Seed Transmission of Tomato yellow leaf curl virus in White Soybean (Glycine max).
ABSTRACT: Tomato yellow leaf curl virus (TYLCV) infection of the common bean (Phaseolus vulgaris) has been reported, but soybean (Glycine max) has not previously been identified as a TYLCV host. Five cultivars of white soybean were agro-inoculated using an infectious TYLCV clone. At 30 days post-inoculation, they showed infection rates of 25% to 100%. Typical TYLCV symptoms were not observed in any inoculated plants. To examine whether TYLCV was transmitted in soybean seeds, DNA was isolated from bundles of five randomly selected seeds from TYLCV-inoculated soybean plants and amplified with a TYLCV-specific primer set. With the exception of one bundle, all bundles of seeds were verified to be TYLCV-infected. Virus dissemination was also confirmed in three of the 14 bunches. Viral replication was also identified in seeds and seedlings. This is the first report demonstrating that soybean is a TYLCV host, and that TYLCV is a seed-transmissible virus in white soybean.
Project description:Tomato yellow leaf curl virus (TYLCV) is one of the most well-known tomato-infecting begomoviruses and transmitted by Bemisia tabaci. Seed transmission has previously been reported for some RNA viruses, but TYLCV has not previously been described as a seed-borne virus. In 2013 and 2014, without whitefly-mediated transmission, TYLCV was detected in young tomato plants germinated from fallen fruits produced from TYLCV-infected tomato plants in the previous cultivation season. In addition, TYLCV-Israel (TYLCV-IL) was also detected in seeds and their seedlings of TYLCV-infected tomato plants that were infected by both viruliferous whitefly-mediated transmission and agro-inoculation. The seed infectivity was 20-100%, respectively, and the average transmission rate to seedlings was also 84.62% and 80.77%, respectively. TYLCV-tolerant tomatoes also produced TYLCV-infected seeds, but the amount of viral genome was less than seen in TYLCV-susceptible tomato plants. When tomato plants germinated from TYLCV-infected seeds, non-viruliferous whiteflies and healthy tomato plants were placed in an insect cage together, TYLCV was detected from whiteflies as well as receiver tomato plants six weeks later. Taken together, TYLCV-IL can be transmitted via seeds, and tomato plants germinated from TYLCV-infected seeds can be an inoculum source of TYLCV. This is the first report about TYLCV seed transmission in tomato.
Project description:BACKGROUND: Tomato yellow leaf curl virus (TYLCV) is a member of the family Geminiviridae, genus Begomovirus. To test the infectivity of TYLCV in tomato plants, an improved protocol for inoculation of in vitro-cultured tomato plants was developed. RESULTS: A TYLCV isolate was cloned, sequenced and used to construct a 1.8-mer infectious clone. Three weeks old microshoots of TYLCV-susceptible tomato plants were inoculated with Agrobacterium tumefaciens harboring the infectious clone for the TYLCV isolate. After two weeks, the TYLCV symptoms started to appear on the in vitro-inoculated plants and the symptoms became more severe and pronounced eight weeks post-inoculation. The method was used efficiently to uncover the resistance mechanism against TYLCV in Solanum habrochaites accession LA 1777, a wild tomato known for its high resistance to whitefly and TYLCV. CONCLUSIONS: The reported in vitro-inoculation method can be used to screen tomato genotypes for their responses to TYLCV under controlled conditions and it will be a useful tool for better understanding of the TYLCV biology in tomato plants.
Project description:Tomato yellow leaf curl virus (TYLCV) causes great losses in tomato production. In addition to tomato, TYLCV infects many crops or weeds as alternative hosts. These alternative hosts may serve as reservoirs for TYLCV survival and spread. Here, we tested the capability of cultivated, flue-cured tobacco to act as a reservoir host plant for TYLCV. TYLCV DNA was detected in nine flue-cured tobacco cultivars inoculated with an infectious TYLCV clone, although no visible symptoms developed on TYLCV-infected tobacco plants. The percentage of whiteflies with viral DNA increased with an increasing acquisition access period (AAP) and reached 100% after a 12 h AAP on infected tobacco plants. Using infected tobacco plants as virus resources, TYLCV was capable of being transmitted to tobacco and tomato plants by whiteflies, and typical symptoms of TYLCV infection were observed on infected tomato plants but not on infected tobacco plants. Our results suggest that flue-cured tobacco can serve as a reservoir host plant for TYLCV and may play an important role in the spread of TYLCV epidemics in China.
Project description:Since 1997 two distinct geminivirus species, Tomato yellow leaf curl Sardinia virus (TYLCSV) and Tomato yellow leaf curl virus (TYLCV), have caused a similar yellow leaf curl disease in tomato, coexisted in the fields of southern Spain, and very frequently doubly infected single plants. Tomatoes as well as experimental test plants (e.g., Nicotiana benthamiana) showed enhanced symptoms upon mixed infections under greenhouse conditions. Viral DNA accumulated to a similar extent in singly and doubly infected plants. In situ tissue hybridization showed TYLCSV and TYLCV DNAs to be confined to the phloem in both hosts, irrespective of whether they were inoculated individually or in combination. The number of infected nuclei in singly or doubly infected plants was determined by in situ hybridization of purified nuclei. The percentage of nuclei containing viral DNA (i.e., 1.4% in tomato or 6% in N. benthamiana) was the same in plants infected with either TYLCSV, TYLCV, or both. In situ hybridization of doubly infected plants, with probes that discriminate between both DNAs, revealed that at least one-fifth of infected nuclei harbored DNAs from both virus species. Such a high number of coinfected nuclei may explain why recombination between different geminivirus DNAs occurs frequently. The impact of these findings for epidemiology and for resistance breeding concerning tomato yellow leaf curl diseases is discussed.
Project description:Tomato yellow leaf curl virus (TYLCV), a monopartite begomovirus (family Geminiviridae) is responsible for heavy yield losses for tomato production around the globe. In Oman at least five distinct begomoviruses cause disease in tomato, including TYLCV. Unusually, TYLCV infections in Oman are sometimes associated with a betasatellite (Tomato leaf curl betasatellite [ToLCB]; a symptom modulating satellite). RNA interference (RNAi) can be used to develop resistance against begomoviruses at either the transcriptional or post-transcriptional levels.A hairpin RNAi (hpRNAi) construct to express double-stranded RNA homologous to sequences of the intergenic region, coat protein gene, V2 gene and replication-associated gene of Tomato yellow leaf curl virus-Oman (TYLCV-OM) was produced. Initially, transient expression of the hpRNAi construct at the site of virus inoculation was shown to reduce the number of plants developing symptoms when inoculated with either TYLCV-OM or TYLCV-OM with ToLCB-OM to Nicotiana benthamiana or tomato. Solanum lycopersicum L. cv. Pusa Ruby was transformed with the hpRNAi construct and nine confirmed transgenic lines were obtained and challenged with TYLCV-OM and ToLCB-OM by Agrobacterium-mediated inoculation. For all but one line, for which all plants remained symptomless, inoculation with TYLCV-OM led to a proportion (?25%) of tomato plants developing symptoms of infection. For inoculation with TYLCV-OM and ToLCB-OM all lines showed a proportion of plants (?45%) symptomatic. However, for all infected transgenic plants the symptoms were milder and virus titre in plants was lower than in infected non-transgenic tomato plants.These results show that RNAi can be used to develop resistance against geminiviruses in tomato. The resistance in this case is not immunity but does reduce the severity of infections and virus titer. Also, the betasatellite may compromise resistance, increasing the proportion of plants which ultimately show symptoms.
Project description:Several recent studies have reported on the role of mitogen-activated protein kinase (MAPK3) in plant immune responses. However, little is known about how MAPK3 functions in tomato (Solanum lycopersicum L.) infected with tomato yellow leaf curl virus (TYLCV). There is also uncertainty about the connection between plant MAPK3 and the salicylic acid (SA) and jasmonic acid (JA) defense-signaling pathways. The results of this study indicated that SlMAPK3 participates in the antiviral response against TYLCV. Tomato seedlings were inoculated with TYLCV to investigate the possible roles of SlMAPK1, SlMAPK2, and SlMAPK3 against this virus. Inoculation with TYLCV strongly induced the expression and the activity of all three genes. Silencing of SlMAPK1, SlMAPK2, and SlMAPK3 reduced tolerance to TYLCV, increased leaf H2O2 concentrations, and attenuated expression of defense-related genes after TYLCV infection, especially in SlMAPK3-silenced plants. Exogenous SA and methyl jasmonic acid (MeJA) both significantly induced SlMAPK3 expression in tomato leaves. Over-expression of SlMAPK3 increased the transcript levels of SA/JA-mediated defense-related genes (PR1, PR1b/SlLapA, SlPI-I, and SlPI-II) and enhanced tolerance to TYLCV. After TYLCV inoculation, the leaves of SlMAPK3 over-expressed plants compared with wild type plants showed less H2O2 accumulation and greater superoxide dismutase (SOD), peroxidase (POD), catalase (CAT), and ascorbate peroxidase (APX) activity. Overall, the results suggested that SlMAPK3 participates in the antiviral response of tomato to TYLCV, and that this process may be through either the SA or JA defense-signaling pathways.
Project description:Bemisia tabaci, the whitefly vector of Tomato yellow leaf curl virus (TYLCV), seriously reduces tomato production and quality. Here, we report the first evidence that infection by TYLCV alters the host preferences of invasive B. tabaci B (Middle East-Minor Asia 1) and Q (Mediterranean genetic group), in which TYLCV-free B. tabaci Q preferred to settle on TYLCV-infected tomato plants over healthy ones. TYLCV-free B. tabaci B, however, preferred healthy tomato plants to TYLCV-infected plants. In contrast, TYLCV-infected B. tabaci, either B or Q, did not exhibit a preference between TYLCV-infected and TYLCV-free tomato plants. Based on gas chromatography-mass spectrometry (GCMS)analysis of plant terpene volatiles, significantly more β-myrcene, thymene, β-phellandrene, caryophyllene, (+)-4-carene, and α-humulene were released from the TYLCV-free tomato plants than from the TYLCV-infected ones. The results indicate TYLCV can alter the host preferences of its vector Bemisia tabaci B and Q.
Project description:Long non-coding RNAs (lncRNAs) play an important role in regulating many biological processes. In this study, tomato seeds were first irradiated by neutrons. Eight tomato mutants were then selected and infected by Tomato yellow leaf curl virus (TYLCV). RNA sequencing followed by bioinformatics analyses identified 1,563 tomato lncRNAs. About half of the lncRNAs were derived from intergenic regions, whereas antisense lncRNAs accounted for 35%. There were fewer lncRNAs identified in our study than in other studies identifying tomato lncRNAs. Functional classification of 794 lncRNAs associated with tomato genes showed that many lncRNAs were associated with binding functions required for interactions with other molecules and localized in the cytosol and membrane. In addition, we identified 19 up-regulated and 11 down-regulated tomato lncRNAs by comparing TYLCV infected plants to non-infected plants using previously published data. Based on these results, the lncRNAs identified in this study provide important resources for characterization of tomato lncRNAs in response to TYLCV infection.
Project description:It has already been demonstrated that a betasatellite associated with cotton leaf curl Multan virus (CLCuMB) can be used as a plant and animal gene delivery vector to plants. To examine the ability of CLCuMB as a tool to transfer coat protein genes of HIV-1 to plants, two recombinant CLCuMB constructs in which the CLCuMB ?C1 ORF was replaced with two HIV-1 genes fractions including a 696 bp DNA fragment related to the HIV-1 p24 gene and a 1501 bp DNA fragment related to the HIV-1 gag gene were constructed. Gag is the HIV-1 coat protein gene and p24 is a component of the particle capsid. Gag and p24 are used for vaccine production. Recombinant constructs were inoculated to Nicotiana glutinosa and N. benthamiana plants in the presence of an Iranian isolate of Tomato yellow leaf curl virus (TYLCV-[Ab]) as a helper virus. PCR analysis of inoculated plants indicated that p24 gene was successfully replicated in inoculated plants, but the gag gene was not. Real-time PCR and ELISA analysis of N. glutinosa and N. benthamiana plants containing the replicative forms of recombinant construct of CLCuMB/p24 indicated that p24 was expressed in these plants. This CLCuMB-based expression system offers the possibility of mass production of recombinant HIV-1 p24 protein in plants.
Project description:CRISPR/Cas systems confer molecular immunity against phages and conjugative plasmids in prokaryotes. Recently, CRISPR/Cas9 systems have been used to confer interference against eukaryotic viruses. Here, we engineered Nicotiana benthamiana and tomato (Solanum lycopersicum) plants with the CRISPR/Cas9 system to confer immunity against the Tomato yellow leaf curl virus (TYLCV). Targeting the TYLCV genome with Cas9-single guide RNA at the sequences encoding the coat protein (CP) or replicase (Rep) resulted in efficient virus interference, as evidenced by low accumulation of the TYLCV DNA genome in the transgenic plants. The CRISPR/Cas9-based immunity remained active across multiple generations in the N. benthamiana and tomato plants. Together, our results confirmed the efficiency of the CRISPR/Cas9 system for stable engineering of TYLCV resistance in N. benthamiana and tomato, and opens the possibilities of engineering virus resistance against single and multiple infectious viruses in other crops.