Beet Western Yellows Research Articles

Posterior midgut and hindgut are both sites of acquisition of Cucurbit aphid-borne yellows virus in Myzus persicae and Aphis gossypii.

Members of the family Luteoviridae ('luteovirids') rely strictly on aphid vectors for plant-to-plant transmission. This interaction operates according to a persistent and circulative manner, which implies that the virions are being endocytosed and exocytosed across two epithelial barriers (alimentary tract and accessory salivary glands) in the vector's body. In several luteovirid-aphid vector species combinations, the route of virions in the insect has been investigated ultrastructurally by transmission electron microscopy (TEM). Here, we used TEM to follow the route of Cucurbit aphid-borne yellows virus (CABYV; genus Polerovirus) in its two efficient vector species, Myzus persicae and Aphis gossypii. We demonstrated that CABYV particles are acquired from the gut lumen to the haemocoel through two different sites in both aphid species, i.e. the posterior midgut (as for Beet western yellows virus in M. persicae) and the hindgut (as for Barley yellow dwarf virus complex in cereal aphids). This 'dual' tissue specificity of CABYV represents an original situation among viruses in the family Luteoviridae examined so far by TEM. A variety of virion-containing structures (e.g. clathrin-coated and tubular vesicles, endosome-like bodies) are found in intestinal cells of both types in both aphids. Release of virus particles from midgut and hindgut cells into the haemolymph was confirmed by immunotrapping using CABYV-specific antibodies. In accessory salivary glands, transport of CABYV virions across the cells was similar in each aphid species, and occurred by a transcytosis mechanism involving formation of tubular and coated vesicles before release of free virions in the salivary canal.

Incidence of Viruses Infecting Winter Oilseed Rape (Brassica napus ssp. oleifera) in Iran

AbstractOilseed rape (Brassica napus ssp. oleifera) is an important crop in Iran. Of 581 oilseed rape samples tested by DAS‐ELISA and immunoelectron microscopy during a preliminary survey from 1999 to 2001, 14.45% were infected with Beet western yellows virus (BWYV, Luteovirus), 12.9% with Cauliflower mosaic virus (CaMV, Caulimovirus) and 9.3% with Turnip mosaic virus (TuMV, Potyvirus). Cucumber mosaic virus (4.6%) and Tomato spotted wilt virus (0.51%) also occurred, but to a lesser extent. The occurrence of BWYV, TuMV and CaMV in oilseed rape crops could have important consequences for rape seed production in Iran and infected plants could also serve as sources of virus for other susceptible vegetable crops.

A Survey of Viruses Affecting Faba Bean (Vicia faba) in Tunisia Includes First Record of Soybean dwarf virus.

A survey was conducted in April 2003 to identify viruses infecting faba bean (Vicia faba L.) in six regions (Beja, Bizerte, Cap-bon, Le Kef, Siliana, and Zaghouan) in Tunisia. A total of 292 faba bean samples with symptoms of viral infection (leaf rolling, yellowing, and mosaic) were collected. The samples were tested at the virology laboratory of the International Center for Agricultural Research in the Dry Areas (ICARDA), Syria, for 11 viruses using the tissue-blot immunoassay procedure (3). Specific rabbit polyclonal antisera were used to test for Chickpea chlorotic dwarf virus (CpCDV) (provided by H. J. Vetten, BBA, Braunschweig, Germany), Alfalfa mosaic virus (AMV), Bean yellow mosaic virus (BYMV), Broad bean mottle virus (BBMV), Broad bean stain virus (BBSV), Cucumber mosaic virus (CMV), and Pea seedborne mosaic virus (PSbMV) (ICARDA, Aleppo, Syria). In addition, four specific monoclonal antibodies were used to detect Bean leaf roll virus (BLRV) (4B10) (2), Beet western yellows virus (BWYV) (ATCC PVAS-647; American Type Culture Collection, Manassas, VA), Faba bean necrotic yellows virus (FBNYV) (3-2E9) (1), and Soybean dwarf virus (SbDV) (ATCC PVAS-650). Serological tests showed that BBMV, a beetle-transmitted and seedborne virus identified in 23.3% (68 samples) of the samples tested, was the most common. BLRV, FBNYV, BWYV, BYMV, SbDV, and PSbMV were detected in 56, 33, 31, 10, 5, and 1 sample(s) of 292 samples tested, respectively. AMV, BBSV, CMV, and CpCDV were not detected in any samples tested. In Tunisia, BLRV, BWYV, BYMV, FBNYV, and PSbMV have previously been reported in faba bean (4), but to our knowledge, this is the first record of SbDV affecting faba bean in Tunisia, where it was detected in two fields in the Cap-bon Region. In sodium dodecyl sulfate-polyacrylamide gel electrophoresis followed by western blots, extracts from SbDV-infected plants were observed to contain 23-kDa structural proteins, which reacted strongly with SbDV monoclonal antibodies. Transmission tests showed that the samples, which reacted with SbDV monoclonal antibodies, were transmitted to faba bean plants by the pea aphid (Acyrthosiphon pisum Harris) in a persistent manner. To our knowledge, this is the first report of SbDV naturally infecting faba bean in Tunisia and it could cause a serious problem to other leguminous crops grown in Tunisia, such as French bean and peas, which are hosts for the virus.

First report of Faba bean necrotic yellows virus (FBNYV) infecting chickpea (Cicer arietinum) and faba bean (Vicia faba) crops in Sudan

A total of 45 faba bean samples ( Vicia faba ) and 130 chickpea samples ( Cicer arietinum ) were collected from plants with symptoms that suggested a viral infection (leaf rolling, yellowing and stunting) during the period 1999– 2001 at the Shambat and Hudeiba Research Stations, Sudan. These samples were tested, using the tissue-blot immunoassay procedure (Makkouk & Comeau, 1994), at the Virology Laboratory of ICARDA, Syria. The antisera used included both monoclonal and polyclonal antibodies. The monoclonal antibodies were ATCC PVAS647 (American Type Culture Collection, Manassas, VA, USA), used to test for Beet western yellows virus (BWYV, genus Polerovirus , family Luteoviridae ); ATCC PVAS650, used to test for Soybean dwarf virus (SbDV, family Luteoviridae ); 4B10, used to test for Bean leaf roll virus (BLRV, family Luteoviridae ; Katul, 1992); and 3-2E9 and 2-5H9, used to test for Faba bean necrotic yellows virus (FBNYV, genus Nanovirus , family Nanoviridae ) – the latter monoclonal is FBNYV-specific (Franz et al ., 1996). A polyclonal antibody (provided by H.J. Vetten, BBA, Braunschweig, Germany) was used to test for Chickpea chlorotic dwarf virus (CpCDV, genus Mastrevirus , family Geminiviridae ). Laboratory tests showed that CpCDV, a leafhopper-transmitted virus, was the most abundant virus in chickpea; however, FBNYV, an aphid-transmitted virus, was the most common virus in faba bean samples. FBNYV has been reported to infect food legumes in many countries in western Asia and North Africa, and reaches epidemic levels in some countries, including Egypt (Makkouk et al ., 1994). CpCDV was detected in 77·0% of chickpea and 15·5% of faba bean samples tested, whereas FBNYV was detected in 95·5% of faba bean and 6·2% of chickpea samples tested. Only two chickpea samples tested positive for BWYV, while SbDV and BLRV were not detected in any of the samples. Based on the reaction with the FBNYV-specific monoclonal (2-5H9) and the characteristic symptoms of leaf rolling, stunting and yellowing of leaves, with necrosis of the leaf periphery in faba bean and yellowing and stunting in chickpea, this is the first record of FBNYV affecting faba bean and chickpea in Sudan.

First record of Chickpea chlorotic dwarf virus (CpCDV) affecting faba bean (Vicia faba) crops in Egypt

A field visit was conducted in March 2002 to identify viruses infecting the faba bean ( Vicia faba ) crop in four governorates (Fayoum, Beni Suef, El-Nobareia and Bihera) in Egypt. A total of 71 faba bean samples with symptoms of viral infection (leaf rolling, yellowing and mosaic) were collected. These were tested at the Virology Laboratory of ICARDA, Syria, for seven viruses using the tissue-blot immunoassay procedure (Makkouk & Comeau, 1994). Specific rabbit polyclonal antisera were used to test for Chickpea chlorotic dwarf virus (CpCDV, genus Mastrevirus , family Geminiviridae ; provided by H. J. Vetten, BBA, Braunschweig, Germany); for Broad bean stain virus (BBSV, genus Comovirus , family Comoviridae ); for Bean yellow mosaic virus (BYMV, genus Potyvirus , family Potyviridae ) and for Pea seed borne mosaic virus (PSbMV, genus Potyvirus , family Potyviridae ). In addition, three specific monoclonal antibodies were used to detect Faba bean necrotic yellows virus (FBNYV, genus Nanovirus , family Nanoviridae ) (Franz et al ., 1996); Bean leaf roll virus (BLRV, family Luteoviridae ) (4B10; Katul, 1992) and Beet western yellows virus (BWYV, genus Polerovirus , family Luteoviridae ) (ATCC PVAS-647; American Type Culture Collection, Manassas, VA, USA). Serological tests showed that BYMV, an aphid-transmitted and seedborne virus that was identified in 89% of samples tested, was the most common virus. In most of the fields surveyed, BYMV symptoms were noted to occur at high levels (80–100% infection). PSbMV was detected in nine samples (one from Fayoum and eight from Beni Suef); FBNYV was detected in 11 samples (all from Beni Suef). CpCDV, a leafhopper-transmitted virus, was, by contrast, detected in only two samples collected from the El-Nobareia governorate showing leaf rolling, yellowing and stunting. BWYV, BLRV and BBSV were not detected in any of the samples tested. In Egypt, FBNYV, BYMV and PSbMV have previously been reported in faba bean (Makkouk et al ., 1994), but this is the first record of CpCDV affecting faba bean in Egypt. CpCDV is the only member of the Geminivirus that is reported to naturally infect faba bean.

Effects of point mutations in the major capsid protein of beet western yellows virus on capsid formation, virus accumulation, and aphid transmission.

Point mutations were introduced into the major capsid protein (P3) of cloned infectious cDNA of the polerovirus beet western yellows virus (BWYV) by manipulation of cloned infectious cDNA. Seven mutations targeted sites on the S domain predicted to lie on the capsid surface. An eighth mutation eliminated two arginine residues in the R domain, which is thought to extend into the capsid interior. The effects of the mutations on virus capsid formation, virus accumulation in protoplasts and plants, and aphid transmission were tested. All of the mutants replicated in protoplasts. The S-domain mutant W166R failed to protect viral RNA from RNase attack, suggesting that this particular mutation interfered with stable capsid formation. The R-domain mutant R7A/R8A protected approximately 90% of the viral RNA strand from RNase, suggesting that lower positive-charge density in the mutant capsid interior interfered with stable packaging of the complete strand into virions. Neither of these mutants systemically infected plants. The six remaining mutants properly packaged viral RNA and could invade Nicotiana clevelandii systemically following agroinfection. Mutant Q121E/N122D was poorly transmitted by aphids, implicating one or both targeted residues in virus-vector interactions. Successful transmission of mutant D172N was accompanied either by reversion to the wild type or by appearance of a second-site mutation, N137D. This finding indicates that D172 is also important for transmission but that the D172N transmission defect can be compensated for by a "reverse" substitution at another site. The results have been used to evaluate possible structural models for the BWYV capsid.

Incidence of three viruses in vegetable brassica plantings and associated wild radish weeds in south-west Australia

During the spring and summer of 2001, surveys of 50 plantings of vegetable brassicas (broccoli, cabbage and cauliflower) and associated wild radish (Raphanus raphanistrum) weeds were done to determine the occurrence of three aphid-borne viruses, Beet western yellows virus (BWYV), Cauliflower mosaic virus (CaMV) and Turnip mosaic virus (TuMV). Plantings at 15 widely separated farms were sampled in the Perth metropolitan and Manjimup regions of south-west Australia. None of the three viruses was detected in vegetable brassica plantings at Manjimup and neither were CaMV or TuMV detected in the Perth metropolitan region. BWYV was only detected at a low incidence (2% infection) in one planting of cabbage at Carabooda. In contrast, populations of wild radish from one farm in Perth and five out of six farms at Manjimup were infected with BWYV at incidences up to 65%. One population of wild radish was also infected with TuMV at an incidence of 2%. Additional keywords : Cauliflower mosaic virus, Turnip mosaic virus, Beet western yellows virus, surveys, virus reservoirs, control.

The incidence of viruses in wild Brassica rapa ssp. sylvestris in southern England

SummaryUsing enzyme‐linked immunosorbent assays, the frequency of occurrence of six viruses was determined in Brassica rapa ssp. sylvestris collected from two Thameside sites (Abingdon and Culham) in Oxfordshire and one near the Avon (Claverton) in Bath & North East Somerset. During 2000–2001, the viruses detected were: Beet western yellows virus (genus Polerovirus) (BWYV), Cauliflower mosaic virus (genus Caulimovirus) (CaMV), Turnip crinkle virus (genus Carmovirus) (TCV), Turnip rosette virus (genus Sobemovirus) (TRoV), and Turnip yellow mosaic virus (genus Tymovirus) (TYMV). BWYV and TYMV were the most frequently detected viruses at the Oxford shire sites, both as single infections (20/1743 and 66/1743 respectively) and as dual infections (7/1743). Turnip mosaic virus (genus Potyvirus) (TuMV) was not detected in the field‐grown plants assayed from any of the sites. There was a highly significant (x2[1]=30.07, P<0.001) difference in the proportion of plants at each Oxfordshire site in which one or more viruses were detected, and essentially the same pattern of virus infection was observed in tests on B. rapa from the site near Claverton. At least one representative isolate of each detected virus was tested for its morphological and serological effects on glasshouse‐grown individuals from different half‐sib families of B. rapa from both Oxfordshire sites. Except for TRoV, where there was a large difference in the frequency of successful infection in B. rapa from the two locations (1/15 vs 11/15), no clear evidence of resistance or immunity to challenge was observed, although tolerance (virus invasion without symptoms) was frequent. Fewer of the plants from Abingdon were infected than those from Culham, when mechanically challenged with TRoV, but the two B. rapa populations were not otherwise consistently different, either in their infectibility by this virus or in their responses to challenge. However, with TCV, viral antigen concentration was closely linked to the severity of disease and the B. rapa from both Oxfordshire sites segregated into two classes: those with symptoms and most viral antigen, and those without symptoms and least viral antigen. These results suggest that generic risk assessments cannot be made due to differences in the way distinct B. rapa populations react to virus challenge.

Solution Structure of a Luteoviral P1–P2 Frameshifting mRNA Pseudoknot

A hairpin-type messenger RNA pseudoknot from pea enation mosaic virus RNA1 (PEMV-1) regulates the efficiency of programmed −1 ribosomal frameshifting. The solution structure and 15N relaxation rates reveal that the PEMV-1 pseudoknot is a compact-folded structure composed almost entirely of RNA triple helix. A three nucleotide reverse turn in loop 1 positions a protonated cytidine, C 10, in the correct orientation to form an A ( n−1) ·C +·G-C n major groove base quadruple, like that found in the beet western yellows virus pseudoknot and the hepatitis delta virus ribozyme, despite distinct structural contexts. A novel loop 2–loop 1 A·U Hoogsteen base-pair stacks on the C 10 +·G 28 base-pair of the A 12·C 10 +·G 28-C 13 quadruple and forms a wedge between the pseudoknot stems stabilizing a bent and over-rotated global conformation. Substitution of key nucleotides that stabilize the unique conformation of the PEMV-1 pseudoknot greatly reduces ribosomal frameshifting efficacy.

Involvement ofBeet western yellows virus,Cauliflower mosaic virus, andTurnip mosaic virusin Internal Disorders of Stored White Cabbage

ABSTRACT Experiments over two growing seasons clearly showed that Turnip mosaic virus (TuMV) infection was associated with internal necrosis (sunken necrotic spots 5 to 10 mm in diameter) and Beet western yellows virus (BWYV) infection was associated with collapse of leaf tissue at the margins (tipburn) in heads of stored white cabbage (Brassica oleracea var. capitata). Virtually no tipburn was seen in cv. Polinius, whereas cv. Impala was affected severely. Internal necrotic spots were seen in both cultivars. BWYV appeared to interact with TuMV. Plants infected with both viruses showed a lower incidence of external symptoms and had less internal necrosis than plants infected with TuMV alone. Cauliflower mosaic virus (CaMV) did not induce significant amounts of internal necrosis or tipburn, but did, in most cases, exacerbate symptoms caused by TuMV and BWYV. BWYV-induced tipburn worsened significantly during storage. Post-transplanting inoculation with TuMV induced more internal necrosis than pre-transplant inoculation. There was a significant association between detection of TuMV just prior to harvest and subsequent development of internal necrotic spots. Individually, all three viruses significantly reduced the yield of cv. Polinius, whereas only BWYV and CaMV treatments reduced the yield of cv. Impala.

Virus-induced gene silencing in transgenic plants expressing the minor capsid protein of Beet western yellows virus.

Transgenic Nicotiana benthamiana expressing the minor coat protein P74 of the phloem-limited Beet western yellows virus (BWYV) exhibited an unusual spatial pattern of post-transcriptional gene silencing (PTGS) when infected with BWYV or related viruses. Following infection, transgenic P74 and its mRNA accumulated to only low levels, 21 to 23 nucleotide RNAs homologous to the transgene appeared, and the transgene DNA underwent methylation. The infecting viral RNA, however, was not subject to significant silencing but multiplied readily and produced P74 in the phloem tissues, although the P74 encoded by the transgene disappeared from the phloem as well as the nonvascular tissues.

P0 of beet Western yellows virus is a suppressor of posttranscriptional gene silencing.

Higher plants employ a homology-dependent RNA-degradation system known as posttranscriptional gene silencing (PTGS) as a defense against virus infection. Several plant viruses are known to encode proteins that can suppress PTGS. Here we show that P0 of beet western yellows virus (BWYV) displays strong silencing suppressor activity in a transient expression assay based upon its ability to inhibit PTGS of green fluorescent protein (GFP) when expressed in agro-infiltrated leaves of Nicotiana benthamiana containing a GFP transgene. PTGS suppressor activity was also observed for the P0s of two other poleroviruses, cucurbit aphid-borne yellows virus and potato leafroll virus. P0 is encoded by the 5'-proximal gene in BWYV RNA but does not accumulate to detectable levels when expressed from the genome-length RNA during infection. The low accumulation of P0 and the resulting low PTGS suppressor activity are in part a consequence of the suboptimal translation initiation context of the P0 start codon in viral RNA, although other factors, probably related to the viral replication process, also play a role. A mutation to optimize the P0 translation initiation efficiency in BWYV RNA was not stable during virus multiplication in planta. Instead, the P0 initiation codon in the progeny was frequently replaced by a less efficient initiation codon such as ACG, GTG, or ATA, indicating that there is selection against overexpression of P0 from the viral genome.

The incidence of viruses in wild Brassica nigra in Dorset (UK)

SummaryUsing enzyme‐linked immunosorbent assays, the frequency of occurrence of six viruses was determined in Brassica nigra collected from five coastal sites in Dorset, spanning approximately 24 km. During 1998–2000, the viruses detected were: Turnip mosaic virus (genus Potyvirus) (TuMV), Turnip yellow mosaic virus (genus Tymovirus) (TYMV), Turnip crinkle virus (genus Carmovirus) (TCV), Turnip rosette virus (genus Sobemovirus) (TRoV), Beet western yellows virus (genus Polerovirus) (BWYV) and Cauliflower mosaic virus (genus Caulimovirus) (CaMV). Multiple infections were detected in some individuals (48/447). TuMV was detected infrequently over the three‐year period (5/597). A representative isolate of each virus was tested for its effects on glasshouse‐grown individuals from different half‐sib families of B. nigra from four of the sites. Whether inoculated manually or via aphids (Myzus persicae), TuMV caused a rapid (within 10 days) lethal systemic necrosis in the B. nigra seedlings except when they were near flowering at the time of inoculation. Each of the other viruses invaded systemically but were not lethal. Indeed, BWYV systemically invaded 13/19 glasshouse‐grown B. nigra seedlings but did not produce any visible symptoms. Otherwise, the isolates tested differed in their pathogenicity and in the symptoms they produced in infected B. nigra. With TYMV or TCV viral antigen concentration was closely linked to pathogenicity; for TRoV or CaMV, there was little or no difference in virus concentration between plants with and without symptoms. Substantial and reproducible differences were observed in sensitivity/susceptibility among B. nigra genotypes from different sites in Dorset challenged with the same virus isolate.

First record of Beet western yellows virus, Chickpea chlorotic dwarf virus and Faba bean necrotic yellows virus affecting faba bean (Vicia faba) crops in Iraq

A survey to identify virus diseases affecting faba bean(Vicia faba) in Iraq was conducted during April 2000.Around 4400 samples were collected from 36 faba beanfields distributed in five governorates (ten in Baghdad,nine in Al-Anbar, three in Diyala, five in At-Tamim andnine in Ninawa). These samples were tested for thepresence of 12 different viruses by tissue-blot immu-noassay (Lin et al., 1990), using a battery of polyclonaland monoclonal antibodies. Rabbit polyclonal antiseraspecific for Chickpea chlorotic dwarf virus (CCDV)(Horn et al., 1993); Broad bean true mosaic virus(BBTMV) (from H. Rohloff, BBA, Braunschweig,Germany); Broad bean wilt virus (BBWV); Pea enationmosaic virus (PEMV) (from IPO, Wageningen, theNetherlands); Alfalfa mosaic virus (AMV); Bean yellowmosaic virus (BYMV); Broad bean mottle virus(BBMV); Broad bean stain virus (BBSV); Cucumbermosaic virus (CMV); and Pea seed-borne mosaic virus(PSbMV) (from Virology Laboratory, ICARDA,Aleppo, Syria) were used, in addition to a monoclonalspecific for Faba bean necrotic yellows virus (FBNYV)(Franz et al., 1996) and a broad-spectrum legumeluteovirus monoclonal (5G4) (Katul, 1992). To identifyspecific luteoviruses, samples that tested positive usingmonoclonal 5G4 were tested against five furthermonoclonals: Beet western yellows virus (BWYV)(ATCC PVAS-647); Soybean dwarf virus (SbDV) (ATCCPVAS-650); Potato leaf roll virus (PLRV) (ATCC PVAS-649); and Bean leaf roll virus (BLRV) (4B10) and 6F9,which reacts with either BLRV or SbDV (Katul, 1992).BYMV was the most abundant virus in faba bean inIraq, identified in 68% of test samples. In 50% of thefields surveyed, BYMV was identified at extremely highlevels (80–100% infection). BWYV was detected in 135samples collected from 15 fields from all five governor-ates. Two fields in the Ninawa governorate were highlyinfected with BWYV (62 and 96%). CCDV wasdetected in eight samples collected from three fields(two in El-Anbar and one in Ninawa), and FBNYV wasdetected in seven samples collected from two fields inNinawa governorate.This is the first record of BWYV, CCDV and FBNYVaffecting faba bean in Iraq. Other viruses such as AMV,BBWVand CMV were detected in fewer than 1% of thesamples tested. BBMV, BBSV, BBTMV, PEMV andPSbMV were not detected in any of the samples.

A major quantitative trait locus for resistance to Turnip Yellows Virus (TuYV, syn. beet western yellows virus, BWYV) in rapeseed

AbstractTurnip yellows virus (TuYV) is responsible for a recognizable loss of yield in European winter oilseed rape cultivation. To map genes involved in TuYV resistance, a double haploid population was established by crossing a resynthesized rapeseed line (R54) as donor for TuYV resistance with an elite rapeseed line (‘Express’). Resistance was determined with 10 plants per line by double antibody sandwich‐enzyme‐linked immunosorbent assay. After screening 17 primer combinations (Pstl/Msel and EcoRI/Msel), 143 amplified fragment length polymorphism markers were mapped to 20 linkage groups representing 15 chromosomes of the rapeseed genome. Quantitative trait loci (QTL) were mapped using the composite interval mapping approach. As a result, one major quantitative trait locus was found on linkage group MS17, explaining up to 50% of the phenotypic variation. Because no other factors displaying a significant effect on the expression of resistance could be identified, a simple mode of inheritance for TuYV resistance is suggested, thus enabling marker‐assisted selection during rapeseed breeding.

Molecular dynamics of the frame-shifting pseudoknot from beet western yellows virus: the role of non-Watson-Crick base-pairing, ordered hydration, cation binding and base mutations on stability and unfolding

Molecular dynamics simulations of the frame-shifting pseudoknot from beet western yellows virus (BWYV, NDB file UR0004) were performed with explicit inclusion of solvent and counterions. In all, 33 ns of simulation were carried out, including 10 ns of the native structure with protonation of the crucial cytosine residue, C8(N3+). The native structure exhibited stable trajectories retaining all Watson-Crick and tertiary base-pairs, except for fluctuations or transient disruptions at specific sites. The most significant fluctuations involved the change or disruption of hydrogen-bonding between C8(N3+) and bases G12, A25, and C26, as well as disruption of the water bridges linking C8(N3+) with A25 and C26. To increase sampling of rare events, the native simulation was continued at 400 K. A partial, irreversible unfolding of the molecule was initiated by slippage of C8(N3+) relative to G12 and continued by sudden concerted changes in hydrogen-bonding involving A23, A24, and A25. These events were followed by a gradual loss of stacking interactions in loop 2. Of the Watson-Crick base-pairs, only the 5′-terminal pair of stem 1 dissociated at 400 K, while the trans sugar-edge/sugar-edge A20·G4 interaction remained surprisingly stable. Four additional room-temperature simulations were carried out to obtain insights into the structural and dynamic effects of selected mutations. In two of these, C8 was left unprotonated. Considerable local rearrangements occurred that were not observed in the crystal structure, thus confirming N3-protonation of C8 in the native molecule. We also investigated the effect of mutating C8(N3+) to U8, to correlate with experimental and phylogenetic studies, and of changing the G4·C17 base-pair to A4·U17 to weaken the trans sugar-edge interaction between positions 4 and 20 and to test models of unfolding. The simulations indicate that the C8·G12·C26 base-triple at the junction is the most labile region of the frame-shifting pseudoknot. They provide insights into the roles of the other non-Watson-Crick base-pairs in the early stages of unfolding of the pseudoknot, which must occur to allow readthrough of the message by the ribosome. The simulations revealed several critical, highly ordered hydration sites with close to 100 % occupancies and residency times of individual water molecules of up to 5 ns. Sodium cation coordination sites with occupancies above 50 % were also observed.

Mechanical transmission of poleroviruses

Previously, transmission of poleroviruses has relied solely on the use of their aphid vectors. Biolistic inoculation allowed for the first time the mechanical transmission of Beet western yellows virus (BWYV) and Potato leafroll virus (PLRV) to several host plants. Inoculation with purified preparations and viral RNA extracts of PLRV resulted in 30–50% systemically infected Nicotiana occidentalis P1 plants and 15–30% infected Nicotiana clevelandii plants. Particle bombardment was also used successfully to infect N. clevelandii plants with in vitro RNA transcripts of full-length cDNA of BWYV.

Incidence of virus infection in experimental plots, commercial crops, and seed stocks of cool season crop legumes

Experimental plots of cool season crop legumes growing at diverse locations in Western Australia were inspected for plants with suspect virus symptoms over 4 growing seasons (1994, 1997, 1998, 1999), and plant samples were tested for infection with alfalfa mosaic (AMV), bean yellow mosaic (BYMV), cucumber mosaic (CMV), and pea seed-borne mosaic (PSbMV) viruses. All 4 viruses were detected in faba bean (Vicia faba); BYMV, CMV, and PSbMV in field pea (Pisum sativum); AMV, CMV, and PSbMV in lentil (Lens culinaris); and AMV and CMV in chickpea (Cicer arietinum). Among minor crop species, AMV, BYMV, and CMV were found in narbon bean (V. narbonensis) and grass pea (Lathyrus sativus); BYMV and CMV in dwarf chickling (L. cicera); BYMV in bitter vetch (V. e r v i l i a ) and L. clymenum; and AMV in fenugreek (Trigonella foenum-graecum). Incidences of individual viruses varied widely from site to site but plot infection sometimes reached 100%. Symptom severity varied widely with virus–crop combination. In large-scale surveys of commercial crops of field pea and faba bean over 2 (1998, 1999) and 3 (1994, 1998, 1999) growing seasons, respectively, randomly collected samples from each crop were tested for presence of AMV, BYMV, CMV, and PSbMV. In 1999 they were also tested for beet western yellows virus (BWYV). All 5 viruses were detected in both species. BWYV was found in 35% of faba bean and 56% of the field pea crops sampled in 1999, with incidences of infection in individual crops up to 40% and 49%, respectively. PSbMV was found in 42% and BYMV in 18% of field pea crops in 1999. In individual crops, highest infection incidences of BYMV and PSbMV detected were 31% for BYMV in faba bean in 1998 and 9% for PSbMV in field pea in 1999. CMV and AMV incidences in both species never exceeded 7% of crops or 4% of plants within individual crops. Infection by 2 different viruses within individual crops was common, even 3 were sometimes found. Cultivars infected with most viruses were Fiesta and Fiord for faba bean, and Dundale, Laura, and Magnet for field pea. BYMV was detected in the crop tested of dwarf chickling. In tests on seed samples from Western Australia of 30 commercial seed stocks of field pea, 11 of faba bean, and 50 of chickpea, PSbMV was detected in 11, 1, and 1, respectively; CMV in 1, 1, and 3; BYMV in 3, 1, and 0; and AMV in 0, 0, and 1. This appears to be the first record of seed transmission of CMV in pea and faba bean. Seed samples from Victoria were also found to contain viruses: PSbMV in pea and AMV in lentil. Widespread infection with viruses in evaluation plots and commercial crops of cool season crop legumes is a cause for concern, especially where individual crop incidences are high and 2 or more viruses are present. Sowing of infected seed stocks leads to introduction of randomly dispersed sources of virus infection within the crop sown, resulting in spread of infection and yield losses. Appropriate control measures are discussed.

A century of plant virus management in the Salinas Valley of California, ‘East of Eden’

The mild climate of the Salinas Valley, CA lends itself well to a diverse agricultural industry. However, the diversity of weeds, crops and insect and fungal vectors also provide favorable conditions for plant virus disease development. This paper considers the incidence and management of several plant viruses that have caused serious epidemics and been significant in the agricultural development of the Salinas Valley during the 20th century. Beet curly top virus (BCTV) almost destroyed the newly established sugarbeet industry soon after its establishment in the 1870s. A combination of resistant varieties, cultural management of beet crops to provide early plant emergence and development, and a highly coordinated beet leafhopper vector scouting and spray programme have achieved adequate control of BCTV. These programmes were first developed by the USDA and still operate. Lettuce mosaic virus was first recognized as causing a serious disease of lettuce crops in the 1930s. The virus is still a threat but it is controlled by a lettuce-free period in December and a seed certification programme that allows only seed lots with less than one infected seed in 30 000 to be grown. ‘Virus Yellows’ is a term used to describe a complex of yellows inducing viruses which affect mainly sugarbeet and lettuce. These viruses include Beet yellows virus and Beet western yellows virus. During the 1950s, the complex caused significant yield losses to susceptible crops in the Salinas Valley. A beet-free period was introduced and is still used for control. The fungus-borne rhizomania disease of sugarbeet caused by Beet necrotic yellow vein virus was first detected in Salinas Valley in 1983. Assumed to have been introduced from Europe, this virus has now become widespread in California wherever beets are grown and crop losses can be as high as 100%. Movement of infested soil and beets accounts for its spread throughout the beet-growing regions of the United States. Control of rhizomania involves several cultural practices, but the use of resistant varieties is the most effective and is necessary where soils are infested. Rhizomania-resistant varieties are now available that perform almost as well as the non-resistant varieties under non-rhizomania conditions. Another soil-borne disease termed lettuce dieback, caused by a tomato bushy stunt-like tombusvirus, has become economically limiting to romaine and leaf lettuce varieties. The virus has no known vector and it seems to be moved through infested soil and water. Heavy rains in the past 4 years have caused flooding of the Salinas River and lettuce fields along the river have been affected severely by dieback. Studies are now in progress to characterize this new virus and identify sources of resistance. Agriculture in the Salinas Valley continues to grow and diversify, driven by demands for ‘clean’, high quality food by the American public and for export. The major aspects of plant virus control, including crop-free periods, breeding for resistance, elimination of inoculum sources, and vector control will continue to be vital to this expansion. Undoubtedly, the advances in crop production through genetic manipulation and advances in pest management through biological control will eventually become an important part of agricultural improvement.

First Record of Faba bean necrotic yellows virus and Beet western yellows virus Infecting Faba Bean in Tunisia.

A survey was conducted in April 2000 to identify viruses infecting faba bean (Vicia faba L.) in seven regions (Beja, Bizerte, Cap-bon, Jendouba, Kairouan, Siliana, and Zaghouan) of Tunisia. Around 4,000 random samples were collected from 34 fields (100 to 200 samples per field). Samples were tested by tissue-blot immunoassay procedure (2) at the Virology Laboratory of INRAT for the presence of 14 different viruses. All antisera used were specific, including those for Faba bean necrotic yellows virus (FBNYV) (1) and Beet western yellows virus (BWYV) (ATCC-PVAS-647). Broad bean mottle, a beetle-transmitted and seedborne virus, was the most abundant with an average incidence of 2.3%. The highest disease incidence occurred with BWYV, which was in 20 and FBNYV in 4 of 98 random samples from a field in the Siliana region; and with BWYV in 7 and BBMV in 21 of 120 samples from a field in the Jendouba region. Other viruses such as broad bean stain, cucumber mosaic, and bean leaf roll were detected rarely (<1%). This is the first record of FBNYV and BWYV infecting faba bean in Tunisia. BWYV was detected in 1.2% of the samples tested collected from 20 fields comprising all seven regions. FBNYV was detected in 0.9% of the samples tested collected from 13 fields comprising all except the Capbon region.