Tobacco Mosaic Virus Accumulation Research Articles

Unveiling novel anti-viral mechanisms of ε-poly-l-lysine on tobacco mosaic virus-infected Nicotiana tabacum through microRNA and transcriptome sequencing

MicroRNAs (miRNAs) play important roles in plant defense against various pathogens. ε-poly-l-lysine (ε-PL), a natural anti-microbial peptide produced by microorganisms, effectively suppresses tobacco mosaic virus (TMV) infection. To investigate the anti-viral mechanism of ε-PL, the expression profiles of miRNAs in TMV-infected Nicotiana tabacum after ε-PL treatment were analyzed. The results showed that the expression levels of 328 miRNAs were significantly altered by ε-PL. Degradome sequencing was used to identify their target genes. Integrative analysis of miRNAs target genes and gene-enriched GO/KEGG pathways indicated that ε-PL regulates the expression of miRNAs involved in critical pathways of plant hormone signal transduction, host defense response, and plant pathogen interaction. Subsequently, virus induced gene silencing combined with the short tandem targets mimic technology was used to analyze the function of these miRNAs and their target genes. The results indicated that silencing miR319 and miR164 reduced TMV accumulation in N. benthamiana, indicating the essential roles of these miRNAs and their target genes during ε-PL-mediated anti-viral responses. Collectively, this study reveals that microbial source metabolites can inhibit plant viruses by regulating crucial host miRNAs and further elucidate anti-viral mechanisms of ε-PL.

Dicer-like2b suppresses the wiry leaf phenotype in tomato induced by tobacco mosaic virus.

Dicer-like (DCL) proteins are principal components of RNA silencing, a major defense mechanism against plant virus infections. However, their functions in suppressing virus-induced disease phenotypes remain largely unknown. Here, we identified a role for tomato (Solanum lycopersicum) DCL2b in regulating the wiry leaf phenotype during defense against tobacco mosaic virus (TMV). Knocking out SlyDCL2b promoted TMV accumulation in the leaf primordium, resulting in a wiry phenotype in distal leaves. Biochemical and bioinformatics analyses showed that 22-nt virus-derived small interfering RNAs (vsiRNAs) accumulated less abundantly in slydcl2b mutants than in wild-type plants, suggesting that SlyDCL2b-dependent 22-nt vsiRNAs are required to exclude virus from leaf primordia. Moreover, the wiry leaf phenotype was accompanied by upregulation of Auxin Response Factors (ARFs), resulting from a reduction in trans-acting siRNAs targeting ARFs (tasiARFs) in TMV-infected slydcl2b mutants. Loss of tasiARF production in the slydcl2b mutant was in turn caused by inhibition of miRNA390b function. Importantly, silencing SlyARF3 and SlyARF4 largely restored the wiry phenotype in TMV-infected slydcl2b mutants. Our work exemplifies the complex relationship between RNA viruses and the endogenous RNA silencing machinery, whereby SlyDCL2b protects the normal development of newly emerging organs by excluding virus from these regions and thus maintaining developmental silencing.

CASC3 Biomolecular Condensates Restrict Turnip Crinkle Virus by Limiting Host Factor Availability

The exon-junction complex (EJC) plays a role in post-transcriptional gene regulation and exerts antiviral activity towards several positive-strand RNA viruses. However, the spectrum of RNA viruses that are targeted by the EJC or the underlying mechanisms are not well understood. EJC components from Arabidopsis thaliana were screened for antiviral activity towards Turnip crinkle virus (TCV, Tombusviridae). Overexpression of the accessory EJC component CASC3 inhibited TCV accumulation > 10-fold in Nicotiana benthamiana while knock-down of endogenous CASC3 resulted in a > 4-fold increase in TCV accumulation. CASC3 forms cytoplasmic condensates and deletion of the conserved SELOR domain reduced condensate size 7-fold and significantly decreased antiviral activity towards TCV. Mass spectrometry of CASC3 complexes did not identify endogenous stress granule or P-body markers and CASC3 failed to co-localize with an aggresome-specific dye suggesting that CASC3 condensates are distinct from well-established membraneless compartments. Mass spectrometry and bimolecular fluorescence complementation assays revealed that CASC3 sequesters Heat shock protein 70 (Hsp70-1) and Glyceraldehyde-3-phosphate dehydrogenase (GAPDH), two host factors with roles in tombusvirus replication. Overexpression of Hsp70-1 or GAPDH reduced the antiviral activity of CASC3 2.1-fold and 2.8-fold, respectively, and suggests that CASC3 inhibits TCV by limiting host factor availability. Unrelated Tobacco mosaic virus (TMV) also depends on Hsp70-1 and CASC3 overexpression restricted TMV accumulation 4-fold and demonstrates that CASC3 antiviral activity is not TCV-specific. Like CASC3, Auxin response factor 19 (ARF19) forms poorly dynamic condensates but ARF19 overexpression failed to inhibit TCV accumulation and suggests that CASC3 has antiviral activities that are not ubiquitous among cytoplasmic condensates.

The Virus-Induced Transcription Factor SHE1 Interacts with and Regulates Expression of the Inhibitor of Virus Replication (IVR) in N Gene Tobacco.

The transcription factor SHE1 was induced by tobacco mosaic virus (TMV) infection in tobacco cv. Samsun NN (SNN) and SHE1 inhibited TMV accumulation when expressed constitutively. To better understand the role of SHE1 in virus infection, transgenic SNN tobacco plants generated to over-express SHE1 (OEx-SHE1) or silence expression of SHE1 (si-SHE1) were infected with TMV. OEx-SHE1 affected the local lesion resistance response to TMV, whereas si-SHE1 did not. However, si-SHE1 allowed a slow systemic infection to occur in SNN tobacco. An inhibitor of virus replication (IVR) was known to reduce the accumulation of TMV in SNN tobacco. Analysis of SHE1 and IVR mRNA levels in OEx-SHE1 plants showed constitutive expression of both mRNAs, whereas both mRNAs were less expressed in si-SHE1 plants, even after TMV infection, indicating that SHE1 and IVR were associated with a common signaling pathway. SHE1 and IVR interacted with each other in four different assay systems. The yeast two-hybrid assay also delimited sequences required for the interaction of these two proteins to the SHE1 central 58-79% region and the IVR C-terminal 50% of the protein sequences. This suggests that SHE is a transcription factor involved in the induction of IVR and that IVR binds to SHE1 to regulate its own synthesis.

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.

Pseudomonas aeruginosa isolate PM1 effectively controls virus infection and promotes growth in plants.

A bacterial isolate PM1 obtained from the rhizosphere of healthy plants was identified as Pseudomonas aeruginosa by biochemical characteristics and 16S rRNA gene sequence (GenBank ID OL321133.1). It induced resistance in Nicotiana tabacum cv. Xanthi-nc and Cyamopsis tetragonoloba, against Tobacco mosaic virus (TMV) and Sunn-hemp rosette virus (SRV), respectively. Foliar treatment with isolate PM1 curbed TMV accumulation in susceptible N. tabacum cv. White Burley. PM1 was more effective as a foliar than a root/soil drench treatment, evident through a comparative decrease in ELISA values, and reduced viral RNA accumulation. Foliar and soil drench treatment with PM1 resulted in a disease index of 48 and 86 per cent, and a control rate of 48.9 and 8.5 per cent, respectively. PM1 exhibited phosphate solubilization, produced siderophores, auxins, HCN, and ammonia, all important plant growth-promoting traits. Foliar treatment with PM1 enhanced growth in tobacco, while its volatiles significantly promoted seedling growth in C. tetragonoloba. Of the several metabolites produced by the isolate, many are known contributors to induction of systemic resistance, antibiosis, and growth promotion in plants. Soluble metabolites of PM1 were less effective in inducing antiviral resistance in N. tabacum cv. Xanthi-nc in comparison with its broth culture. PM1 and its metabolites were antagonistic to Gram-positive Bacillus spizizenii and Staphylococcus aureus, and fungi Fusarium oxysporum, Aspergillus niger, and Rhizopus stolonifer. Its volatiles were inhibitory to F. oxysporum and R. stolonifer. Thus, PM1 exhibited considerable potential for further evaluation in plant virus control and production of diverse metabolites of use in agriculture and medicine.

The curative activity of some arylidene dihydropyrimidine hydrazone against Tobacco mosaic virus infestation

A series of novel pyrimidine-based derivative compounds were synthesized and evaluated for antiviral activity against Tobacco mosaic virus (TMV). All chemical structures of the prepared compounds were identified and characterized by IR, NMR, and elemental analysis. Under greenhouse conditions, the preliminary bioassays revealed that all compounds exhibited anti-TMV activity ranging from 23.5 % to 90.1% at 50 µg/mL concentration, indicating potential antiviral efficacy. Among the new compounds, the benzylidene derivative 3a was the most potent and demonstrated the highest inhibition effect of 90%, with a significant reduction in the TMV accumulation level of 91.4%. Furthermore, the curative activity of 3a was associated with a considerable decrease in H2O2 and MDA (nonenzymatic oxidative stress markers) compared to the TMV treatment. In addition, significant increases in the transcriptional levels of pathogenesis-related proteins (PR-1, PR-3, PR-4, and PR-5) at 4 and 6 dpi were shown to induce systemic resistance against TMV. Furthermore, molecular docking investigation demonstrated that the benzylidene derivative 3a was strongly lodged in the binding sites of TMV-coat protein at position 2.Consequently, the obtained results suggest that the new 3a compound could be applied as a safe control agent against plant viral infection.

Foliar Applications of Bacillus subtilis HA1 Culture Filtrate Enhance Tomato Growth and Induce Systemic Resistance against Tobacco mosaic virus Infection

The application of microbial products as natural biocontrol agents for inducing systemic resistance against plant viral infections represents a promising strategy for sustainable and eco-friendly agricultural applications. Under greenhouse conditions, the efficacy of the culture filtrate of Bacillus subtilis strain HA1 (Acc# OM286889) for protecting tomato plants from Tobacco mosaic virus (TMV) infection was assessed. The results showed that the dual foliar application of this culture filtrate (HA1-CF) 24 h before and 24 h after TMV inoculation was the most effective treatment for enhancing tomato plant development, with substantial improvements in shoot and root parameters. Furthermore, compared to non-treated plants, HA1-CF-treated tomato had a significant increase in total phenolic and flavonoid contents of up to 27% and 50%, respectively. In addition, a considerable increase in the activities of reactive oxygen species scavenging enzymes (PPO, SOD, and POX) and a significant decrease in non-enzymatic oxidative stress markers (H2O2 and MDA) were reported. In comparison to untreated control plants, all HA1-CF-treated plants showed a significant reduction in TMV accumulation in systemically infected tomato leaves, up to a 91% reduction at 15 dpi. The qRT-PCR results confirmed that HA1-CF stimulated the transcription of several defense-related tomato genes (PR-1, PAL, CHS, and HQT), pointing to their potential role in induced resistance against TMV. GC–MS analysis showed that phenol, 2,4-bis (1,1-dimethylethyl)-, Pyrrolo [1,2-a] pyrazine-1,4-dione, hexahydro-3-(2-methylpropyl)- and eicosane are the primary ingredient compounds in the HA1-CF ethyl acetate extract, suggesting that these molecules take part in stimulating induced systemic resistance in tomato plants. Our results imply that HA1-CF is a potential resistance inducer to control plant viral infections, a plant growth promoter, and a source of bioactive compounds for sustainable disease management.

Trichoderma hamatum Strain Th23 Promotes Tomato Growth and Induces Systemic Resistance against Tobacco Mosaic Virus.

Trichoderma hamatum strain Th23, isolated from tomato roots, was molecularly identified using phylogenetic analysis based on ITS, tef1, and rpb2 gene sequences and evaluated for its efficiency in suppressing tobacco mosaic virus (TMV) infection for the first time. Under greenhouse conditions, the application of Th23 promoted tomato growth with significant increases in shoot and root parameters as well as improved total chlorophyll content. Compared to the nontreated tomato plants, the soil pretreatment of tomato plants 48 h before TMV inoculation produced a significant reduction in the TMV accumulation level by 84.69% and enhanced different growth parameters. In contrast, TMV had a deleterious impact on fresh and dry matter accumulation and inhibited photosynthetic capacity. Furthermore, the protective activity of Th23 was associated with a significant increase in reactive oxygen species scavenging enzymes (PPO, CAT, and SOD) as well as decreased nonenzymatic oxidative stress markers (H2O2 and MDA) compared to the TMV treatment at 15 days post-viral inoculation (dpi). In addition, considerable increases in the transcriptional levels of polyphenolic genes (HQT and CHS) and pathogenesis-related proteins (PR-1 and PR-7) were shown to induce systemic resistance against TMV. Consequently, the ability of T. hamatum strain Th23 to promote plant growth, induce systemic resistance, and boost innate immunity against TMV infestation supported the incorporation of Th23 as a potential biocontrol agent for managing plant viral infections. To the best of our knowledge, this is the first report of the antiviral activity of T. hamatum against plant viral infection.

First Report of Protective Activity of Paronychia argentea Extract against Tobacco Mosaic Virus Infection

The widespread use of chemical control agents and pesticides for plant-pathogen control has caused many human health and environmental issues. Plant extracts and biocontrol agents have robust antimicrobial activity against different plant pathogens. However, their antiviral activities are still being investigated. In the present study, the methanol extract of Paronychia argentea was characterized and evaluated for its protective activity against the tobacco mosaic virus (TMV) infection in tomato plants under greenhouse conditions at 21 days post-inoculation. The results showed that the foliar application of P. argentea extract (10 µg/mL) enhanced tomato plant growth, resulting in significant increases in shoot and root parameters and total chlorophyll contents. Moreover, a significant reduction in TMV accumulation level in P. argentea-treated plants of 77.88% compared to non-treated plants was reported. Furthermore, induction of systemic resistance with significant elevation in production of antioxidant enzymes (PPO, CAT, and SOD) and transcriptional levels of the pathogenesis-related proteins (PR-1 and PR-7) and polyphenolic genes (CHS and HQT) were also observed. Out of 16 detected compounds, HPLC analysis revealed that the most abundant polyphenolic compounds found in P. argentea extract were gallic acid (5.36 µg/mL), kaempferol (7.39 µg/mL), quercetin (7.44 µg/mL), ellagic acid (7.89 µg/mL), myricetin (8.36 µg/mL), and ferulic acid (8.69 µg/mL). The findings suggest that the use of P. argentea extract as an effective and safe source for the production of bioactive compounds may offer a solution for a promising approach for the management of plant viral infections. To the best of our knowledge, this is the first report of the protective activity of P. argentea extract against plant viral diseases.

Antiviral activity and tobacco-induced resistance, mediated by essential oil nano-emulsions from Zataria multiflora and Satureja bakhtiarica

There has been a growing interest in extracting essential oils from aromatic plants as potential sources of biorational pesticides. In the present study, essential oil emulsions of Satureja bakhtiarica (SEO), Oliveria decumbens (OEO), Trachyspermum ammi (TEO) and Zataria multiflora (ZEO) were prepared. Their major chemical compositions were characterized and their antiviral effects were evaluated against two important plant viruses: Cucumber mosaic virus (CMV) and Tobacco mosaic virus (TMV). SEO and ZEO indicated higher antiviral activity in comparison with OEO and TEO (p < 0.05). In systemic bioassay, the pre-treatment test of ZEO and SEO showed a reduced level of TMV accumulation in TMV-infected tobacco, with rates of 26.75% and 23.97%, respectively. Also, the two essential oils had mechanisms by which plant resistance acted against plant viruses. These mechanisms involved the activities of several defense-related enzymes (PAL and POD) and genes (ICS1, NPR1, and PR-1a). These results indicate that the essential oils of Zataria multiflora and Satureja bakhtiarica have the potential to be used as sources of effective antiviral agents and eco-friendly pesticide.

BLB8, an antiviral protein from Brevibacillus laterosporus strain B8, inhibits Tobacco mosaic virus infection by triggering immune response in tobacco

Tobacco mosaic virus (TMV) is one of destructive plant viruses, causing serious economic losses in the world. Using antiviral proteins or elicitors to inhibit viral infection or promote plant immunity is one of the efficient strategies against TMV. Our previous study identified that the fermentation broth of Brevibacillus laterosporus strain B8 showed strong antiviral activity against TMV. However, the active antiviral ingredient is still unclear. Here, BLB8 (B. laterosporus strain B8 protein, BLB8), an antiviral protein from B. laterosporus strain B8 was isolated and characterized. BLB8 showed protective, inactive and curative effects against TMV, and the inhibition rate reached up to 63%, 83% and 55%, respectively. BLB8 infiltrated around the infection site of the recombinant virus TMV-GFP inhibited the systemic extend and movement of TMV. Pretreatment of the bottom leaves with BLB8 inhibited the spread and accumulation of TMV in upper systemic leaves. Furthermore, BLB8 caused hypersensitive response (HR) in a dose-dependent way, promoted H2 O2 accumulation, and induced the expression of defense-relative genes in Nicotiana benthamiana. The antiviral protein BLB8 from B. laterosporus strain B8 effectively inhibits TMV infection in inactivation, protective and curative effects through triggering plant immunity in tobacco. Therefore, the present study provides a new antiviral agent for prevention and control of viral disease. © 2021 Society of Chemical Industry.

DsRNA Molecules From the Tobacco Mosaic Virus p126 Gene Counteract TMV-Induced Proteome Changes at an Early Stage of Infection.

Exogenous application of double-stranded RNA (dsRNA) in the tobacco–Tobacco mosaic virus (TMV) pathosystem was shown previously to induce resistance against TMV providing an alternative approach to transgenesis. In the present study, we employed proteomics technology to elucidate the effect of TMV on tobacco as well as the effect of exogenous application of TMV p126 dsRNA molecules (dsRNAp126) at an early stage of the tobacco–TMV interaction. The proteome of tobacco leaf at 15 min post inoculation (mpi) in the presence or absence of dsRNAp126 molecules was studied. Thirty-six tobacco proteins were differentially accumulated in TMV-infected vs. healthy tobacco leaf tissue. The identified main differential TMV-responsive proteins were found to be involved in photosynthesis, energy metabolism, stress, and defense responses. Most of the virus-induced changes in the tobacco leaf proteome were not observed in the leaves treated with dsRNAp126 + TMV. The results indicated that the protein changes induced by TMV infection were counteracted by the exogenous application of dsRNAp126 molecules. Moreover, using small RNA sequencing, we showed that the exogenously applied dsRNAp126 was efficiently processed in tobacco as early as 15 min post application (mpa) to produce small interfering RNAs (siRNAs); the dicing pattern was not affected by the presence of TMV. The presence of dsRNAp126 reduced TMV p126 RNA abundance suggesting virus titer reduction via a sequence-specific mechanism, since a non-homologous dsRNA did not protect from TMV infection nor affect TMV accumulation.

Transcriptomic and Functional Analyses Indicate Novel Anti-viral Mode of Actions on Tobacco Mosaic Virus of a Microbial Natural Product ε-Poly-l-lysine.

Novel anti-viral natural product ε-poly-l-lysine (ε-PL) produced by Streptomyces is a homopolymer of l-lysine, of which the underlying molecular mode of action remains to be further elucidated. In this study, ε-PL induced significant fragmentation of tobacco mosaic virus (TMV) virions and delayed the systemic infection of TMV-GFP as well as wild-type TMV in plants. ε-PL treatment also markedly inhibited RNA accumulation of TMV in tobacco BY-2 protoplasts. The results of RNA-seq indicated that the agent induced significantly differential expression of genes that are associated with defense response, stress response, autophagy, and ubiquitination. Among them, 15 critical differential expressed genes were selected for real-time quantitative PCR validation. We further demonstrated that ε-PL can induce host defense responses by assessing the activity of several defense-related enzymes in plants. Our results provided valuable insights into molecular anti-viral mode of action for ε-PL, which is expected to be applied as a novel microbial natural product against plant virus diseases.

Induction of Plant Resistance against Tobacco Mosaic Virus Using the Biocontrol Agent Streptomyces cellulosae Isolate Actino 48

Viral plant diseases represent a serious problem in agricultural production, causing large shortages in the production of food crops. Eco-friendly approaches are used in controlling viral plant infections, such as biocontrol agents. In the current study, Streptomyces cellulosae isolate Actino 48 is tested as a biocontrol agent for the management of tobacco mosaic virus (TMV) and inducing tomato plant systemic resistance under greenhouse conditions. Foliar application of a cell pellet suspension of Actino 48 (2 × 107 cfu. mL−1) is performed at 48 h before inoculation with TMV. Peroxidase activity, chitinase activity, protein content, and the total phenolic compounds are measured in tomato leaves at 21 dpi. On the other hand, the TMV accumulation level and the transcriptional changes of five tomato defense-related genes (PAL, PR-1, CHS, PR-3, and PR-2) are studied. Treatment with Actino 48 before TMV inoculation (48 h) induced tomato plants to increase their levels of peroxidase and chitinase enzymes. Furthermore, a significant increase in the concentration of total phenolic compounds was observed in Actino 48 and TMV-treated tomato plants compared to TMV-treated tomato plants alone. Treatment with Actino 48 reduced the TMV accumulation level (53.8%) compared to treatment with the virus alone. Actino 48 induced plant growth, where the fresh and dry weights of tomato plants increased. Additionally, significant increases of the PAL, PR-1, CHS, and PR-3 transcripts were observed. On the other hand, a higher induction of PR-2 was only observed in TMV-treated tomato plants. In conclusion, S. cellulosae isolate Actino 48 can be used as a biocontrol agent for the reduction of symptoms and severity of TMV.

The Phytochemical, Antifungal, and First Report of the Antiviral Properties of Egyptian Haplophyllum tuberculatum Extract.

In this study, ethanol whole plant extract (WPE) of Haplophyllum tuberculatum was characterized and tested for its antifungal and antiviral activities against Fusarium culmorum, Rhizoctonia solani and tobacco mosaic virus (TMV). High Performance Liquid Chromatography (HPLC) analysis showed that the main phytochemical constituents of H. tuberculatum WPE were resveratrol (5178.58 mg/kg), kaempferol (1735.23 mg/kg), myricetin (561.18 mg/kg), rutin (487.04 mg/kg), quercetin (401.04 mg/kg), and rosmarinic acid (387.33 mg/kg). By increasing H. tuberculatum WPE at concentrations of 1%, 2%, and 3%, all of the fungal isolates were suppressed compared to the two positive and negative controls. Under greenhouse conditions, WPE-treated Chenopodium amaranticolor plants strongly inhibited TMV infection and significantly reduced TMV accumulation levels when compared to non-treated plants. Moreover, the induction of systemic resistance with significant increases in the transcriptional levels of the pathogenesis-related protein-1 (PR-1), chalcone synthase (CHS), and hydroxycinnamoyl-CoA quinate transferase (HQT) genes for treated plants were noticed at 3 and 5 days post-inoculation (dpi) for both assays. To the best of our knowledge, this is the first reported observation of the antiviral activity of H. tuberculatum extract against plant viral infections. Finally, the results obtained suggest that H. tuberculatum WPE can be considered a promising source of both antifungal and antiviral substances for practical use and for developing plant-derived compounds for the effective management of plant diseases.

Plant SNAREs SYP22 and SYP23 interact with Tobacco mosaic virus 126 kDa protein and SYP2s are required for normal local virus accumulation and spread

Viral proteins often interact with multiple host proteins during virus accumulation and spread. Identities and functions of all interacting host proteins are not known. Through a yeast two-hybrid screen an Arabidopsis thaliana Qa-SNARE protein [syntaxin of plants 23 (AtSYP23)], associated with pre-vacuolar compartment and vacuolar membrane fusion activities, interacted with Tobacco mosaic virus (TMV) 126 kDa protein, associated with virus accumulation and spread. In planta, AtSYP23 and AtSYP22 each fused with mCherry, co-localized with 126 kDa protein-GFP. Additionally, A. thaliana and Nicotiana benthamiana SYP2 proteins and 126 kDa protein interacted during bimolecular fluorescence complementation analysis. Decreased TMV accumulation in Arabidopsis plants lacking SYP23 and in N. benthamiana plants subjected to virus-induced gene silencing (VIGS) of SYP2 orthologs was observed. Diminished TMV accumulation during VIGS correlated with less intercellular virus spread. The inability to eliminate virus accumulation suggests that SYP2 proteins function redundantly for TMV accumulation, as for plant development.

A Conserved Carboxylesterase Inhibits Tobacco mosaic virus (TMV) Accumulation in Nicotiana benthamiana Plants

A carboxylesterase (CXE) or carboxylic-ester hydrolase is an enzyme that catalyzes carboxylic ester and water into alcohol and carboxylate. In plants, CXEs have been implicated in defense, development, and secondary metabolism. We discovered a new CXE gene in Nicotiana benthamiana that is related to virus resistance. The transcriptional level of NbCXE expression was significantly increased after Tobacco mosaic virus (TMV) infection. Transient over-expression of NbCXE inhibited TMV accumulation in N. benthamiana plants. Conversely, when the NbCXE gene was silenced with a Tobacco rattle virus (TRV)-based gene silencing system, TMV RNA accumulation was increased in NbCXE-silenced plants after infection. NbCXE protein was shown to interact with TMV coat protein (CP) in vitro. Additionally, the expressions of host defense-related genes were increased in transient NbCXE-overexpressed plants but decreased in NbCXE silenced N. benthamiana plants. In summary, our study showed that NbCXE is a novel resistance-related gene involved in host defense responses against TMV infection.

Cancer Theranostic Applications of Albumin-Coated Tobacco Mosaic Virus Nanoparticles

Nanotechnology holds great promise in cancer drug delivery, and of particular interest are theranostic approaches in which drug delivery and imaging are integrated. In this work, we studied and developed the plant virus tobacco mosaic virus (TMV) as a platform nanotechnology for drug delivery and imaging. Specifically, a serum albumin (SA)-coated TMV formulation was produced. The SA coating fulfils two functions: SA provides a stealth coating for enhanced biocompatibility; it also acts as a targeting ligand enabling efficient tumor accumulation of SA-TMV versus TMV in mouse models of breast and prostate cancer. We demonstrate drug delivery of the chemotherapy doxorubicin (DOX); TMV-delivered DOX outperformed free DOX, resulting in significant delayed tumor growth and increased survival. Furthermore, we demonstrated the ability of SA-coated TMV loaded with chelated Gd(DOTA) for magnetic resonance imaging detection of tumors. In the future, we envision the application of such probes as theranostic, where first imaging is performed to assess whether the nanoparticles are effective at targeting a particular patient tumor. If targeting is confirmed, the therapeutic would be added and treatment can begin. The combination of imaging and therapy would allow to monitor disease progression and therefore inform about the effectiveness of the drug delivery approach.

Tobacco mosaic virus-directed reprogramming of auxin/indole acetic acid protein transcriptional responses enhances virus phloem loading

Vascular phloem loading has long been recognized as an essential step in the establishment of a systemic virus infection. In this study, an interaction between the replication protein of tobacco mosaic virus (TMV) and phloem-specific auxin/indole acetic acid (Aux/IAA) transcriptional regulators was found to modulate virus phloem loading in an age-dependent manner. Promoter expression studies show that in mature tissues TMV 126/183-kDa-interacting Aux/IAAs predominantly express and accumulate within the nuclei of phloem companion cells (CCs). Furthermore, CC Aux/IAA nuclear localization is disrupted upon infection with an interacting virus. In situ analysis of virus spread shows that the inability to disrupt Aux/IAA CC nuclear localization correlates with a reduced ability to load into the vascular tissue. Subsequent systemic movement assays also demonstrate that a virus capable of disrupting Aux/IAA localization is significantly more competitive at moving out of older plant tissues than a noninteracting virus. Similarly, CC expression and overaccumulation of a degradation-resistant Aux/IAA-interacting protein was found to inhibit TMV accumulation and phloem loading selectively in flowering plants. Transcriptional expression studies demonstrate a role for Aux/IAA-interacting proteins in the regulation of salicylic and jasmonic acid host defense responses as well as virus-specific movement factors, including pectin methylesterase, that are involved in regulating plasmodesmata size-exclusion limits and promoting virus cell-to-cell movement. Combined, these findings indicate that TMV directs the reprogramming of auxin-regulated gene expression within the vascular phloem of mature tissues as a means to enhance phloem loading and systemic spread.