Foliar Application of Rhizobium leguminosarum bv. viciae Strain 33504-Borg201 Promotes Faba Bean Growth and Enhances Systemic Resistance Against Bean Yellow Mosaic Virus Infection.

The bean yellow mosaic virus (BYMV) is one of the most serious economic diseases affecting faba bean crop production. Rhizobium spp., well known for its high nitrogen fixation capacity in legumes, has received little study as a possible biocontrol agent and antiviral. Under greenhouse conditions, foliar application of molecularly characterized Rhizobium leguminosarum bv. viciae strain 33504-Borg201 to the faba bean leaves 24 h before they were infected with BYMV made them much more resistant to the disease while also lowering its severity and accumulation. Furthermore, the treatment promoted plant growth and health, as evidenced by the increased total chlorophyll (32.75 mg/g f.wt.) and protein content (14.39 mg/g f.wt.), as well as the improved fresh and dry weights of the plants. The protective effects of 33504-Borg201 greatly lowered the levels of hydrogen peroxide (H2O2) (4.92 µmol/g f.wt.) and malondialdehyde (MDA) (173.72 µmol/g f.wt.). The antioxidant enzymes peroxidase (1.58 µM/g f.wt.) and polyphenol oxidase (0.57 µM/g f.wt.) inhibited the development of BYMV in plants treated with 33504-Borg201. Gene expression analysis showed that faba bean plants treated with 33504-Borg201 had higher amounts of pathogenesis-related protein-1 (PR-1) (3.28-fold) and hydroxycinnamoyl-CoA quinate hydroxycinnamoyltransferase (4.13-fold) than control plants. These findings demonstrate the potential of 33,504-Borg201 as a cost-effective and eco-friendly method to protect faba bean plants against BYMV. Implementing this approach could help develop a simple and sustainable strategy for protecting faba bean crops from the devastating effects of BYMV.

Synthesis of Tungsten Oxide, Iron Oxide, and Copper-Doped Iron Oxide Nanocomposites and Evaluation of Their Mixing Effects with Cyromazine against Spodoptera littoralis (Boisduval).

Nanotechnology research is emerging as a cutting-edge technology, and nanocomposites have played a significant role in pest control. Therefore, the present study focuses on the synthesis of tungsten oxide (WO3), iron oxide (magnetic nanoparticle, MNP), and copper-doped iron oxide (MNP-Cu) nanocomposites and explores the different effects of their binary combinations with the insecticide cyromazine against Spodoptera littoralis. The synthesized nanoparticles were characterized by transmission electron microscopy, X-ray diffraction, Fourier-transform infrared spectroscopy, and Raman spectroscopy. None of the tested nanomaterials showed any toxicity against the different stages of S. littoralis. Larval and pupal durations increased with increasing cyromazine and nanomaterial concentrations. The longest larval and pupal durations were recorded under treatment with the mixture of cyromazine (100 mg/L) + MNP-Cu (500 mg/L); the survival periods were 23.5 and 15.6 days, compared with 10.8 and 7.7 days in the control, respectively. The percentages of pupation and adult emergence were negatively affected by all treatments. Among the 500 mg/L nanomaterial combinations, only cyromazine (25 mg/L) and WO3 (500 mg/L) resulted in adult emergence (at a rate of 27.3%). Some abnormalities in the S. littoralis stages were observed following treatment with the tested materials. The glutathione S-transferase and alpha-esterase enzyme activities in S. littoralis were significantly increased after treatment with cyromazine, followed by cyromazine/MNP-Cu combinations. The quantitative polymerase chain reaction (Q-PCR) data showed that all treated insects had a higher immune response than the control. Finally, mixes of nanocomposites and cyromazine may be suggested as viable alternatives for S. littoralis management.

Antiviral Activity of Biosynthesized Silver Nanoparticles from Pomegranate (Punica granatum L.) Peel Extract against Tobacco Mosaic Virus.

Tobacco mosaic virus (TMV) is a major pathogen affecting tomato plants worldwide. The efficacy of silver nanoparticles (Ag-NPs) mediated by Punica granatum biowaste peel extract in mitigating the negative impact of TMV infection on tomato growth and oxidative stress was investigated through scanning electron microscopy (SEM), transmission electron microscopy (TEM), UV-Visible (UV-Vis) spectrophotometer, X-ray Diffraction (XRD), dynamic light scattering (DLS), zeta potential, energy-dispersive X-ray spectroscopy (EDX), and Fourier-transform infrared spectra (FTIR). Results of SEM analysis of green Ag-NPs revealed the presence of condensed spherical or round NPs with diameters ranging between 61 and 97 nm. TEM confirmed the SEM results and showed round-shaped Ag-NPs with an average size of 33.37 ± 12.7 nm. The elemental analysis (EDX) of prepared Ag-NPs revealed the presence of elemental Ag as a major peak (64.43%) at 3-3.5 KeV. The FTIR revealed several functional groups on the prepared Ag-NPs, for which three treatment strategies for Ag-NP applications were evaluated in the greenhouse study and compared to inoculated TMV and control plants: pre-infection treatment (TB), post-infection treatment (TA), and dual treatment (TD). The results showed that the TD strategy is the most effective in improving tomato growth and reducing viral replication, whereas all Ag-NP treatments (TB, TA, and TD) were found to significantly increase expression of the pathogenesis-related (PR) genes PR-1 and PR-2, as well as polyphenolic compounds, HQT, and C4H genes compared to control plants. In contrast, the flavonoid content of tomato plants was not affected by the viral infection, while the phenolic content was significantly reduced in the TMV group. Furthermore, TMV infection led to a significant increase in oxidative stress markers MDA and H2O2, as well as a reduction in the enzymatic activity of the antioxidants PPO, SOD, and POX. Our results clearly showed that the application of Ag-NPs on TMV-infected plants reduces virus accumulation, delays viral replication in all treatments, and greatly enhances the expression of the CHS gene involved in flavonoid biosynthesis. Overall, these findings suggest that treatment with Ag-NPs may be an effective strategy to mitigate the negative impact of TMV infection on tomato plants.

Foliar Application of Nanoclay Promotes Potato (Solanum tuberosum L.) Growth and Induces Systemic Resistance against Potato Virus Y.

Potato virus Y (PVY) is one of the most harmful phytopathogens. It causes big problems for potatoes and other important crops around the world. Nanoclays have been extensively studied for various biomedical applications. However, reports on their interactions with phytopathogens, particularly viral infections, are still limited. In this study, the protective activity of Egyptian nanoclay (CE) and standard nanoclay (CS) against PVY was evaluated on potato (Solanum tuberosum L.) plants. Their physicochemical and morphological properties were examined with scanning electron microscopy (SEM), transmission electron microscopy (TEM), Fourier-transform infrared spectroscopy (FTIR), and energy dispersive spectrometer (EDS). SEM and TEM analyses revealed that CE has a spherical and hexagonal structure ranging from 20 to 80 nm in size, while CS has boulder-like and tubular structures of about 320 nm in size. FTIR and EDS showed that both nanoclay types have different functional groups and contain many vital plant nutrients that are necessary for every stage and process of the plant, including development, productivity, and metabolism. Under greenhouse conditions, a 1% nanoclay foliar application enhanced potato growth, reduced disease symptoms, and reduced PVY accumulation levels compared with non-treated plants. Significant increases in levels of antioxidant enzymes (PPO and POX) and considerable decreases in oxidative stress markers (MDA and H2O2) were also reported. Moreover, a significant increase in the transcriptional levels of defense-related genes (PAL-1, PR-5, and CHI-2) was observed. All experiment and analysis results indicate that the CE type is more effective than the CS type against PVY infection. Based on these results, the foliar applications of nanoclay could be used to manage plant viral infections in a way that is both effective and environmentally friendly. To our knowledge, this is the first report of the antiviral activity of the foliar application of nanoclay against PVY infection.

Induction of Systemic Resistance to Tobacco mosaic virus in Tomato through Foliar Application of Bacillus amyloliquefaciens Strain TBorg1 Culture Filtrate.

The application of microbe-derived products as natural biocontrol agents to boost systemic disease resistance to virus infections in plants is a prospective strategy to make agriculture more sustainable and environmentally friendly. In the current study, the rhizobacterium Bacillus amyloliquefaciens strain TBorg1 was identified based on 16S rRNA, rpoB, and gyrA gene sequences, and evaluated for its efficiency in conferring protection of tomato from infection by Tobacco mosaic virus (TMV). Under greenhouse circumstances, foliar sprays of TBorg1 culture filtrate (TBorg1-CF) promoted tomato growth, lowered disease severity, and significantly decreased TMV accumulation in systemically infected leaves of treated plants relative to untreated controls. TMV accumulation was reduced by 90% following the dual treatment, applied 24 h before and after TMV infection. Significant increases in levels of total soluble carbohydrates, proteins, and ascorbic acid were also found. In addition, a significant rise in activities of enzymes capable of scavenging reactive oxygen species (PPO and POX), as well as decreased levels of non-enzymatic oxidative stress markers (H2O2 and MDA) were observed, compared to untreated plants. Enhanced systemic resistance to TMV was indicated by significantly increased transcript accumulation of polyphenolic pathway (C4H, HCT, and CHI) and pathogenesis-related (PR-1 and PR-5) genes. Out of the 15 compounds identified in the GC-MS analysis, 1,2-benzenedicarboxylic acid mono(2-ethylhexyl) ester and phenol, 2,4-bis(1,1-dimethylethyl), as well as L-proline, N-valeryl-, and heptadecyl ester were present in the highest concentrations in the ethyl acetate extract of TBorg1-CF. In addition, significant amounts of n-hexadecanoic acid, pyrrolo [1,2-a] pyrazine-1,4-dione hexahydro-3-(2-methylpropyl)-, nonane, 5-butyl-, and eicosane were also detected. These compounds may act as inducers of systemic resistance to viral infection. Our findings indicate that the newly isolated B. amyloliquefaciens strain TBorg1 could be a potentially useful rhizobacterium for promoting plant growth and a possible source of biocontrol agents for combating plant virus infections.

Quantitative detection of induced systemic resistance genes of potato roots upon ethylene treatment and cyst nematode, Globodera rostochiensis, infection during plant-nematode interactions.

Potato cyst nematodes caused by Globodera rostochiensis, are quarantine-restricted pests causing significant yield losses to potato growers. The phytohormone ethylene play significant roles in various plant-pathogen interactions, however, the molecular knowledge of how ethylene influences potato-nematode interaction is still lacking. Precise detection of potato-induced genes is essential for recognizing plant-induced systemic resistance (ISR). Candidate genes or PR- proteins with putative functions in modulating the response to potato cyst nematode stress were selected and functionally characterized. Using real-time polymerase chain reaction (RT-PCR), we measured the quantified expression of four pathogenesis-related (PR) genes, PR2, PR3, peroxidase, and polyphenol oxidase. The activation of these genes is intermediate during the ISR signaling in the root tissues. Using different ethylene concentrations could detect and induce defense genes in infected potato roots compared to the control treatment. The observed differences in the gene expression of treated infected plants are because of different concentrations of ethylene treatment and pathogenicity. Besides, the overexpressed or suppressed of defense- related genes during developmental stages and pathogen infection. We concluded that ethylene treatments positively affected potato defensive genes expression levels against cyst nematode infection. The results emphasize the necessity of studying molecular signaling pathways controlling biotic stress responses. Understanding such mechanisms will be critical for the development of broad-spectrum and stress-tolerant crops in the future.

Biodiversity of Pathogenic and Toxigenic Seed-Borne Mycoflora of Wheat in Egypt and Their Correlations with Weather Variables.

Surveillance investigations for pathogenic and toxigenic fungi are important to refine our understanding of their epidemiology and help in predicting their outbreaks. During 2019, 198 samples of wheat grains were collected from 25 wheat-growing governorates in Egypt to detect and identify seed-borne mycoflora in vitro. Forty-four fungal species belonging to 20 genera were identified. Molecular data for these fungi were analyzed to construct a phylogenetic tree. Occurrence and biodiversity indicators were calculated. Two prevalent pathogens (average incidence > 40%) were Alternaria alternata and Cladosporium spp. Ustilago tritici was present in only seven of the 25 governorates, and less abundant than Tilletia tritici, the causal agent of stinking smut. Sinai governorate recorded the greatest species diversity, while the greatest species richness was in Qena and Sohag governorates. Canonical correspondence analysis of data for 20 fungal genera with temperature, relative humidity, precipitation, wind speed or solar radiation revealed that relative humidity was the most influential weather variable. It showed that occurrence and distribution of the 20 genera corresponded well with three out of four Egyptian climatic regions: Mediterranean, semi-arid, and arid. Knowing pathogen occurrence and distribution in Egypt is the first step to developing future disease management strategies to limit yield losses and improve food security. Despite this study being conducted on the wheat-growing areas in Egypt, our findings are useful for other wheat-growing countries that share the same climatic conditions. The correlation between a given fungus and the climatic variables can be useful in other ecosystems.

Rhizophagus irregularis and Rhizoctonia solani Differentially Elicit Systemic Transcriptional Expression of Polyphenol Biosynthetic Pathways Genes in Sunflower.

Plant roots are exposed to penetration by different biotrophic and necrotrophic fungi. However, plant immune responses vary, depending on the root-penetrating fungus. Using qRT-PCR, changes over time in the systemic transcriptional expression of the polyphenol biosynthesis-related genes were investigated in sunflower plants in response to colonization with Rhizophagus irregularis and/or infection with Rhizoctonia solani. The results demonstrated that both fungi systemically induced the transcriptional expression of most of the addressed genes at varying degrees. However, the inducing effect differed according to the treatment type, plant organ, targeted gene, and time stage. The inducing effect of R. irregularis was more prevalent than R. solani in the early stages. In general, the dual treatment showed a superior inducing effect over the single treatments at most of the time. The hierarchical clustering analysis showed that cinnamate-4-hydroxylase was the master expressed gene along the studied time period. The cell wall lignification was the main plant-defensive-mechanism induced. In addition, accumulations of chlorogenic acid, flavonoids, and anthocyanins were also triggered. Moreover, colonization with R. irregularis improved the plant growth and reduced the disease severity. We can conclude that the proactive, rather than curative, colonization with R. irregularis is of great importance, owing to their protective and growth-promoting roles, even if no infection occurred.

Arbuscular Mycorrhizal Fungi Trigger Transcriptional Expression of Flavonoid and Chlorogenic Acid Biosynthetic Pathways Genes in Tomato against Tomato Mosaic Virus.

Tomato mosaic disease, caused by Tomato Mosaic Virus (ToMV), is one of the most destructive diseases which results in serious crop losses. Research investigations dealing with the biocontrol activity of arbuscular mycorrhizal fungi (AMF) against this viral disease are limited. In this study, the biocontrol activity of AMF on tomato plants infected with ToMV was evaluated in the greenhouse. In addition, their impacts on the transcriptional expression levels of thirteen genes controlling the phenylpropanoid, flavonoid and chlorogenic acid biosynthetic pathways were also investigated using quantitative real-time PCR. Transcriptional expressions of the majority of the studied genes were up-regulated by mycorrhizal colonization in the presence of ToMV, particularly PAL1 and HQT, suggesting their pathogen-dependent inducing effect. Under greenhouse conditions, a significant reduction in the disease severity and incidence, as well as the viral accumulation level was observed as a response to the mycorrhizal colonization of the infected plants. Moreover, the evaluated growth parameters, photosynthetic pigments, and flavonoid content were significantly enhanced by AMF colonization. The obtained results demonstrated the protective role of AMF in triggering the plant immunity against ToMV in a pathogen-dependent manner. Beside their protective and growth-promotion activities, AMF are characterized by low-cost and environment-friendly properties which support their possible use for control of tomato mosaic disease.