Inactivation Of Tobacco Mosaic Virus Research Articles

Inactivation of tobacco mosaic virus using gamma irradiation and its potential modes of action

Gamma irradiation is a non-thermal processing technique used to disinfect harmful microorganisms in agriculture. This technology has been shown to be an effective method to control bacterial and fungal plant pathogens. However, its effect on viral plant pathogen is less understood. Gamma irradiation was evaluated for the inactivation of tobacco mosaic virus (TMV). TMV infectivity has gradually decreased following irradiation in a dose-dependent manner and virus was completely inactivated at a dose over 40 kGy. Transmission electron microscopy revealed that increased gamma irradiation disrupts the virion structure and degrades viral proteins, which results in TMV inactivation. The mechanisms, through which gamma irradiation inactivates TMV, can be directly associated with the damage to the virus constituents.

Co-digestion of tobacco waste with different agricultural biomass feedstocks and the inhibition of tobacco viruses by anaerobic digestion

Tobacco is widely planted across the world especially in China, which means that a large amount of tobacco waste needs to be treated. This study investigated the biogas fermentation of tobacco stalks co-digested with different biomass feedstocks and the inactivation of Tobacco mosaic virus (TMV), Cucumber mosaic virus (CMV) by anaerobic digestion. Results showed that the maximum methane yield of tobacco stalks at 35°C was 0.163m3CH4⋅kgVS−1, which was from the co-digestion of tobacco stalks, wheat stalks and pig manure. The largest VS removal rate of tobacco stalks was 59.10%. Proven by indicator paper stripe, half-leaf lesion and RT-PCR, CMV could be inactivated by mesophilic and thermophilic anaerobic digestion, whereas TMV could be only inactivated by thermophilic anaerobic digestion over 20days. These results suggested that using tobacco stalks as feedstock for anaerobic digestion and applying the digested residue and slurry to Solanaceae crop land are feasible.

Inactivation of Tobacco mosaic virus in soil by Pseudomonas putida A3-m strain to prevent virus mosaic disease

Previous studies showed that Pseudomonas putida A3-m exhibited a strong antiviral activity against Tobacco mosaic virus (TMV) in tobacco. However, its potential effect on the amount of TMV in the soil remains unknown. In this study, all plants in two treatments and the control did not have significant differences in terms of height and fresh weight at 20 and 30 days after transplanting. And the effect of A3-m on soil TMV community in tobacco rhizosphere was investigated through real-time reverse transcriptase-polymerase chain reaction. The results showed that the amount of TMV in the A3-m treated soil initially decreased significantly compared with the control whose amount maintained at certain levels. The severity rate of TMV disease was significantly lower in P. putida A3-m treatment before the seedlings were transplanted (referred to as the early treatment) compared with that in P. putida A3-m treatment after the seedlings were transplanted (referred to as the late treatment) and in the control. These data show that the early treatment of the soil benefit for protecting the plants against TMV infection. Thus, P. putidaA3-m can be utilized as a biological control for plant protection against viral pathogens through the inactivation of TMV in the soil. Key word: Tobacco mosaic virus, Pesudomonas putida A3-m, inactivation, soil.

Effective Thermophilic Composting of Crop Residues for Inactivation of Tobacco Mosaic Virus

An effective thermophilic composting bioreactor, in which a homogenous distribution of temperature was maintained at 63-65°C by the addition of a bioavailable carbon and low mixing, was developed. The bioreactor operated on a mixture of tomato plant residues-wood shavings-municipal solid waste compost infected with tobacco mosaic virus (TMV). The initial C: N ratio and moisture content of the compost mixture were adjusted to 30:1 and 60%, respectively. The composting process was successful in destroying the tobacco mosaic virus. The results showed that the ability of the untreated virus (inoculum) to infect tobacco plants (150 LL L-1) was much higher than its ability to infect tomato plants (22 LL L-1). The TMV completely lost its ability to infect the leaves of susceptible hosts (tobacco and tomato plants) after 96 hrs of controlled thermophilic (63-65 oC) composting (or 126 h from the start of the composting process). Semilog plots of the ratio of the infection ability of the surviving virus to that of the initial inoculum (as measured by the number of local lesions) were developed. The decimal reduction time (the time necessary to reduce the infection ability of TMV by 1-log or 90%) was found to be 62.4 and 109.7 hrs for tobacco and tomato plants, respectively. The relatively short time required for complete inactivation of TMV in this study was achieved as a result of the extension of the thermophilic stage and maintaining a constant high temperature with a uniform temperature distribution by the continuous addition of the proper amount of bioavailable carbon (used cooking oil) and low mixing.

Interaction between the tobacco mosaic virus movement protein and host cell pectin methylesterases is required for viral cell-to-cell movement.

Virus-encoded movement protein (MP) mediates cell-to-cell spread of tobacco mosaic virus (TMV) through plant intercellular connections, the plasmodesmata. The molecular pathway by which TMV MP interacts with the host cell is largely unknown. To understand this process better, a cell wall-associated protein that specifically binds the viral MP was purified from tobacco leaf cell walls and identified as pectin methylesterase (PME). In addition to TMV MP, PME is recognized by MPs of turnip vein clearing virus (TVCV) and cauliflower mosaic virus (CaMV). The use of amino acid deletion mutants of TMV MP showed that its domain was necessary and sufficient for association with PME. Deletion of the PME-binding region resulted in inactivation of TMV cell-to-cell movement.

Mechanism of inactivation of tobacco mosaic virus with ozone

The inactivation mechanism of tobacco mosaic virus (TMV) in a phosphate buffer (pH 6.9) by ozone was studied. We previously reported that the damage of naked TMV-RNA occurred at the guanine moiety of RNA (Shinriki et al., Biochim. biophys. Acta 655, 323, 1981). In this paper, we clarified the mode of the inactivation of TMV by using tritium-labeled TMV (TMV ∗) prepared by the reconstitution of tritium-labeled TMV-RNA (TMV-RNA ∗) and coat protein of TMV. It was found that the amount of extracted TMV-RNA ∗ from ozone-treated TMV ∗ decreased with the advance of ozonization, and that there was good correlation between the loss of infectivity and the decrease of recovery of TMV-RNA ∗. When TMV lost its infectivity due to ozone, tryptophan and tyrosine of the coat protein were also degraded by ozone. Polyacrylamide gel electrophoretic analysis of the substance produced during ozonization showed that the coat protein subunits were aggregated with each other and cross-linked with TMV-RNA ∗. From these results, it was concluded that the inability of uncoating is the major cause of the TMV inactivation by ozone.

Morphological changes in tobacco mosaic virus after exposure to space.

Tobacco mosaic virus exposed to space during the Gemini 12 flight showed gaps in the virus coat and fragmented into smaller segments. These changes were similar to those produced by ultraviolet radiation in the laboratory and support the conclusion that solar ultraviolet radiation is a major cause of inactivation of tobacco mosaic virus in space.

Efficiency of nitrous acid as an inactivating and mutagenic agent of intact tobacco mosaic virus and its isolated nucleic acid.

When tobacco mosaic virus (TMV) and its isolated nucleic acid (TMV-RNA) were treated with nitrous acid, the nucleic acid was inactivated six times faster than the intact virus. Inactivation of both the infectious entities was exponential with treatment time to 0.1% level of survival. Eight different mutant phenotypes were scored after inactivation of TMV and TMV-RNA to 50, 10, 1.0, and 0.1% survival levels. Significantly more mutants in relation to unaltered isolates were induced at all levels of survival upon nitrous acid treatment of TMV than of TMV-RNA. Furthermore, the proportion of two specific mutant phenotypes was significantly greater in treated TMV than in treated TMV-RNA. No qualitative differences, however, were observed between the mutational spectra of nitrous acid-treated TMV and TMV-RNA. These results indicate that, in the intact virus, the viral capsid protects some of the sites involved in lethality; thus, proportionately more mutants are induced on nitrous acid treatment of TMV versus TMV-RNA.

Inactivation of purified plant viruses and their nucleic acids by photosensitizing dyes.

Abstract The reaction between tobacco mosaic virus (TMV ), alfalfa mosaic virus (AMV), cucumber mosaic virus (CMV), their ribonucleic acids (RNA), and three photosensitizing dyes was investigated. Optimal conditions for the inactivation of TMV by toluidine blue and acridine orange were: presence of light, pH 9, and 0.01 ionic strength. Inactivation of TMV by neutral red was optimal at pH 6, but this dye was less effective than the other two dyes. TMV-RNA was more sensitive to inactivation than was the whole virus. Inactivation of TMV by toludine blue at a low concentration followed a slow first-order reaction after an instantaneous decrease. At a high dye concentration, inactivation was first order over 98% of the range, followed by a period of decreased inactivation which also seemed to be first order. Inocula containing high concentrations of toluidine blue were less infectious than inocula from which dye had been removed. Thus dye at high concentrations seemed to affect not only the virus but also the host. TMV could be photosensitized when free toluidine blue was removed from the suspension prior to illumination, indicating that dye bound to the virus was responsible for inactivation. The capacity of TMV to bind acridine orange was reduced by suboptimal ionic strength of the suspension, but not by suboptimal pH; whereas infectivity was affected by both suboptimal pH and ionic strength. Binding of TMV to dyes was also indicated by a shift of maximal absorption of virus-treated dyes and dye-treated virus. No morphological changes of photosensitized TMV were visible under the electron microscope. AMV, AMV-RNA, CMV, and CMV-RNA were more rapidly inactivated than TMV or TMV-RNA. As with TMV, inactivation of these viruses was more gradual in the dark than in the light. CMV and CMV-RNA could also be inactivated by exposure to light without treatment with photoreactive dyes.

The photodynamic inactivation of tobacco mosaic virus and its ribonucleic acid by acridine orange

Acridine orange-sensitized photoinactivation of tobacco mosaic virus and tobacco mosaic virus-ribonucleic acid has been studied in detail. The intact virus is only partially inactivated by acridine orange even at 10 −5 M acridine orange. Tobacco mosaic virus-ribonucleic acid inactivation by acridine orange has been studied from the standpoint of the ribonucleic acid-dye complexes (Complex I and Complex II), and the ternary complex of ribonucleic acid-dye and metal ions (Complex III). Tobacco mosaic virus-ribonucleic acid infectivity was destroyed with first-order kinetics by low levels of acridine orange (Complex II). “Complex III”, formed in the presence of paramagnetic metal ions, was resistant to photodynamic inactivation, the protection by metal ions being linear with respect to concentration of the metal ions. A comparison of the photosensitization by methylene blue and acridine orange with respect to tobacco mosaic virus-ribonucleic acid and free guanosine showed that, while acridine orange was only 5–6 times less active than methylene blue with ribonucleic acid, it was considerably less efficient in destroying guanosine. Sedimentation studies with acridine orange-inactivated tobacco mosaic virus-ribonucleic acid showed that mild conditions leading to measurable loss of infectivity involve no depolymerization of tobacco mosaic virus-ribonucleic acid which sets in only under more severe conditions. The results have been discussed from the standpoint of the mechanisms of photodynamic inactivation.

Tudies on the inactivation of tobacco mosaic virus by ultraviolet light.

Abstract— The irradiation of TMV with u.v. light of 2537 Å wavelength results in the binding of protein subunits to the RNA. These bound subunits are stable towards warm sodium dodecyl‐sulfate; however, the binding is not covalent since the subunits are removed by 66% acetic acid, guanidine hydrochloride, or phenol. Approximately one protein subunit is bound per lethal biological ‘hit’. The virus and the nucleic acid extracted from irradiated virus show virtually identical rates of inactivation.

The relationship between inactivation of tobacco mosaic virus by X-rays and breakage of nucleic acid

The analytical ultracentrifuge, with u.v. absorption optics, has been used to demonstrate and measure bond breakage in the main chain of RNA strands extracted from X-irradiated TMV. When the whole virus was irradiated in solution at 0°, the rate of loss of intact RNA strands was about equal to the rate of loss of viral activity. However, the reduction in yield of extracted RNA that accompanied increasing X-ray dose, together with the probability that longer strands would be selectively lost, cast considerable doubt on the significance of this result. When TMV was irradiated while frozen at about —50° or while in centrigugal pellets with little water present, the correlation between reduction in yield and X-ray dose disappeared. Those data indicate a D 0 for strand breakages of 4.0·10 5 R as compared to an average measured D 0 of 1.8·10 5 R for virus inactivation. Thus, breakage proceeds 40% as rapidly as does inactivation. Also, 40% of all the ionizations in TMV RNA induced by radiation may be expected to occur among backbone chain electrons. If it is assumed 1 that any ionization within the nucleic acid complement of a virus inactivates the virus, then the present data indicate that ionizations within the backbone chain of TMV RNA break main chain bonds and cleave the strand with an efficiency of close to 100%. By analogy, then, it is suggested that ionizations elsewhere in the nucleic acid also are effective in breaking bonds.

Recovery of infectious nucleic acid from tobacco mosaic virus inactivated by antibody.

ALTHOUGH the combination of antibody with tobacco mosaic virus is reversible1, it does not necessarily follow that dissociating the antibody restores infectivity. Inactivation could be permanent despite the reversibility of the antibody - virus reaction. Several reports have indicated, however, that antibody inactivation of tobacco mosaic virus is not entirely irreversible.

A Comparative Study of Antibody Inactivation of Tobacco Mosaic Virus

Summary Five strains of tobacco mosaic virus (TMV) were inactivated by three different antisera. All strains are related antigenically and fall into at least three distinct groups. These relationships are in basic harmony with those previously uncovered (1, 2). The masked and common strain of TMV can be distinguished by the inactivation method. The data suggest that the amino acids histidine and methionine probably are not involved in the antigenic structure associated with the nfectivity of Holmes' ribgrass.

The Effect of Developing Embryos on Plant Viruses

The theory that inactivation of tobacco mosaic virus by developing embryos prevents its seed transmission was tested by growing tomato embryos in media containing the virus. In many trials, using a variety of techniques, no evidence of any virus�inactivating activity was detected.

The reaction of tobacco mosaic virus with formaldehyde: III. Kinetics of the Loss of Infectivity

The experiments reported here were designed to elucidate the nature of the chemical group on tobacco mosaic virus nucleoprotein which causes the biological inactivation of the virus when it reacts with formaldehyde. The methods of chemical kinetics were used to determine the specific reaction rate constants for the inactivation of the virus by 2% formaldehyde at several pH values within the range 4.1 to 8.4 and at 30°, 35°, and 40°. The reaction rate was found to be only slightly pH dependent, indicating that the critical group does not dissociate within the pH range studied. The reaction did not vary with ionic strength. A plot of the Arrhenius equation for the velocity constants obtained at pH values of 5.3, 6.8, and 7.8 gave an energy of activation of 19,500 calories per mole, which is the approximate value found for a reaction involving a single chemical bond. The energy of activation was invariable with pH. The rate constant was dependent on formaldehyde concentration when 2% virus solutions were allowed to react with 1%, 2%, 4%, and 7% formaldehyde solutions at pH 6.6 and 30°. These results could be interpreted on the basis of the inactivation's being caused by the reaction of a single formaldehyde molecule at one site on the virus characteristic particle. Prior treatment of the virus nucleoprotein with iodine, diazotized sulfanilic acid, acetic anhydride, and propionaldehyde failed to produce any effect on the rate constants for the subsequent formaldehyde inactivation process, indicating that the sulfhydryl, phenolic hydroxy, and protein amino groups are probably not the ones which cause the inactivation when they react with formaldehyde. Since changes in pH did not appreciably affect the rate constant value between pH 4.1 and pH 8.4, certain other groups can be eliminated from consideration on the basis of their pK values. Proton donor groups with pK values less than 9 and proton acceptor groups with pK values greater than about pK 3 to 4 can be excluded. The amide groups are probably not fundamentally involved, since they are reported to react more readily in acid medium. If this is true, then the inactivation rate constant would increase with increasing acidity, which is contrary to the results of this work. It has been shown by other investigators that tryptophan in tobacco mosaic virus probably does not react with formaldehyde in the pH range employed here. On the basis of the foregoing considerations, it is probable that the groups most likely involved in the formaldehyde inactivation of tobacco mosaic virus are the beta hydroxy groups of serine and threonine, the hydroxy group of ribose, and the amino group on the pyrimidine and purine rings in the nucleic acid.

Mechanism of Inactivation of Tobacco Mosaic Virus by X-rays

INACTIVATION of tobacco mosaic virus by X-rays has been studied by Gowen and Price, by Lea and associates1 and by us2. The fraction surviving is an exponential function of dose with D 0 about 2.3 × 105 r. This value corresponds to a target 3000 A. long and about 40 A. in diameter2.

Inactivation of Tobacco Mosaic Virus by Rabbit Antiserum

Summary Quantitative precipitin analysis of rabbit antiserum to TMV revealed antibody-antigen ratios which plotted as a curve, and suggested a ten-fold change in R from 0.2 at equivalence to 2.0 in extreme antibody excess. An explanation was offered for this large ratio change in which it was assumed that all of the antigenic sites on the virus particle were not satisfied in antibody excess, because of steric hindrance in the precipitates. The virus was shown to be specifically and rapidly inactivated by antibody. Through the serum dilution range examined, specific inactivation was a function of the antibody-antigen ratio. Virus aggregation occurred, but accounted for little or none of the reduced lesion count. The extent of aggregation, at any one ratio, was a function of the total concentration of the reactants. Virus-antibody aggregates were infectious if they contained one or more viable particles. The inactivation data together with the UV multiplicity studies further suggested the possibility that only a minute fraction of the total viral population was infectious. An hypothesis, based on the interpretation of the quantitative precipitin reaction, was presented to explain the inactivation results. It was assumed that the same conditions for steric hindrance which operated in the precipitin reaction were encountered during inactivation to prevent an antibody from combining at a specific virus site necessary to initiate a lesion.

Inativação do vírus do mosaico comum do fumo pelo filtrado de culturas de Trichoderma sp

Trichoderma sp. grown in liquid medium produced a substance which has caused up to 90% reduction in the infection capacity of the tobacco mosaic virus, measured in number of local lesions on Nicotiana glutinosa half leaf inoculations. Under the present experiments the fungus has been grown in liquid medium and the filtrates used in inactivation tests on the virus. From the fact that the liquid filtrate showed inactivation power after only two days of culture it is concluded that the inactivation is of the nature of a secretion of the fungus into the liquid medium. The inactivation proceeded within 2 minutes after mixture of the culture extract with the virus, no better results were obtained if the mixtures allowed to stand for a longer period of time. If cultures were grown in the light in the laboratory the inactivation power decreased after the second day as was found by Weindling (8), for the effect on Rhizoctonia sp. If grown in the dark the inactivator continues to increase in concentrations for at least 22 days. In the dark no spores formation could be observed. Physiological activity associated with spores formation may be the reason for the loss of inactivator in day light grown cultures where spores start to be formed after the second day of growth. Extraction of the inactivator with chloroform was attempted according to Weindling's method. The inactivator could never be completely extracted since the treated filtrate kept showing inactivation on tobacco mosaic virus. This suggests that the inactivator for the virus may be a different from the one found by Weindling against Rhizoctonia sp. By ultracentrifugation at 35,000 r.p.m. no inactivator could be obtained from the filtrate. Using Takahashi's acetone method a whitish precipitate could be obtained which showed an inactivation of tobacco mosaic virus.

The ultraviolet light and photosensitized inactivation of tobacco mosaic virus.

The quantum yield for the inactivation of tobacco mosaic virus has been determined at 253.7 mmicro and found to be 4.3 x 10(-6). The possible significance of the observed one-hit process of inactivation has been discussed in terms of the kinetics and the rupture of model substances including nucleic acid. The ultraviolet light inactivation, which proceeds independent of oxygen, occurs without change in physicochemical properties, with the possible exception of an enhanced sensitivity to thermal denaturation. The photosensitized inactivation of virus by acriflavine has been found to proceed parallel with the destruction of the dye. The action was found to be dependent upon adsorbed dye, and the inactivation is enhanced by the presence of oxygen.