Transmission of tomato spotted wilt tospovirus by Thripstabaci populations originating from leek

The transmission of tomato spotted wilt tospovirus (TSWV) by Thrips tabaci collected from leek was studied using the petunia local-lesion leaf-disc assay. After an acquisition-access period of 72 h given to newborn larvae up to 8 h old, the efficiency of transmission by adults was determined in three inoculation-access periods of 48 h. This efficiency varied for six T. tabaci populations from 0.7 to 11.6% in experiments using the Greek TSWV isolate GR-04. Males were more efficient transmitters than females (19 out of 176 versus five out of 494). Frankliniella occidentalis transmitted the same virus with a higher efficiency (34.8%). The transmission rate differed also among TSWV isolates, as shown in tests with four T. tabaci using two isolates. The virus was more efficiently acquired from infected leaf material of Datura stramonium than from that of Emilia sonchifolia . Plants of the latter species were more susceptible than Nicotiana tabacum in thrips transmission tests.     

The Efficiency by Which Thrips tabaci Populations Transmit Tomato spotted wilt virus Depends on Their Host Preference and Reproductive Strategy

ABSTRACT Arrhenotokous and thelytokous populations of Thrips tabaci from tobacco or leek plants were evaluated for their ability to transmit Tomato spotted wilt virus (TSWV) and for their host preference. Transmission efficiencies were comparatively studied using leaf disks of Petunia hybrida, Datura stramonium, and Nicotiana tabacum cv. Basmas. Adults of arrhenotokous populations collected on infected tobacco plants in the field were efficient transmitters (up to 48.5% transmission) and remained so when maintained on tobacco for several generations. Arrhenotokous T. tabacipopulations from leek plants were poor transmitters (up to 3.1% transmission), whereas no transmission was obtained with thelytokous populations from leek. All populations could infest leek, however none of the arrhenotokous and thelytokous populations from leek plants was able to infest tobacco. TSWV could be acquired by both first and second larval instars of a T. tabacipopulation from tobacco. However, the transmission by adults decreased with the age at which the virus was acquired by larvae. The highest efficiencies (61% of males and 51% of females transmitted) were obtained when newborn (0- to 24-h old) larvae acquired the virus. The majority of thrips started to transmit after becoming adult and rates were positively correlated with the temperature at which the thrips were kept. The median latent period values found for adults decreased with increasing temperature. The median acquisition access period (AAP50) of the population was 41 min, whereas the AAP(50) was 65 min for males and 35 min for females. The median inoculation access period of males was 246 and 365 min on tobacco and petunia, respectively, and 96 and 345 min for females. The results show that T. tabaci forms a complex in terms of host preference, reproductive strategy, and ability to transmit TSWV. The transmission parameters show that the thrips of arrhenotokous populations infesting tobacco are highly efficient vectors.     

Acquisition of Tomato spotted wilt virus by Adults of Two Thrips Species

ABSTRACT Only larval thrips that acquire Tomato spotted wilt virus (TSWV), or adults derived from such larvae, transmit the virus. Nonviruliferous adults can ingest virus particles while feeding on TSWV-infected plants, but such adult thrips have not been shown to transmit TSWV. Immunofluorescence microscopy was used to show that thrips 1, 5, 10, and 20 days after adult emergence (DAE) fed on TSWV-infected plants acquired TSWV with virus replication and accumulation occurring in both epithelial and muscle cells of Frankliniella fusca (tobacco thrips [TT]) and F. occidentalis (western flower thrips [WFT]), as indicated by immunodetection of the nonstructural (NSs) protein encoded by the small RNA and the nucleocapsid (N) protein, respectively. Adult WFT acquired TSWV more efficiently than TT. There was no significant effect of insect age on TSWV acquisition by TT. In contrast, acquisition by adult WFT at 1 and 5 DAE was higher than acquisition at 10 and 20 DAE. Subsequent transmission competence of adult cohorts was studied by vector transmission assays. All adult thrips tested that had an acquisition access period as an adult were unable to transmit the virus. These results indicate the susceptibility of adult TT and WFT to infection of midgut cells by TSWV and subsequent virus replication and confirm earlier studies that adult thrips that feed on virus-infected plants do not transmit the virus. The role of a tissue barrier in TSWV movement and infection from midgut muscle cells to the salivary glands is discussed.     

The proportion of viruliferous individuals in field populations of Frankliniella occidentalis: Implications for tomato spotted wilt virus epidemics in tomato

Tomato spotted wilt tospovirus (TSWV) infected plants and western flower thrips (Frankliniella occidentalis Perg., WFT) adult population densities were monitored during 1993 and 1994 in field tomatoes in Northeastern Spain. The proportion of viruliferous WFT adults in field populations was quantified. A significant association has been found between early population densities of WFT adult thrips and final TSWV incidence for early transplanted tomato crops. In contrast, for late transplanted tomato crops, whereas similar high final disease incidences of TSWV could be attained, very low WFT adult population densities were always detected. The significantly higher infectious potential of WFT populations collected during the early growth stages of late transplanted tomatoes could be relevant for the TSWV incidences attained in spite of the low thrips numbers detected.     

Impeded Thrips Transmission of Defective Tomato spotted wilt virus Isolates

Two defective RNA-containing isolates (Pe-1 and 16-2) and an envelope-deficient (env ) isolate of Tomato spotted wilt virus (TSWV) were tested for their transmissibility by Frankliniella occidentalis. The Pe-1 isolate contained a truncated L RNA segment that barely interfered with symptom expression and replication of the wild-type (wt) L RNA segment. This isolate was transmitted with an efficiency of 51%, a value comparable to that found for wt TSWV (54%). Isolate 16-2, which contained a genuine defective interfering L RNA as concluded from its ability to suppress wt L RNA synthesis and attenuation of symptom expression, was not transmitted at all. The midguts of all larvae that ingested Pe-1 became infected, whereas limited midgut infections were found in 24% of the larvae that ingested 16-2. This difference in infection could be explained by the presence of a low number of infectious units in the inoculum ingested from plants as demonstrated in infection experiments and verified by northern blot analysis. The env isolate failed to infect the midgut after ingestion and could not be transmitted by any thrips stage. This isolate also cannot infect primary thrips cell cultures. Taken together, these results suggest that the envelope of TSWV contains the determinants required for binding and subsequent infection of thrips cells.     

The competence of four thrips species to transmit and replicate four tospoviruses

The tospoviruses Tomato spotted wilt virus (TSWV), Tomato chlorotic spot virus (TCSV), Groundnut ringspot virus (GRSV) and Chrysanthemum stem necrosis virus (CSNV) are well-known pathogens on tomato in Brazil. The thrips species Frankliniella occidentalis , F. schultzei , Thrips tabaci and T. palmi were studied for their competence to transmit these tospoviruses. Frankliniella occidentalis transmitted all four tospoviruses with different efficiencies. Frankliniella schultzei transmitted TCSV, GRSV and CSNV. Although F. schultzei has been reported as a vector of TSWV, the F. schultzei population in the present study did not transmit the TSWV isolate used. A population of T. tabaci known to transmit Iris yellow spot virus (onion isolate) did not transmit any of the studied tospoviruses, and nor did T. palmi . Replication of these tospoviruses could be demonstrated by ELISA, not only in the thrips species that could transmit them, but also in those that could not. The results strongly suggest that competence to transmit is regulated not only by the initial amount of virus acquired and replication, but also by possible barriers to virus circulation inside the thrip's body.     

Dynamics of Tomato spotted wilt virus Replication in the Alimentary Canal of Two Thrips Species

ABSTRACT Transmission of Tomato spotted wilt virus (TSWV) is dependent on virus uptake in the midgut prior to virus movement to the salivary glands. Replication of TSWV in the alimentary canal of tobacco thrips (TT, Frankliniella fusca) and western flower thrips (WFT, F. occidentalis) was investigated by immunolocalization of the nonstructural protein (NSs) encoded by the small RNA of TSWV and fluorescence microscopy. Analysis of cohorts during development from larva to adults following virus acquisition by first instar larva indicated that virus replication followed a specific time-course pattern in the foregut, regions of the midgut, salivary glands, and ligaments between the midgut and salivary glands. Initial virus replication occurred only in epithelial cells of midgut-1 but, upon infection of muscle cells, the virus moved to the midgut-2, foregut, midgut-3, and salivary glands. The ligaments between the midgut and salivary glands appeared to be a route for virus to invade the salivary glands. No virus replication was observed in the hindgut, Malpighian tubules, or tubular salivary glands. The dynamics of TSWV replication, as measured by NSs accumulation, were similar in both TT and WFT.     

Tomato spotted wilt virus Infection Improves Host Suitability for Its Vector Frankliniella occidentalis

ABSTRACT The effect of Tomato spotted wilt virus (TSWV) infection on plant attractiveness for the western flower thrips (Frankliniella occidentalis) was studied. Significantly more thrips were recovered on infected than were recovered on noninfected pepper (Capsicum annuum) plants in different preference tests. In addition, more offspring were produced on the virus-infected pepper plants, and this effect also was found for TSWV-infected Datura stramonium. Thrips behavior was minimally influenced by TSWV-infection of host plants with only a slight preference for feeding on infected plants. Offspring development was positively affected since larvae hatched earlier from eggs and subsequently pupated faster on TSWV-infected plants. These results show a mutualistic relationship between F. occidentalis and TSWV.     

Interaction of Tomato Spotted Wilt Tospovirus (TSWV) Glycoproteins with a Thrips Midgut Protein, a Potential Cellular Receptor for TSWV

ABSTRACT Interactions between viral and cellular membrane fusion proteins mediate virus penetration of cells for many arthropod-borne viruses. Electron microscope observations and circumstantial evidence indicate insect acquisition of tomato spotted wilt virus (TSWV) (genus Tospovirus, family Bunyaviridae) is receptor mediated, and TSWV membrane glycoproteins (GP1 and GP2) serve as virus attachment proteins. The tospoviruses are plant-infecting members of the family Bunyaviridae and are transmitted by several thrips species, including Frankliniella occidentalis. Gel overlay assays and immunolabeling were used to investigate the putative role of TSWV GPs as viral attachment proteins and deter mine whether a corresponding cellular receptor may be present in F. occidentalis. A single band in the 50-kDa region was detected with murine monoclonal antibodies (MAbs) to the TSWV-GPs when isolated TSWV or TSWV-GPs were used to overlay separated thrips proteins. This band was not detected when blots were probed with antibody to the non-structural protein encoded by the small RNA of TSWV or the TSWV nucleocapsid protein, nor were proteins from nonvector insects labeled. Anti-idiotype antibodies prepared to murine MAbs against GP1 or GP2 specifically labeled a single band at 50 kDa in Western blots and the plasmalemma of larval thrips midguts. These results support the putative role of the TSWV GPs as viral attachment proteins and identified potential cellular receptor(s) in thrips.     

Replication of Tomato spotted wilt virus After Ingestion by Adult Thrips setosus is Restricted to Midgut Epithelial Cells

ABSTRACT If acquisition access feeding (AAF) is first given after adult eclosion, none of the nine thrips species able to serve as tospovirus vectors can become infective. The previous cellular investigations of this phenomenon, carried out only in Frankliniella occidentalis, suggested that infectivity was prevented because the type member of the tospoviruses, Tomato spotted wilt virus (TSWV), was unable to enter the midgut of adult thrips. The present study extends a cellular view of tospovirus-thrips interactions to a species other than the western flower thrips, F. occidentalis. Our findings show that TSWV enters and replicates within the midgut of adult Thrips setosus, but does not infect cells beyond the midgut epithelia. After AAF as adult, TSWV replicated in T. setosus midgut cells as indicated by significant increases in nucleocapsid (N) protein detected by double-antibody sandwich enzyme-linked immunosorbent assay, and the presence of inclusions containing the S RNA-encoded nonstructural and N proteins revealed by microscopic observations. Electron microscopic observations of adult insects showed that no infection occurred in cells beyond the midgut epithelia, and insects subsampled from the same cohorts could not transmit TSWV. In contrast, electron microscopy observations of larval T. setosus revealed that TSWV infected the midgut and muscle cells, and adult insects developing from these cohorts had infected salivary glands and were able to transmit TSWV. Mature virions were observed only in the salivary glands of adults developing from infected larvae. Our findings suggest that the barrier to infectivity in T. setosus adults differs from that shown for F. occidentalis adults.     

Role of weed hosts and the western flower thrips, Frankliniella occidentalis, in epidemiology of Tomato spotted wilt virus in the Çukurova region of Turkey

Surveys of thrips and Tomato spotted wilt virus (TSWV) on weeds were conducted in the eastern Mediterranean region of Turkey during the years 2004–2006. Thrips species were collected by vigorously shaking weedy plants into a white container for 15 sec. Plant material collected during field surveys and plants which were used for mechanical inoculation of TSWV, were tested by DAS–ELISA. The weed species Ranunculus muricatus, Melilotus officinalis, Sinapis arvensis and Portulaca oleracea were used for the virus transmission experiments in an enclosed high plastic tunnel and in cage experiments. Western flower thrips, Frankliniella occidentlis (Pergande) (Thysanoptera: Thripidae), was the most common thrips, inhabiting 80 of the 82 weed species sampled. Adults of F. occidentalis and thrips larvae were significantly more abundant on S. arvensis than on the other weed species sampled (P<0.05). Adult and larval thrips showed peak densities on most weeds in April or May. Summer annual weeds were not good hosts for reproduction of the thrips. A total of 90 samples from 17 plant species belonging to 11 plant families were ELISA positive for TSWV. No TSWV was detected on 65 weed species belonging to 26 plant families. High numbers of plant samples infected by TSWV were obtained in P. oleracea (21 samples) and in R. muricatus (15 samples). In field surveys symptoms of TSWV were detected on only R. muricatus. Incidence of the TSWV on weeds ranged between 5% and 25%. Transmission rates of TSWV by F. occidentalis to the weeds ranged from 33% to 83% in the pepper plastic tunnel and cage experiments.     

Thrips Resistance in Pepper and Its Consequences for the Acquisition and Inoculation of Tomato spotted wilt virus by the Western Flower Thrips

ABSTRACT Different levels of thrips resistance were found in seven Capsicum accessions. Based on the level of feeding damage, host preference, and host suitability for reproduction, a thrips susceptible and a resistant accession were selected to study their performance as Tomato spotted wilt virus (TSWV) sources and targets during thrips-mediated virus transmission. Vector resistance did not affect the virus acquisition efficiency in a broad range of acquisition access periods. Inoculation efficiency was also not affected in short inoculation periods, but was significantly lower on plants of the thrips resistant accession during longer inoculation access periods. Under the experimental conditions used, the results obtained show that transmission of TSWV is little affected by vector resistance. However, due to a lower reproduction rate on resistant plants and a lower preference of thrips for these plants, beneficial effects of vector resistance might be expected under field conditions.     

Incidence of weed reservoirs and vectors of tomato spotted wilt tospovirus on southern Tasmanian lettuce farms

Thrips species and tomato spotted wilt virus (TSWV) alternate weed hosts were surveyed on two lettuce farms in southern Tasmania during 1994 and 1995. Only one known vector species, Thrips tabaci, was found at either site, comprising on average 36.8% of the total monthly catch. A major peak of thrips activity in the summer corresponded with an increase of disease in autumn harvested lettuce. Two thrips species new for Tasmania were recorded, Pseudanaphothrips achaetus and Tenothrips frici . Infection patterns within the crop indicated that localized weed infestations were the most likely reservoir of virus. ELISA testing showed that TSWV was present in a range of dicotyledonous weed species, although usually infecting only a low percentage of the plants. Arctotheca calendula appeared to be the single most important reservoir host species at one property, whilst this species and Sonchus oleraceus, Malva sylvestris, Brassica rapa ssp. silvestris, Erodium moschatum and Trifolium sp. were probably the most important reservoirs at the other property. Two new natural TSWV host species were recorded, Erodium moschatum and Brassica rapa ssp. silvestris . The property with the highest incidence of TSWV-infected lettuce had a relatively higher proportion of virus-infected weeds but less thrips activity during the infection period.     

Seasonal abundance of the western flower thripsFrankliniella occidentalis in the arava valley of Israel

Flowers of 22 plant species grown in the Arava region of Israel were sampled at periodic intervals to study the seasonal abundance ofFrankliniella occidentalis (Pergande) (Thysanoptera: Thripidae). Samples were collected from the most common cut-flower and vegetable plant species in this area during 1997 and from pepper in 1996–1998. Western flower thrips (WFT) was detected on 19 of the plant species surveyed, and it reproduced on numerous plant species. WFT counts peaked in late February–April and in October–December. The peak density of WFT during March in our survey corresponded to the peak flowering of cut-flower plants (during spring) and sweet pepper (during autumn). Thrips population was less abundant during autumn than in spring and was very low during summer. The possible reasons for fluctuations in thrips densities are abiotic factors (temperature and relative humidity), and availability of flowering plants. The high temperature (>40°C) and low humidity (∼20%) in summer caused a drastic reduction in WFT populations, although some flowering plant species were available in this season. WFT population remained low until October, when adults appeared in large numbers on flowering pepper. We assume that the thrips had survived the hottest months by aestivation and migrated to the plants when air temperature decreased. In most samples from the cut-flower fields, 84.8% were adult WFT; only during the period from the end of January to March, did the proportion of immature stages in the thrips population increase to 31.6%. Females always outnumbered males (71% females). In pepper fields during the first cropping month the thrips population included 96% adults; from November onwards, the proportions of adults and immatures were almost equal; and females outnumbered males. Organic pepper fields were not infested with thrips because of the presence of the predatory bugOrius albidipennis (Fieber). Contribution from the Agricultural Research Organization. No. 534/00, 2000 series. http://www.phytoparasitica.org posting July 16, 2002.     

Assessing the susceptibility of chrysanthemum cultivars to tomato spotted wilt virus

Three methods were compared to assess the susceptibility of vegetatively propagated chrysanthemum to tomato spotted wilt tospovirus (TSWV): mechanical and thrips-mediated inoculation of whole plants, and a leaf-disc assay. As symptom expression was often poor or even absent, TSWV infections and subsequent susceptibility to TSWV were determined by ELISA. All 15 chrysanthemum cultivars tested were susceptible to TSWV, irrespective of their degree of vector resistance (based on feeding-scar damage rates). Thrips-mediated inoculation using different numbers of thrips revealed that 100% infection was obtained when plants were challenged by six thrips per plant, whereas 80 and over 50%, respectively, of the plants became infected when inoculated by a single male or female thrips. However, false negatives were scored even after intensive sampling because of erratic, cultivar-specific and time-dependent virus distribution after inoculation in the plants. Labour-intensive samplings and long incubation periods could be overcome by a readily applicable leaf-disc assay. This assay was as reliable as thrips-mediated inoculation of whole plants, and its use is therefore favoured to assess chrysanthemum cultivars for TSWV susceptibility.     

Characterization of a vector-non-transmissible isolate of Tomato spotted wilt virus

The characteristics of a thrips‐non‐transmissible isolate of Tomato spotted wilt virus (TSWV), designated TSWV‐M, were compared with those of a thrips‐transmissible isolate, designated TSWV‐T. TSWV‐M showed a narrower host range than TSWV‐T. Adult thrips failed to transmit TSWV‐M, although the vector acquired the virus during the larval stages. TSWV‐M was detected by RT‐PCR in adult thrips bodies, but not in thrips heads, suggesting that loss of thrips transmissibility was the result of the absence of virus in adult thrips salivary glands. Whereas N (nucleoprotein), NSs (non‐structural protein) and G_C (the C‐terminal portion of the glycoprotein precursor protein) were present in similar amounts in leaf tissue from TSWV‐M‐ or TSWV‐T‐infected plants, G_N (the N‐terminal portion of the glycoprotein precursor protein) was present at much lower amounts in TSWV‐M‐ than in TSWV‐T‐infected plants. SDS‐PAGE and immunoblotting analysis of TSWV‐M and TSWV‐T virion preparations with G_N‐ and G_C‐specific antibodies revealed similar amounts of the G_N and G_C glycoproteins in TSWV‐T virions, but lower amounts of G_N than G_C in TSWV‐M virions. This resulted in a statistically significant reduction in the G_N/G_C ratio in TSWV‐M virions. In affinoblots, the G_C and G_N glycoproteins of TSWV‐M exhibited weak binding with lectins showing affinity for N‐linked oligosaccharide structures. Sequence analysis of M RNA (medium segment of the TSMV genome) revealed no deletions or frameshift mutations in the G_N/G_C precursor of TSWV‐M. However, five amino acid changes were detected in the G_N/G_C precursor. A single, relatively conservative amino acid substitution (V→I) was observed in the NSm protein. Sequence analysis of S RNA (small portion of the TSMV genome) revealed a large intergenic region with no changes in the N protein and with three amino acid changes in the NSs protein.     

Overwintering of Frankliniella fusca (Thysanoptera: Thripidae) on Winter Annual Weeds Infected with Tomato spotted wilt virus and Patterns of Virus Movement Between Susceptible Weed Hosts

ABSTRACT Overwintering of tobacco thrips, Frankliniella fusca, was investigated on common winter annual host plants infected with Tomato spotted wilt virus (TSWV). Populations of tobacco thrips produced on TSWV-infected plants did not differ from those produced on healthy plants, whereas populations varied greatly among host plant species. The mean per plant populations of F. fusca averaged 401, 162, and 10 thrips per plant on Stellaria media, Scleranthus annuus, and Sonchus asper, respectively, during peak abundance in May. Adult F. fusca collected from plant hosts were predominately brachypterous throughout the winter and early spring, but macropterous forms predominated in late spring. Weed hosts varied in their ability to serve as overwintering sources of TSWV inoculum. Following the initial infection by TSWV in October 1997, 75% of Scleranthus annuus and Stellaria media retained infection over the winter and spring season, whereas only 17% of Sonchus asper plants remained infected throughout the same interval. Mortality of TSWV-infected Sonchus asper plants exceeded 25%, but mortality of infected Stellaria media and Scleranthus annuus did not exceed 8%. TSWV transmission by thrips produced on infected plants was greatest on Stellaria media (18%), intermediate on Scleranthus annuus (6%), and lowest on Sonchus asper (2%). Very few viruliferous F. fusca were recovered from soil samples collected below infected wild host plants. Vegetative growth stages of Stellaria media, Sonchus asper, and Ranunculus sardous were more susceptible to F. fusca transmission of TSWV than flowering growth stages, whereas both growth stages of Scleranthus annuus were equally susceptible. In a field study to monitor the spatial and temporal patterns of virus movement from a central source of TSWV-infected Stellaria media to adjacent plots of R. sardous, the incidence of infection in R. sardous plots increased from <1% in March to >42% in June 1999. Infection levels in the Stellaria media inoculum source remained high throughout the experiment, averaging nearly 80% until June 1999 when all Stellaria media plants had senesced. Dispersal of TSWV from the inoculum source extended to the limits of the experimental plot (>37 m). Significant directional patterns of TSWV spread to the R. sardous plots were detected in April and May but not in June. R. sardous infections were detected as early as March and April, suggesting that overwintering inoculum levels in an area can increase rapidly during the spring in susceptible weed hosts prior to planting of susceptible crops. This increase in the abundance of TSWV inoculum sources occurs at a time when vector populations are increasing rapidly. The spread of TSWV among weeds in the spring serves to bridge the period when overwintered inoculum sources decline and susceptible crops are planted.     

The host range of Polymyxa betae in Britain

The host range of Polymyxa betae on common arable weed species in Britain was determined by growing plants in naturally infested soil and examining their root systems for the presence of resting spores (cystosori). Of the 24 species tested, only Atriplex patula and Chenopodium album of the Chenopodiaceae, and Silene alba of the Caryophyllaceae, were found to be heavily infected. S. alba is a newly recorded host species for Polymyxa. The host specificity of isolates of P. betae from Beta vulgaris, C. album and A. patula was investigated by observing which of 11 test plants could be infected by the isolates obtained from this soil. Three main biotypes of P. betae appeared to be distinguishable: one which was able to infect all chenopodiaceous species; one which had a narrower host range; and one which was able to infect S. alba. The role of weed species in the epidemiology of rhizomania is discussed.     

RAPID SEROLOGICAL DETECTION OF TOMATO SPOTTED WILT VIRUS IN INDIVIDUAL THRIPS BY SQUASH-BLOT ASSAY FOR USE IN EPIDEMIOLOGICAL STUDIES

Squash blotting on nitrocellulose membranes was used to detect tomato spotted wilt virus (TSWV) in individual Frankliniella occidentalis adult thrips using a specific polyclonal antiserum. This method offers a simple and reliable procedure to test a large number of thrips and was less time-consuming than alternative techniques available. A 95% agreement was found between the results of squash-blot and plant transmission assay. Based on this agreement, this technique was used to study the relative number of viruliferous thrips in F. occidentalis field populations collected from a farm in Catalonia where TSWV causes important economic losses. The percentages of viruliferous individuals detected from a total of 1509 collected thrips increased from 0% to 2% in the different surveys conducted in the 1993 growing season. Squash-blot proved to be a rapid and inexpensive technique to screen TSWV viruliferous thrips from field populations. The results obtained by this method could be used together with other epidemiological data to forecast TSWV-induced diseases.     

Effects of Temperature and Host on the Generation of Tomato Spotted Wilt Virus Defective Interfering RNAs

ABSTRACT The generation of defective interfering (DI) RNA molecules of tomato spotted wilt tospovirus (TSWV) was studied by serially passaging in-ocula from plant to plant under different controlled conditions. DI RNAs were generated at higher rates in plants at 16 degrees C than in plants incubated at higher temperatures. Another factor promoting the TSWV DI RNA generation was the use of high virus concentrations in the inocula. The solanaceous species Capsicum annuum, Datura stramonium, Lycopersicon esculentum, Nicotiana benthamiana, and N. rustica supported the generation of DI RNAs, whereas the virus recovered from the inoculated composite species, Emilia sonchifolia, remained free of any DI RNA under all conditions tested. This study resulted in a strategy to maintain DI RNA-free TSWV isolates, as well as in an efficient way to produce a large population of different DI RNA species. A single DI RNA species usually became dominant in an isolate after a few rounds of serial inoculations. The possible mechanisms involved in TSWV DI RNA generation under different inoculation circumstances are discussed.