Tomato Yellow Leaf Curl Geminivirus Research Articles

Expression of an Entire Bacterial Operon in Plants

Multigene expression is required for metabolic engineering, i.e. coregulated expression of all genes in a metabolic pathway for the production of a desired secondary metabolite. To that end, several transgenic approaches have been attempted with limited success. Better success has been achieved by transforming plastids with operons. IL-60 is a platform of constructs driven from the geminivirus Tomato yellow leaf curl virus. We demonstrate that IL-60 enables nontransgenic expression of an entire bacterial operon in tomato (Solanum lycopersicum) plants without the need for plastid (or any other) transformation. Delivery to the plant is simple, and the rate of expressing plants is close to 100%, eliminating the need for selectable markers. Using this platform, we show the expression of an entire metabolic pathway in plants and delivery of the end product secondary metabolite (pyrrolnitrin). Expression of this unique secondary metabolite resulted in the appearance of a unique plant phenotype disease resistance. Pyrrolnitrin production was already evident 2 d after application of the operon to plants and persisted throughout the plant's life span. Expression of entire metabolic pathways in plants is potentially beneficial for plant improvement, disease resistance, and biotechnological advances, such as commercial production of desired metabolites.

Accurate and sensitive diagnosis of geminiviruses through enrichment, high-throughput sequencing and automated sequence identification

Existing diagnostic techniques used to identify plant-infecting DNA viruses and their associated molecules are often limited in their specificity and can be challenged by samples containing multiple viruses. We adapted a simple method of amplifying circular viral DNA and, in combination with high-throughput sequencing and bioinformatic analysis, used it as a virus diagnostic method. We validated this diagnostic method with a plant sample infected with a tomato yellow leaf curl geminivirus infectious clone and also compared PCR- and high-throughput-sequencing diagnostics on a geminivirus-infected field sample, showing that both methods gave similar results. Finally, we analyzed infected field samples of pepper from Mexico and tomato from India using this approach, demonstrating that it is both sensitive and capable of simultaneously identifying multiple discrete DNA viruses and subviral DNA elements in densely infected samples.

우리나라 토마토에 발생한 토마토황화잎말림바이러스(Tomato yellow leaf curl geminivirus)의 초간편 Virion Capture(VC)/PCR 진단법

국내에서 최근 발견된 Tomato yellow leaf curl virus(TYLCV)가 토마토에 발생하여 큰 피해가 발생하였다. TYLCV의 피해 발생 예방을 위하여 농업현장의 연구지도연구기관에서 쉽고 간편하게 사용할 수 있는 조기 정밀 유전자 진단기술 개발은 매우 중요하다. TYLCV에 대해 특이적 프라이머를 제작하고 특별한 핵산분리 기술이나 도구가 필요 없는 빠르고 정확하며 경제적인 VC/PCR 진단법을 이용한 유전자 진단법을 개발하였다. TYLCV 진단용 프라이머를 22개 제작하여 감염주 및 건전 토마토의 전체 핵산을 이용해 특이성 검정으로 PCR용으로 9점을 선발하였고, VC/PCR 진단법 용으로 9종을 선발하였다. 이러한 두 가지 진단법에 모두 특이적인 프라이머를 6종을 선발한 후 TLCV로 알려진 다른 Geminivirus와의 특이성 검정결과로 총 4종이 최종 선발하였다. 이들 중 Deng(540,541) 프라이머는 Ceminivirus를 진단할 수 있는 degenerate 프라이머로 VC/PCR진단법이 개발 되었다. Tomato yellow leaf curl virus (TYLCV), a newly reported Geminivirus from tomato, generated recently large economic losses in Korea. Development of a fast and precise genetic diagnosis technique for detecting TYLCV which Agricultural research and extension services can utilize easy and handy is very important to prevent yield losses. Virion Capture (VC)/PCR is a simple, accurate and economical genetic detection method without any works or commercial kits for the extraction of the nucleic acid from the infected plants. Primers of twenty two for detection of TYLCV were designed and tested with extracted total DNA or crude sap from tomato leaf infected with TYLCV and healthy plant. Nine primers for total DNA using conventional PCR and another 9 primers for VC/PCR were selected eventually. Primers of six having same specificity were selected from the two methods and tested with other Geminivirus, Tobacco leaf curl virus (TLCV) by VC/PCR. Finally specific primers of four were selected for detection of TYLCV using VC/PCR, and Deng (540, 541), a degenerate primer for Geminivirus reported in 1996, was also developed for VC/PCR.

Potentiality of Methylation-sensitive Amplification Polymorphism (MSAP) in Identifying Genes Involved in Tomato Response to Tomato Yellow Leaf Curl Sardinia Virus

Plants have developed several genetically regulated defense strategies against a wide range of biotic stresses (viruses, fungi, bacteria, nematodes). There is experimental evidence that DNA methylation and post-transcriptional gene silencing (PTGS) are strongly associated and have evolved as plant defense mechanisms against invasion by foreign nucleic acids, including viruses. Recent studies have shown that PTGS is activated in plants infected by the geminivirus Tomato yellow leaf curl Sardinia virus (TYLCSV). To compare the DNA methylation profiles in TYLCSV-infected vs mock-inoculated tomato (Solanum lycopersicum syn. Lycopersicon esculentum) plants, a methylation-sensitive amplification polymorphism (MSAP) methodology was applied. The pattern of methylation was assayed at different stages of infection (1, 7, and 14 days) after inoculation by the natural vector Bemisia tabaci. Thirty-four polymorphic fragments were identified, of which ten were sequenced. The majority (eight out of ten) of polymorphisms generated by changes in methylation after TYLCSV infection belong to expressed portions of the tomato genome, mostly involved in defense and stress responses. Although the differences in expression levels during infection (in all cases examined except one) could be detected at least at one time-point, no clear correlation could be found between overexpression or underexpression and changes in methylation profiles. Our data demonstrate that TYLCSV induces changes in the methylation status of the tomato genome and that MSAP might contribute in identifying genes involved in plant–virus interactions.

Interaction with host SGS3 is required for suppression of RNA silencing by tomato yellow leaf curl virus V2 protein

The V2 protein of tomato yellow leaf curl geminivirus (TYLCV) functions as an RNA-silencing suppressor that counteracts the innate immune response of the host plant. The host-cell target of V2, however, remains unknown. Here we show that V2 interacts directly with SlSGS3, the tomato homolog of the Arabidopsis SGS3 protein (AtSGS3), which is known to be involved in the RNA-silencing pathway. SlSGS3 genetically complemented an AtSGS3 mutation and restored RNA silencing, indicating that SlSGS3 is indeed a functional homolog of AtSGS3. A point mutant of V2 that is unable to bind SlSGS3 also lost its ability to suppress RNA silencing, suggesting a correlation between the V2-SlSGS3 interaction in planta and the suppressor activity of V2.

Solution Structure, Divalent Metal and DNA Binding of the Endonuclease Domain from the Replication Initiation Protein from Porcine Circovirus 2

Circoviruses are the smallest circular single-stranded DNA viruses able to replicate in mammalian cells. Essential to their replication is the replication initiator, or Rep protein that initiates the rolling circle replication (RCR) of the viral genome. Here we report the NMR solution three-dimensional structure of the endonuclease domain from the Rep protein of porcine circovirus type 2 (PCV2), the causative agent of postweaning multisystemic wasting syndrome in swine. The domain comprises residues 12–112 of the full-length protein and exhibits the fold described previously for the Rep protein of the representative geminivirus tomato yellow leaf curl Sardinia virus. The structure, however, differs significantly in some secondary structure elements that decorate the central five-stranded β-sheet, including the replacement of a β-hairpin by an α-helix in PCV2 Rep. The identification of the divalent metal binding site was accomplished by following the paramagnetic broadening of NMR amide signals upon Mn2+ titration. The site comprises three conserved acidic residues on the exposed face of the central β-sheet. For the 1:1 complex of the PCV2 Rep nuclease domain with a 22mer double-stranded DNA oligonucleotide chemical shift mapping allowed the identification of the DNA binding site on the protein and aided in constructing a model of the protein/DNA complex.

Suppressor of RNA silencing encoded by Tomato yellow leaf curl virus-Israel

The Israeli isolate of Tomato yellow leaf curl geminivirus (TYLCV-Is) is a major tomato pathogen, causing extensive crop losses both in the New and Old World. Surprisingly, however, little is known about the molecular mechanisms of TYLCV-Is interactions with tomato cells. Here, we have identified a TYLCV-Is protein, V2, which acts as a suppressor of RNA silencing and which is unrelated to presently known viral suppressors. Specifically, V2, but not other proteins of TYLCV-Is, inhibited RNA silencing of a reporter transgene, GFP. This inhibition elevated the cellular levels of the GFP transcript and the GFP protein, but it had no apparent effect on the accumulation of GFP-specific short interfering RNAs (siRNAs), suggesting that TYLCV-Is V2 targets a step in the RNA silencing pathway which is subsequent to the Dicer-mediated cleavage of dsRNA. Visualization of the sub-cellular localization of TYLCV-Is V2 in plant protoplasts and tissues showed that this protein is associated with cytoplasmic strands and inclusion bodies in the cortical regions of the cell.

DNA Helicase Activity Is Associated with the Replication Initiator Protein Rep of Tomato Yellow Leaf Curl Geminivirus

The Rep protein of tomato yellow leaf curl Sardinia virus (TYLCSV), a single-stranded DNA virus of plants, is the replication initiator essential for virus replication. TYLCSV Rep has been classified among ATPases associated with various cellular activities (AAA+ ATPases), in superfamily 3 of small DNA and RNA virus replication initiators whose paradigmatic member is simian virus 40 large T antigen. Members of this family are DNA- or RNA-dependent ATPases with helicase activity necessary for viral replication. Another distinctive feature of AAA+ ATPases is their quaternary structure, often composed of hexameric rings. TYLCSV Rep has ATPase activity, but the helicase activity, which is instrumental in further characterization of the mechanism of rolling-circle replication used by geminiviruses, has been a longstanding question. We present results showing that TYLCSV Rep lacking the 121 N-terminal amino acids has helicase activity comparable to that of the other helicases: requirements for a 3' overhang and 3'-to-5' polarity of unwinding, with some distinct features and with a minimal AAA+ ATPase domain. We also show that the helicase activity is dependent on the oligomeric state of the protein.

Spread of Tomato yellow leaf curl virus in Sicily: Partial Displacement of Another Geminivirus Originally Present

The geminivirus Tomato yellow leaf curl virus (TYLCV) was reported for the first time in Italy in 2002. We have followed its spread in Sicily, where Tomato yellow leaf curl Sardinia virus (TYLCSV), another tomato-infecting geminivirus, is endemic and has been causing severe crop losses since 1989. The presence of the two viruses was monitored in the main tomato growing area, the Ragusa province, analyzing samples with yellow leaf curling symptoms. At first (spring–summer 2002) both viruses were always found in mixed infections, but in 2003 and 2004 18–35% of plants were found infected by TYLCV alone and 8–28% by TYLCSV alone, with 41–69% carrying both viruses. TYLCV has spread quickly in the area, demonstrating, as in other parts of the world, its high virulence and invasiveness; however it has not, so far, completely displaced TYLCSV. An infectious clone of TYLCV from Sicily (TYLCV-IT) was sequenced. The nucleotide sequence was 97% identical to other TYLCV strains of the ‘severe’ type, found in many countries worldwide.

Ultraviolet-Deficient Greenhouse Environment Affects Whitefly Attraction and Flight-Behavior

The effect of a UV-deficient environment on the attraction and dispersal behavior of whiteflies, Bemisia argentifolii (Bellows & Perring), and on the transmission efficiency of the whitefly-borne tomato yellow leaf curl geminivirus, was tested under field conditions and through controlled experiments. We found that the rate of tomato yellow leaf curl virus-disease spread to tomato plants grown under walk-in tunnels covered with regular greenhouse plastic sheets increases sharply with time, whereas the virus infection-rate under UV-absorbing sheets proceeds at a very slow pace. Average number of whiteflies trapped under regular plastic sheet tunnels was significantly higher than numbers trapped in UV-absorbing plastic sheet tunnels. Similarly, the average number of whiteflies trapped on yellow-sticky traps placed on the outside walls of tunnels covered with regular plastic was higher than the number trapped on the outside walls of tunnels covered with UV-absorbing plastic sheets. No differences were found in the whitefly's ability to transmit tomato yellow leaf curl virus under the two types of plastic covers. Whitefly dispersal pattern under the two types of plastic covers was examined using a release-recapture experiment. In each type of walk-in tunnel we established a grid of yellow-sticky traps forming two concentric circles: an inner and an external. Under UV-absorbing tunnels, significantly higher numbers of whiteflies were captured on the internal circle of traps than the external circle. The fraction of whiteflies that were captured on theexternalcirclewasmuchhigherunderregularcovers,whencomparedwithUV-absorbingcovers, suggesting that filtration of UV light hindered the ability of whiteflies to disperse inside these plastic tunnels. Our results indicate that the mechanisms by which UV-deficiency protects covered crops from insect infestation and spread of viruses are that the lack of UV interferes with insect flight orientation; and that the lack of UV radiation alters the normal behavior of the invading insects, resulting in reduced dispersal activity.

The effect of thiamethoxam, a second generation neonicotinoid insecticide, in preventing transmission of tomato yellow leaf curl geminivirus (TYLCV) by the whitefly Bemisia tabaci (Gennadius)

The effect of thiamethoxam, a new neonicotinoid insecticide, in preventing transmission of tomato yellow leaf curl geminivirus (TYLCV) by the whitefly Bemisia tabaci has been studied by means of experimental transmissions to tomato seedlings. Source plants were foliar-treated while test plants were both foliar- and drench-treated at the six-leaf stage. Doses of 50 ppm a.i. for the foliar and of 7 mg a.i. per plant for the drench application were evaluated. Drench application to test plants provided a good level of protection from TYLCV infections in all tested conditions (from 1 to 22 days after treatment application); foliar application resulted in a prompt but short-lasting protection (<8 days). Foliar treatment of source plants made infected tomatoes totally ineffective as a virus source for at least 8 days. The proportion of viruliferous whiteflies surviving the acquisition on treated plants appeared similar to that of insects fed on untreated plants, suggesting that thiamethoxam activity in preventing TYLCV transmission by B. tabaci is simply due to its killing activity, and antifeeding or repellent actions can be excluded. Viruliferous whiteflies exposed to thiamethoxam-treated plants stopped feeding before acquiring enough virus to subsequently inoculate plants.

Tomato yellow leaf curl geminivirus (TYLCV-Is) is transmitted among whiteflies (Bemisia tabaci) in a sex-related manner.

Tomato yellow leaf curl virus (TYLCV) is the name given to a complex of geminiviruses infecting tomato cultures worldwide. TYLCV is transmitted by a single insect species, the whitefly Bemisia tabaci. Herein we show that a TYLCV isolate from Israel (TYLCV-Is) can be transmitted among whiteflies in a sex-dependent manner, in the absence of any other source of virus. TYLCV was transmitted from viruliferous males to females and from viruliferous females to males but not among insects of the same sex. Transmission took place when insects were caged in groups or in couples, in a feeding chamber or on cotton plants, a TYLCV nonhost. The recipient insects were able to efficiently inoculate tomato test plants. Insect-to-insect virus transmission was instrumental in increasing the number of whiteflies capable of infecting tomato test plants in a whitefly population. TYLCV was present in the hemolymph of whiteflies caged with viruliferous insects of the other sex; therefore, the virus follows, at least in part, the circulative pathway associated with acquisition from infected plants. Taken as a whole, these results imply that a plant virus can be sexually transmitted from insect to insect.

Correlation Between Whitefly (Homoptera: Aleyrodidae) Feeding Behavior and Transmission of Tomato Yellow Leaf Curl Virus

The feeding behavior of the whitefly Bemisia tabaci (Gennadius) was monitored using the electrical penetration graph (EPG) technique during the transmission process of tomato yellow leaf curl geminivirus (TYLCV). The behavior of individual viruliferous whiteflies was recorded on two-leaf stage tomato test plants (Lycopersicon esculentum Mill ‘Riofuego’). A total of 213 whitefly individuals was recorded on single test plants during an inoculation access period that ranged from 3.5 to 14 h. Recordings were classified into 4 categories depending of the waveforms observed: group I, including only waveform C (pathway), was associated with a residual 2.4% TYLCV transmission efficiency; group II, showing stylet pathway and a single E(pd)1 waveform, was associated with a 7.4% transmission efficiency; group III, showing stylet pathway and a single E(pd)1 + E(pd)2 waveform, achieved a 23.4% transmission efficiency; and finally group IV, showing pathway followed by several E(pd)1 + E(pd)2 waveforms, was associated with the highest transmission efficiency (37.5%). A total of 16 different behavioral variables was introduced into a stepwise-backward logistic regression model to determine the variables most related to TYLCV inoculation. Among them, the total duration of E(pd)1 was the most significant variable associated with virus inoculation by B. tabaci (P = 0.002, positive relationship). In addition, the regression analysis indicated a strong positive relationship (P = 0.005) between transmission efficiency and total number of E(pd)1 (t = 4.39, β = 0.45 ± 0.10, r = 0.87), and total duration of waveform E(pd)1 (P = 0.05) (t = 4.00, β = 0.02 ± 0.004, r = 0.94). The minimum phloem contact E(pd)1 threshold period observed for successful inoculation of TYLCV by B. tabaci on tomato plants was 1.8 min. Implications of these results for persistent virus transmission by whiteflies are discussed.

First Report of Tomato Yellow Leaf Curl Geminivirus in Morocco.

In July 1998, shortened internodes, reduced leaf size, leaf curling, and bushy plants were observed in tomato crops in the coastal region near Casablanca, Morocco. The symptoms were similar to those described for tomato yellow leaf curl virus (TYLCV) disease. During September, the disease was present in more than 130 ha of outdoor and protected crops. Economic losses ranged between 20 and 100%. Similar symptoms were observed in tomato crops in the northeastern region of Morocco. During the same period, high populations of Bemisia tabaci (Gennadius) were observed on tomato, sweet pepper, and several weeds, including Sonchus oleraceus L., Chenopodium sp., Malva parviflora L., and Datura stramonium L. Attempts to control whiteflies with insecticides generally were unsuccessful. Using a pair of degenerate primers (3) that are expected to amplify a region of the A component of begomoviruses between the intergenic conserved nonanucleotide sequence and the first 5' quarter of the capsid protein gene, an expected size product of 500 bp was obtained from six tomato leaf samples collected near Casablanca: four were obtained from symptomatic plants and two from apparently symptomless plants. A product of the same size was obtained from a sample of tomato infected with a TYLCV isolate from Reunion Island, France (2), but not from a sample from symptomless tomato plants from Reunion. The polymerase chain reaction (PCR) product obtained from one symptomatic sample from Morocco was cloned and sequenced (2). The resulting sequence (EMBL no. AJ133491) was at least 95.2% identical to sequences of TYLCV isolates from the Dominican Republic (EMBL no. AF024715), Cuba (EMBL no. AJ223505), Israel (EMBL no. X15656; EMBL no. X76319 for the mild clone), Spain (EMBL no. AF071228), and Reunion (EMBL no. AJ010790). Based on sequence comparisons, the tomato leaf sample was infected by a geminivirus isolate belonging to the Israeli species of TYLCV and was most closely related to the Dominican Republic isolate (98.4% nucleotide identity). In symptomless plants from which a 500-bp PCR product was obtained, infection may have been in an early stage. This is consistent with the fact that in the 2-ha field from which tomato samples were collected, only 4% of the plants exhibited TYLCV symptoms at the time of collection, whereas at least 70% of plants showed symptoms later, requiring the farmer to abandon the crop. The presence of the Eastern Mediterranean TYLCV species in a Western Mediterranean country is not a new observation; this virus has been described in Spain and Portugal (1). In Morocco, TYLCV symptoms were observed during the 1996 to 1997 growing season, following importation of grafted tomato plants from the Netherlands, by a farmer from the Casablanca region.

Transmission of Tomato Yellow Leaf Curl Geminivirus to Imidacloprid Treated Tomato Plants by the Whitefly Bemisia tabaci(Homoptera: Aleyrodidae)

The ability of whiteflies [ Bemisia tabaci (Gennadius), B biotype] to transmit tomato yellow leaf curl geminivirus (TYLCV, Israeli isolate) to imidacloprid-treated and untreated tomato plants was investigated. Viruliferous whiteflies were caged with treated tomato plants held in a nethouse. Insect mortality and tomato infection was assessed during the summer and the winter seasons, after insecticide application. In summer, insects that were given access to tomato plants 3 and 11 d after insecticide treatment died within 80 min. This period increased to 150 min 18 d after treatment. The insecticide lost its potency 25 d after application. In winter the lethal effect of the insecticide lasted longer than in summer. In summer as well as in winter, 70% of the tomato plants caged with viruliferous whiteflies 3 d after insecticide treatment became infected. These figures increased to 80% 11 d after treatment and to nearly 100% 18 d after treatment. Insecticide-treated plants were as prone to infection as nontreated plants. Approximately 48 min of access to a treated tomato plant was sufficient for a single viruliferous insect to inoculate the virus with an efficiency rate of 20% or more. Transmission efficiency was similar to that achieved on nontreated plants. Therefore, viruliferous insects had enough time to inoculate TYLCV to imidacloprid-treated plants before they died.

A GroEL Homologue from Endosymbiotic Bacteria of the WhiteflyBemisia tabaciIs Implicated in the Circulative Transmission of Tomato Yellow Leaf Curl Virus

Evidence for the involvement of aBemisia tabaciGroEL homologue in the transmission of tomato yellow leaf curl geminivirus (TYLCV) is presented. A ∼63-kDa protein was identified inB. tabaciwhole-body extracts using an antiserum raised against aphidBuchneraGroEL. The GroEL homologue was immunolocalized to a coccoid-shaped whitefly endosymbiont. The 30 N-terminal amino acids of the whitefly GroEL homologue showed 80% homology with that from different aphid species and GroEL fromEscherichia coli. Purified GroEL fromB. tabaciexhibited ultrastructural similarities to that of the endosymbiont from aphids andE. coli.In vitroligand assays showed that tomato yellow leaf curl virus (TYLCV) particles displayed a specific affinity for theB. tabaci63-kDa GroEL homologue. Feeding whiteflies anti-BuchneraGroEL antiserum before the acquisition of virions reduced TYLCV transmission to tomato test plants by >80%. In the haemolymph of these whiteflies, TYLCV DNA was reduced to amounts below the threshold of detection by Southern blot hybridization. Active antibodies were recovered from the insect haemolymph suggesting that by complexing the GoEL homologue, the antibody disturbed interaction with TYLCV, leading to degradation of the virus. We propose that GroEL ofB. tabaciprotects the virus from destruction during its passage through the haemolymph.

Detection of a Geminivirus Infecting Sweet Potato in the United States

In 1994, a sweet potato sample showing leaf curl symptoms was collected from the field in Louisiana. When graft-inoculated, Ipomoea nil cv. Scarlett O'Hara reacted with severe leaf distortion and chlorosis symptoms. I. aquatica reacted with a bright yellow mottle. The virus isolated was designated the United States isolate of sweet potato leaf curl virus (SPLCV-US). It was transmitted to I. nil by the sweet potato whitefly, Bemisia tabaci biotype B. DNA probes prepared with component A of pepper Huasteco geminivirus, with an isolate of bean golden mosaic geminivirus from Guatemala, with an isolate of tomato mottle geminivirus from Florida, and with an isolate of tomato yellow leaf curl geminivirus from the Dominican Republic (TYLCV-DR) hybridized with a 2.6-kb DNA band present in DNA extracts from plants infected with SPLCV-US. Probes prepared with the B component of these geminiviruses did not hybridize with these DNA extracts. We were unable to amplify SPLCV-US DNA products by polymerase chain reaction (PCR) in quantities that could be visualized by ethidium bromide staining. However, Southern blots from amplifications with primers AV494/AC1048 revealed PCR products of approximately 600 bp and 550 bp when hybridized with the TYLCV-DR probe. These results were consistently obtained from infected I. cordatotriloba and less consistently from I. aquatica or I. setosa. Fibrillar inclusions were occasionally seen, and granular aggregates of viruslike particles were observed in the nucleus of infected I. cordatotriloba. These results suggest that the virus isolated from sweet potato with leaf curl symptoms belongs to the geminivirus group.

Invisible invaders: Insect-transmitted viruses threaten agriculture

The vast movement of people and agricultural products between distant geographical regions has created unprecedented opportunities for introducing plant viruses and the insects that carry them (vectors) to new areas. Outbreaks of new viruses may be favored in these agroecosystems by crop susceptibility, the presence of particular weeds and certain agricultural practices. In some cases, conditions in these ecosystems may be ideal for the emergence of altered plant viruses and new virus/vector relationships. This may result in the appearance of insect-transmitted plant viruses in crops and regions where they have not been seen before. Because plant viruses and their insect vectors are intimately linked, the status of both must be considered in formulating strategies to prevent or slow their introduction, as well as to manage any invasions. To illustrate these points we highlight two situations that could threaten California agriculture. First, a devastating plant virus, tomato yellow leaf curl geminivirus, is not present in California, but an insect (the silverleaf whitefly) that transmits it is present. Second, the brown citrus aphid is not present in California, but a citrus virus (citrus tristeza closterovirus) that this insect efficiently spreads, is present.

Evidence for Transovarial Transmission of Tomato Yellow Leaf Curl Virus by Its Vector, the WhiteflyBemisia tabaci

The whiteflyBemisia tabaciis the only vector of the tomato yellow leaf curl geminivirus (TYLCV). The insect transmits the virus in a persistent-circulative manner. TYLCV DNA was detected by polymerase chain reaction and by Southern blot hybridization in progeny (eggs, first and second instars, adults) of single viruliferous whiteflies that developed on eggplant or on cotton (two TYLCV nonhost plants). Furthermore, TYLCV DNA was present in the progeny of insects that had acquired the virus through the egg. The adult progeny of the viruliferous insects and their own progeny were able to infect tomato test plants, producing typical disease symptoms. Ovaries and maturing eggs of viruliferous insects contained viral DNA, as did eggs laid by viruliferous insects maintained on an artificial diet. Eggs laid by nonviruliferous whiteflies on cotton plants previously caged with viruliferous insects did not acquire viral DNA from the plant. Hence, TYLCV can be transmitted through the egg for at least two generations. In the absence of an available plant host, the whitefly may serve as a reservoir of the virus between growing seasons.

Solanum nigrum : an indigenous weed reservoir for a tomato yellow leaf curl geminivirus in southern Spain

A tomato yellow leaf curl geminivirus (TYLCV-AL), was first identified in tomato plants in Almeria, southern Spain in 1992. This virus is transmitted by the tobacco whitefly, Bemisia tabaci (Gennadius), and is presently infecting tomato crops throughout the south eastern region of Spain. Solanum nigrum, collected from a field in south east Spain and exhibiting leaf curl symptoms, was squash blotted onto nylon membrane and gave a positive signal when hybridised to a TYLCV-Is DNA probe. Laboratory tests showed B. tabaci to transmit TYLCV-AL from infected tomato plants to S. nigrum seedlings. The virus could then be acquired by B. tabaci and transmitted back from infected S. nigrum plants to tomato, inducing typical TYLCV symptoms. These results indicate the importance of S. nigrum as a weed host/reservoir for a TYLCV and its possible role in the spread of this virus within Europe.