Tomato Bushy Stunt Research Articles

A single amino acid substitution in the ORF1 of cymbidium ringspot virus determines the accumulation of two satellite RNAs

Tombusviruses may support the replication of satellite (sat) RNAs. In particular, two satRNAs, sat L and Cymsat RNAs, are replicated by carnation Italian ringspot (CIRV) and tomato bushy stunt (TBSV) virus, but not by cymbidium ringspot virus (CymRSV) in vitro transcripts unless they contain a poly(A) tail at the 3′ end. Conversely, the replication of both satRNAs was supported by virus particles or viral RNA of the original CymRSV inoculum even in the absence of the poly(A) tail. Sequence and mutational analyses revealed that the full-length infectious CymRSV clone contains one relevant sequence variation in the ORF 1-encoded protein (p33) compared with the original inoculum, i.e. a Ser19 TCC codon instead of a Phe19 TTC codon, which inhibited the replication of sat L and Cymsat RNAs. It is suggested that this amino acid is contained in a domain essential for the replication of some subviral RNAs.

Chenopodium necrosis: a distinctive strain of tobacco necrosis virus isolated from river water

SUMMARYA distinctive strain of tobacco necrosis virus (TNV) of unknown source was repeatedly isolated from water of the River Avon (Warwickshire) and two of its tributaries (R. Swift and R. Alne) using a technique developed for the concentration and isolation of water‐borne bacteriophages. The same strain was isolated from the rivers Cam and Thames and from Lake Esthwaite (Cumbria) together with tomato bushy stunt virus.The TNV strain, designated Chenopodium necrosis (TNV‐CN) was mechanically transmissible to C. amaranticolor and C. quinoa in both of which it caused local lesions and systemic infection. TNV‐CN caused no infection when inoculated to tobacco (Nicotiana tabacum cv. White Burley) plants.The virus was not adsorbed to soil, could be isolated from leachate of soil in which systemically infected C. quinoa were grown and C. quinoa plants became infected when grown in soil watered with suspensions of the virus.The virus was not transmitted by Myzus persicae but was vectored by the zoospores of a lettuce isolate of Olpidium brassicae.TNV‐CN was infective after 10 min at 85 °C., 3 wk at 20 °C and when diluted to 10‐8 but not 10‐9. Purified virus preparations contained c. 26 nm isometric virus particles.TNV‐CN contained single‐stranded RNA (mol. wt 1·5 × 106) and one protein (mol. wt c. 26·4 × 103) which co‐electrophoresed in polyacrylamide gels with the protein of the D strain of TNV (TNV‐D). Analytical centrifugation of TNV‐CN indicated a single component virus with the same sedimentation coefficient (s20, w= 115S) and buoyant density (1·385) in a CsCl gradient as those of TNV‐D. TNV‐CN and TNV‐D were indistinguishable serologically.

Internal structural anisotropy of spherical viruses studied with magnetic birefringence.

Six so called spherical viruses (four plant and two animal) are shown to exhibit magnetically induced birefringence in solution. They must therefore be magnetically and optically anisotropic. This is attributed to static structural anisotropy of the interiors as neither natural shape nor field-induced deformations are likely causes. Thus at least part of these virus cores have a symmetry differing from that of their capsids. An estimate of the average orientation of the RNA bases is given for the plant viruses: turnip yellow mosaic, bromegrass mosaic, tomato bushy stunt and turnip crinkle. The packing geometry of the nucleic acid/protein cores of adenovirus and probably influenza virus are anisotropic but to an extent that cannot be quantified.

Specificity of trapping of plant viruses on antibody-coated electron microscope grids

Conditions suitable for the serological trapping of different plant viruses on electron microscope grids coated with the respective antisera, have been defined. The viruses studied are turnip yellow mosaic, tomato bushy stunt and cauliflower mosaic viruses, and five strains of tobacco mosaic virus. Since the extent of serological cross-reactivity between the TMV strains was known, it was possible to determine how closely related serologically two strains of a virus have to be in order to be trapped on grids coated with the heterologous antiserum. The findings show that optimal conditions for the specific trapping of virions must be determined empirically for each particular virus.

Physiochemical and serological properties of a virus from saguaro cactus

The molecular weight of a spherical virus from saguaro cactus (SV) was estimated to be 7.9 × 10 6 from a sedimentation coefficient of 118 S and a diffusion coefficient of 1.22 × 10 −7 cm 2/sec. SV was found to contain 17.2% RNA, based on its phosphorus content of 1.66% and the nucleotide composition of its nucleic acid (29% guanylic acid, 24% adenylic acid, 21% cytidylic acid, and 27% uridylic acid). A single RNA component, a major protein component (90–95%), and two minor protein components were detected by electrophoresis in polyacrylamide gels and their molecular weights were estimated at 1.4 × 10 6, 38,900, 90,200, and 29,400, respectively. At pH 4, 5, 6, and 7, SV had three electrophoretic components, at pH 8 only two. The three components at pH 7 had similar amino acid compositions. SV was dissociated by low levels of sodium dodecyl- sulphate (0.05% at pH 7 and 0.10% at pH 5) and is probably held together by protein-RNA interactions. SV was not related serologically to sowbane mosaic, southern bean mosaic, tobacco necrosis, carnation mottle, turnip crinkle, tomato bushy stunt, cucumber necrosis, or carnation ringspot viruses.

Specificity and Site of in vitro Acquisition of Tobacco Necrosis Virus by Zoospoores of Olpidium brassicae

SUMMARY Four single-sporangium isolates of Olpidium brassicae were inoculated on 14 different plant species to determine their ability to infect and reproduce in each host and their ability to transmit two tobacco necrosis virus isolates. Two O. brassicae isolates, one from lettuce and one from tomato, were able to reproduce in most hosts and transmitted both tobacco necrosis virus isolates to all hosts. By contrast, a mustard isolate reproduced in only six species and was not a vector of tobacco necrosis virus. An oat isolate reproduced in six species (three in common with the mustard) and was a poor vector, transmitting one tobacco necrosis virus isolate to only two hosts and not transmitting the other tobacco necrosis virus isolate to any host. Zoospores of the different fungus isolates were mixed with suspensions of virus, washed, negatively stained, and examined in the electron microscope. Virus adsorbed tightly to the surface membranes (plasmalemma of the body and axonemal sheath) of zoospores of isolates that transmitted it, but not to those of the non-vector (mustard) isolate. Zoospores of the poor vector (oat) isolate adsorbed fewer particles of the tobacco necrosis virus isolate they transmitted than did those of good vectors and did not adsorb the other tobacco necrosis virus isolate. Most, but not all, of the observed specificity of transmission of tobacco necrosis virus seems to be associated with the ability or inability of zoospores to adsorb the virus on their surfaces. None of these isolates adsorbed particles of turnip yellow mosaic, tomato bushy stunt or cucumber necrosis viruses. O. brassicae zoospores also adsorbed satellite virus particles. Zoospores of Olpidium cucurbitacearum adsorbed particles of cucumber necrosis virus, but not of tobacco necrosis virus, to their surfaces. Thus, in vitro acquisition consisted of a tight adsorption of virus to the zoospore surface membranes in each of the three known instances of this type of relationship.

Physical, chemical, and serological studies on carnation mottle virus

Preparations of carnation mottle virus purified at pH 7.4 contained a single component. The virus had an isoelectric point of 5.2 and was slowly degraded at pH 6.2 to 6.4 in 0.02 M sodium phosphate buffers. The virus contained 1.89% phosphorus and 20% RNA. The nucleic acid consisted of 30% adenylic acid, 27% guanylic acid, 19% cytidylic acid, and 24% uridylic acid. The protein consisted of about 243 amino acid residues and had a molecular weight of 26,200. The molecular weight of the virus calculated from the sedimentation and diffusion coefficients was 7.7 × 10 6. Carnation mottle virus was not serologically related to sowbane mosaic, turnip crinkle, tomato bushy stunt, or cucumber necrosis viruses.

Amino acid sequence of the f 2 coat protein

The effect of the anionic detergent sodium dodecyl sulfate (SDS) on a number of simple isometric RNA viruses and empty capsids was tested. Some viruses showed extreme sensitivity, e.g., cucumber mosaic (CMV), brome mosaic (BMV) and alfalfa mosaic (AMV) virus, some extreme resistance, e.g., turnip yellow mosaic (TYMV) and tomato bushy stunt (TBSV) virus, and some intermediate resistance, e.g., southern bean mosaic virus (SBMV) and bacteriophage f2, to dissociation into components by this detergent. In the viruses most sensitive to SDS, virion dissociation is apparently caused by disruption of the electrostatic protein-RNA interactions which are responsible for stabilizing the virions. It is proposed that dodecyl sulfate (DS−) ions bind by means of the hydrocarbon chain to specific binding sites on the virion so that the sulfate groups are near lysine- or arginine-phosphate interaction points; these interactions are neutralized and the phosphates are repulsed, resulting in virion dissociation. Based on this hypothesis several predictions were made, tested experimentally and found valid: (1) Virions become more resistant to SDS as the contribution of the protein-protein interactions to virus stability increases. This was confirmed by testing the sensitivities of a number of viruses, including those mentioned above, to SDS. (2) Capsids devoid of nucleic acid are less sensitive to SDS than the respective intact virions. The behavior of TYMV, BMV and bacteriophage f2 virions and capsids with SDS confirmed this prediction. (3) Positive and neutral detergents do not cause dissociation of the SDS-sensitive viruses. (4) Reassembly of viruses stabilized by protein-RNA interactions is inhibited by SDS but not by positive detergents. Predictions (3) and (4) were confirmed by appropriate experiments with CMV or BMV and SDS, dodecyltrimethylammonium chloride (DTAC) and Triton-X-100. Measurements of amounts of detergent bound at low SDS concentrations showed that CMV and BMV have much greater affinitives than TYMV for DS− ions. It is suggested that SDS is useful as a probe for protein-RNA interactions and that relative sensitivity to SDS could be used in categorizing viruses according to stabilizing interactions.

Studies of pelargonium leaf curl virus

SUMMARYPelargonium leaf curl virus (PLCV) was compared with nine other viruses with isometric particles: broad bean mottle, carnation mottle, carnation Italian ringspot (CIRV), cymbidium ringspot, raspberry ringspot, tobacco necrosis, tomato bushy stunt (TBSV), turnip crinkle and a virus from Trifolium repens.PLCV was serologically related to CIRV and TBSV, although cross‐reactions between the latter two viruses occurred only with sera from rabbits given one more injection of antigen than usual. Considerable antigenic differences were found among seven isolates of PLCV, and between these isolates and CIRV and TBSV.Cymbidium ringspot virus was serologically closely related to the virus from Trifolium repens, and both were similar but serologically unrelated to the PLCV‐CIRV‐TBSV group. No other serological relationships were found among the ten viruses. Serologically related viruses gave similar zones after centrifugation in sucrose density‐gradients and, usually similar sedimentation coefficients.The different viruses infected twenty‐three to forty‐four out of sixty plant species tested. Carnation mottle and turnip crinkle viruses were not eliminated from plants grown at 37d̀C. for 4–6 weeks, whereas the other viruses usually were. The properties of the viruses in vitro were broadly similar.

Partial amino acid composition of tomato spotted wilt virus, (Strain E)

The amino acid composition of a preparation of strain E of tomato spotted wilt virus (TSWV) has been determined with the aid of a Moore and Stein automatic amino acid analyser with the following results (amino acid nitrogen is expressed as percentage of total nitrogen at 44% recovery): aspartic 2.34, threonine 1.72, serine 2.44, glutamic 2.71, glycine 2.38, alanine 2.95, valine 2.00, isoleucine 2.07, leucine 3.11, tyrosine 1.06, phenylalanine 1.66, lysine 4.05, histidine 1.50, amido-N 6.27, arginine 5.94, proline 2.01. In comparison with published results of other plant viruses TSWV was found to contain twice as much lysine, 1.8 times as much histidine, but less than half the amount of threonine as the average content of the viruses of tobacco mosaic, turnip yellow mosiac, and tomato bushy stunt. When compared on the basis of the same percentage recovery of amino acid N, lysine, histidine, and amido N are also considerably higher in the virus than published figures give for plant leaf proteins.

A nucleotide triplet code for amino acids

We have recently devised a nucleotide triplet code which meets one of the requirements for being the correct solution to the “coding problem” in biology (Woese, 1961). This code was designed to, and does, predict the nucleotide composition of each of the six ribonucleic acid viruses, tobacco mosaic, tomato bushy stunt, southern bean mosaic, turnip yellow, cucumber, and polio, from their particular amino acid compositions. Because the protein and nucleic acid compositions of the six viruses vary considerably, it is probable that the code, while not unique, is one of a relatively small number of closely related codes, all of which fit the experimental data (Yčas, 1960). In the present communication we shall develop this code further.

Biophysical studies of the interactions between the viruses of tobacco mosaic and tomato bushy stunt and their rabbit antisera

Light scattering procedures have been employed in the study of the interactions between the viruses of tobacco mosaic (TMV) and bushy stunt (BSV) and their rabbit antisera. These procedures are particularly suited to kinetic studies and to quantitative titrations of virus and of antiserum. The bushy stunt virus system in the region of antigen excess has also been studied by analytical centrifugation and by electron microscopy. The observed distributions of small complexes are consistent with the predictions of the Goldberg theory. In the zone of equivalence ( A G ∼ 1 ) the TMV-antibody complexes occur as loose bundles of from 7 to 14 virus rods. Complexes in the BSV-antibody system appear as loose spherical aggregates of 30 to 60 virus particles. The data for the antibody-combining properties of BSV illustrate the likely behavior of other small plant and animal viruses.

Studies on the ability of light to counteract the inactivating action of ultraviolet radiation on plant viruses.

Summary: Of seven plant viruses tested, all except tobacco mosaic showed the phenomenon of photoreactivation, i.e. plants exposed to visible light after inoculation with preparations partially inactivated by ultraviolet radiation produced more local lesions than plants kept in darkness. Five strains of tobacco mosaic virus, which differed widely in their pathogenicity and other properties, were tested, but none showed the phenomenon. Of the six viruses that did, potato showed it much the most strongly, tomato bushy stunt and a tobacco necrosis virus the least; cabbage black ringspot, cucumber mosaic and tobacco ringspot were intermediate. Photoreactivation does not occur immediately after plants are inoculated. With irradiated preparations of potato virus X, most particles need about 30 min. to reach the state in which their ability to infect is affected by light; once in this state, exposure to daylight for 15 min. gives almost complete photoreactivation. Some irradiated particles respond to light of 80 f.c., but others need brighter light; no additional response occurs when light intensity is increased above 600 f.c. The sensitive state of potato virus X persists for about 1 hr. in plants kept in darkness, after which the particles seem to be permanently inactivated. With antibodies and enzymes, a constant amount of absorbed energy decreases the activity of a unit weight by a given fraction. This rule does not apply to plant viruses, which lose infectivity with less absorbed energy than the rule predicts. If viruses are more sensitive because they contain nucleic acid, their sensitivity is not a direct function of their content of nucleic acid. Potato virus X has the same nucleic acid content as strains of tobacco mosaic virus, but is inactivated by less absorbed energy and individual strains of tobacco mosaic virus also differ by factors of two in the amount of radiation needed to decrease their infectivity by a given fraction.

HEAT‐THERAPY OF VIRUS‐INFECTED PLANTS

Virus‐free plants were produced from parents systemically infected with the following five viruses: tomato bushy stunt, carnation ring spot, cucumber mosaic, tomato aspermy and Abutilon variegation. The leaves formed while the infected plants were kept at 36°C. were free from symptoms, and test plants inoculated from these remained uninfected. When cuttings were taken from the infected plants at the end of the treatment most grew into healthy plants. The treated plants themselves usually developed symptoms after varying lengths of time at 20°C, but some that before treatment were infected with tomato aspermy, cucumber mosaic or Abutilon variegation viruses, remained permanently healthy.The same method failed to cure plants infected with tomato spotted wilt, potato virus X and tobacco mosaic virus, although it decreased their virus content. Heat‐therapy seems not to be correlated with the thermal inactivation end point of the virus in vitro.

SOME EFFECTS OF HIGH TEMPERATURE ON THE SUSCEPTIBILITY OF PLANTS TO INFECTION WITH VIRUSES

When plants were kept at 36°C. for some time before inoculation, their susceptibility to infection by five mechanically transmissible viruses was greatly increased. When kept at 36° after inoculation, fewer local lesions were produced than at lower temperatures, but the effects of the post‐inoculation treatment differed with different viruses. Tomato spotted wilt and tobacco mosaic viruses multiply in plants at 36°, and the post‐inoculation treatment reduced the local lesions they caused to numbers that varied between 10 and 90% of the control; these two viruses also have large thermal coefficients of heat inactivation. By contrast, tobacco necrosis, tomato bushy stunt and cucumber mosaic viruses, were much affected by post‐inoculation treatment, lesion formation being completely prevented by exposure to 36° for a day or more. These three viruses appear not to multiply in plants at 36°, and although they have high thermal inactivation points, they have small temperature coefficients of thermal inactivation.The extent to which lesion formation was affected by pre‐ or post‐inoculation exposure of plants to 36° depended not only on the length of the treatment, but also on the physiological condition of the plants.The symptoms of infected plants changed considerably if kept at 36°. At 36°Nicotiana glutinosa, inoculated with tobacco mosaic virus, gave chlorotic local lesions instead of necrotic ones, and became systemically infected. When systemically infected plants were brought to ordinary glasshouse temperature, the infected tissues all collapsed and died in a day.

The application of electron microscopy to the study of plant viruses in unpurified plant extracts.

SUMMARY: Rods of variable lengths occurred in sap from plants infected with tobacco mosaic, cucumber mosaic, potato X, potato Y, henbane mosaic, tobacco etch, and cabbage blackring viruses; the first two were about 15 mμ. wide and appeared rigid, the others were about 10 mμ. wide and apparently flexible. Sap from plants infected with tomato bushy stunt, tobacco ringspot and two tobacco necrosiss viruses contained spherical particles about 26 mμ. in diameter; two particles, one about 18 mμ. and the other about 37 mμ. in diameter occurred in sap from plants infected with a third tobacco necrosis virus. No specific particles were identified in sap from plants infected with tomato spotted wilt, potato leaf roll, cauliflower mosaic, tomato aspermy, sugar beet mosaic and sugar beet yellows viruses. Serologically related strains of any one virus were morphologically indistinguishable, but this has little diagnostic value because so also were some unrelated viruses.

THE INFECTION OF PLANTS BY VIRUSES THROUGH ROOTS

Roots of young tomato plants became infected when inoculated with tomato bushy stunt, tobacco mosaic, and potato X viruses. Root infections also occurred when these viruses were added to soil or culture solutions in which plants were growing.The viruses were sometimes localized around their initial entry points in roots; sometimes they invaded the root system but not the shoots, and sometimes they produced full systemic infection of roots and shoots. In some experiments, but not all, systemic infections were more frequent when the upper tap root or superficial roots were inoculated than when fibrous roots were inoculated.In both tomato and potato, virus X spread from diseased to healthy plants sharing the same culture solution, if their roots were in contact, but not otherwise. Infection of the roots of potato plants by inoculation, produced only one plant with virus‐infected haulms, although several had infected tubers.

Over het zuiveren van plantenviren

Some methods of virus purification are discussed. A distinction has been made between the purification, which can be obtained in a rather simple way (e.g. of the viruses of tobacco mosaic, tomato bushy stunt and turnip yellow mosaic) and that, which require a more complicated manipulation (e.g. of the potato virus X).