Yellow Dwarf Virus Research Articles

Sequencing and validation of reference genes to analyze endogenous gene expression and quantify yellow dwarf viruses using RT-qPCR in viruliferous Rhopalosiphum padi.

The bird cherry-oat aphid (Rhopalosiphum padi), an important pest of cereal crops, not only directly sucks sap from plants, but also transmits a number of plant viruses, collectively the yellow dwarf viruses (YDVs). For quantifying changes in gene expression in vector aphids, reverse transcription-quantitative polymerase chain reaction (RT-qPCR) is a touchstone method, but the selection and validation of housekeeping genes (HKGs) as reference genes to normalize the expression level of endogenous genes of the vector and for exogenous genes of the virus in the aphids is critical to obtaining valid results. Such an assessment has not been done, however, for R. padi and YDVs. Here, we tested three algorithms (GeNorm, NormFinder and BestKeeper) to assess the suitability of candidate reference genes (EF-1α, ACT1, GAPDH, 18S rRNA) in 6 combinations of YDV and vector aphid morph. EF-1α and ACT1 together or in combination with GAPDH or with GAPDH and 18S rRNA could confidently be used to normalize virus titre and expression levels of endogenous genes in winged or wingless R. padi infected with Barley yellow dwarf virus isolates (BYDV)-PAV and BYDV-GAV. The use of only one reference gene, whether the most stably expressed (EF-1α) or the least stably expressed (18S rRNA), was not adequate for obtaining valid relative expression data from the RT-qPCR. Because of discrepancies among values for changes in relative expression obtained using 3 regions of the same gene, different regions of an endogenous aphid gene, including each terminus and the middle, should be analyzed at the same time with RT-qPCR. Our results highlight the necessity of choosing the best reference genes to obtain valid experimental data and provide several HKGs for relative quantification of virus titre in YDV-viruliferous aphids.

Expression Comparisons of Pathogenesis-Related (PR) Genes in Wheat in Response to Infection/Infestation by Fusarium, Yellow dwarf virus (YDV) Aphid-Transmitted and Hessian Fly

Expression profiles of ten pathogenesis-related (PR) genes during plant defense against Fusarium, Yellow dwarf virus (YDV) aphid-transmitted and Hessian fly (Hf) were compared temporally in both resistant and susceptible genotypes following pathogen infection or insect infestation. Quantitative real-time PCR (qRT-PCR) revealed that PR1, PR2, PR3, PR5, PR6, PR8, PR9, and PR15 appeared to be induced or suppressed independently in response to Fusarium, YDV aphid-transmitted or Hf during the interactions. The PR gene(s) essential to defense against one organism may play little or no role in defense against another pathogen or pest, suggesting the alternative mechanisms may be involved in different interactions of wheat-Fusarium, wheat-YDV aphid-transmitted and wheat-Hf. However, strong up- or down-regulation of PR12 and PR14 encoding low molecular membrane acting protein, defensin and lipid transfer protein (LTP), respectively, had been detected after either pathogen infection or insect infestation, therefore showed broad responses to pathogens and insects. It was postulated that low molecular proteins such as defensins and LTPs might play a role in the early stages of pathogenesis in the signaling process that informs plants about the attack from biotic stresses. In addition, a synergistic action between different PR genes might exist in plants to defense certain pathogens and insects on the basis of comprehensive expression profiling of various pathogenesis-related genes revealed by qRT-PCR in this study.

Population Growth and Damage Caused by Rhopalosiphum padi (L.) (Hemiptera, Aphididae) on Different Cultivars and Phenological Stages of Wheat.

Among the aphids associated with wheat and other winter cereals, Rhopalosiphum padi (L.) is currently the predominant species in the wheat growing region of southern Brazil. The damage caused by this aphid occurs by direct feeding and/or by the transmission of pathogenic viruses, such as the Barley/Cereal yellow dwarf virus. In order to estimate the direct damage caused by R. padi on wheat, we evaluated the population growth of this aphid during the tillering and elongation stages and its effects on grain yield components. The experiment was conducted in a screenhouse with three wheat cultivars (BRS Guabiju, BRS Timbaúva, and Embrapa 16). The effect of a period of 16days, starting from an infestation of 40 aviruliferous aphids/plant, was evaluated and compared to non-infested plants. In both stages, the population growth of R. padi was lower on the BRS Timbaúva. Although infestation caused a reduction in the grain yield of the three cultivars, this effect was lower for BRS Timbaúva. The cultivar Embrapa 16 supported higher infestations and was more tolerant to damage than the BRS Guabiju.

Psathyrostachys huashanica, a potential resource for resistance to Barley yellow dwarf virus-GAV

Barley/cereal yellow dwarf viruses (B/CYDVs) are recognized as serious pathogens of wheat. It is critical to find diverse sources of resistance to reduce yield losses caused by B/CYDVs. Psathyrostachys huashanica Keng, a perennial wheat-relative species, has been regarded as a potential germplasm resource because of its excellent performance on resistance to drought, salinity, wheat stripe rust and the take-all fungus. In this work, the susceptibility to BYDV-GAV of P. huashanica was evaluated following inoculation using reverse transcription-polymerase chain reaction (RT-PCR) and double antibody sandwich-enzyme linked immunosorbent assay (DAS-ELISA). Additionally, an aphid-transmission test was conducted for further verification of virus infection. The results indicated that BYDV-GAV could infect P. huashanica but accumulated to much lower level in inoculated leaves of P. huashanica than in the leaves of resistant wheat line Zhong 5 and the susceptible wheat line 7182. However, no BYDV-GAV particles could be detected in new leaves of inoculated P. huashanica. In this study, we first demonstrated that P. huashanica is highly resistant to BYDV-GAV and possibly has a negative effect on the movement of the viral particles from inoculated leaves to new leaves.

Virus resistance of Australian pea (Pisum sativum) varieties

Australian pea varieties were evaluated for virus resistance using spreader plots sown with Pea seed-borne mosaic virus (PSbMV)-infected seed and aphid inoculations with Bean leafroll virus (BLRV). Natural infections by Bean yellow mosaic virus (BYMV) and Soybean dwarf virus (SbDV) allowed also screening for these viruses. Complete PSbMV resistance was limited to a few varieties, but BYMV resistance was relatively frequent. No consistent ranking of partial PSbMV resistance or PSbMV seed transmission rates were found. Selection for partial resistance to PSbMV is therefore not a practical breeding objective, while incorporating complete resistance is relatively easy. A wide range of resistance was found for BLRV and confirmed in field and greenhouse tests against local BLRV strains in Syria, but infection levels varied over years and no complete resistance was identified. Genotypes with a high level of partial resistance to BLRV also appeared to show resistance to the closely related SbDV.

The complete nucleotide sequence of the genome of Barley yellow dwarf virus-RMV reveals it to be a new Polerovirus distantly related to other yellow dwarf viruses.

The yellow dwarf viruses (YDVs) of the Luteoviridae family represent the most widespread group of cereal viruses worldwide. They include the Barley yellow dwarf viruses (BYDVs) of genus Luteovirus, the Cereal yellow dwarf viruses (CYDVs) and Wheat yellow dwarf virus (WYDV) of genus Polerovirus. All of these viruses are obligately aphid transmitted and phloem-limited. The first described YDVs (initially all called BYDV) were classified by their most efficient vector. One of these viruses, BYDV-RMV, is transmitted most efficiently by the corn leaf aphid, Rhopalosiphum maidis. Here we report the complete 5612 nucleotide sequence of the genomic RNA of a Montana isolate of BYDV-RMV (isolate RMV MTFE87, Genbank accession no. KC921392). The sequence revealed that BYDV-RMV is a polerovirus, but it is quite distantly related to the CYDVs or WYDV, which are very closely related to each other. Nor is BYDV-RMV closely related to any other particular polerovirus. Depending on the gene that is compared, different poleroviruses (none of them a YDV) share the most sequence similarity to BYDV-RMV. Because of its distant relationship to other YDVs, and because it commonly infects maize via its vector, R. maidis, we propose that BYDV-RMV be renamed Maize yellow dwarf virus-RMV (MYDV-RMV).

EURALLIVEG: ESTABLISHMENT OF A EUROPEAN CORE COLLECTION BY CRYOPRESERVATION AND VIRUS ELIMINATION IN GARLIC

A European project named EURALLIVEG (Vegetative Allium, Europe’s Core Collection, safe & sound) was performed under Council Regulation EC 870/2004 from 2007 until 2011. It was coordinated by IPK. The main focus of the project was the development of a European integrated Allium Core collection provided by national collections of Germany, Czech Republic, Poland, Italy, France, and Nordic countries. This collection was designed as a part of the European Genebank Integrated System (AEGIS) of the European Cooperative Program for Plant Genetic Resources (ECPGR). The project consists of five work packages: documentation, molecular screening, cryopreservation, virus elimination, and coordination. Main aim was identification of a core collection of the 200 most important garlic accessions. This collection was cryopreserved using vitrification, which is the most economic way for long term storage. A Cryobanks Network was organized by three project partners. This system ensures safe storage of material by adopting a safety duplicate strategy. The documentation of the collection used passport and characterization data, pictures and other information. In addition genetic fingerprinting with AFLP markers was conducted. In the project, the garlic accessions were introduced into meristem culture for virus elimination. Of the viruses present in garlic, Onion Yellow Dwarf (OYDV) and Leek Yellow Stripe (LYSV) Viruses are most harmful. Therefore, because of the impossibility to do all treatments and tests on all germplasm, it was decided to test the major part of the collection on OYDV and LYSV only. However, a specific part with higher priority (backbone subset) was examined for five viruses. Virus detection used ELISA testing.

The geographical distribution of Yellow dwarf viruses and their aphid vectors in Australian grasslands and wheat

This article reviews and analyzes the literature on Yellow dwarf viruses (YDVs) in Australia, examining the range of environmental and climatic factors that explain the observed geographical distribution of the virus and its vectors. BYDV-PAV, vectored mainly by the aphid Rhopalosiphum padi, is the most prevalent YDV species in wheat and grasslands across all states, except Queensland. BYDV-RMV, vectored mainly by Rhopalosiphum maidis, dominates in Queensland grasslands, with very low incidence in wheat. Queensland experiences higher rainfall and warmer temperatures than southern Australia. Across Australia disease incidence in wheat is generally low (around 10 %) and varies from year to year, with the highest incidence found on occasion in Western Australia (up to 52 %) and the lowest in Queensland (<1 %). Across Australia there is a much higher virus incidence and more variation in YDV species present in grasslands than in wheat, although in general BYDV-PAV still dominates. An overview of the differences between the YDV species in terms of symptoms, impacts, frequency, transmission rates and geographical distribution is necessary to appreciate the implications of virus spread across Australia, as well as the risks from the interaction of YDV with more recently introduced wheat pathogens. This overview is set in the context of a changing climate, with a discussion of the possible implications of anthropogenic climate change for future epidemics. For example, increasing temperatures in the future may result in more rapid transmission of the virus in the cooler months than at present, with implications for winter crops such as wheat, where YDV currently does most damage. Also, there is potential for the spread of BYDV-RMV further south, as changes in climatic conditions alter both the transmission potential of the virus as well as the vectoring potential by the aphids R. padi and R. maidis. Finally, critical knowledge gaps are identified, highlighting a need for ongoing seasonal monitoring of the virus and vectors to support the use of simulation models to predict the incidence of YDVs in near real-time.

Agrobacterium-mediated infection of whole plants by yellow dwarf viruses

Barley yellow dwarf virus-PAV (BYDV-PAV) and cereal yellow dwarf virus-RPV (CYDV-RPV) are only transmitted between host plants by aphid vectors and not by mechanical transmission. This presents a severe limitation for the use of a reverse genetics approach to analyze the effects of mutations in these viruses on plant infection and aphid transmission. Here we describe the use of agroinfection to infect plants with BYDV-PAV and CYDV-RPV. The cDNAs corresponding to the complete RNA genomes of BYDV-PAV and CYDV-RPV were cloned into a binary vector under the control of the cauliflower mosaic virus 35S promoter and the nopaline synthase transcription termination signal. The self-cleaving ribozyme from hepatitis virus D was included to produce a transcript in planta with a 3′ terminus identical to the natural viral RNA. ELISA and RT-PCR analysis showed that the replicons of BYDV-PAV and CYDV-RPV introduced by Agrobacterium into Nicotiana benthamiana and N. clevelandii gave rise to a local infection in the infiltrated mesophyll cells. After several weeks systemic infection of phloem tissue was detected, although no systemic symptoms were observed. Three heterologous virus silencing suppressors increased the efficiency of agroinfection and accumulation of BYDV-PAV and CYDV-RPV in the two Nicotiana species. The progeny viruses purified from infiltrated tissues were successfully transmitted to oat plants by aphids, and typical yellow dwarf symptoms were observed. This study reports the first agroinfection of eudicot plants using BYDV-PAV and CYDV-RPV.

Biomarker discovery from the top down: Protein biomarkers for efficient virus transmission by insects (Homoptera: Aphididae) discovered by coupling genetics and 2‐D DIGE

Yellow dwarf viruses cause the most economically important virus diseases of cereal crops worldwide and are vectored by aphids. The identification of vector proteins mediating virus transmission is critical to develop sustainable virus management practices and to understand viral strategies for circulative movement in all insect vectors. Previously, we applied 2-D DIGE to an aphid filial generation 2 population to identify proteins correlated with the transmission phenotype that were stably inherited and expressed in the absence of the virus. In the present study, we examined the expression of the DIGE candidates in previously unstudied, field-collected aphid populations. We hypothesized that the expression of proteins involved in virus transmission could be clinically validated in unrelated, virus transmission-competent, field-collected aphid populations. All putative biomarkers were expressed in the field-collected biotypes, and the expression of nine of these aligned with the virus transmission-competent phenotype. The strong conservation of the expression of the biomarkers in multiple field-collected populations facilitates new and testable hypotheses concerning the genetics and biochemistry of virus transmission. Integration of these biomarkers into current aphid-scouting methodologies will enable rational strategies for vector control aimed at judicious use and development of precision pest control methods that reduce plant virus infection.

Demonstrating an integrated pest management strategy in autumnsown wheat to arable farmers

Six crops of autumnsown wheat managed under integrated pest management (IPM) were compared to six adjacent crops grown under the participating farmers current pest management practices in demonstration trials in Canterbury during the 200809 and 200910 seasons Farmer training with a focus on monitoring and beneficial predator identification was carried out The presence and abundance of key pests and diseases (slugs aphids yellow dwarf virus (YDV)) and beneficial predators was determined Carabid beetles and other beneficial insects capable of contributing to pest control were present in the arable cropping systems There was an increasing trend in the number of beneficial organisms a reduction in pests and a 50 reduction in the number of insecticides applied in the IPM managed crops There were negligible YDV and crop yield differences between the two approaches IPM adoption at these farms was very high after these participatory trials

Maize Debris Increases Barley Yellow Dwarf Virus Severity in North Carolina Winter Wheat

In the eastern United States, wheat (Triticum aestivum L.) is often planted with minimal or no tillage into maize (Zea mays L.) residues. We conducted a field experiment in the North Carolina Piedmont to compare the effects of three maize residue treatments (unchopped, chopped, and removed) on Fusarium head blight (FHB) in two winter wheat cultivars. While FHB levels were too low for meaningful comparisons, severe epidemics of barley/cereal yellow dwarf virus (YDV) did develop in 2 yr out of 3. In those 2 yr, YDV symptoms of discoloration and stunting were greater (P ≤ 0.001), and yield was lower (P ≤ 0.01), in plots with maize residue than in plots without maize residue. In the third year, when planting was late because of a severe fall drought, no YDV epidemic developed, and there were no differences in wheat yield due to maize residue treatment (P = 0.25). In the first 2 yr, leaf samples from all plots were assayed for viruses using a multiplexed reverse transcription polymerase chain reaction (RT‐PCR) method. The most common YDV serotypes were MAV, PAV, and RPV, which were each detected in at least 46 and 74% of samples in the 2 yr, respectively. Our finding of greater YDV severity in association with surface residue is consistent with the reported aphid preference for high‐intensity yellow colors, which we hypothesize attracted aphids preferentially to residue‐covered plots in the fall. Our results support a recommendation of seed or seedling insecticide treatment when planting wheat into heavy unincorporated maize residue in the U.S. Piedmont.

Conditions surrounding the outbreak of yellow dwarf virus in autumn/wintersown cereals in Canterbury during 2005

To understand the factors contributing to a severe outbreak of yellow dwarf virus (YDV) in autumn/wintersown wheat in Canterbury in 2005 records of winged cerealinhabiting yellow dwarf virustransmitting aphids caught in the 75 m Lincoln suction trap and the degree day accumulations of ambient temperatures at Lincoln (from 1982 to 2006) were examined as measures of virus infection resulting from primary and secondary aphid infestation respectively The results strongly suggest that infestation by winged aphids into cereal crops was higher than usual throughout winter in 2005 The unusually warm May to August (especially July and August) 2005 enabled a greater than usual aphid buildup (ie through increased development and reproduction) in the crop at this time spreading YDV The unusually high numbers of flying aphids trapped in late spring were the result of additional aphid buildup in cereal crops in September and October The results emphasise the need to include measures of secondary aphid infestation and virus infection in future YDV forecasts

Barley yellow dwarf virus and Cereal yellow dwarf virus Quantification by Real-Time Polymerase Chain Reaction in Resistant and Susceptible Plants

ABSTRACT Reliable detection and quantification of barley and cereal yellow dwarf viruses (YDVs) is a critical component in managing yellow dwarf diseases in small grain cereal crops. The method currently used is enzyme-linked immunosorbent assay (ELISA), using antisera against the coat proteins that are specific for each of the various YDVs. Recently, quantitative real-time reverse-transcription polymerase chain reaction (Q-RT-PCR) has been used to detect bacterial and viral pathogens and to study gene expression. We applied this technique to detect and quantify YDVs using primers specific for Barley yellow dwarf virus-PAV (BYDV-PAV) and Cereal yellow dwarf virus-RPV (CYDV-RPV) coat protein genes because of the higher sensitivity of RT-PCR and the advantage of using a real-time PCR instrument. This Q-RT-PCR was used to detect BYDV and CYDV, and to examine disease development in a resistant wheatgrass, a resistant wheat line, a susceptible wheat line, and a susceptible oat line. BYDV-PAV and CYDV-RPV were detected as early as 2 and 6 h, respectively, in susceptible oat compared with detection by ELISA at 4 and 10 days postinoculation. BYDV-PAV RNA accumulated more rapidly and to a higher level than CYDV-RPV RNA in both oat and wheat, which may account for PAV being more prevalent and causing more severe viral disease than CYDV. Q-RT-PCR is reproducible, sensitive, and has the potential to be used for examining yellow dwarf disease and as a rapid diagnostic tool for YDVs.

Farmer survey of yellow dwarf viruses in autumnsown cereals in Canterbury

A postal survey of 700 Canterbury arable farmers was undertaken during August/September 1998 to ascertain the importance of yellow dwarf viruses (YDV) in autumn sown cereal crops The survey also sought to document farmers practices for YDV management and the use to which they put the aphid monitoring and virus forecasting system provided by Crop and Food Research as well as their opinions on its effectiveness The response rate was 24 Most farmers had experienced YDV although for many it was not a recent problem Many farmers employed more than one strategy for YDV management the most common being to sow late or apply preventative insecticides to control aphid vectors of these viruses The monitoring information had been seen by over 80 of respondents and was judged as a good or satisfactory use of levy funds by over 92 Information from the aphid monitoring virus forecasting system was used as an aid in the decision making process for YDV management

Measures of spatial heterogeneity for species occurrence or disease incidence with finite‐counts

We propose two types of indices with finite‐count correction to measure the spatial heterogeneity of binary characteristics of organisms, such as occurrence or non‐occurrence of organisms and infected or non‐infected plants. We consider the following two examples: plant occurrence in a grassland community, and yellow dwarf disease infection in a rice field. For the grassland community, N quadrats comprising n cells of equal area, were set at random sites in a grassland, and the occurrence of a given species A in each of n cells was recorded. For disease infection, N quadrats, each consisting of n rice plants, were set at random sites in a paddy field, and the number of plants infected with yellow dwarf virus in each quadrat were counted. In these examples, since the number of cells in a quadrat is finite, neither occurrence nor incidence increase infinitely, unlike the number of aphids on a maize leaf. The first category of index belongs to the mean : variance ratio type. The estimated index value for occurrence (or incidence) is the same as that for non‐occurrence (or non‐incidence). The second category belongs to the k‐type of a negative binomial distribution. If some random plants die or recover from the disease then the expected value of the second type of index does not change. For n→∞, the current indices approach the mean : variance ratio and the inverse of k in a negative binomial distribution, respectively. This indicates that these indices are suitable for binary data sets.

The cytogenetics of a Triticum turgidum x Psathyrostachys juncea hybrid and its backcross derivatives

Psathyrostachys juncea (2n = 2x = 14, NN), a source of barley yellow dwarf (BYDV) virus resistance with tolerance to drought and salinity, has been successfully hybridized in its autotetraploid form (2n = 4x = 28, NNNN) as the pollen parent to durum wheat (Triticum turgidum L.). The 2n = 4x = 28 (ABNN) F1 hybrid has a mean meiotic metaphase-I configuration of 20.29 univalents + 0.29 ring bivalents + 3.36 rod bivalents + 0.14 trivalents. Spike length, internode length, glume awn length and lemma awn length, as well as the general spike morphology of the F1 hybrid, are intermediate with those of the two parents. Pollinating the ABNN F1 hybrid has given backcross (BC)-I derivatives of an amphiploid (AABBNN) that expresses limited self-fertility. BC-2 derivatives have been obtained from these plants. Direct transfers of useful genes from Ps. juncea to wheat would require substantial genetic manipulation strategies. Both conventional and novel methodologies, which may complement each other, and so facilitate reaching an agricultural objective end point, are addressed.

Purification and Identification of a Rhabdovirus from Thunbergia alata

A subgroup 2 rhabdovirus was isolated in south-east Queensland from black-eyed Susan (Thunbergia alata) with symptoms of vein yellowing, vein clearing and leaf distortion. Bacilliform particles accumulated in the perinuclear space of infected plants and measured 69 +/- 7 x 161 +/- 8 nm in unfixed preparations. The virus was not transmitted mechanically. Purified preparations of the Thunbergia alata rhabdovirus (TaRV) contained four major proteins with molecular weights of 80 kD, 48 kD, 40 kD and 35 kD, similar to those of datura yellow vein virus (DYVV), a newly described rhabdovirus from Australia. The 80 kD protein was identified as the viral glycoprotein. In immunoblots, the two largest proteins of TaRV reacted strongly with antiserum to DYVV, but were serologically distinct from sonchus yellow net, cereal chlorotic mottle, potato yellow dwarf and lettuce necrotic yellows viruses. TaRV is considered to be a strain of DYVV.

Effects of barley yellow dwarf and wheat spindle streak mosaic viruses on cold hardiness of cereals

Effects of barley yellow dwarf and wheat spindle streak mosaic viruses on cold hardiness of cereals

Sugar and fructosan accumulation in virus‐infected plants: rapid testing by circular‐paper chromatography

SUMMARYThe effects of virus infection on the sugar and fructosan contents of plants were studied by circular‐paper chromatography. Fructose and glucose in sugar‐beet leaves were greatly increased, and sucrose slightly, by infection with beet yellows virus (BYV): sugar content was increased more by an avirulent than by a virulent strain. Yellow‐net virus decreased the sugar concentration, whereas yellow‐net mild yellows virus increased it. Sugar accumulated in leaves of magnesium‐deficient plants, which also showed symptoms resembling virus yellows.Carrot motley dwarf virus increased fructose, glucose and sucrose concentrations in carrot leaves, but in petioles and roots sucrose increased while fructose and glucose decreased: more sugar accumulated in plants infected late than early. Applying menazon to the seed to control aphids increased the sucrose concentration of the roots.Cereal yellow dwarf virus (CYDV) increased fructose, glucose, sucrose and four fructosans in leaves of oats, an avirulent strain more than a virulent one. Virulent strains of BYV and CYDV apparently decrease photosynthesis more than avirulent ones.