Turnip mosaic virus (TuMV), a species of the genus Potyvirus, occurs worldwide. Seventy-six isolates of TuMV were collected from around the world, mostly from Brassica and Raphanus crops, but also from several non-brassica species. Host tests grouped the isolates into one or other of two pathotypes ; Brassica (B) and Brassica-Raphanus (BR). The nucleotide sequences of the first protein (P1) and coat protein (CP) genes of the isolates were determined. One-tenth of the isolates were found to have anomalous and variable phylogenetic relationships as a result of recombination. The 5h-terminal 300 nt of the P1 gene of many isolates was also variable and phylogenetically anomalous, whereas the 380 nt 3h terminus of the CP gene was mostly conserved. Trees calculated from the remaining informative parts of the two genes of the non-recombinant sequences by neighbour-joining, maximum-likelihood and maximum-parsimony methods were closely similar, and so these parts of the sequences were concatenated and trees calculated from the resulting 1150 nt. The isolates fell into four consistent groups ; only the relationships of these groups with one another and with the outgroup differed. The ' basal-B ' cluster of eight B-pathotype isolates was most variable, was not monophyletic, and came from both brassicas and non-brassicas from southwest and central Eurasia. Closest to it, and forming a monophyletic subgroup of it in most trees, and similarly variable, was the ' basal-BR ' group of eight BR pathotype Eurasian isolates. The third and least variable group, the ' Asian-BR ' group, was of 22 BR-pathotype isolates, all from brassicas, mostly Raphanus, and all from east Asia mostly Japan. The fourth group of 36 isolates, the ' world-B ' group, was from all continents, most were isolated from brassicas and most were of the B-pathotype. The simplest of several possible interpretations of the trees is that TuMV originated, like its brassica hosts, in Europe and spread to the other parts of the world, and that the BR pathotype has recently evolved in east Asia.

The complete nucleotide sequences of rice blackstreaked dwarf virus (RBSDV) genome segments 8 ($8) and 7 ($7) were determined, and were found to have high sequence identities to the corresponding maize rough dwarf virus (MRDV) genome segments. RBSDV $8 and $7 consisted of 1927 and 2193 nucleotides, respectively. RBSDV $8 had a single long open reading frame (ORF)encoding 591 amino acids. RBSDV $7 had two nonoverlapping ORFs encoding 362 (ORF1) and 309 (ORF2) amino acids. The two ORFs of RBSDV $7 were inserted separately into an Escherichia coil expression vector (pKK223-3). When they were expressed in E. coli cells, the products of both ORFs migrated identically at an apparent M r of 40K. High nucleotide sequence identity was observed between RBSDV $7 and MRDV $6 (85 %), and between the terminal regions of RBSDV $8 and MRDV $7. In addition, RBSDV $7 and MRDV $6 showed 91% (ORF1 product) and 85% (ORF2 product) amino acid sequence identities.

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Plant reoviruses have segmented genomes consisting of 10-12 dsRNAs. They have been classified into subgroups 1, 2 and 3 based on vector species, particle morphology, and number of segmented genome. 7) Genome structure of wound tumor virus (WTV) genome segments 4-12 and rice dwarf virus (RDV) genome segments 9 and 10 have been determined.'~-4).s~,15).1`f2') From these studies, RDV and wTV, both belong to the same subgroup 1, have been shown to have some structural features in common. For example, both RDV and wTV have the same conserved tetranucleotides at both 5' and 3' termini and have the segment-specific inverted repeat at the terminal regions.1 19>,21) Furthermore, the comparable genes encoded by the genome segments of RDV and WTV were found to have a significant amino acid sequence homology.2'~.s~ These results indicated comparative analyses of the genome structure would elucidate some insight on function of the genes and thus on mechanisms involved in transcription, replication, and gene expression of the plant reoviruses. Rice black-streaked dwarf virus (RBSDV), a member of the subgroup 2, has a distinct double-shelled particle and 10 genome segments."'~ This is the largest subgroup among three subgroups consisting of 9 viruses. 7 However, none is known about the genome structure of the viruses in the subgroup 2. Since we are interested in whether the genes encoded by the RBSDV have some structural features common to other plant reoviruses, we have sequenced the genome segment 10 and a deduced amino acid sequence is presented. This is the first report on the genome structure of the plant reovirus subgroup 2. Materials and methods. RNA extraction. RBSDV was purified from the infected maize leaves and sheeth as previously described.The genome dsRNAs were extracted from the purified virus by phenol-SDS after proteinase-K treatment as previously described.' ~~ ssRNA contaminants were removed by 2M LiCI fractionation.1 (DNA synthesis and molecular cloning. The cDNAs were synthesized from the denatured genome dsRNAs as described elsewhere.>> The cDNAs were inserted into bacterial plasmid pBR 322 at Pst I site after homopolymeric tailing. And E. toll strain HB101 was transformed with the recombinant DNAs according to CaCl method described by Maniatis et al.13? Sequence analyses. cDNA was subeloned into M-13 phage mp18 and mp19" and sequenced by dideoxy chain termination methods 7 using a DEAZA sequencing kit (Takara Shuzou). Direct RNA sequence was done by reverse transcription and dideoxy chain termination method using viral transcripts or genomic RNA as a template.J~ 2 ' Results and discussion. A clone pRB C3 had a insert of about 1800 nucleotides. This clone reacted specifically with the genome segment 10 by a

By polyacrylamide gel electrophoresis (PAGE) and nucleic acid hybridization, the extent of genomic relatedness between rice black-streaked dwarf Fijivirus (RBSDV) and maize rough dwarf Fijivirus (MRDV) was investigated. It was previously reported that RBSDV S7 and S8 correspond to MRDV S6 and S7, respectively, based on nucleotide sequences (Marzachi et al., Virology, 1991). However, present comparison of the genomic segments of the two viruses in polyacrylamide gel electrophoresis together with the hybridization studies with the cDNA probes showed correspondence of the two virus segments as RBSDV S7-MRDV S7 and RBSDV S8-MRDV S8. Hybridization studies using cDNAs to all the RBSDV segments except for S3 as a probe showed that extent of sequence relatedness was found to differ among the segments. Partial cDNA clones to RBSDV S1, S4, S5and S6 reacted substantially weaker to MRDV genome than those to the other segments tested. And the cDNAs to the RBSDV S5 and S6 were found to be useful to differentiate the two viruses.