Beet Necrotic Yellow Vein Virus Isolates Research Articles

Generation of specific antibody against recombinant major coat protein of Beet necrotic yellow vein virus and its application for detection of rhizomania disease

Beet necrotic yellow vein virus (BNYVV) is one of the main contributors to economic losses in sugar beet production. The present study generated a polyclonal antibody that detects BNYVV. The conserved genomic region of all BNYVV isolates gene encoding the viral major coat protein (BNYVV-CP) was amplified, cloned, sequenced, and expressed in E. coli strain BL21 (DE3). The expressed protein was purified under native conditions by affinity chromatography. The purified BNYVV CP was used as a recombinant antigen to raise polyclonal antibodies in rabbits. Purified anti-CP-IgG detected the BNYVV-CP recombinant protein and BNYVV in infected sugar beet by indirect-ELISA, dot immunobinding assay, and western blot analysis blotting. The serological assays strongly proved the sensitivity and specificity of anti-BNYVV-CP-IgG against BNYVV. These results suggest that the generated anti-BNYVV-CP-IgG polyclonal antibodies can be used as reliable and quick means for the BNYVV virus detection under field conditions.

Türkiye’de şeker pancarı üretim alanlarında enfeksiyon oluşturan Beet necrotic yellow vein virus izolatlarının p31 geninin moleküler karakterizasyonu

Beet necrotic yellow vein virus (BNYVV), which is known as "the agent of rhizomania disease" and transmitted by Polymyxa betae, is the most destructive and prevalent virus species in sugar beet fields in Turkey and in the world. In this study, seventeen BNYVV-infested soil samples were selected according to their geographic locations. Firstly, BNYVV was propogated as bait plant technique. 
 P31 protein encoded by BNYVV RNA-4 is associated with efficient vector transmission and virulence. In this study, nucleotide sequences of p31 coding region of 12 different isolates were obtained. The BLAST and phylogenetic analysis divided BNYVV isolates into two different p31 groups as Group II and Group III. The Group II consisted of IGR-6, IGR-9 and ERC-52 isolates and found to have high nucleotide identity with Kas2 isolate (99.76%-99.88%). Also, GZP-27, ELZ-44, SMS-61, EDR-125, BRS-148, CNK-150, KAS-281, KYS-524 and ANK-617 which were divided into Group III had the highest similarity (99.88%-100%) with IV isolate (Italy).

Resistance in Beta vulgaris L. subsp. maritima (L.) Thell. to the Rz1-breaking strain of rhizomania

Rhizomania, caused by Beet necrotic yellow vein virus (BNYVV) and transmitted by Polymyxa betae, is considered the most important disease of sugar beet and has spread to almost all production areas of the world. The most efficient way to control this soil-borne disease is through the cultivation of rhizomania-resistant varieties, which has proven successful since the 1980s. In the 2002–2003 season, sugar beet fields in the Imperial Valley, California, planted with a resistant commercial variety were observed with severe rhizomania symptoms suggesting resistance conferred by the most widely deployed gene, Rz1, was compromised. BNYVV isolates were identified to which all resistant check varieties became infected and tested positive for the virus (RB-BNYVV, Rz1-resistance breaking strain of Beet necrotic yellow vein virus). For continuing development of BNYVV and RB-BNYVV resistant germplasm, a highly important reservoir of resistance factors is the Beta vulgaris subsp. maritima (BVM) accession collection. We evaluated 404 BVM accessions under BNYVV and RB-BNYVV field and greenhouse conditions to identify and select rhizomania resistant accessions useful for sugar beet germplasm improvement. Accessions from Denmark, Ireland, and Portugal showed the highest level of rhizomania resistance as characterized by foliar yellowing due to rhizomania, rhizomania disease index, and percent resistant beets. The low bolting tendency of Denmark and Ireland accessions suggest these for pre-breeding donor material.

Distribution of resistance-breaking isolates of beet necrotic yellow vein virus differing in virulence in sugar beet fields in Turkey

ABSTRACTRhizomania, caused by Beet necrotic yellow vein virus (BNYVV), is one of the most devastating diseases on sugar beet throughout the world. A total of 235 BNYVV-infested soil samples taken from 29 provinces during the growing seasons of 2004–2005 and 2010–2011 were investigated for resistance-breaking properties of BNYVV isolates. BNYVV resistant (Rz1, Rz1 + R2, Rz1 + C48 + minor genes) and susceptible (rz1) sugar beet cultivars were grown in the BNYVV-infested soils by the bait plant technique, and virus infection was evaluated based on disease symptom and DAS-ELISA. Of the soil samples tested, 57.4% produced infection in the roots of sugar beet cultivar (cv.) containing the Rz1 gene, 51.9% in cv. carrying the Rz1 + Rz2 and 18.9% in cv. carrying the Rz1 + C48 + minor genes. Also, the presence of Beet soil-borne virus (BSBV) was evaluated by TAS-ELISA. Additionally, the presence of BNYVV RNA-5 component was investigated by RT–PCR using the primers specific for p26 coding region. Our results revealed that BNYVV isolates compromising resistance genes are highly common and widespread in sugar beet production areas in Turkey, and it seems unlikely that BSBV and/or RNA-5 were directly involved in RB-event.

P25 pathogenicity factor deletion mutants of beet necrotic yellow vein virus occurring in sugar beet fields in Turkey

Beet necrotic yellow vein virus (BNYVV) causes rhizomania which is one of the most devastating diseases in sugar beet fields worldwide. BNYVV symptoms were observed in Rz1-resistant sugar beet plants in a number of fields in Aksaray and Konya Provinces during surveys in 2011. Seven soil samples, taken from problematic areas in these provinces and known to be infested with populations of Polymyxa betae Keskin carrying BNYVV, were used in bait plant and molecular studies. Initially, the presence of other soil-borne viruses was investigated by using ELISA and RT-PCR. It was determined that BNYVV was in co-infections with pomoviruses, namely beet soil-borne virus and/or beet virus Q. The partial RNA-3 segment of BNYVV was amplified by RT-PCR, and the nucleotide sequences involving the p25 coding region were obtained. The deduced amino acid (aa) sequences of the p25 protein indicated that four BNYVV populations had ‘ACHG’ or ‘VHHG’ residues at positions 67–70. In addition, three populations containing three nucleotide deletions, not shifting the reading frame but affecting the aa at position 67 (-DHG) or 68 (D-HG), were identified. A deleted form of the p25 was identified in both the A- and J-type BNYVV populations. The pathogenicities of these BNYVV populations in the sugar beet genotypes rz1 and Rz1 were investigated under controlled conditions. All seven BNYVV isolates were capable of overcoming Rz1 resistance. These results seem to indicate that mutants of BNYVV with a deletion are aggressive, as are the isolates without deletion. Resistant beet cultivars will most likely be more productive in the future if these isolates become widespread in Turkey.

Identification of strain types of some Beet necrotic yellow vein virus isolates determined in Northern and Central Parts of Turkey

Beet necrotic yellow vein virus (BNYVV), the agent of rhizomania disease, causes severe economic losses in sugar beet fields in all over the world. The virus is transmitted by a plasmodiophorid vector, Polymyxa betae Keskin. Twenty soil samples, collected from sugar beet fields in northern and central parts of Turkey during surveys in 2004 and 2005 and known to be infested with viruliferous cultures of P. betae carrying BNYVV, were selected and used in this study. Sample selection was made according to symptom expression of beet seedlings in preliminary bait plant tests and locations of the soil samples that accurately represent the region from which they were taken. Total RNAs were extracted from sugar beet plants grown in these soils and used to amplify RNA-2 (nt. 19-1088) and RNA-3 (nt. 50-1268) of BNYVV by reverse transcription polymerase chain reaction (RT-PCR) method. Restriction fragment length polymorphism (RFLP) analysis of PCR-amplified products showed that most of BNYVV isolates studied were A-type strain, however, two isolates did not exactly match the band profile of A-type strain. Additionally, the presence of BNYVV RNA-5 component was investigated by RT-PCR using the primers specific for P26 coding region. Four samples belonging to three provinces were found to be involving RNA-5 segment (20%).

The widespread occurrences of Beet soil borne virus and RNA-5 containing Beet necrotic yellow vein virus isolates in sugar beet production areas in Turkey

A total of 632 soil samples were collected from different sugar beet fields in 32 provinces during 2005, 2010 and 2011 growing seasons. Incidences of Beet necrotic yellow vein virus (BNYVV), Beet soil-borne virus (BSBV) and their vector Polymyxa betae were determined by bait plant tests using ELISA and root staining techniques. The percentages of BNYVV and BSBV infested soil samples were 38 and 49.8 %, respectively. The study showed that P. betae was very common (87.7 %) in the sampled sugar beet fields. Additionally, the presence of BNYVV RNA-5 component was investigated by RT-PCR using the primers specific for p26 coding region. Out of 240 BNYVV-infected samples, 85 were selected according to their geographic origin, and 62.4 % of them belonging to 29 provinces were found to contain the RNA-5 segment. The partial nucleotide sequences of RNA-5 of BNYVV isolates from Igdir, Edirne, Ankara and Konya provinces of Turkey were obtained. Nucleotide sequencing and phylogenetic analyses showed that RNA-5 of Turkish BNYVV isolates is closer to the J type BNYVV isolates than the P type isolates. These results also indicated that BNYVV populations containing RNA-5 is highly widespread in sugar beet production areas in Turkey.

Deep sequencing-based transcriptome profiling reveals comprehensive insights into the responses of Nicotiana benthamiana to beet necrotic yellow vein virus infections containing or lacking RNA4.

Background Beet necrotic yellow vein virus (BNYVV), encodes either four or five plus-sense single stranded RNAs and is the causal agent of sugar beet rhizomania disease, which is widely distributed in most regions of the world. BNYVV can also infect Nicotiana benthamiana systemically, and causes severe curling and stunting symptoms in the presence of RNA4 or mild symptoms in the absence of RNA4.ResultsConfocal laser scanning microscopy (CLSM) analyses showed that the RNA4-encoded p31 protein fused to the red fluorescent protein (RFP) accumulated mainly in the nuclei of N. benthamiana epidermal cells. This suggested that severe RNA4-induced symptoms might result from p31-dependent modifications of the transcriptome. Therefore, we used next-generation sequencing technologies to analyze the transcriptome profile of N. benthamiana in response to infection with different isolates of BNYVV. Comparisons of the transcriptomes of mock, BN3 (RNAs 1+2+3), and BN34 (RNAs 1+2+3+4) infected plants identified 3,016 differentially expressed transcripts, which provided a list of candidate genes that potentially are elicited in response to virus infection. Our data indicate that modifications in the expression of genes involved in RNA silencing, ubiquitin-proteasome pathway, cellulose synthesis, and metabolism of the plant hormone gibberellin may contribute to the severe symptoms induced by RNA4 from BNYVV.ConclusionsThese results expand our understanding of the genetic architecture of N. benthamiana as well as provide valuable clues to identify genes potentially involved in resistance to BNYVV infection. Our global survey of gene expression changes in infected plants reveals new insights into the complicated molecular mechanisms underlying symptom development, and aids research into new strategies to protect crops against viruses.

CHARACTERIZATION OF BEET NECROTIC YELLOW VEIN VIRUS INFECTING SUGAR BEET IN LITHUANIA

Beet necrotic yellow vein virus (BNYVV) was shown to occur in sugar beets in three of six regions surveyed in Lithuania. Characterizing the coat protein (CP) and p25 genes by RT-PCR and sequencing showed the presence of A and B types of BNYVV isolates. The p25 protein of Lithuanian BNYVV isolates contained a distinct amino acid tetrad (residues 67-70) with AYHR associated with a B type isolate and AHHG with two A type isolates. These sequence features were consistent with phylogenetic grouping of isolates based on CP and p25 gene sequences. This is the first characterization of BNYVV isolates in Lithuania.

Analysis of the Resistance-Breaking Ability of Different Beet necrotic yellow vein virus Isolates Loaded into a Single Polymyxa betae Population in Soil

The genome of most Beet necrotic yellow vein virus (BNYVV) isolates is comprised of four RNAs. The ability of certain isolates to overcome Rz1-mediated resistance in sugar beet grown in the United States and Europe is associated with point mutations in the pathogenicity factor P25. When the virus is inoculated mechanically into sugar beet roots at high density, the ability depends on an alanine to valine substitution at P25 position 67. Increased aggressiveness is shown by BNYVV P type isolates, which carry an additional RNA species that encodes a second pathogenicity factor, P26. Direct comparison of aggressive isolates transmitted by the vector, Polymyxa betae, has been impossible due to varying population densities of the vector and other soilborne pathogens that interfere with BNYVV infection. Mechanical root inoculation and subsequent cultivation in soil that carried a virus-free P. betae population was used to load P. betae with three BNYVV isolates: a European A type isolate, an American A type isolate, and a P type isolate. Resistance tests demonstrated that changes in viral aggressiveness towards Rz1 cultivars were independent of the vector population. This method can be applied to the study of the synergism of BNYVV with other P. betae-transmitted viruses.

Evidence for the presence of Beet necrotic yellow vein virus types A and B in Poland

Beet necrotic yellow vein virus (BNYVV), which causes a viral disease of sugar beet known as “rhizomania”, is widely spread throughout the sugar beet growing areas of Poland. The analysis of the molecular variability of two separate regions, namely the part of coat protein (CP) and triple gene block (TGB) genes of the BNYVV genome, revealed very low level of nucleotide sequence diversity among the isolates (98–100% sequence identity). Sequence alignments of BNYVV isolates from Poland and other isolates available to data showed also high level of nucleotide sequence identity among these isolates. Based on multiplex RT-PCR and phylogenetic analysis we distinguished types A and B of BNYVV. The sequence identity matrix between type A and B revealed 92–93% similarity. Mainly, BNYVV type A is widespread in Poland whereas type B has rarely been found. These results suggest that Polish isolates probably originated from Southern Europe rather then from Germany or France where BNYVV type B is a dominating one.

Molecular characterization of three soil-borne sugar beet-infecting viruses based on the coat protein gene.

SUMMARY Sugar beet (Beta vulgaris L. ssp. vulgaris) hosts the soil-borne viruses Beet necrotic yellow vein virus (BNYVV), Beet soil-borne virus (BSBV), Beet soil-borne mosaic virus (BSBMV) and Beet virus Q (BVQ). Only a few isolates of these viruses have been characterized at the coat protein (CP) gene level. In this work, CP gene sequences were determined of three isolates each of BNYVV, BSBV and BVQ from different parts of Poland (Wielkopolska, Mazowsze and Dolny Slask). Phylogenetic analysis showed that Polish BNYVV isolates belong to the A and B type groups. The CP gene of the three BSBV isolates had 98-100% identity with comparable sequences available in GenBank. CP genes of the three BVQ isolates were almost identical to one another (9899%) and contained three additional nucleotides, resulting in a further amino acid residue (arginine) at position 86, as reported by Lennefors et al. (2005).

Beet soil-borne mosaic virus RNA-3 is replicated and encapsidated in the presence of BNYVV RNA-1 and -2 and allows long distance movement in Beta macrocarpa

Beet soil-borne mosaic virus (BSBMV) and Beet necrotic yellow vein virus (BNYVV) belong to the Benyvirus genus. BSBMV has been reported only in the United States, while BNYVV has a worldwide distribution. Both viruses are vectored by Polymyxa betae and possess similar host ranges, particle number and morphology. BNYVV and BSBMV are not serologically related but they have similar genomic organizations. Field isolates usually consist of four RNA species but some BNYVV isolates contain a fifth RNA. RNAs 1 and 2 are essential for infection and replication while RNAs 3 and 4 play important roles in plant and vector interactions, respectively. Nucleotide and amino acid analyses revealed that BSBMV and BNYVV are sufficiently different to be classified as two species. Complementary base changes found within the BSBMV RNA-3 5′ UTR made it resemble to BNYVV 5′ RNA-3 structure whereas the 3′ UTRs of both species were more conserved. cDNA clones were obtained, and allowed complete copies of BSBMV RNA-3 to be trans-replicated, trans-encapsidated by the BNYVV viral machinery. Long-distance movement was observed indicating that BSBMV RNA-3 could substitute BNYVV RNA-3 for systemic spread, even though the p29 encoded by BSBMV RNA-3 is much closer to the RNA-5-encoded p26 than to BNYVV RNA-3-encoded p25. Competition occurred when BSBMV RNA-3-derived replicons were used together with BNYVV-derived RNA-3 but not when the RNA-5-derived component was used. Exploitation of the similarities and divergences between BSBMV and BNYVV should lead to a better understanding of molecular interactions between Benyviruses and their hosts.

Breaking of Beet necrotic yellow vein virus resistance in sugar beet is independent of virus and vector inoculum densities

Beet necrotic yellow vein virus (BNYVV) is transmitted by Polymyxa betae to sugar beet, causing rhizomania disease. Resistance-breaking strains of BNYVV, overcoming single (Rz1) or double (e.g. Rz1 + Rz2) major resistance genes in sugar beet have been observed in France and recently in the USA and Spain. To demonstrate if resistance-breaking is dependent on inoculum density, the inoculum concentration of BNYVV and P. betae in soil samples where resistance-breaking had been observed was estimated using the most probable number (MPN) method. The MPN-values obtained displayed highly significant differences with respect to the virus concentration in various soils and did not correlate with the ability to overcome resistance. Virus quantification in susceptible plants demonstrated that soils containing resistance-breaking isolates of BNYVV did not produce higher virus concentrations. The MPN assay was repeated with Rz1 + Rz2 partially-resistant sugar beets to see if the resistance-breaking is concentration-dependent. There was no correlation between soil dilution and increased virus concentration in Rz1 + Rz2 plants produced by BNYVV resistance-breaking strains. Determination of the absolute P. betae concentration by ELISA demonstrated that all resistance-breaking soil samples contained elevated concentrations. However, the calculation of the proportion of viruliferous P. betae did not show a positive correlation with the resistance-breaking ability. Finally resistance-breaking was studied with susceptible, Rz1 and Rz1 + Rz2 genotypes and standardised rhizomania inoculum added to sterilised soil. Results from these experiments supported the conclusion that resistance-breaking did not correlate with virus concentration or level of viruliferous P. betae in the soil.

Suppression of Resistance-Breaking Beet necrotic yellow vein virus Isolates by Beet oak-leaf virus in Sugar Beet.

Rhizomania, a serious disease of sugar beet (Beta vulgaris), is caused by Beet necrotic yellow vein virus (BNYVV). Resistance allele Rz1 has been widely incorporated into commercial cultivars. Recently, resistance-breaking isolates of BNYVV (RB-BNYVV) were identified and characterized. When the occurrence of RB-BNYVV was surveyed throughout the sugar-beet-growing areas in the United States, most soil samples contained Beet oak-leaf virus (BOLV) as well. BNYVV and BOLV often occurred in the same field and sometimes in the same sugar beet plant. The possibility of interactions between these two Polymyxa betae-transmitted sugar beet viruses was tested. Plants grown in soils infested with aviruliferous P. betae or carrying RB-BNYVV and BOLV, alone and in combination, were compared with plants grown in noninfested soil for differences in plant fresh weight and virus content as measured by enzyme-linked immunosorbent assay (ELISA). Rz1 and Rz2 resistance genes that condition resistance to BNYVV did not confer resistance to BOLV. BNYVV ELISA values were significantly higher in single infections than in mixed infections with BOLV in both the rhizomania-resistant and -susceptible cultivars. In contrast, ELISA values of BOLV were not significantly different between single and mixed infections in both the rhizomania-resistant and -susceptible cultivars. Results indicate that BOLV may suppress BNYVV in mixed infections. Soils infested with P. betae significantly reduced fresh weight of sugar beet seedlings regardless of whether they were with or without one or both viruses or resistance genes.

Identification of amino acids of the beet necrotic yellow vein virus p25 protein required for induction of the resistance response in leaves of Beta vulgaris plants

The RNA3-encoded p25 protein of beet necrotic yellow vein virus (BNYVV) is responsible for the production of rhizomania symptoms of sugar beet roots (Beta vulgaris subsp. vulgaris). Here, it was found that the presence of the p25 protein is also associated with the resistance response in rub-inoculated leaves of sugar beet and wild beet (Beta vulgaris subsp. maritima) plants. The resistance phenotype displayed a range of symptoms from no visible lesions to necrotic or greyish lesions at the inoculation site, and only very low levels of virus and viral RNA accumulated. The susceptible phenotype showed large, bright yellow lesions and developed high levels of virus accumulation. In roots after Polymyxa betae vector inoculation, however, no drastic differences in virus and viral RNA accumulation levels were found between plants with susceptible and resistant phenotypes, except at an early stage of infection. There was a genotype-specific interaction between BNYVV strains and two selected wild beet lines (MR1 and MR2) and sugar beet cultivars. Sequence analysis of natural BNYVV isolates and site-directed mutagenesis of the p25 protein revealed that 3 aa residues at positions 68, 70 and 179 are important in determining the resistance phenotype, and that host-genotype specificity is controlled by single amino acid changes at position 68. The mechanism of the occurrence of resistance-breaking BNYVV strains is discussed.

Distribution and Molecular Characterization of Resistance-Breaking Isolates of Beet necrotic yellow vein virus in the United States.

Beet necrotic yellow vein virus (BNYVV) is the causal agent of rhizomania in sugar beet (Beta vulgaris). The virus is transmitted by the plasmodiophorid Polymyxa betae. The disease is controlled primarily by the use of partially resistant cultivars. During 2003 and 2004 in the Imperial Valley of California, partially resistant sugar beet cultivars with Rz1 allele seemed to be compromised. Field trials at Salinas, CA have confirmed that Rz1 has been defeated by resistance-breaking isolates. Distinct BNYVV isolates have been identified from these plants. Rhizomania-infested sugar beet fields throughout the United States were surveyed in 2004-05. Soil surveys indicated that the resistance-breaking isolates not only existed in the Imperial Valley and San Joaquin Valley of California but also in Colorado, Idaho, Minnesota, Nebraska, and Oregon. Of the soil samples tested by baited plant technique, 92.5% produced infection with BNYVV in 'Beta 6600' (rz1rz1rz1), 77.5% in 'Beta 4430R' (Rz1rz1), 45.0% in 'Beta G017R' (Rz2rz2), and 15.0% in 'KWS Angelina' (Rz1rz1+Rz2rz2). Analyses of the deduced amino acid sequence of coat protein and P-25 protein of resistance-breaking BNYVV isolates revealed the high percentage of identity with non-resistance-breaking BNYVV isolates (99.9 and >98.0%, respectively). The variable amino acids in P-25 proteins were located at the residues of 67 and 68. In the United States, the two amino acids found in the non-resistance-breaking isolates were conserved (AC). The resistance-breaking isolates were variable including, AF, AL, SY, VC, VL, and AC. The change of these two amino acids cannot be depended upon to differentiate resistance-breaking and non-resistance-breaking isolates of BNYVV.

Sequence variation within Beet necrotic yellow vein virus p25 protein influences its oligomerization and isolate pathogenicity on Tetragonia expansa

The p25 protein encoded by Beet necrotic yellow vein virus (BNYVV) RNA-3 is a pathogenicity determinant that has been implicated in symptom exacerbation on Chenopodiaceae hosts. Several p25 variants exist within natural isolates and p25 sequence variation may influence the degree of pathogenicity of such BNYVV isolates. Expression of p25 from natural A- and P-type isolates in the background of B-type BNYVV cDNA clones gave symptom discrepancies when compared to B-type p25 expression. Such pathogenicity fluctuation was not due to a different subcellular localization of p25 but was correlated with the nature of the tetrad motif present between amino acid residues 67–70, as well as with the capacity of p25 to self-associate and to activate transcription in a yeast one-hybrid system. Our data suggest that the complete sequence of p25 is required for its functions and the identified sequence variations may contribute to correct folding of the protein.

Partial nucleotide sequence analysis of Turkish isolates of Beet necrotic yellow vein virus (BNYVV) RNA‐3

A survey was carried out to detect Beet necrotic yellow vein virus (BNYVV) in soil samples using RT‐PCR and bait plant techniques from the sugar beet production area of Tokat, Turkey in 2001. More than 80% of the soil samples analyzed were found to be contaminated with the virus. The partial nucleotide sequence of cDNA corresponding to RNA‐3 of BNYVV isolates were analyzed for six different regions of Tokat province. All isolates were assigned to type A strains based on RFLP analysis and DNA sequences. Sequence comparison revealed differences at amino acid positions 35, 68, 71 and 179 of the P25 coding region amongst Turkish isolates. Additionally, all Turkish isolates were compared with Japanese, French, Kazakh, Italian and Belgian isolates.

Susceptibility and resistance of Beta vulgaris subsp. maritima to foliar rub-inoculation with Beet necrotic yellow vein virus

Four lines (designated MR0, MR1, MR2, and M8) from 13 accessions of Beta vulgaris subsp. maritima were selected on the basis of phenotypes produced after foliar rub-inoculation with Beet necrotic yellow vein virus (BNYVV). The susceptible phenotype developed bright yellow local lesions, whereas the resistant phenotype had symptoms ranging from no visible lesions to necrotic lesions at the inoculation site. MR1 and MR2 lines had a resistant phenotype depending on the isolate and the MR0 line was susceptible to all isolates of BNYVV tested. The M8 line was highly susceptible; the virus spread systemically and caused severe stunting. These plant lines will be useful for distinguishing BNYVV isolates having different pathogenicities, especially those controlled by RNA3 and/or RNA5.