Wheat Dwarf Virus Research Articles

Agroinfection of wheat: Inoculation of in vitro grown seedlings and embryos

Various inoculation procedures of in vitro grown wheat seedlings with Agrobacterium tumefaciens carrying a wheat dwarf virus (WDV) dimer led to 13–46% agroinfection. Deliberate wounding of seedlings during or before inoculation gave higher agroinfection rates than in seedlings without wounding. Seedlings from 1–4 days old were equally receptive to inoculations in the meristematic regions. Vacuum infiltration of the Agrobacterium inoculum into excised cultured wheat embryos gave up to 35% agroinfection (with 4 ωg 1 −1 acetosyringone). WDV was detectable in seedlings by the ELISA assay 4 days after inoculation and there was good agreement between the ELISA and DNA dot blot assays of virus infection. Agrobacterium was found to deliver DNA (WDV) to the three wheat varieties tested and to Aegilops speltoides, Triticum monococcum and T. durum.

Replication of wheat dwarf virus DNA in protoplasts and analysis of coat protein mutants in protoplasts and plants

The replication of wheat dwarf virus (WDV) in protoplasts derived from a Triticum monococcum suspension cell system was investigated. The production of circular viral double-stranded DNA (dsDNA) forms consistent with the replication of the viral genome was observed. In comparison to whole plants, the production of viral single-stranded DNA (ssDNA) was reduced, possibly due to only low levels of viral coat protein being produced in the protoplasts. Mutations introduced into the viral coat protein open reading frame (ORF) did not affect the ability of the viral DNA to replicate, and a deletion of ca. 400 bp was tolerated. However, these mutations abolished the infectivity of the viral genome when agroinoculated onto wheat plants, providing evidence that, contrary to the case for the bipartite geminiviruses, the coat protein is essential for infection by WDV.

Agroinfection of wheat: A comparison of Agrobacterium strains

Fifteen strains of Agrobacterium were investigated for their ability to agroinfect wheat dwarf virus into 4-day-old wheat seedlings. Viral infection was determined by ELISA. The greatest proportionof plants infected by any strain was 55%, achieved by the Agrobacterium rhizogenes strain Ar2629. In general, A. rhizogenes strains were found to give a higher proportion of agroinfected plants than A. tumefaciens strains. Nopaline types of A. tumefaciens were superior to octopine types in their ability to transfer DNA to wheat plants, but some strains ofthe latter type did infect to a low frequency. Seedling size at inoculation had no effect on the number of plants that developed a viral infection, although seedlings with shoots less than 1.0 cm long were not likely to survive damage caused by the inoculation procedure. The experiment was replicated over time and no significant variation was found between the three sets of data obtained.

Uptake and transient expression of chimeric genes in seed-derived embryos.

Uptake of DNA in dry and viable embryos of wheat by imbibition in DNA solution was detected by monitoring the transient expression of chimeric genes. Gene expression vectors used in this study contained a neomycin phosphotransferase (NPT) II reporter gene fused to various promoters. Some of the chimeric "neo" genes were shown to yield reproducibly NPT II activity in germinating embryos. This NPT II activity was increased markedly when the neo genes were carried by a vector capable of autonomous replication. Dimers of wheat dwarf virus, a monopartite gemini virus, were thus shown to be effective in amplifying the transient expressed NPT II activity in embryos of several cereals. These and other observations indicate that the observed transient expression really results from DNA uptake and expression in plant embryo cells and is not due to contaminating microorganisms.

A single wheatgrass chromosome reduces the concentration of barley yellow dwarf virus in wheat

SUMMARYSeedlings of a series of addition or substitution lines of wheat containing different Thinopyrum intermedium chromosomes were inoculated with the PAV and RPV serotypes of barley yellow dwarf virus (BYDV). Reduced virus titres in infected plants were ascribed to a single pair of homoeologous group 7 chromosomes from Th. intermedium in the disomic addition lines L1 and TAF 2. The group 7 chromosome is associated with red pigmentation of coleoptiles, which was also observed in two lines ditelosomic for the α arm of the chromosome. However, when infected with the PAV serotype of BYDV, the ditelosomic lines had normal virus titres and it is concluded that potential determinants of BYDV resistance are located on the β arm of the Group 7 chromosome.

Nucleotide sequence of the geminivirus chloris striate mosaic virus

The genome of chloris striate mosaic virus (CSMV) comprises a single circular DNA as determined by analyses on virion single-stranded (ss) DNA and virus-specific covalently closed circular (ccc) DNA isolated from infected plants. The nucleotide sequence of CSMV DNA was determined from cccDNA and the data were accommodated into one DNA circle of 2750 nucleotides. Comparison of the nucleotide sequence with those of maize streak virus (MSV), wheat dwarf virus (WDV), and digitaria streak virus (DSV) showed 49, 47, and 48% DNA homology, respectively. The sequence has four potential open reading frames for proteins of >10,000 mol wt, two in the viral (+) sense and two in the complementary (−) sense. Three of these potential coding regions have homologous counterparts, by comparison of the amino acid sequences, among the open reading frames reported for MSV, WDV, and DSV. CSMV encapasidates primer molecules able to prime the synthesis in vitro of a complementary strand to virion DNA, initiating this reaction at one site on the genome. The CSMV primer comprising approximately 88 nucleotides was located within the smaller of two intergenic or noncoding regions.

Characterization of a Subgenomic DNA Isolated from Triticum Aestivum Plants Infected with Wheat Dwarf Virus

Summary A subgenomic DNA has been isolated from wheat tissue infected with a Swedish isolate of wheat dwarf virus and cloned. Restriction endonuclease analysis and nucleotide sequence determination indicated that the subgenomic DNA (1472 nucleotides) was derived from the genomic DNA (2749 nucleotides) by two separate deletions. The subgenomic DNA had lost open reading frames (ORFs) encoding the virus coat protein and a putative protein of M r 10146, but retained an ORF and an open reading region encoding putative proteins of M r 30156 and 17292, respectively, and two structural features thought to be important for virus DNA replication i.e. a potentially stable stem-loop structure containing the conserved TAATATTAC sequence and a putative primer initiation site.

Priming of Complementary DNA Synthesis in Vitro by Small DNA Molecules Tightly Bound to Virion DNA of Wheat Dwarf Virus

Summary DNA isolated from purified preparations of wheat dwarf virus (WDV) has been shown to contain tightly bound small DNA molecules which can act as primers for the synthesis of full-length complementary DNA in vitro. The small DNA molecules are bound in the terminating intergenic region of the WDV genome between the end of an open reading frame encoding a putative protein of M r 17292 and a conserved ‘A-T’ box containing putative transcriptional polyadenylation signals. Evidence that the small DNA molecules contain ribonucleotides at their 5′ termini is presented and their possible role in the priming of virus DNA synthesis in vivo is discussed.

Agroinfection of Triticum aestivum with Cloned DNA of Wheat Dwarf Virus

Summary A head-to-tail dimer of cloned DNA from a Swedish isolate of wheat dwarf virus (WDV) was integrated between the T-DNA border sequences of a broad host range binary vector and transferred into cells of wheat seedlings using an Agrobacterium-mediated delivery system. Two-thirds of the inoculated plants developed a systemic infection. Extracts of infected leaves were shown to contain the virus double-stranded (supercoiled, open circular and linear) and single-stranded DNA forms of unit genome length and the virus capsid polypeptide. The results demonstrate the infectivity of a previously sequenced clone of WDV DNA.

Detection of a non-structural protein of M r 11 000 encoded by the virion DNA of maize streak virus

A polypeptide of approximately 11 000 daltons (11 kDa protein) encoded by an open reading frame (10.9 ORF) from the virion sense of maize streak virus (MSV) DNA has been detected among the products of in vitro translation reactions programmed with RNA from infected maize plants and also in total protein extracts from infected leaves. The 11 kDa protein has not been detected in virions and is therefore proposed to have a nonstructural role.Viral DNA with an additional in-frame translation stop codon in the 10.9 ORF was not infectious when transmitted to maize plants via Agrobacterium tumefaciens "agroinfection", suggesting that the 10.9 ORF may be essential for virus function. Computer comparison data show that equivalent ORFs in wheat dwarf virus (WDV) and digitaria streak virus (DSV) have some sequences in common with the 10.9 ORF of MSV. Further-more, the absence of similar sequences in geminiviruses which infect dicotyledonous plants suggests that the 11 kDa protein and its putative homologs in WDV and DSV have a function necessary only for those geminiviruses which infect the Gramineae.The significance of the 11 kDa protein in relation to expression of the virion sense DNA of MSV is discussed.

Agroinfection and nucleotide sequence of cloned wheat dwarf virus DNA

Cloned DNA of the geminivirus wheat dwarf virus (WDV) was successfully used to infect seedling wheat plants. The clone was derived from circular double-stranded viral DNA isolated from naturally infected tissue. The initiation of infection was mediated by Agrobacterium tumefaciens using cloned dimeric WDV genomes in a binary Agrobacterium vector. The WDV DNA which comprised the infectious clone was sequenced and is compared with the published sequence of a Swedish isolate of the same virus. The results confirm that the single WDV genome component of 2.75 kb carries all the information necessary for production of viral symptoms, virus particles and viral double- and single-stranded DNA forms.

Infectivity and complete nucleotide sequence of the genome of a South African isolate of maize streak virus.

The complete infectious genome of a South African isolate of the geminivirus maize streak (MSV-S) has been cloned, characterized, and sequenced. Using an A. tumefaciens Ti plasmid delivery system, the cloned -2.7 kb single circular MSV component was shown to be necessary and sufficient for infection of maize. Based on sequence analysis of the infectious clone, MSV-S is highly homologous to the previously characterized Kenyan and Nigerian isolates. While the genomic organization of MSV-S has elements in common with each of these previously characterized isolates, it is identical to neither and its analysis addresses the discrepancies between them. The result is a somewhat simplified and unified picture of the viral genome, the structural organization of which is essentially identical to that of wheat dwarf virus.

The nucleotide sequence of a geminivirus from Digitaria sanguinalis

The encapsidated single-stranded circular DNA of a geminivirus isolated from Digitaria sanguinalis has been sequenced. The data obtained are consistent with there being one DNA circle of 2701 nucleotides. Comparison of the nucleotide sewuence with those of maize streak virus (MSV) and wheat dwarf virus showed 64 and 47% DNA homology, respectively. The sequence has four potential coding regions for proteins of >10 kDa, two in the viral (+) sense and two in the complementary (−) sense. Each of these potential coding regions has a highly homologous counterpart among the seven open reading frames previously described for MSV. Virion DNA contained, in addition to the circular single-stranded DNA, a population of small DNA molecules similar to those associated with MSV particles. A comparison with MSV DNA of the region complementary to these small DNA molecules revealed conserved sequences, which may have a role in defining the limits of these primer-like molecules.

Sequence homology between the coat proteins of DNA and RNA plant viruses

The coat proteins of the leafhopper transmitted gemini viruses, viz., maize streak virus and wheat dwarf virus that contain a single circle of DNA in their genomes are shown to bear good amino acid sequence homology with the coat protein of an RNA plant virus, the satellite tobacco necrosis virus. It is suggested that for icosahedral assemblage, the coat protein architecture transcends genomic preferences.

Spread of barley yellow dwarf virus and relative importance of local aphid vectors in central Washington

SUMMARYData from bioassays of field collected aphids, barley indicator plants exposed to natural conditions, and various types of aphid traps were used to describe the spread of barley yellow dwarf virus (BYDV) in wheat and barley near Prosser, Washington. Bioassays were also used to assess the relative importance of local vector species.Of alate aphids collected from grain in the 1982 and 1983 fall migration seasons, 3.4–14–5% transmitted BYDV. Data from concurrent and post‐migration assays of resident aphids (apterae and nymphs) reflected an increase in the proportion of infected plants in the field. Maximum increase in the percentage of viruliferous aphids occurred in late November and December of 1982 and November of 1983. The 1982 increase occurred after aphid flights had ceased for the year, suggesting active secondary spread. Collections in pitfall traps and infected trap plants from November to February confirmed aphid activity and virus spread. Rhopalosiphum padi was the most important vector in central Washington in 1982 and 1983 because of its abundance and relative BYDV transmission efficiency. Metopolophium dirhodum was more winter‐hardy than R. padi and equal to R. padi in its efficiency as a vector; however, it was not as abundant as R. padi except during the mild winter of 1982–83, when it was a major contributor to secondary spread. Sitobion avenae may be important in years when it is abundant, but it was only a quarter as efficient as R. padi. Rhopalosiphum maidis was a much less efficient vector than R. padi and it only reached high populations in late autumn barley.

The nucleotide sequence of cloned wheat dwarf virus DNA

Restriction analysis and cloning of virus-specific double-stranded DNA isolated from plants infected with wheat dwarf virus (WDV) indicated that the virus genome, like that of maize streak virus (MSV), consists of a single DNA circle. The complete nucleotide sequence of cloned WDV DNA (2749 nucleotides) has been determined. Comparison of the potential coding regions in WDV DNA with those in the DNA of two strains of MSV suggests that these viruses encode at least two functional proteins, the coat protein read in the virion (+) DNA sense and a composite protein, formed from two open reading regions, in the complementary (-) DNA sense. Although WDV and MSV are serologically unrelated their coat proteins showed 35% direct amino acid sequence and their DNAs showed 46% nucleotide sequence homology. There was too little homology between the DNAs of WDV and those of two geminiviruses with bipartite genomes, cassava latent virus (CLV) and tomato golden mosaic virus (TGMV), to align the sequences. However comparison of the amino acid sequences of predicted proteins of WDV, MSV, TGMV and CLV revealed clear relationships between these viruses and suggested that the monopartite and the bipartite geminiviruses have a common ancestral origin. Four inverted repeat sequences which have the potential to form hairpin structures of deltaG >/= -14 kcal/mol were detected in WDV DNA. The sequence TAATATTAC present in the loop of one of these hairpins is conserved in similar putative structures in MSV DNA and in both DNA components of CLV and TGMV and may function as a recognition sequence for a protein involved in virus DNA replication.

The influence of preinfection cold hardening and disease development period on the interaction between barley yellow dwarf virus and cold stress tolerance in wheat

Cold hardness and ice encasement tolerance of 'Fredrick' and 'Norstar' winter wheats as affected by infection with barley yellow dwarf virus (BYDV) were determined during inoculation, disease development periods, and low-temperature growth. Plants were either prehardened to cold, or warm grown before infection; two disease development periods (DDP) were utilized. A long DDP induced greater pathogenesis and greater hardiness reduction than a short DDP. The effect of virus infection on the final level of hardiness of prehardened plants was generally greater than on that of nonprehardened plants. Viral infection reduced hardiness up to 3.5 °C in 'Fredrick' wheat, but reductions of 6–10 °C below hardiness potential were recorded after certain environmental regimes allowing disease development. Ice tolerance was reduced by BYDV infection in early low-temperature growth but was increased by infection after 4 months at low temperature. This increase in survival was associated with higher dry matter content in infected than in noninfected plants.

Identification of barley yellow dwarf virus and cereal aphid Infestations In winter wheat by aerial photography

In 1976 distinctive patterns were identified on aerial photographs of winter wheat crops in East Anglia. The patterns consisted of numerous dark foci which were found to be associated with sooty moulds (mostly Cladosporium spp.) in the crops. Both green and mature crops were photographed in 1977 and the patterns were matched with crop inspection data. These suggested that barley yellow dwarf virus (BYDV) was the cause of the patterns and that sooty moulds secondarily invaded the infected plants. A different type of focal pattern appeared to be associated with damage caused by aphid feeding. All crops showing the pattern associated with BYDV were sown before 13 October but there was no relationship with sowing date for the crops damaged directly by aphids. Yield losses within foci were estimated by sampling and the total crop areas affected were calculated using an image analysis computer: yield losses were estimated to range from 10% (BYDV) to 2% (aphid feeding). The epidemiology of BYDV is discussed and reasons are suggested for the development and appearance of the foci.

Interaction between barley yellow dwarf virus and rust in wheat, barley and oats, and the effects on grain yield and quality

SUMMARYIn three separate experiments, the upper leaf surface of the fifth formed leaf of wheat cv. Highbury, the fourth and fifth leaves of barley cv. Julia and the third and fourth leaves of oat cv. Mostyn were inoculated in a spore settling tower with wheat brown rust (Puccinia recondita f. sp. tritici), barley brown rust (P. hordei) or oat crown rust (P. coronata f. sp. avenae), respectively. Fewer pustules developed on distal portions of leaves of plants infected with barley yellow dwarf virus (BYDV) than on similar portions of leaves from virus‐free plants. There were no significant differences in the number of pustules on proximal leaf portions. In barley and oats, the number of pustules on distal leaf portions was negatively correlated with the amount of yellowing of the leaf areas scored. In wheat, symptoms of BYDV were mild and leaves were little affected by yellowing. The latent period of rust on wheat and oats was not affected by BYDV. In barley, BYDV reduced the latent period of rust on leaf 5, but not on leaf 4, and reduced it on proximal, but not distal, leaf portions.In other experiments, BYDV reduced the yield of wheat and oats by 44% and 66%, respectively, while BYDV‐infected barley was almost sterile. The appropriate rust reduced the yield of wheat, barley and oats by 33%, 13% and 86%, respectively. When infected with both BYDV and rust, yield of wheat and oats was reduced by 63% and 91%, respectively. Neither BYDV nor rust affected the percentage crude protein content of wheat grain, nor did rust affect that of barley. In oats, BYDV and rust each significantly increased crude protein of grain, but rust infection of BYDV‐infected plants tended to reduce it.

Crop loss assessment studies on the effects of barley yellow dwarf virus in wheat in Victoria

The effect of barley yellow dwarf virus (BYDV) infection on grain yield of wheat, cv. Olympic, was determined over 3 years at three sites in Victoria in field plots inoculated with infective Rhopalosiphum padi (L.). Inoculation before tillering lowered grain yields by 9-79 %, whereas inoculation at early stem extension lowered yields by only 6-9 %. There was a linear relationship between the percentage of plants infected with BYDV at an early stage of growth and grain yield at all sites. Other components of yield affected included numbers of tillers, numbers of heads with sterile terminal spikelets, grain weight per head and weight of individual grains.