phase, the biochemical basis of the induced systemic resistance (ISR) under influence of PAA was elucidated by performing non-targeted whole metabolomics through GC/MS analysis. Tomato plants were treated with PAA and fungal pathogen in various combinations. Exposure to PAA and subsequent pathogen challenge extensively re-modulated tomato metabolic networks along with defense related pathways. In addition, various phenylpropanoid precursors were significantly up-regulated in treatments receiving PAA. This work suggests that ISR elicitor released from B. fortis IAGS162 contributes to resistance against fungal pathogens through dynamic reprogramming of plant pathways that are functionally correlated with defense responses.

Structural Elucidation and Toxicity Assessment of Degraded Products of Aflatoxin B1 and B2 by Aqueous Extracts of Trachyspermum ammi.

In this study aqueous extract of seeds and leaves of Trachyspermum ammi were evaluated for their ability to detoxify aflatoxin B1 and B2 (AFB1; 100 µg L(-1) and AFB2; 50 µg L(-1)) by in vitro and in vivo assays. Results indicated that T. ammi seeds extract was found to be significant (P < 0.05) in degrading AFB1 and AFB2 i.e., 92.8 and 91.9% respectively. However, T. ammi leaves extract proved to be less efficient in degrading these aflatoxins, under optimized conditions i.e., pH 8, temperature 30°C and incubation period of 72 h. The structural elucidation of degraded toxin products by LCMS/MS analysis showed that eight degraded products of AFB1 and AFB2 were formed. MS/MS spectra showed that most of the products were formed by the removal of double bond in the terminal furan ring and modification of lactone group indicating less toxicity as compared to parent compounds. Brine shrimps bioassay further confirmed the low toxicity of degraded products, showing that T. ammi seeds extract can be used as an effective tool for the detoxification of aflatoxins.

Mass spectrometric identification and toxicity assessment of degraded products of aflatoxin B1 and B2 by Corymbia citriodora aqueous extracts.

This study explores the detoxification potential of Corymbia citriodora plant extracts against aflatoxin B1 and B2 (AFB1; 100 µg L(-1) and AFB2; 50 µg L(-1)) in In vitro and In vivo assays. Detoxification was qualitatively and quantitatively analyzed by TLC and HPLC, respectively. The study was carried out by using different parameters of optimal temperature, pH and incubation time period. Results indicated that C. citriodora leaf extract(s) more effectively degrade AFB1 and AFB2 i.e. 95.21% and 92.95% respectively than C. citriodora branch extract, under optimized conditions. The structural elucidation of degraded toxin products was done by LCMS/MS analysis. Ten degraded products of AFB1 and AFB2 and their fragmentation pathways were proposed based on molecular formulas and MS/MS spectra. Toxicity of these degraded products was significantly reduced as compared to that of parent compounds because of the removal of double bond in the terminal furan ring. The biological toxicity of degraded toxin was further analyzed by brine shrimps bioassay, which showed that only 17.5% mortality in larvae was recorded as compared to untreated toxin where 92.5% mortality was observed after 96hr of incubation. Therefore, our finding suggests that C. citriodora leaf extract can be used as an effective tool for the detoxification of aflatoxins.

Transcriptional modulation of squalene synthase genes in barley treated with PGPR.

Phytosterol contents and food quality of plant produce is directly associated with transcription of gene squalene synthase (SS). In current study, barley plants were treated with different rhizobacterial strains under semi controlled (27 ± 3°C) greenhouse conditions in order to modulate expression of SS gene. Plant samples were analyzed through semi-quantitative PCR to evaluate effect of rhizobacterial application on transcriptional status of SS. Results revealed that among four SS genes (i.e., SSA, SS1, SS2, and SS3), the most expressive gene was SSA; while, SS2 was screened out as the second best induced gene due to Acetobacter aceti. The most efficient bacterial strain which recorded maximum gene expression was A. aceti AC8. Moreover, AC7 was reported as the least efficient bacterial species for inducing SS gene expression. AC8 enhanced the share of SSA and SS2 up to 43 and 31%, respectively. The study also described ribosomal sequence of the most efficient bacterial strain AC8, which was used to determine its phylogenetic relationships with other microbial strains. The study would be helpful to improve quality of plant produce by modulating transcription of SS genes.