A strain of Bean common mosaic necrosis virus (BCMNV) from Idaho was identified by enzyme-linked immunosorbent assay using monoclonal antibodies and determined to be similar to the NL-3 D strain (of Drifjhout) by reaction of differential bean cultivars. However, this BCMNV strain (designated NL-3 K) caused earlier and more severe symptoms on bean plants representing host groups 0, 4, and 5. The nucleotide sequence encoding the predicted polyprotein of NL-3 K was 9,893 nucleotides (nt) in length, yielding a peptide with a molecular size of 362.1 kDa compared with a 9,626-nt, 350.9-kDa polyprotein for NL-3 D. Sequence analysis of the putative P1 protein suggests that the NL-3 K strain is a recombinant between NL-3 D and the Russian strain (RU1) of Bean common mosaic virus. The P1 protein of NL-3 K consisted of 415 amino acids compared with 317 for NL-3 D. The first 114 predicted amino acids of the NL-3 K P1 region were 98% identical with RU1. The remaining 301 amino acids of the protein shared only 34% identity with RU1 but were 98% identical with NL-3 D. Primers were designed that flanked the recombination point in the P1 coding sequence of NL-3 K. An amplicon of the expected size was produced by reverse-transcriptase polymerase chain reaction of total nucleic acid extracts of bean plants inoculated with NL-3 K, but not from those with NL-3 D or RU1. The increased symptom severity on selected common bean lines induced by NL-3 K suggests that the P1 gene may play a significant role in pathogenicity and virulence.
The 3'-terminal nucleotide sequences of thirteen authenticated strains of bean common mosaic virus (BCMV) and one strain of bean common mosaic necrosis virus (BCMNV) were obtained. The regions sequenced included the coat protein coding sequence and 3'-end non-coding region. These data, combined with sequence information from other legume-infecting potyviruses and the Potyviridae were used for phylogenetic analysis. Evidence is provided for delineation of BCMNV as distinct from BCMV and the inclusion of azuki mosaic, dendrobium mosaic, blackeye cowpea mosaic, and peanut stripe viruses as strains of BCMV. This relationship defines the members of the BCMV and BCMNV subgroups. These data also provide a basis upon which to define virus strains, in combination with biological data. Other aspects and implications of legume-infecting potyvirus phylogenetics are discussed.
Little cherry disease, one of the major viral diseases of sweet cherry (Prunus avium) worldwide, is associated with either of two closteroviruses, Little cherry virus 1 (LChV-1) and Little cherry virus 2 (LChV-2). Two sets of primers corresponding to a portion of the replicase gene of LChV-1 and LChV-2 were used in one-tube reverse-transcription polymerase chain reactions to detect these viruses in total RNA extracts of field-collected sweet cherry tissues. LChV-1 and LChV-2 were detected both alone and in combination in five sweet cherry orchards in Washington State. Sequence analysis of a 240-nucleotide (nt) fragment of the replicase open reading frame (ORF)1b and a 232-nt fragment from a portion of ORF8 and the 3′ untranslated region (UTR) of LChV-1 indicated that North American (NA) isolates shared 90 to 99% nucleotide identity in both genome segments analyzed. In contrast, comparisons of NA isolates to two Eurasian isolates of LChV-1 indicated shared nucleotide identities of 79 to 82% in the replicase fragment and 89 to 90% in the ORF8/3′UTR fragment. Sequence variation in the replicase region did not affect detection of LChV-1 in 12 isolates using the replicase-specific primers reported here. This article represents the first report of LChV-1 and LChV-2 in sweet cherry in Washington.
The geographic distribution of Iris yellow spot virus (IYSV, Genus Tospovirus, Family Bunyaviridae) in onion (Allium cepa L.) crops in the western United States has increased with the most recent report in Colorado (1,4). Furthermore, the incidence of IYSV has increased significantly in onion crops in the Treasure Valley of southern Idaho and eastern Oregon, where the disease was first detected in the United States (1,2). Surveys of onion seed crops in Washington during the past 2 years showed the presence of plants with symptoms characteristic of IYSV infection, including distinct diamond-shaped chlorotic or necrotic lesions, as well as indistinct circular to irregular, chlorotic or necrotic lesions of various sizes on the scapes of flowering plants. To date, symptomatic plants have been observed in five seed crops in Washington, at incidences ranging from <1% to approximately 20% in individual seed crops. Enzyme-linked immunosorbent assays carried out directly on symptomatic onion samples collected in July 2002, and on Nicotiana benthamiana plants mechanically inoculated with sap from these symptomatic plants, did not detect the presence of IYSV. In late July 2003, symptomatic plants were collected from an onion seed crop in Grant County and tested for IYSV infection by reverse transcription-polymerase chain reaction (RT-PCR). Total nucleic acid was extracted from symptomatic areas of the scapes with the procedure described by Presting et al. (3). Primers specific to the nucleocapsid (NP) gene of IYSV were designed based on sequences in GenBank: 5′-TCA GAA ATC GAG AAA CTT-3′ and 5′-TAA TTA TAT CTA TCT TTC TTG G-3′ (sense and antisense polarity, respectively). The RT-PCR assay produced an amplicon of the expected size (approximately 700 bp) that was cloned and sequenced. Comparison with the GenBank IYSV gene sequences showed 98% sequence identity of the NP gene. In August 2003, symptoms of IYSV infection were observed in two onion bulb crops, each located within 2 miles of the symptomatic onion seed crop in Grant County. The presence of IYSV in these crops was confirmed by RT-PCR with cloning and sequencing of the amplicon, as described for the seed crop samples. To our knowledge, this is the first confirmation of IYSV in onion bulb and seed crops in Washington, where 16,000 to 18,000 acres of onion bulb crops and 700 to 900 acres of onion seed crops are grown annually (USDA National Agricultural Statistics Service). The increase in prevalence of IYSV in the Pacific Northwest highlights the need for additional research to clarify the epidemiology of this potentially significant pathogen and to develop a regional management program for iris yellow spot. References: (1) J. M. Hall et al. Plant Dis. 77:952, 1993. (2) J. W. Moyer et al. (Abstr.) Phytopathology 93(suppl.):S115, 2003. (3) G. G. Presting et al. Phytopathology 85:436, 1995. (4) H. F. Schwartz et al. Plant Dis. 86:560, 2002.
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