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Publications

2020
Sade, D. ; Sade, N. ; Brotman, Y. ; Czosnek, H. Tomato yellow leaf curl virus (TYLCV)-resistant tomatoes share molecular mechanisms sustaining resistance with their wild progenitor Solanum habrochaites but not with TYLCV-susceptible tomatoes. Food Security under Climate Change 2020, 295, 110439. Publisher's VersionAbstract
The wild tomato species Solanum habrochaites (Sh) has been used as a source for tomato yellow leaf curl virus (TYLCV) resistance in a breeding program to generate a TYLCV-resistant tomato line. Susceptible (S) and resistant (R) lines have been developed through this program. We compared the behavior of R, S and Sh tomato plants upon infection to find out whether the resistant phenotype of R plants originated from Sh. Results showed that mechanisms involving sugar-signaling (i.e., LIN6/HT1), water channels (i.e., TIP1;1), hormone homeostasis (i.e., ABA and SA) and urea accumulation were shared by S. habrochaites and R plants, but not by S. habrochaites and S tomatoes. This finding supports the hypothesis that these mechanisms were introgressed in the R genotype from the wild tomato progenitor during breeding for TYLCV resistance. Hence, identification of genes contributing to resistance to biotic stress from wild tomato species and their introgression into domestic plants ensures tomato supply and food security.
Kliot, A. ; Johnson, R. S. ; MacCoss, M. J. ; Kontsedalov, S. ; Lebedev, G. ; Czosnek, H. ; Heck, M. ; Ghanim, M. A proteomic approach reveals possible molecular mechanisms and roles for endosymbiotic bacteria in begomovirus transmission by whiteflies. Gigascience 2020, 9. Publisher's VersionAbstract
Many plant viruses are vector-borne and depend on arthropods for transmission between host plants. Begomoviruses, the largest, most damaging and emerging group of plant viruses, infect hundreds of plant species, and new virus species of the group are discovered each year. Begomoviruses are transmitted by members of the whitefly Bemisia tabaci species complex in a persistent-circulative manner. Tomato yellow leaf curl virus (TYLCV) is one of the most devastating begomoviruses worldwide and causes major losses in tomato crops, as well as in many agriculturally important plant species. Different B. tabaci populations vary in their virus transmission abilities; however, the causes for these variations are attributed among others to genetic differences among vector populations, as well as to differences in the bacterial symbionts housed within B. tabaci.Here, we performed discovery proteomic analyses in 9 whitefly populations from both Middle East Asia Minor I (MEAM1, formerly known as B biotype) and Mediterranean (MED, formerly known as Q biotype) species. We analysed our proteomic results on the basis of the different TYLCV transmission abilities of the various populations included in the study. The results provide the first comprehensive list of candidate insect and bacterial symbiont (mainly Rickettsia) proteins associated with virus transmission.Our data demonstrate that the proteomic signatures of better vector populations differ considerably when compared with less efficient vector populations in the 2 whitefly species tested in this study. While MEAM1 efficient vector populations have a more lenient immune system, the Q efficient vector populations have higher abundance of proteins possibly implicated in virus passage through cells. Both species show a strong link of the facultative symbiont Rickettsia to virus transmission.
Gorovits, R. ; Sobol, I. ; Akama, K. ; Chefetz, B. ; Czosnek, H. Pharmaceuticals in treated wastewater induce a stress response in tomato plants. Sci Rep 2020, 10, 1856.Abstract
Pharmaceuticals remain in treated wastewater used to irrigate agricultural crops. Their effect on terrestrial plants is practically unknown. Here we tested whether these compounds can be considered as plant stress inducers. Several features characterize the general stress response in plants: production of reactive oxygen species acting as stress-response signals, MAPKs signaling cascade inducing expression of defense genes, heat shock proteins preventing protein denaturation and degradation, and amino acids playing signaling roles and involved in osmoregulation. Tomato seedlings bathing in a cocktail of pharmaceuticals (Carbamazepine, Valporic acid, Phenytoin, Diazepam, Lamotrigine) or in Carbamazepine alone, at different concentrations and during different time-periods, were used to study the patterns of stress-related markers. The accumulation of the stress-related biomarkers in leaf and root tissues pointed to a cumulative stress response, mobilizing the cell protection machinery to avoid metabolic modifications and to restore homeostasis. The described approach is suitable for the investigation of stress response of different crop plants to various contaminants present in treated wastewater.
2019
Ghosh, S. ; Kanakala, S. ; Lebedev, G. ; Kontsedalov, S. ; Silverman, D. ; Alon, T. ; Mor, N. ; Sela, N. ; Luria, N. ; Dombrovsky, A. ; et al. Transmission of a New Polerovirus Infecting Pepper by the Whitefly Bemisia tabaci. Journal of Virology 2019, 93. Publisher's VersionAbstract
Many animal and plant viruses depend on arthropods for their transmission. Virus-vector interactions are highly specific, and only one vector or one of a group of vectors from the same family is able to transmit a given virus. Poleroviruses (Luteoviridae) are phloem-restricted RNA plant viruses that are exclusively transmitted by aphids. Multiple aphid-transmitted polerovirus species commonly infect pepper, causing vein yellowing, leaf rolling, and fruit discoloration. Despite low aphid populations, a recent outbreak with such severe symptoms in many bell pepper farms in Israel led to reinvestigation of the disease and its insect vector. Here we report that this outbreak was caused by a new whitefly (Bemisia tabaci)-transmitted polerovirus, which we named Pepper whitefly-borne vein yellows virus (PeWBVYV). PeWBVYV is highly (>95%) homologous to Pepper vein yellows virus (PeVYV) from Israel and Greece on its 5' end half, while it is homologous to African eggplant yellows virus (AeYV) on its 3' half. Koch’s postulates were proven by constructing a PeWBVYV infectious clone causing the pepper disease, which was in turn transmitted to test pepper plants by B. tabaci but not by aphids. PeWBVYV represents the first report of a whitefly-transmitted polerovirus.IMPORTANCE The high specificity of virus-vector interactions limits the possibility of a given virus changing vectors. Our report describes a new virus from a family of viruses strictly transmitted by aphids which is now transmitted by whiteflies (Bemisia tabaci) and not by aphids. This report presents the first description of polerovirus transmission by whiteflies. Whiteflies are highly resistant to insecticides and disperse over long distances, carrying virus inoculum. Thus, the report of such unusual polerovirus transmission by a supervector has extensive implications for the epidemiology of the virus disease, with ramifications concerning the international trade of agricultural commodities.
Kliot, A. ; Kontsedalov, S. ; Lebedev, G. ; Czosnek, H. ; Ghanim, M. Combined infection with Tomato yellow leaf curl virus and Rickettsia influences fecundity, attraction to infected plants and expression of immunity-related genes in the whitefly Bemisia tabaci. Journal of General Virology 2019, 100, 721-731. Publisher's VersionAbstract
We have recently shown that Rickettsia, a secondary facultative bacterial symbiont that infects the whitefly B. tabaci is implicated in the transmission of Tomato yellow leaf curl virus (TYLCV). Infection with Rickettsia improved the acquisition and transmission of the virus by B. tabaci adults. Here we performed a transcriptomic analysis with Rickettsia-infected and uninfected B. tabaci adults before and after TYLCV acquisition. The results show a dramatic and specific activation of the immune system in the presence of Rickettsia before TYLCV acquisition. However, when TYLCV was acquired, it induced massive activation of gene expression in the Rickettsia uninfected population, whereas in the Rickettsia-infected population the virus induced massive down-regulation of gene expression. Fitness and choice experiments revealed that while Rickettsia-infected whiteflies are always more attracted to TYLCV-infected plants, this attraction is not always beneficiary for their offspring. These studies further confirm the role of Rickettsia in many aspects of B. tabaci interactions with TYLCV, and possibly serves as an important factor in the dissemination of the virus. © 2019 The Authors.
Ghosh, S. ; Kanakala, S. ; Lebedev, G. ; Kontsedalov, S. ; Silverman, D. ; Alon, T. ; Mor, N. ; Sela, N. ; Luria, N. ; Dombrovsky, A. ; et al. Transmission of a new polerovirus infecting pepper by the whitefly Bemisia tabaci. Journal of Virology 2019, 93. Publisher's VersionAbstract
Many animal and plant viruses depend on arthropods for their transmission. Virus-vector interactions are highly specific, and only one vector or one of a group of vectors from the same family is able to transmit a given virus. Poleroviruses (Luteoviridae) are phloem-restricted RNA plant viruses that are exclusively transmitted by aphids. Multiple aphid-transmitted polerovirus species commonly infect pepper, causing vein yellowing, leaf rolling, and fruit discoloration. Despite low aphid populations, a recent outbreak with such severe symptoms in many bell pepper farms in Israel led to reinvestigation of the disease and its insect vector. Here we report that this outbreak was caused by a new whitefly (Bemisia tabaci)-transmitted polerovirus, which we named Pepper whitefly-borne vein yellows virus (PeWBVYV). PeWBVYV is highly (95%) homologous to Pepper vein yellows virus (PeVYV) from Israel and Greece on its 5ʹ end half, while it is homologous to African eggplant yellows virus (AeYV) on its 3ʹ half. Koch’s postulates were proven by constructing a PeWBVYV infectious clone causing the pepper disease, which was in turn transmitted to test pepper plants by B. tabaci but not by aphids. PeWBVYV represents the first report of a whitefly-transmitted polerovirus. IMPORTANCE The high specificity of virus-vector interactions limits the possibility of a given virus changing vectors. Our report describes a new virus from a family of viruses strictly transmitted by aphids which is now transmitted by whiteflies (Bemisia tabaci) and not by aphids. This report presents the first description of polerovirus transmission by whiteflies. Whiteflies are highly resistant to insecticides and disperse over long distances, carrying virus inoculum. Thus, the report of such unusual polerovirus transmission by a supervector has extensive implications for the epidemiology of the virus disease, with ramifications concerning the international trade of agricultural commodities. © 2019 American Society for Microbiology. All Rights Reserved.
Bar, L. ; Czosnek, H. ; Sobol, I. ; Ghanim, M. ; Hariton Shalev, A. Downregulation of dystrophin expression in pupae of the whitefly Bemisia tabaci inhibits the emergence of adults. Insect Mol Biol 2019, 28, 662-675.Abstract
The whitefly Bemisia tabaci is a major pest to agriculture. Adults are able to fly for long distances and to colonize staple crops, herbs and ornamentals, and to vector viruses belonging to several important taxonomic groups. During their early development, whiteflies mature from eggs through several nymphal stages (instars I to IV) until adults emerge from pupae. We aim at reducing whitefly populations by inhibiting the emergence of adults from nymphs. Here we targeted dystrophin, a conserved protein essential for the development of the muscle system in humans, other animals and insects. We have exploited the fact that whitefly nymphs developing on tomato leaves feed from the plant phloem via their stylets. Thus, we delivered dystrophin-silencing double-stranded RNA to nymphs developing on leaves of tomato plantlets with their roots bathing in the silencing solution. Downregulation of dystrophin expression occurred mainly in pupae. Dystrophin silencing induced also the downregulation of the dystrophin-associated protein genes actin and tropomyosin, and disrupted F-actin. Most significantly, the treatment inhibited the emergence of adults from pupae, suggesting that targeting dystrophin may help to restrain whitefly populations. This study demonstrates for the first time the important role of dystrophin in the development of a major insect pest to agriculture.
2018
Dahan-Meir, T. ; Filler-Hayut, S. ; Melamed-Bessudo, C. ; Bocobza, S. ; Czosnek, H. ; Aharoni, A. ; Levy, A. A. Efficient in planta gene targeting in tomato using geminiviral replicons and the CRISPR/Cas9 system. Plant J 2018, 95, 5-16.Abstract
Current breeding relies mostly on random mutagenesis and recombination to generate novel genetic variation. However, targeted genome editing is becoming an increasingly important tool for precise plant breeding. Using the CRISPR-Cas system combined with the bean yellow dwarf virus rolling circle replicon, we optimized a method for targeted mutagenesis and gene replacement in tomato. The carotenoid isomerase (CRTISO) and phytoene synthase 1 (PSY1) genes from the carotenoid biosynthesis pathway were chosen as targets due to their easily detectable change of phenotype. We took advantage of the geminiviral replicon amplification as a means to provide a large amount of donor template for the repair of a CRISPR-Cas-induced DNA double-strand break (DSB) in the target gene, via homologous recombination (HR). Mutagenesis experiments, performed in the Micro-Tom variety, achieved precise modification of the CRTISO and PSY1 loci at an efficiency of up to 90%. In the gene targeting (GT) experiments, our target was a fast-neutron-induced crtiso allele that contained a 281-bp deletion. This deletion was repaired with the wild-type sequence through HR between the CRISPR-Cas-induced DSB in the crtiso target and the amplified donor in 25% of the plants transformed. This shows that efficient GT can be achieved in the absence of selection markers or reporters using a single and modular construct that is adaptable to other tomato targets and other crops.
2017
Gorovits, R. ; Czosnek, H. The involvement of heat shock proteins in the establishment of Tomato Yellow Leaf Curl Virus infection. Frontiers in Plant Science 2017, 8. Publisher's VersionAbstract
Tomato yellow leaf curl virus (TYLCV), a begomovirus, induces protein aggregation in infected tomatoes and in its whitefly vector Bemisia tabaci. The interactions between TYLCV and HSP70 and HSP90 in plants and vectors are necessity for virus infection to proceed. In infected host cells, HSP70 and HSP90 are redistributed from a soluble to an aggregated state. These aggregates contain, together with viral DNA/proteins and virions, HSPs and components of the protein quality control system such as ubiquitin, 26S proteasome subunits, and the autophagy protein ATG8. TYLCV CP can form complexes with HSPs in tomato and whitefly. Nonetheless, HSP70 and HSP90 play different roles in the viral cell cycle in the plant host. In the infected host cell, HSP70, but not HSP90, participates in the translocation of CP from the cytoplasm into the nucleus. Viral amounts decrease when HSP70 is inhibited, but increase when HSP90 is downregulated. In the whitefly vector, HSP70 impairs the circulative transmission of TYLCV; its inhibition increases transmission. Hence, the efficiency of virus acquisition by whiteflies depends on the functionality of both plant chaperones and their cross-talk with other protein mechanisms controlling virus-induced aggregation. © 2017 Gorovits and Czosnek.
Pan, L. - L. ; Chen, Q. - F. ; Zhao, J. - J. ; Guo, T. ; Wang, X. - W. ; Hariton-Shalev, A. ; Czosnek, H. ; Liu, S. - S. Clathrin-mediated endocytosis is involved in Tomato yellow leaf curl virus transport across the midgut barrier of its whitefly vector. Virology 2017, 502, 152-159.Abstract
Tomato yellow leaf curl virus (TYLCV) is a begomovirus transmitted by the whitefly Bemisia tabaci. The circulative translocation of the virus in the insect is known in its broad line. However, transit of TYLCV from the digestive tract into the haemolymph is poorly understood. We studied the involvement of clathrin in this process by disrupting the clathrin-mediated endocytosis and the endosome network using inhibitor feeding, antibody blocking and dsRNA silencing. We monitored the quantities of TYLCV in the whitefly and virus transmission efficiency. Following endocytosis and endosome network disruption, the quantity of virus was higher in the midgut relative to that of the whole insect body, and the quantity of virus in the haemolymph was reduced. The transmission efficiency of TYLCV by the treated insects was also reduced. These findings indicate that clathrin-mediated endocytosis and endosomes play an important role in the transport of TYLCV across the whitefly midgut.
Gorovits, R. ; Moshe, A. ; Amrani, L. ; Kleinberger, R. ; Anfoka, G. ; Czosnek, H. The six Tomato yellow leaf curl virus genes expressed individually in tomato induce different levels of plant stress response attenuation. Cell Stress Chaperones 2017, 22, 345-355.Abstract
Tomato yellow leaf curl virus (TYLCV) is a begomovirus infecting tomato plants worldwide. TYLCV needs a healthy host environment to ensure a successful infection cycle for long periods. Hence, TYLCV restrains its destructive effect and induces neither a hypersensitive response nor cell death in infected tomatoes. On the contrary, TYLCV counteracts cell death induced by other factors, such as inactivation of HSP90 functionality. Suppression of plant death is associated with the inhibition of the ubiquitin 26S proteasome degradation and with a deactivation of the heat shock transcription factor HSFA2 pathways (including decreased HSP17 levels). The goal of the current study was to find if the individual TYLCV genes were capable of suppressing HSP90-dependent death and HSFA2 deactivation. The expression of C2 (C3 and CP to a lesser extent) caused a decrease in the severity of death phenotypes, while the expression of V2 (C1 and C4 to a lesser extent) strengthened cell death. However, C2 or V2 markedly affected stress response under conditions of viral infection. The downregulation of HSFA2 signaling, initiated by the expression of C1 and V2, was detected in the absence of virus infection, but was enhanced in infected plants, while CP and C4 mitigated HSFA2 levels only in the infected tomatoes. The dependence of analyzed plant stress response suppression on the interaction of the expressed genes with the environment created by the whole virus infection was more pronounced than on the expression of individual TYLCV genes.
Czosnek, H. ; Hariton-Shalev, A. ; Sobol, I. ; Gorovits, R. ; Ghanim, M. The Incredible Journey of Begomoviruses in Their Whitefly Vector. Viruses 2017, 9.Abstract
Begomoviruses are vectored in a circulative persistent manner by the whitefly The insect ingests viral particles with its stylets. Virions pass along the food canal and reach the esophagus and the midgut. They cross the filter chamber and the midgut into the haemolymph, translocate into the primary salivary glands and are egested with the saliva into the plant phloem. Begomoviruses have to cross several barriers and checkpoints successfully, while interacting with would-be receptors and other whitefly proteins. The bulk of the virus remains associated with the midgut and the filter chamber. In these tissues, viral genomes, mainly from the tomato yellow leaf curl virus (TYLCV) family, may be transcribed and may replicate. However, at the same time, virus amounts peak, and the insect autophagic response is activated, which in turn inhibits replication and induces the destruction of the virus. Some begomoviruses invade tissues outside the circulative pathway, such as ovaries and fat cells. Autophagy limits the amounts of virus associated with these organs. In this review, we discuss the different sites begomoviruses need to cross to complete a successful circular infection, the role of the coat protein in this process and the sites that balance between virus accumulation and virus destruction.
2016
Ghanim, M. ; Czosnek, H. Interactions between the Whitefly Bemisia Tabaci and begomoviruses: Biological and genomic perspectives; Management of Insect Pests to Agriculture: Lessons Learned from Deciphering their Genome, Transcriptome and Proteome; 2016; pp. 181-200. Publisher's VersionAbstract
Begomoviruses are an emerging group of plant viruses, exclusively transmitted by the whitefly Bemisia tabaci in a persistent-circulative manner. Despite the economic importance of both, very little is known about begomovirus-whitefly interactions. Specific topics of interest that have been a subject of intensive research during the last decade include the route of the virus in the insect organs and cells, the influence of the virus on the insect’s behavior and transcriptome, the proteins that mediate begomovirus translocation and the role of bacterial symbionts in this phenomenon. These topics are summarized and discussed in this chapter. © Springer International Publishing Switzerland 2016.
Czosnek, H. ; Ghanim, M. Management of insect pests to agriculture: Lessons learned from deciphering their genome, transcriptome and proteome; Management of Insect Pests to Agriculture: Lessons Learned from Deciphering their Genome, Transcriptome and Proteome; 2016; pp. 1-290. Publisher's VersionAbstract
Thanks to the application of new technologies such as whole-genome sequencing, analysis of transcriptome and proteome of insect pest to agriculture, great progress has been made in understanding the life style, reproduction, evolution and nuisance to crops caused by insect pests such as aphids, planthoppers, and whiteflies. We believe that time has come to summarize progress and to have a glance over the horizon. In this Book experts in the field discuss novel means to increase the different kinds of resistances of plants to better limit the effects of pest, to understand and disturb the hormonal regulation of embryogenesis, molting, metamorphosis and reproduction, to determine the function of insect genes in diverse processes such as metabolism, interaction with plants, virus transmission, development, and adaptation to a changing environment. The knowledge presented here is discussed with the aim of further improving control strategies of insect pestsman";mso-hansi-theme-font:minor-bidi;mso-bidi-theme-font:minor-bidi; mso-ansi-language:NL;mso-fareast-language:NL;mso-bidi-language:AR-SA">. © Springer International Publishing Switzerland 2016.
Shalev, A. H. ; Sobol, I. ; Ghanim, M. ; Liu, S. - S. ; Czosnek, H. The whitefly Bemisia tabaci Knottin-1 gene is implicated in regulating the quantity of tomato yellow leaf curl virus ingested and transmitted by the insect. Viruses 2016, 8. Publisher's VersionAbstract
The whitefly Bemisia tabaci is a major pest to agricultural crops. It transmits begomoviruses, such as Tomato yellow leaf curl virus (TYLCV), in a circular, persistent fashion. Transcriptome analyses revealed that B. tabaci knottin genes were responsive to various stresses. Upon ingestion of tomato begomoviruses, two of the four knottin genes were upregulated, knot-1 (with the highest expression) and knot-3. In this study, we examined the involvement of B. tabaci knottin genes in relation to TYLCV circulative transmission. Knottins were silenced by feeding whiteflies with knottin dsRNA via detached tomato leaves. Large amounts of knot-1 transcripts were present in the abdomen of whiteflies, an obligatory transit site of begomoviruses in their circulative transmission pathway; knot-1 silencing significantly depleted the abdomen from knot-1 transcripts. Knot-1 silencing led to an increase in the amounts of TYLCV ingested by the insects and transmitted to tomato test plants by several orders of magnitude. This effect was not observed following knot-3 silencing. Hence, knot-1 plays a role in restricting the quantity of virions an insect may acquire and transmit. We suggest that knot-1 protects B. tabaci against deleterious effects caused by TYLCV by limiting the amount of virus associated with the whitefly vector. © 2016 by the authors; licensee MDPI, Basel, Switzerland.
Gorovits, R. ; Fridman, L. ; Kolot, M. ; Rotem, O. ; Ghanim, M. ; Shriki, O. ; Czosnek, H. Tomato yellow leaf curl virus confronts host degradation by sheltering in small/midsized protein aggregates. Virus Res 2016, 213, 304-313.Abstract
Tomato yellow leaf curl virus (TYLCV) is a begomovirus transmitted by the whitefly Bemisia tabaci to tomato and other crops. TYLCV proteins are endangered by the host defenses. We have analyzed the capacity of the tomato plant and of the whitefly insect vector to degrade the six proteins encoded by the TYLCV genome. Tomato and whitefly demonstrated the highest proteolytic activity in the fractions containing soluble proteins, less-in large protein aggregates; a significant decrease of TYLCV proteolysis was detected in the intermediate-sized aggregates. All the six TYLCV proteins were differently targeted by the cytoplasmic and nuclear degradation machineries (proteases, ubiquitin 26S proteasome, autophagy). TYLCV could confront host degradation by sheltering in small/midsized aggregates, where viral proteins are less exposed to proteolysis. Indeed, TYLCV proteins were localized in aggregates of various sizes in both host organisms. This is the first study comparing degradation machinery in plant and insect hosts targeting all TYLCV proteins.
Moshe, A. ; Gorovits, R. ; Liu, Y. ; Czosnek, H. Tomato plant cell death induced by inhibition of HSP90 is alleviated by Tomato yellow leaf curl virus infection. Mol Plant Pathol 2016, 17, 247-60.Abstract
To ensure a successful long-term infection cycle, begomoviruses must restrain their destructive effect on host cells and prevent drastic plant responses, at least in the early stages of infection. The monopartite begomovirus Tomato yellow leaf curl virus (TYLCV) does not induce a hypersensitive response and cell death on whitefly-mediated infection of virus-susceptible tomato plants until diseased tomatoes become senescent. The way in which begomoviruses evade plant defences and interfere with cell death pathways is still poorly understood. We show that the chaperone HSP90 (heat shock protein 90) and its co-chaperone SGT1 (suppressor of the G2 allele of Skp1) are involved in the establishment of TYLCV infection. Inactivation of HSP90, as well as silencing of the Hsp90 and Sgt1 genes, leads to the accumulation of damaged ubiquitinated proteins and to a cell death phenotype. These effects are relieved under TYLCV infection. HSP90-dependent inactivation of 26S proteasome degradation and the transcriptional activation of the heat shock transcription factors HsfA2 and HsfB1 and of the downstream genes Hsp17 and Apx1/2 are suppressed in TYLCV-infected tomatoes. Following suppression of the plant stress response, TYLCV can replicate and accumulate in a permissive environment.
2015
Pakkianathan, B. C. ; Kontsedalov, S. ; Lebedev, G. ; Mahadav, A. ; Zeidan, M. ; Czosnek, H. ; Ghanim, M. Replication of Tomato Yellow Leaf Curl Virus in Its Whitefly Vector, Bemisia tabaci. J Virol 2015, 89, 9791-803.Abstract
UNLABELLED: Tomato yellow leaf curl virus (TYLCV) is a begomovirus transmitted exclusively by the whitefly Bemisia tabaci in a persistent, circulative manner. Replication of TYLCV in its vector remains controversial, and thus far, the virus has been considered to be nonpropagative. Following 8 h of acquisition on TYLCV-infected tomato plants or purified virions and then transfer to non-TYLCV-host cotton plants, the amounts of virus inside whitefly adults significantly increased (>2-fold) during the first few days and then continuously decreased, as measured by the amounts of genes on both virus DNA strands. Reported alterations in insect immune and defense responses upon virus retention led us to hypothesize a role for the immune response in suppressing virus replication. After virus acquisition, stress conditions were imposed on whiteflies, and the levels of three viral gene sequences were measured over time. When whiteflies were exposed to TYLCV and treatment with two different pesticides, the virus levels continuously increased. Upon exposure to heat stress, the virus levels gradually decreased, without any initial accumulation. Switching of whiteflies between pesticide, heat stress, and control treatments caused fluctuating increases and decreases in virus levels. Fluorescence in situ hybridization analysis confirmed these results and showed virus signals inside midgut epithelial cell nuclei. Combining the pesticide and heat treatments with virus acquisition had significant effects on fecundity. Altogether, our results demonstrate for the first time that a single-stranded DNA plant virus can replicate in its hemipteran vector. IMPORTANCE: Plant viruses in agricultural crops are of great concern worldwide. Many of them are transmitted from infected to healthy plants by insects. Persistently transmitted viruses often have a complex association with their vectors; however, most are believed not to replicate within these vectors. Such replication is important, as it contributes to the virus's spread and can impact vector biology. Tomato yellow leaf curl virus (TYLCV) is a devastating begomovirus that infects tomatoes. It is persistently transmitted by the whitefly Bemisia tabaci but is believed not to replicate in the insect. To demonstrate that TYLCV is, in fact, propagative (i.e., it replicates in its insect host), we hypothesized that insect defenses play a role in suppressing virus replication. We thus exposed whitefly to pesticide and heat stress conditions to manipulate its physiology, and we showed that under such conditions, the virus is able to replicate and significantly influence the insect's fecundity.
Moshe, A. ; Belausov, E. ; Niehl, A. ; Heinlein, M. ; Czosnek, H. ; Gorovits, R. The Tomato yellow leaf curl virus V2 protein forms aggregates depending on the cytoskeleton integrity and binds viral genomic DNA. Sci Rep 2015, 5 9967.Abstract
The spread of Tomato yellow leaf curl virus (TYLCV) was accompanied by the formation of coat protein (CP) aggregates of increasing size in the cytoplasm and nucleus of infected tomato (Solanum lycopersicum) cells. In order to better understand the TYLCV-host interaction, we investigated the properties and the subcellular accumulation pattern of the non-structural viral protein V2. CP and V2 are the only sense-oriented genes on the virus circular single-stranded DNA genome. Similar to CP, V2 localized to cytoplasmic aggregates of increasing size and as infection progressed was also found in nuclei, where it co-localized with CP. V2 was associated with viral genomic DNA molecules, suggesting that V2 functions as a DNA shuttling protein. The formation and the 26S proteasome-mediated degradation of V2 aggregates were dependent on the integrity of the actin and microtubule cytoskeleton. We propose that the cytoskeleton-dependent formation and growth of V2 aggregates play an important role during TYLCV infection, and that microtubules and actin filaments are important for the delivery of V2 to the 26S proteasome.