Tobacco Mosaic Virus Particles Research Articles

Novel flavonol derivatives containing benzoxazole as potential antiviral agents: design, synthesis, and biological evaluation.

A series of flavonol derivatives containing benzoxazole were designed and synthesized, and the structures of all the target compounds were determined by nuclear magnetic resonance (NMR) and high-resolution mass spectrometry (HRMS). The structure of X2 was further confirmed by single crystal X-ray diffraction analysis. The results of the bioactivity tests showed that some of the target compounds possessed excellent antiviral activity against tobacco mosaic virus (TMV) in vivo. In particular, the median effective concentration (EC50) values for the curative and protective activities of X17 against TMV were 127.6 and 101.2μg/mL, respectively, which were superior to those of ningnanmycin (320.0 and 234.6μg/mL). The results of preliminary mechanism study indicated that X17 had a strong binding affinity for TMV coat protein (TMV-CP), which might hinder the self-assembly and replication of TMV particles. In addition, X17 was able to effectively inhibit tobacco leaf membrane lipid peroxidation and facilitate the removal of O2- from the body, thereby improving the disease resistance of tobacco plants. Therefore, the design and synthesis of flavonol derivatives containing benzoxazole provides value for the development of new antiviral drugs.

Myricetin derivatives containing the benzoxazinone moiety discovered as potential anti-tobacco mosaic virus agents

A series of myricetin derivatives containing benzoxazinone were designed and synthesized. The structures of all compounds were characterized by NMR and HRMS. The structure of Y4 had been confirmed by single-crystal X-ray diffraction analysis. The test results of EC50 values of tobacco mosaic virus (TMV) suggested that Y8 had the best curative and protective effects, with EC50 values of 236.8, 206.0 μg/mL, respectively, which were higher than that of ningnanmycin (372.4, 360.6 μg/mL). Microscale thermophoresis (MST) experiments demonstrated that Y8 possessed a strong binding affinity for tobacco mosaic virus coat protein (TMV-CP), with a dissociation constant (Kd) value of 0.045 μM, which was superior to the ningnanmycin (0.700 μM). The findings of molecular docking studies revealed that Y8 interacted with multiple amino acid residues of TMV-CP through the formation of non-covalent bonds, which had an effect on the self-assembly of TMV particles. The malondialdehyde (MDA) and superoxide dismutase assay (SOD) content assays also fully verified that Y8 could stimulate the plant immune system and enhance disease resistance by reducing MDA content and increasing SOD content. In summary, myricetin derivatives containing benzoxazinone could be considered to further research and development as novel antiviral agents.

Raman-dielectrophoresis goes viral: towards a rapid and label-free platform for plant virus characterization

An innovative spectroscopic method that allows to chemically and structurally characterize viruses directly in suspension within few minutes was developed. A library of five different plant viruses was obtained combining dielectrophoresis (DEP), performed with a device specifically designed to capture and agglomerate virus particles, and Raman spectroscopy to provide a chemical fingerprint of virions. The tested viruses, purified from infected plants, were chosen for their economic impact on horticultural crops and for their different morphological and structural features. Using the Raman-DEP device, specific profiles for each virus were successfully obtained, relying on chemical differences occurring even with genetically similar viruses belonging to the same taxonomic species and morphologically indiscernible by transmission electron microscopy (TEM). Moreover, we investigated the potentiality of Raman-DEP to follow dynamic changes occurring upon heat treatment of tobacco mosaic virus (TMV) particles. Raman peak deviations linked to TMV coat protein conformation were observed upon treatment at temperatures equal or higher than 85°C, substantiating the rod-to-spherical shape transitions observed by TEM and the concomitant drastic loss of infectivity following plant inoculation. Overall, the Raman-DEP method can be useful for the characterization of virus (nano)particles, setting the basis to create a database suitable for the study of viruses or virus derived-nanoparticles relevant for the agricultural, medical, or biotechnological fields.

Soft Sub-Structured Multi-Material Biosensor Hydrogels with Enzymes Retained by Plant Viral Scaffolds.

An all-soft multi-material combination consisting of a hydrogel based on poly(ethylene glycol) (PEG) coated with spatially defined spots of gelatin methacryloyl (GM) containing selectively addressable viral nanorods is presented, and its basic application as a qualitative biosensor with reporter enzymes displayed on the tobacco mosaic virus (TMV) bioscaffolds within the GM is demonstrated. Biologically inert PEG supports are equipped with GM spots serving as biological matrix for enzymes clustered on TMV particles preventing diffusion out of the gel. For this multi-material combination, i) the PEG-based hydrogel surface is modified to achieve a clear boundary between coated and non-coated regions by introducing either isothiouronium or thiol groups. ii) Cross-linking of the GM spots is studied to achieve anchoring to the hydrogel surface. iii) The enzymes horseradish peroxidase or penicillinase (Pen) are conjugated to TMV and integrated into the GM matrix. In contrast to free enzymes, enzyme-decorated TMVs persist in GM spots and show sustained enzyme activity as evidenced by specific color reaction after 7 days of washing, and for Pen after 22 months after dry storage. Therefore, the integration of enzyme-coupled TMV into hydrogel matrices is a promising and versatile approach to obtaining reusable and analyte-specific sensor components.

Capacitive field-effect biosensor modified with a stacked bilayer of weak polyelectrolyte and plant virus particles as enzyme nanocarriers

This work presents a new approach for the development of field-effect biosensors based on an electrolyte-insulator-semiconductor capacitor (EISCAP) modified with a stacked bilayer of weak polyelectrolyte and tobacco mosaic virus (TMV) particles as enzyme nanocarriers. With the aim to increase the surface density of virus particles and thus, to achieve a dense immobilization of enzymes, the negatively charged TMV particles were loaded onto the EISCAP surface modified with a positively charged poly(allylamine hydrochloride) (PAH) layer. The PAH/TMV bilayer was prepared on the Ta2O5-gate surface by means of layer-by-layer technique. The bare and differently modified EISCAP surfaces were physically characterized by fluorescence microscopy, zeta-potential measurements, atomic force microscopy and scanning electron microscopy. Transmission electron microscopy was used to scrutinize the PAH effect on TMV adsorption in a second system. Finally, a highly sensitive TMV-assisted EISCAP antibiotics biosensor was realized by immobilizing the enzyme penicillinase onto the TMV surface. This PAH/TMV bilayer-modified EISCAP biosensor was electrochemically characterized in solutions with different penicillin concentrations via capacitance-voltage and constant-capacitance methods. The biosensor possessed a mean penicillin sensitivity of 113 mV/dec in a concentration range from 0.1 mM to 5 mM.

Identification of vanillic acid and its new amide derivative from Hyoscyamus niger and their modes of action in controlling tobacco mosaic virus

Plant viruses represented by tobacco mosaic virus (TMV) are hard to control due to the characteristics of wide hosts and difficulties to be inactivated in vitro . The discovery of candidates to damage the virus directly, and elicitors to induce plant resistances are important strategies against TMV. This study aims to identify new candidates from the title plants as biopesticides in controlling TMV. Herein, vanillic acid ( 6 ) and its new amide derivative, nigeroate ( 1 ), and seven known ones were discovered from the seeds of Hyoscyamus niger L. Compound 1 , elucidated by spectral methods, was synthesized by two steps for industrial application in agriculture as a new elicitor. Compared with ningnanmycin, compound 1 appeared higher curative efficacies against TMV with the inhibitory rates of 58.00% and 69.14% at 250 and 500 μg/mL, respectively, while compound 6 displayed higher inactivation and protective efficacies with the inhibitory rates of 47.30–79.71% and 43.28–63.87% at 125–500 μg/mL, respectively. Compound 6 could destroy TMV particles into small fragments dramatically by decreasing the transcription level of TMV-CP gene, while compound 1 just broke the TMV particles slightly, and was no influence on TMV-CP gene. Interestingly, compound 1 induced potent hypersensitive response (HR), and systemic acquired resistance (SAR) activities (53.82% at 500 μg/mL) in tobaccos. The accumulation of H 2 O 2 by activating SOD (superoxide dismutase), CAT (catalase), as well as POD (peroxidase) was the mechanism of compound 1 induced HR. The SAR induced by compound 1 was referred to upregulate SA (salicylic acid) by activating PAL (phenylalanine ammonia-lyase) pathway, ICS (isochorismate synthase) pathway, as well as PR (pathogenesis-related) proteins due to the increasement of PAL enzyme, and PAL , ICS1 , NPR1 , PR1 , and PR2 transcription levels. Thus, vanillic acid ( 6 ) and its new amide derivative ( 1 ) could be developed as antiviral agents and plant immune elicitors for biopesticides in controlling TMV, respectively. • Vanillic acid and its new amide derivative, nigeroate, were identified from Hyoscyamus niger . • Tobacco mosaic virus coat protein is the target for vanillic acid exerting antiviral activity. • Nigeroate was synthesized by two steps for industrial application as an elicitor. • Accumulation of H 2 O 2 and salicylic acid contributed to nigeroate’s systemic resistance. • Defense enzymes, pathogenesis-related, and salicylic acid biosynthesis-related genes manipulated nigeroate’s resistance.

Triplex Immunostrip Assay for Rapid Diagnosis of Tobacco Mosaic Virus, Tobacco Vein Banding Mosaic Virus, and Potato Virus Y.

Mixed virus infection has increasingly become a problem in the production of Solanaceae crops in recent years; therefore, a fast and accurate detection method is needed. In this study, a novel triplex immunostrip assay was developed for the simultaneous detection of tobacco mosaic virus (TMV), tobacco vein banding mosaic virus (TVBMV), and potato virus Y (PVY). The limits of detection of this novel immunostrip reached 200 ppb (ng/ml), 1 ppm (µg/ml), and 2 ppm for TMV, PVY, and TVBMV particles, respectively. Importantly, no cross-reactivity was observed among TMV, TVBMV, and PVY or to a nontarget virus. When the assay was applied to suspected virus-infected tobacco, tomato, and potato samples collected from fields in Southwest China, samples of single or mixed virus infection were successfully identified. In conclusion, the triplex immunostrip assay provides a fast and easy to use on-site detection method for field epidemiological studies of TMV, TVBMV, and PVY, and for managing diseases that are caused by them.

Discovery of Novel α-Methylene-γ-Butyrolactone Derivatives Containing Vanillin Moieties as Antiviral and Antifungal Agents.

On the basis of the structure of nicotlactone A (L1), a series of novel α-methylene-γ-butyrolactone derivatives B1-B43 were designed and synthesized by structure simplification and active fragment replacement strategies, and their antiviral and antifungal activities were evaluated. The bioassay studies indicated that many target compounds possessed good to excellent antiviral activity against tobacco mosaic virus (TMV) and some of these compounds exhibited specific antifungal activities against Valsa mali and Fusarium graminearum. Compound B32 exhibited the best anti-TMV activity (inactivation effect, 88.9%; protection effect, 65.8%; curative effect, 52.8%) in vivo at 500 mg/L, which is significantly higher than that of commercial virucides ribavirin and ningnanmycin. The inhibition effect of compound B32 was also visualized by the inoculation test using green fluorescent protein (GFP)-labeled TMV. The preliminary antiviral mechanism of compound B32 was investigated. Transmission electron microscopy (TEM) showed that compound B32 could destroy the integrity of virus particles. Then, molecular docking and isothermal titration calorimetry (ITC) analysis further demonstrated that compound B32 exhibited a strong binding affinity to the TMV coat protein with a dissociation constant (Kd) of 3.06 μM, superior to ribavirin. Thus, we deduced that compound B32 may interfere with the self-assembly of TMV particles by binding TMV coat protein (CP). In addition, compound B28 showed good in vitro activity against F. graminearum with an inhibition rate of 90.9% at 50 mg/L, which was greater than that of fluxapyroxad (59.1%) but lower than that of the commercial fungicide carbendazim (96.8%). The present study provides support for the application of these α-methylene-γ-butyrolactone derivatives as novel antiviral and antifungal agents in crop protection.

Detection of Acetoin and Diacetyl by a Tobacco Mosaic Virus-Assisted Field-Effect Biosensor

Acetoin and diacetyl have a major impact on the flavor of alcoholic beverages such as wine or beer. Therefore, their measurement is important during the fermentation process. Until now, gas chromatographic techniques have typically been applied; however, these require expensive laboratory equipment and trained staff, and do not allow for online monitoring. In this work, a capacitive electrolyte–insulator–semiconductor sensor modified with tobacco mosaic virus (TMV) particles as enzyme nanocarriers for the detection of acetoin and diacetyl is presented. The enzyme acetoin reductase from Alkalihalobacillus clausii DSM 8716T is immobilized via biotin–streptavidin affinity, binding to the surface of the TMV particles. The TMV-assisted biosensor is electrochemically characterized by means of leakage–current, capacitance–voltage, and constant capacitance measurements. In this paper, the novel biosensor is studied regarding its sensitivity and long-term stability in buffer solution. Moreover, the TMV-assisted capacitive field-effect sensor is applied for the detection of diacetyl for the first time. The measurement of acetoin and diacetyl with the same sensor setup is demonstrated. Finally, the successive detection of acetoin and diacetyl in buffer and in diluted beer is studied by tuning the sensitivity of the biosensor using the pH value of the measurement solution.

Design, synthesis, and bioactivity of ferulic acid derivatives containing an β-amino alcohol

BackgroundPlant diseases caused by viruses and bacteria cause huge economic losses due to the lack of effective control agents. New potential pesticides can be discovered through biomimetic synthesis and structural modification of natural products. A series of ferulic acid derivatives containing an β-amino alcohol were designed and synthesized, and their biological activities were evaluated.ResultBioassays results showed that the EC50 values of compound D24 against Xanthomonas oryzae pv. oryzae (Xoo) was 14.5 μg/mL, which was better than that of bismerthiazol (BT, EC50 = 16.2 μg/mL) and thiodiazole copper (TC, EC50 = 44.5 μg/mL). The in vivo curative and protective activities of compound D24 against Xoo were 50.5% and 50.1%, respectively. The inactivation activities of compounds D2, D3 and D4 against tobacco mosaic virus (TMV) at 500 μg/mL were 89.1, 93.7 and 89.5%, respectively, superior to ningnanmycin (93.2%) and ribavirin (73.5%). In particular, the EC50 value of compound D3 was 38.1 μg/mL, and its molecular docking results showed that compound D3 had a strong affinity for TMV-CP with a binding energy of − 7.54 kcal/mol, which was superior to that of ningnanmycin (− 6.88 kcal /mol).ConclusionsThe preliminary mechanism research results indicated that compound D3 may disrupt the three-dimensional structure of the TMV coat protein, making TMV particles unable to self-assemble, which may provide potential lead compounds for the discovery of novel plant antiviral agents.Graphical

Marine Sesquiterpenes for Plant Protection: Discovery of Laurene Sesquiterpenes and Their Derivatives as Novel Antiviral and Antiphytopathogenic Fungal Agents.

The unreasonable use or long-term use of a single variety of pesticide has led to drug resistance and made the pesticides ineffective. Therefore, the creation of new, efficient, and low-risk pesticides is imminent. Marine natural products play a vital role in serving as new lead compounds. In this work, we realized the efficient preparation of nine marine sesquiterpenes with the Stork-Danheiser reaction as the key step and designed and synthesized a series of their derivatives. The antiviral activity and antifungal activity research showed that these compounds exhibited good to excellent biological activities. Compounds 7b and 8e displayed significantly higher antiviral activities against tobacco mosaic virus (TMV) than ribavirin and could be used as new antiviral candidates. The antiviral mode of action research indicated that compound 8e inhibited the formation of the 20S protein disk by acting on the TMV coat protein and therefore inhibited the assembly of TMV particles. This work provides a new idea for the development of new pesticides based on marine sesquiterpenes.

Isolation of Tobacco Mosaic Virus-Binding Peptides for Biotechnology Applications.

Tobacco mosaic virus (TMV) was the first virus to be discovered and it is now widely used as a tool for biological research and biotechnology applications. TMV particles can be decorated with functional molecules by genetic engineering or bioconjugation. However, this can destabilize the nanoparticles, and/or multiple rounds of modification may be necessary, reducing product yields and preventing the display of certain cargo molecules. To overcome these challenges, we used phage display technology and biopanning to isolate a TMV-binding peptide (TBPT25 ) with strong binding properties (IC50 =0.73 μM, KD =0.16 μM), allowing the display of model cargos via a single mixing step. The TMV-binding peptide is specific for TMV but does not recognize free coat proteins and can therefore be used to decorate intact TMV or detect intact TMV particles in crude plant sap.

1,3,4-Oxadiazole Contained Sesquiterpene Derivatives: Synthesis and Microbiocidal Activity for Plant Disease.

A series of 1,3,4-oxadiazole contained sesquiterpene derivatives were synthesized, and the activity of the target compounds against Xanthomonas oryzae pv. oryzae (Xoo), Xanthomonas axonopodis pv. citri (Xac), and tobacco mosaic virus (TMV) were evaluated. The biological activity results showed that the EC50 values of compounds H4, H8, H11, H12, H14, H16, and H19 for Xac inhibitory activity were 33.3, 42.7, 56.1, 74.5, 37.8, 43.8, and 38.4 μg/ml, respectively. Compounds H4, H8, H15, H19, H22, and H23 had inhibitory effects on Xoo, with EC50 values of 51.0, 43.3, 43.4, 50.5, 74.6, and 51.4 μg/ml, respectively. In particular, the curative and protective activities of compound H8 against Xoo in vivo were 51.9 and 49.3%, respectively. In addition, the EC50 values of the inactivation activity of compounds H4, H5, H9, H10, and H16 against TMV were 69.6, 58.9, 69.4, 43.9, and 60.5 μg/ml, respectively. The results of molecular docking indicated that compound H10 exhibited a strong affinity for TMV-coat protein, with a binding energy of −8.88 kcal/mol. It may inhibit the self-assembly and replication of TMV particles and have an anti-TMV effect, which supports its potential usefulness as an antiviral agent.

Towards Multi-Analyte Detection with Field-Effect Capacitors Modified with Tobacco Mosaic Virus Bioparticles as Enzyme Nanocarriers.

Utilizing an appropriate enzyme immobilization strategy is crucial for designing enzyme-based biosensors. Plant virus-like particles represent ideal nanoscaffolds for an extremely dense and precise immobilization of enzymes, due to their regular shape, high surface-to-volume ratio and high density of surface binding sites. In the present work, tobacco mosaic virus (TMV) particles were applied for the co-immobilization of penicillinase and urease onto the gate surface of a field-effect electrolyte-insulator-semiconductor capacitor (EISCAP) with a p-Si-SiO2-Ta2O5 layer structure for the sequential detection of penicillin and urea. The TMV-assisted bi-enzyme EISCAP biosensor exhibited a high urea and penicillin sensitivity of 54 and 85 mV/dec, respectively, in the concentration range of 0.1–3 mM. For comparison, the characteristics of single-enzyme EISCAP biosensors modified with TMV particles immobilized with either penicillinase or urease were also investigated. The surface morphology of the TMV-modified Ta2O5-gate was analyzed by scanning electron microscopy. Additionally, the bi-enzyme EISCAP was applied to mimic an XOR (Exclusive OR) enzyme logic gate.

Trifluoromethylpyridine thiourea derivatives: design, synthesis and inhibition of the self-assembly of tobacco mosaic virus particles.

Devastating plant virus diseases leading to bad harvests and lower quality of crops have made feeding the beyond seven billion population a huge challenge. Nevertheless, growing resistance and cross resistance of crop protection agents have made this challenge harder. Therefore, an efficient crop protection agent with novel structure and mode of action showing higher efficiency and eco-friendly is urgently needed. The coat protein (CP) of a virus is a potential target for the discovery of new antiviral agents. Antiviral molecules can inhibit infection by obstructing the assembly of virus particles. A series of novel phthalamide-like thiourea derivatives containing trifluoromethylpyridine was designed and synthesized. Most of the compounds exhibited good antiviral activity against tobacco mosaic virus (TMV). Compound 7b showed notable curative, protective and inactivation activities against TMV. Its inactivation half-maximal effective concentration (EC50 ) value (20.5μg mL-1 ) was better even than commercial ningnanmycin (23.2μg mL-1 ). Compound 7b also had stronger TMV-CP binding ability than ningnanmycin and destroyed the external shape of TMV particles and hindered the self-assembly of TMV-CP and TMV-RNA. These phthalamide-like thiourea derivatives especially compound 7b containing trifluoromethylpyridine are potential lead compounds and inhibitors of TMV particle self-assembly. © 2021 Society of Chemical Industry.

Pathological Changes in Sweet Basil (Ocimum basilicum L.) Caused by Mixed Infection of Tobacco Mosaic Virus and Phytoplasma

Ocimum basilicum L. (a medicinal plant known as sweet basil) was investigated and showed disease symptoms, including mosaic mottling, leaf distortion, blistering, witches’ broom, and phyllody. The plants were found to be infected by both tobacco mosaic virus (TMV) and phytoplasma. Positive TMV was indicated by serological identification using direct antibody sandwich enzyme-linked immunosorbent assay (DAS-ELISA). Phytoplasma infection was identified molecularly using the phytoplasma P1/P7 primer pair and the nested primer, R16F2n/R16R2, which successfully generated a DNA band at ~1200 bp. The virus was successfully transmitted mechanically to Nicotiana tabacum, Datura metal, Datura stramonium, and Phaseolus vulgaris. Light and transmission electron microscopy showed pleomorphic phytoplasmas and rod-shaped particles (300 nm) of TMV, respectively. Histopathological and cytopathological studies revealed changes in the cell behavior of the infected leaves of sweet basil due to virus and phytoplasma infection. Hypoplasia and hyperplasia were observed in the mosaic areas of the mesophyll. Alterations in the vascular tissues and degeneration of the organelles were observed in ultrathin sections of the infected mesophyll. The presence of phytoplasma cells and crystalline bodies were indicative of the mixed infection. To our knowledge, this is the first report of a mixed infection of TMV and phytoplasmas in sweet basil.

Analysis of Engineered Tobacco Mosaic Virus and Potato Virus X Nanoparticles as Carriers for Biocatalysts.

Plant virus nanoparticles are promising candidates for the development of novel materials, including nanocomposites and scaffolds/carriers for functional molecules such as enzymes. Their advantages for enzyme immobilization include a modular organization, a robust and programmable structure, and a simple, cost-effective production. However, the activity of many enzymes relies on posttranslational modification and most plant viruses replicate in the cytoplasm, so functional enzymes cannot be displayed on the virus surface by direct coat protein fusions. An alternative display system to present the Trichoderma reesei endoglucanase Cel12A on potato virus X (PVX) using SpyTag/SpyCatcher (ST/SC) technology was recently developed by the authors, which allows the carrier and enzyme to be produced separately before isopeptide conjugation. Although kinetic analysis clearly indicated efficient biocatalyst activity, the PVX carrier interfered with substrate binding. To overcome this, the suitability of tobacco mosaic virus (TMV) was tested, which can also accommodate a larger number of ST peptides. We produced TMV particles displaying ST as a new platform for the immobilization of enzymes such as Cel12A, and compared its performance to the established PVX-ST platform in terms of catalytic efficiency. Although more enzyme molecules were immobilized on the TMV-ST particles, we found that the rigid scaffold and helical spacing significantly affected enzyme activity.

Identification of anti-TMV active flavonoid glycosides and their mode of action on virus particles from Clematis lasiandra Maxim.

Tobacco mosaic virus (TMV) is a disreputable plant pathogen that causes a decline in the quality and yield of various economic crops. Natural products are important potential sources of biopesticides to control TMV. This study focuses on the discovery of anti-TMV active flavonoid glycosides and their mode of action on TMV particles from Clematis lasiandra Maxim. A new benzoyl acylated flavonoid glycoside, kaempferol 3-O-(2''-benzoyl)-β-d-glucopyranosyl-7-O-α-l-rhamnopyranoside (1), and nine known flavonoids (2-10) were identified first from C. lasiandra. The hydroxyl group at C-7, E-p-coumarate at C-6'' in the Glc of C-6, and the glucuronic acid at C-3 were functional groups for the antiviral flavonoid glycosides. Flavonoids 2, 5, and 6 showed higher inactivation efficacies of 64.62% to 82.54% compared with ningnanmycin at 500 μg ml-1 . The protective and curative efficacies for 2 and 5 were 57.44-59.00% and 41.17-43.92% at 500 μg ml-1 , respectively. Compound 5 showed higher TMV systemic resistance with control efficacies of 41.64%, 36.56% and 27.62% at concentrations of 500, 250 and 125 μg ml-1 compared with ningnanmycin in K326 tobaccos, respectively. Compound 5 can directly fracture TMV particles into small fragments combining with the fusion phenomena, and TMV-CP was an important target for 5 to break TMV particles. Flavonoid glycosides from C. lasiandra showed potent antiviral activities against TMV with multiple modes of action including inactivation, protective and curative effects, and inducing systemic resistance. TMV-CP was an important target for active flavonoid glycosides to fracture TMV particles. The results provided evidence that flavonoid glycosides from C. lasiandra have the potential to control TMV.

Detection of plant virus particles with a capacitive field-effect sensor

Plant viruses are major contributors to crop losses and induce high economic costs worldwide. For reliable, on-site and early detection of plant viral diseases, portable biosensors are of great interest. In this study, a field-effect SiO2-gate electrolyte-insulator-semiconductor (EIS) sensor was utilized for the label-free electrostatic detection of tobacco mosaic virus (TMV) particles as a model plant pathogen. The capacitive EIS sensor has been characterized regarding its TMV sensitivity by means of constant-capacitance method. The EIS sensor was able to detect biotinylated TMV particles from a solution with a TMV concentration as low as 0.025 nM. A good correlation between the registered EIS sensor signal and the density of adsorbed TMV particles assessed from scanning electron microscopy images of the SiO2-gate chip surface was observed. Additionally, the isoelectric point of the biotinylated TMV particles was determined via zeta potential measurements and the influence of ionic strength of the measurement solution on the TMV-modified EIS sensor signal has been studied.

(IMCS Second Place Best Paper Award) High-Density Immobilization of Tobacco Mosaic Virus Nanotubes As Enzyme Nanocarriers Onto Field-Effect Biosensor Structures

Introduction Viruses are not only infectious agents, they are also known as promising functional building blocks for application in nano- and biotechnologies. The tobacco mosaic virus (TMV) was the first studied plant virus and is widely distributed; it infects vegetables, like tomato, bell pepper, beans and other members of the family Solanaceae, while it is totally harmless for mammals [1]. It is one of the most studied plant viruses and its genome is completely sequenced, whereby genetical and chemical modification is easy. TMV has a nanotube-like shape with a length of 300 nm, an outer diameter of 18 nm and an inner diameter of 4 nm. Since it possesses a high chemical and physical robustness, it can be integrated with different electronic transducers for bio- and chemical sensing applications [2].Recently, we presented a TMV-based amperometric glucose biosensor [3] and a potentiometric penicillin biosensor [4]. TMV was used as enzyme nanocarrier for the enzyme glucose oxidase and penicillinase, respectively [3,4]. The sensitivity and detection limit of these biosensors, among others, depend on the density of TMVs on the sensor surface. The surface density of the immobilized TMVs is strongly influenced by the electrostatic interactions between the charged TMVs and sensor surface as well as by the inter-TMV-nanotubes repulsion, which could be changed by varying the pH value and the ionic strength of the TMV solution. In this study, we investigated an impact of the pH value and ionic strength of the TMV solution on the surface density of TMV nanotubes immobilized onto Ta2O5-gate capacitive field-effect electrolyte-insulator-semiconductor (EIS) sensors. Materials and Methods TMV particles modified with biotin-linker molecules (TMVBio), which serve as binding sites for streptavidin-conjugated enzymes, have been immobilized onto capacitive field-effect EIS sensors with Ta2O5 as transducer layer, as shown in Fig. 1. Immobilization was performed from TMVBiosolutions with different values of pH between pH 3.0 and pH 9.5 and ionic strength between 0.1 mM and 750 mM. The sensors have been electrochemically characterized before and after TMVBio immobilization by capacitance-voltage- and constant-capacitance methods, respectively. In addition, the density of the immobilized TMVs and morphology of the sensor surface has been investigated by means of scanning electron microscopy (SEM). Results and Conclusions The TMVBio density on the Ta2O5 sensor surface was influenced by varying the pH value (as exemplarily shown in Fig. 1) and ionic strength (not shown) of the TMVBio solution. The amplitude of the field-effect sensor signal correlates well with the density of the immobilized TMVBionanotubes. Thus, optimized conditions for the high-density immobilization of TMVBio nanotubes onto the Ta2O5 surface and thereby enhanced biosensing have been found. Details of the experiments and the obtained results will be presented and discussed. Acknowledgements The authors like to thank Dr. Claudia Koch and Rebecca Hummel, Stuttgart, for scientific and technical support.