The new plant virus family Flexiviridae is described. The family is named because its members have flexuous virions and it includes the existing genera Allexivirus, Capillovirus, Carlavirus, Foveavirus, Potexvirus, Trichovirus and Vitivirus, plus the new genus Mandarivirus together with some related viruses not assigned to any genus. The family is justified from phylogenetic analyses of the polymerase and coat protein (CP) sequences. To help to define suitable molecular criteria for demarcation of species, a complete set of pairwise comparisons was made using the nucleotide (nt) and amino acid (aa) sequences of each fully-sequenced gene from every available accession in the family. Based on the distributions and on inspection of the data, it was concluded that, as a general rule, distinct species have less than ca. 72% identical nt or 80% identical aa between their entire CP or replication protein genes.

Cowpea mild mottle virus (CMMV) has physicochemical properties typical of carlaviruses, but has remained unclassified due to a number of unusual properties, including no serological cross-reaction with l 8 carlaviruses; production of brush-like inclusion bodies in vivo; and the ability to be transmitted by whiteflies (Bermisia tabaci).In this paper we report the use of a carlavirus specific PCR primer to identify CMMV as a member of the carlavirus group. This is confirmed by nucleotide sequence (958 nucleotides) from the 3' terminal region of CMMV RNA which contains a partial open reading frame (ORF) having high similarity with the coat proteins of other carlaviruses. The sequence also contains an 11.7K ORF at the 3' terminus, containing a 'zinc-finger' motif which is unique to carlaviruses.Cowpea mild mottle virus (CMMV) has physicochemical properties which resemble those of members of the carlavirus group, namely the presence of filamentous particles c. 650 nm in length, consisting of a coat protein of 31-33K and a single-stranded RNA of Mr 2.5 x 10 6 [Jeyanandarajah and Brunt, 1993]. However, unlike other members of the carlavirus group which are transmitted non-persistently by aphids [Foster, 1992], CMMV is unusual in that it is transmitted in a non-persistent manner by whiteflies (Bemisia tabaci) [Jeyanandarajah and Brunt 1993]. In addition, CMMV has several other unusual properties compared with other carlaviruses, which include, no serological relationships with 18 recognised members of the carlavirus group, and the induction of unusual brush-like intracellular inclusions in infected plants [Brunt et al., 1983]. It has been suggested that CMMV remain unclassified until the taxonomic significance of these differences have been investigated further [Jeyanandarajah and Brunt, 1993].We report here attempts to develop a rapid and specific identification test for carlaviruses which can also be used to determine whether CMMV and similar unclassified viruses are species of the carlavirus genus. The test is based on the polymerase chain reaction (PCR) using primers to conserved and unique sequences in carlavirus RNAs.The overall genome organisation of carlaviruses is very similar to that of potexviruses with the virus genes of both genera being arranged (from 5' to 3') as replicase, a triple gene block (25K, 12K, and 7K), and coat protein open reading frame, with extensive similarity at the amino acid level between the equivalent proteins of potex-and carlaviruses. However, in addition to these proteins, carlavirus genomes contain an additional protein encoded by an open reading frame

The nucleotide sequences of part of the nuclear inclusion body b (NIb) gene, the complete coat protein gene and the 3h untranslated regions of narcissus latent virus (NLV) and Maclura mosaic virus (MacMV) were determined. Deduced amino acid sequences for the NIb and coat protein genes revealed that NLV and MacMV are closely related. Gel analysis and Western blotting of the coat proteins of NLV and MacMV from infected tissue or purified virus indicated that they have molecular masses of 39n5 kDa and 40 kDa (respectively), whereas estimates from deduced amino acid sequences suggested that they have molecular masses of 32n8 kDa and 34n1 kDa. Comparison of the NIb and coat protein sequences with other viruses showed that NLV and MacMV have close affinities with viruses of the Potyviridae and suggests that they should form a new genus of the family.Narcissus latent virus (NLV) was first described in 1966 (Brunt & Atkey, 1967) as a member of the carlavirus genus. It has been reported to induce very mild leaf chlorosis in the tips of narcissus leaves and affects many commercially important cultivars (Brunt, 1977). NLV has flexuous filamentous particles ca. 657 nm long and ca. 13 nm wide, and a coat protein estimated to be 32n6 kDa (Brunt, 1977), properties suggesting that NLV is a carlavirus. In preliminary tests its coat protein

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The properties of Elderberry latent virus (ELV) and Pelargonium ringspot virus (PelRSV) were compared. The viruses were largely indistinguishable in herbaceous host range and symptomatology, particle morphology, sedimentation coefficient and RNA profiles and size. They were also very closely related serologically with SDI differences in agarose gel double-diffusion tests of 1 to 3. Purified virus particle preparations of each virus contained isometric particles c. 30 nm in diameter that sedimented as a major component with an so2,,, of 112-115s. Purified virus particle preparations contained a major and a minor ssRNA species that In polyacrylamide gel electrophoresis (PAGE) had estimated sizes of c. 3.8 kb and c. 1.6 kb respectively. Plants of Chenopodium quinoa infected with ELV or PelRSV each contained three dsRNA species of c. 3.8, 2.6 and 1.8 kbp, although the smallest of these species was not evident in all preparations. Protein from purified virus particle preparations contained a major polypeptide that, in SDS-PAGE, had an estimated Mr of 40 000 (40K). However, after storage of purified virus particles for 7-1 0 days, protein preparations from PelRSV particles also contained an additional major polypeptide of estimated A( of 37 000 that is probably derived by degradation of the 40K protein; this additional component was not observed in freshly prepared preparations of ELV. Neither virus was found to be related serologically to 16 other viruses with isometric particles and similar properties. These data, together with the recent finding by other researchers that the smallest RNA species is a sub-genomic RNA, suggests that both viruses are members of the genus Carmovirus, and that PelRSV is a minor variant of ELV. However, the taxonomic status of these two viruses is discussed in relation to recent brief reports comparing the nucleotide and amino acid sequences of these two viruses.