Some effects of supplying benzyladenine to leaves and plants inoculated with viruses

With tomato spotted wilt virus in petunia leaf strips, N-6 benzyladenine (BA) was as effective as kinetin in decreasing numbers of local lesions, a result which could not be attributed to an effect on the virus per se. Benzimidazole, adenine and ammonium nitrate were without effect. Benzyladenine was more effective than kinetin when supplied through the petioles of excised whole leaves. Local lesions and infectivity of TSWV in detached leaves of Nicotiana rustica were decreased by supplying BA before and after inoculation. Lesions and infectivity were also decreased in attached leaves when BA was applied 9 days before inoculation. BA supplied to attached leaves after inoculation increased infectivity. Supplying BA to the lower leaves of tomato plants before inoculating with TSWV decreased infectivity of unsprayed, systemically infected tip leaves taken as inoculum; BA supplied after inoculation increased infectivity. Local lesions caused by lucerne mosaic virus in excised leaves of Phaseolus vulgaris were decreased in number by supplying BA. The effects of pre- and post-inoculation sprays of BA are considered in relation to cell metabolism. Since pretreating leaves with kinins did not prevent infection, it is suggested that those which move freely through plants without adverse effects on normal growth may prove of value in increasing the tolerance of plants to virus infection.     

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.     

Engineered resistance to tomato spotted wilt virus in transgenic peanut expressing the viral nucleocapsid gene

The nucleocapsid gene of tomato spotted wilt virus Hawaiian L isolate in a sense orientation, and the GUS and NPTII marker genes, were introduced into peanut (Arachis hypogaea cv. New Mexico Valencia A) using Agrobacterium-mediated transformation. Modifications to a previously defined transformation protocol reduced the time required for production of transformed peanut plants. Transgenes were stably integrated into the peanut genome and transmitted to progeny. RNA expression and production of nucleocapsid protein in transgenic peanut were observed. Progeny of transgenic peanut plants expressing the nucleocapsid gene showed a 10- to 15-day delay in symptom development after mechanical inoculations with the donor isolate of tomato spotted wilt virus. All transgenic plants were protected from systemic tomato spotted wilt virus infection. Inoculated non-transformed control plants and plants transformed with a gene cassette not containing the nucleocapsid gene became systemically infected and displayed typical tomato spotted wilt virus symptoms. These results demonstrate that protection against tomato spotted wilt virus can be achieved in transgenic peanut plants by expression of the sense RNA of the tomato spotted wilt virus nucleocapsid gene     

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.     

Infectivity of Chlamydospores vs Microconidia of Fusarium oxysporum f.sp. lycopersici on Tomato

The effect of nature of inoculum on disease induced by Fusarium oxysporum f.sp. lycopersici on tomato was tested. Chlamydospores produced in soil 30 days after inoculation induced a more severe disease than microconidia indicating a higher inoculum potential of chlamydospores.
The method proposed produces easily an inoculum of F. oxysporum f.sp. lycopersici which infects the plants consistently and induces a relatively high disease severity.     

Chrysanthemum Stem Necrosis Caused by a Possible Novel Tospovirus

Necrotic lesions surrounded by yellow areas on leaves followed by stem necrosis were observed on chrysanthemum plants cultivated in Atibaia, Sào Paulo State, Brazil. The host range, in vitro properties and particle morphology of the causal virus were typical of a Tospovirus. Serological studies demonstrated that the virus differed from tomato spotted wilt. tomato chlorotic spot, groundnut ringspot and inpatiens necrotic spot Tospoviruses. The virus isolate is thus possibly a representativeof a new serogroup of a new virus.     

SOME EFFECTS OF NITROGEN SUPPLY ON THE INFECTION OF TOMATO PLANTS WITH TOMATO SPOTTED WILT VIRUS

The susceptibility of tomato plants to systemic infection by tomato spotted wilt virus was increased by increasing nitrogen supply to levels above that optimal for growth. The virus content, estimated by local-lesions counts, was also raised by increasing nitrogen. The period between inoculation and the appearance of systemic symptoms was decreased by increasing nitrogen to a point slightly greater than the optimal level for growth, but increased by additional applications.
Infected plants receiving more nitrate or ammonium compounds than were needed for optimal growth showed abnormal leaf symptoms and no bronzing. N, P and Mg analyses showed that these symptoms were related primarily to nitrogen content. Such leaves contained more virus than bronzed leaves.     

Mitigation of Tomato Spotted Wilt Virus Infection and Symptom Expression by Water Stress

The effects of water stress on the infection and systemic movement of tomato spotted wilt virus (TSWV) in vegetative tomato plants are examined. Two groups of plants: water stressed (water potential ?1.31 MPa) and well-watered ones (water potential ?0.65 MPa), were mechanically inoculated on the third expanded leaf from the top. Each group was then divided into a well-watered and a water stressed one. TSWV inoculation was effective both on water-stressed and well-watered plants. In all groups, infection was first detected in roots, then in shoot apices and subsequently in the rest of the leaves. Systemic virus infection was somewhat slower in the water stressed plants. Post inoculation stress treatments had a significant effect in reducing infection as well as in attenuating TSWV symptoms. Results from the present work suggest that water stress may be effective in controlling virus concentration and systemic infection symptoms in tomato plants and could thus be used as a tool to study the regulation of development of TSWV infection in this system.     

SOME EFFECTS OF HOST NUTRITION ON THE SUSCEPTIBILITY OF PLANTS TO INFECTION BY CERTAIN VIRUSES

Fertilizer treatments that greatly influenced the growth of tobacco and potato plants in pots had little effect on the number that became infected with potato virus Y when the plants were colonized by equal numbers of infective aphids, though the number was slightly decreased by nitrogen and increased by phosphorus.
The number of local lesions produced on leaves of tobacco and Nicotiana glutinosa by tomato aucuba mosaic and tobacco mosaic viruses was increased by additions of both nitrogen and phosphorus, provided that these also increased growth. The predominant effect of both nutrients in increasing susceptibility was indirect by increasing plant size, but over certain critical ranges both elements also increased the numbers of lesions produced per unit leaf area. Conditions of maximum susceptibility approximated closely to those producing optimal growth, and susceptibility, whether measured by lesions per half-leaf or per unit area, was decreased by a deficiency or excess of either element. Sometimes the addition of nitrogen reduced susceptibility when still increasing plant growth.     

Tactics for management of thrips (Thysanoptera: Thripidae) and tomato spotted wilt virus in tomato

Four studies were conducted in Georgia during spring 1999, 2000, 2001, and 2002 to evaluate various management tactics for reducing thrips and thrips-vectored tomato spotted wilt virus (TSWV) in tomato and their interactions relative to fruit yield. Populations of thrips vectors of TSWV, Frankliniella occidentalis (Pergande) and Frankliniella fusca (Hinds), were determined using flower and sticky trap samples. The management practices evaluated were host plant resistance, insecticide treatments, and silver or metallic reflective mulch. Averaged over all tests, the TSWV-resistant tomato 'BHN444' on silver mulch treatment had the largest effect in terms of reducing thrips and spotted wilt and increasing marketable yield. Of the insecticide treatments tested, the imidacloprid soil treatment followed by early applications of a thrips-effective foliar insecticide treatment provided significant increase in yield over other treatments. Tomato yield was negatively correlated with the number of F. fusca and percentage of TSWV incidence. F. occidentalis per blossom was positively correlated with percentage of TSWV incidence, but not with yield. No significant interactions were observed between cultivar reflective mulch main plot treatments and insecticide subplot treatments; thus, treatment seemed to be additive in reducing the economic impact of thrips-vectored TSWV. Control tactics that manage thrips early in the growing season significantly increased tomato yield in years when the incidence of TSWV was high (>17%).     

Disease intensity of some tomato viroses in Serbia

In this paper we present the data on the disease intensity of the tomato plants grown in glass and plastic-houses, and in the open field. The infection was caused by the following viruses: Tomato mosaic virus (ToMV), Tobacco mosaic virus (TMV), Tomato spotted wilt virus (TSWV), Alfalfa mosaic virus (AMV), Potato virus X (PVX), Potato virus Y (PVY), Tomato black ring virus (TBRV), Tomato ringspot virus (ToRSV), Tomato aspermy virus (TAV), and Cucumber mosaic virus (CMV). These viruses represented most frequent tomato pathogens in Serbia. According to the obtained results, it could be concluded that 92.94% of the tested tomato plants grown in glass and plastic-houses, and 89.82% grown in the open field were infected by one of the above viruses. Most of the plant samples were infected by two or more viruses. The most frequent viruses — tomato pathogens in Serbia were ToMV, PVY and TMV.     

A Survey of Tospoviruses in Bulgaria

A survey of tospoviruses present in Bulgaria was conducted during three vegetation periods. A total of 258 different virus isolates were obtained from tobacco, tomato, weeds and ornamentals in the open fields and from glasshouses. Virus isolates were identified by inoculation to test plants and serology as Tomato spotted wilt virus (TSWV). No indications were obtained that other Tospovirus species were present in Bulgaria. In addition the transmission ability and efficiency of several Bulgarian populations of Thrips tabaci (Lind.) and Frankliniella occidentalis (Perg.) was determined. The experimental results indicated that Bulgarian TSWV isolates are transmitted with high efficiency by F. occidentalis and that only arrhenotokous and not thelytokouse populations of T. tabaci could transmit TSWV.     

Distinct feeding behavior between sexes of shape Frankliniella occidentalis results in higher scar production and lower tospovirus transmission by females

Feeding behavior and scar production of male and female F. occidentalis (Pergande) (Thysanoptera: Thripidae) were studied in relation to transmission of tomato spotted wilt tospovirus (TSWV). Electrical penetration graph (EPG) analysis showed that females feed more frequently and intensively than males. The feeding intensity, reflected by silvery scar production and studied by an image analysis system, demonstrated that females induced more numerous scars than males. At the same time, males transmitted TSWV with a higher efficiency than females, indicating that TSWV transmission and scar production are not positively correlated. Furthermore, males produced significantly more local lesions of TSWV than females. These quantitative differences in scar production and transmission of TSWV can be explained by the lower mobility and higher consumption rate of females. The influence of the sex-ratio on crop damage and virus transmission, and thus to the spread of TSWV, is emphasized.     

Photosynthesis and predisposition of plants to infection with certain viruses

The effects on susceptibility to infection with certain viruses of subjecting plants to various periods of darkness or reduced illumination before and after inoculation were tested. The viruses and hosts used were a tobacco necrosis virus in French bean and tobacco; tomato aucuba mosaic virus in tobacco; and tobacco mosaic and tomato bushy stunt viruses in Nicotiana glutinosa . All the virus-host combinations give necrotic local lesions, and susceptibility was measured by local lesion counts. Susceptibility was consistently increased by pre-inoculation treatments of host plants, whereas post-inoculation treatments had relatively little effect, but most often decreased susceptibility.
Short periods in the dark produced similar responses to longer periods in shade, but the different plants varied in their response to, and tolerance of, darkness. The maximum number of lesions was usually obtained with bean plants kept for only 24 hr. in the dark before inoculation, but with tobacco plants susceptibility increased with increasing time in the dark up to 5 days.
It is suggested that the successful establishment of infection occurs in two stages, the first of which is affected by. the accumulation of photosynthetic products. Whether these products confer resistance by increasing cell turgor or by reacting specifically with virus particles is unknown, but sap from plants in the light possesses no greater virus-inhibiting power than sap from plants kept in the dark.     

A multi-generation analysis of the stability of transgenic virus resistance in doubled-haploid tobacco lines

Plants can be genetically engineered for virus resistance by transformation with a viral gene. We transformed tobacco with the tomato spotted wilt virus (TSWV) nucleocapsid gene from the Hawaiian L isolate in order to obtain TSWV resistant breeding lines. Doubled-haploid lines were produced from primary transgenic plants that were selected for resistance to the virus. Several of these lines showed very high levels of resistance and were symptomless after inoculation with the Hawaiian L isolate of TSWV. The accumulation of only low levels of full-length transgene RNA and protein observed in these lines is consistent with an RNA-mediated mechanism of resistance. The lines that were highly resistant to the Hawaiian L isolate of TSWV were also found to be highly resistant to several other isolates of TSWV, while lines that were only moderately resistant to the Hawaiian L isolate were often susceptible to the other isolates. The highly resistant lines were advanced over several generations by self-pollination. Although these lines were fully homozygous, several lines lost resistance in later generations, indicating that the resistance was unstable. Selection for resistance in these unstable lines did not prevent the occurrence of susceptible progeny in subsequent generations. Therefore, testing over several generations is required to determine the stability of resistance when breeding crops with transgenic virus resistance.     

Infection ofArabidopsis thaliana ecotype columbia by tomato spotted wilt virus

Mechanical inoculation ofArabidopsis thaliana ecotype Columbia with tomato spotted wilt virus led to viral replication and spread as determined by dot blot and ELISA analysis. Severe symptoms were observed three to four weeks post-inoculation. Early symptoms were manifested as chlorotic spots on uninoculated leaves. Later in the infection process, some plants showed complete chlorosis and wilting prior to bolting. Bolts that were developed by infected plants were chlorotic and deformed. These preliminary results suggest thatA. thaliana could become a model system for the genetic analysis of host factors required for the replication of viruses in the family Bunyaviridae, which includes viruses that cause important diseases of both plants and animals.     

The use of abrasives in the transmission of plant viruses

The effect of different abrasives on the transmissibility of several plant viruses was tested. Celite and animal charcoal were as effective as carborundum in increasing the number of lesions produced by a given inoculum: 400-mesh carborundum was the most effective among the different sizes which were tested; this gave a result equivalent to increasing the virus content a hundred times. Some preparations of carborundum and charcoal reduced infectivity.
Uninjured plants resisted infectivity when virus solutions were sprayed over them. Leaves previously nibbed without abrasives developed only few lesions whereas leaves rubbed with abrasives developed large numbers. Three hours after rubbing with abrasives leaves had regained their resistance to sprayed virus solutions.
The effects of rubbing leaves with abrasives are described and their significance discussed.     

Long-term resistance to tomato spotted wilt virus in transgenic tobacco cultivars expressing the viral nucleoprotein gene: greenhouse and field tests

We examined the resistance phenotype of a large number of transgenic tobacco plants originating from 12 commercial (Nicotiana tabacum) cultivars expressing the sense form of the nucleoprotein (N) gene of L3, a Bulgarian isolate of tomato spotted wilt virus (TSWV). The analysis revealed that transgenic plants are completely protected against the homologous L3 isolate of TSWV irrespective of whether or not they contain detectable levels of translational product. The effectiveness of protection against the virus was investigated upon mechanical inoculation under greenhouse conditions and in field trials. Non-segregating resistant lines were selected and the inheritance of the resistance to TSWV was analysed in successive generations (R₃–R₆). Extensive tests under controlled conditions and two-year field trials proved that the resistance to TSWV is stable in different environments and is a stably inherited trait.     

THE EFFECTS OF VARYING THE WATER SUPPLY OF PLANTS ON THEIR SUSCEPTIBILITY TO INFECTION WITH VIRUSES

Increasing the amount of water supplied to plants before they were inoculated with viruses greatly increased their susceptibility to infection; plants that received unlimited water produced 10 or more times as many local lesions as plants that received only enough to prevent wilting. Susceptibility was increased throughout the year, but the full response occurred in 2 weeks in winter and 4 weeks in summer. Plants that received unlimited water for the 2 weeks immediately preceding inoculation were no more susceptible than those that received it during the previous 2 weeks, although the external appearance of the plants differed at the time of inoculation. Varying water supply after inoculation did not affect the numbers of lesions.
The differences in susceptibility to infection produced by differential watering were decreased, but not abolished, by growing plants under shade or by incorporating a diatomaceous earth in the inoculum.
Increasing water produced plants with larger and more succulent leaves; the cuticular layer was thinner, and the palisade tissue was less regularly arranged than in the plants kept dry. The increased susceptibility caused by an abundant water supply may be at least partly due to these structural differences, which allow the leaf to be damaged more easily when rubbed with inocula and so present more entry points for virus particles.     

Cropping system influences Tomato spotted wilt virus disease development,thrips population and yield of tomato (Lycopersicon esculentum)

Yield of tomato is limited by many diseases including Tomato spotted wilt virus disease. This study was conducted in the field at Kenya Agriculture Research Institute Njoro, Kenya, in 2004 and 2006 to determine the effect of intercropping on disease development, thrips population and yield of tomato variety Cal J grown under four intercrop systems involving kale, onion, maize and sole tomato. The experimental design was a Randomised Complete Block Design (RCBD) replicated three times. Disease scores on tomato–maize differed significantly from tomato–kale and tomato–onion in both years of the study. Maize cropping system had a low significant thrips population from the other cropping systems. Tomato–maize intercrop produced the lowest fruit weights and marketable yield in 2004 and 2006, while yield of onion, kale and maize in intercrops were not significantly different from their monocrops. Land equivalent ratio was >1 in all the cropping systems.