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María A. Ayllón, Massimo Turina, Jiatao Xie, Luca Nerva, Shin-Yi Lee Marzano, Livia Donaire, and Daohong Jiang
A summary of this ICTV Report chapter has been published as an ICTV Virus Taxonomy Profile article in the Journal of General Virology, and should be cited when referencing this online chapter as follows:
María A. Ayllón, Massimo Turina, Jiatao Xie, Luca Nerva, Shin-Yi Lee Marzano, Livia Donaire, Daohong Jiang, and ICTV Report Consortium. 2020, ICTV Virus Taxonomy Profile: Botourmiaviridae, Journal of General Virology, (In Press)
The family Botourmiaviridae includes viruses infecting plants and filamentous fungi with positive-sense single-stranded RNA genomes. The genus Ourmiavirus was previously unassigned to a family and was the subject of a previous ICTV Report chapter and Virus Taxonomy Profile (Turina et al., 2017). The family includes four genera: Ourmiavirus, Botoulivirus, Magoulivirus and Scleroulivirus. Members of the genus Ourmiavirus are plant viruses with non-enveloped bacilliform virions composed of a single coat protein. The genome consists of three segments, each one encoding a single protein. Virions possess a unique structure with a series of discrete lengths from 30 to 62 nm. The RNA-directed RNA polymerase has closest similarity to that of invertebrate viruses related to viruses of the family Narnaviridae; the movement protein is similar to the movement proteins of tombusviruses; the coat protein shows limited similarity to the coat proteins of several plant and animal viruses. Members of the other three genera are non-encapsidated fungal viruses with a genome of 2000–3200 nucleotides. Virus full-length genomes contain a unique open reading frame encoding an RNA-directed RNA polymerase with closest similarity to that of ourmiaviruses.
Table 1.Botourmiaviridae. Characteristics of members of family Botourmiaviridae
Ourmia melon virus VE9 (RNA1: EU770623; RNA2:EU770624; RNA3: EU770625), species Ourmia melon virus, genus Ourmiavirus
Ourmiavirus: bacilliform (18 nm × 30–62 nm) with a 23.8 kDa coat protein. Members of other genera are not encapsidated
Positive-sense RNA; monopartite (Botoulivirus, Magoulivirus and Scleroulivirus) (2–3 kb) or tripartate (Ourmiavirus) (2.8; 1.1; 0.97 kb)
Cytoplasmic; virion assembly is coupled to active replication
From genomic RNA; each genomic segment is monocistronic
Plants and fungi
Realm Riboviria, kingdom Orthornavira, phylum Lenarviricota, class Miaviricetes, order Ourlivirales, family Botourmiaviridae, genera Ourmiavirus Botoulivirus, Magoulivirus and Scleroulivirus, each with multiple species
The bacilliform virions of ourmiaviruses constitute a series of particles with conical ends (apparently hemi-icosahedra) and cylindrical bodies 18 nm in diameter. The bodies of the particles are composed of a series of double disks, the most common particle having two disks (particle length 30 nm), a second common particle having three disks (particle length 37 nm) with rarer particles having four disks (particle length 45.5 nm) or six disks (particle length 62 nm) (Figure 1.Botourmiaviridae). There is no envelope (Figure 2.Botourmiaviridae).
Figure 1.Botourmiaviridae. Diagram of virion surface of a member of the genus Ourmiavirus, showing arrangement and number of double disks and conical ends in particles of different length. Each row of five triangles represents a double disk.
Figure 2.Botourmiaviridae. Virion morphology. (A, B, C) Negative-contrast electron micrographs (uranyl acetate) of purified particles of Ourmia melon virus. The bar represents 100 nm. (D, E) Features of the two commonest particle types, enhanced by photographic superimposition.
For members of the genus Ourmiavirus the Mr of virions and their sedimentation coefficients are not known. The buoyant density in CsCl of all particle sizes is 1.375 g cm−3. The particles are stable at pH 7. Thermally, ourmiaviruses are relatively stable; infectivity is retained in crude sap after heating for 10 min at 70 °C but not 80 °C, and is retained after at least one freeze–thaw cycle. The particles are stable after CsCl density gradient centrifugation, treatment with Triton X-100, and treatment with chloroform but not n-butanol.
Ourmiaviruses have a genome comprised of three linear, positive-sense, ssRNA molecules. Members of the other three genera have a monosegmented genome. In Ourmia melon virus (OuMV), the three RNAs are 2814, 1064 and 974 nt, similar to those of other members of the genus (Rastgou et al., 2009). In Botrytis ourmia-like virus (BOLV) (genus Botoulivirus) the RNA is 2903 nt (Donaire et al., 2016); members of the genera Botoulivirus , Magoulivirus and Scleroulivirus have similar or smaller genomes. The genome of Magnaporthe oryzae ourmia-like virus 1 (MOLV1) isolate Guy11, a representative isolate of the genus Magoulivirus, is polyadenylated at the 3′-end (Illana et al., 2017); this is not observed for members of other genera.
The single structural protein or coat protein (CP) of Ourmia melon virus is 23.8 kDa and is encoded by RNA3.
In members of the family Botourmiaviridae, each genomic RNA has one ORF (Figure 3.Botourmiaviridae). RNA1 of Ourmiavirus or the unique RNA of the other three genera encodes a protein carrying the highly conserved core domain GDD typical of an RNA-directed RNA polymerase (RdRP) (Rastgou et al., 2009, Donaire et al., 2016, Illana et al., 2017, Marzano et al., 2016, Marzano and Domier 2016). Ourmia melon virus encodes a 23.8 kDa CP (encoded by RNA3) and the two non-structural proteins RdRP (97.5 kDa, encoded by RNA1) and the movement protein (MP, 31.6 kDa, encoded by RNA2) (Crivelli et al., 2011). Similarly-sized proteins are predicted to be encoded by members of the species, Epirus cherry virus and Cassava virus C (Rastgou et al., 2009).
Figure 3.Botourmiaviridae. Schematic genome organization of representative isolates of the four genera of the family Botourmiaviridae showing the size of each RNA and the positions and coding capacity of the ORFs. CP, coat protein; MP, movement protein; RdRP, RNA-directed RNA Polymerase.
A protein fusion of the CP to GFP localizes specifically to the nucleolus (Rossi et al., 2014) but there is no direct evidence of the CP in the nucleus during infection (Rossi et al., 2015). Synthesis of CP from actively replicating RNA3 is necessary for both virion assembly and systemic infection of the host (Crivelli et al., 2011). There is no evidence for the presence of subgenomic RNAs or the production of additional proteins by ribosomal readthrough or frameshifting. The MP may undergo post-translational modification. Alanine scanning mutagenesis of conserved residues in the MP showed their importance in determining symptoms, movement, and formation of tubular structures that may play a role in cell-to-cell movement (Margaria et al., 2016). Details of replication are not known except that CP interferes with the plant silencing defense only in the context of virus infection (Rossi et al., 2015).
The 81.88 kDa RdRP of Botrytis ourmia-like virus is encoded by the monosegmented genomic RNA (Donaire et al., 2016), and predicted to be similar or smaller for members of the other three genera. Mycoviruses can survive inside their fungal hosts without capsid or movement proteins, hence, it is assumed that members of genera Botoulivirus, Magoulivirus and Scleroulivirus may require only the RdRP for their replication.
Virions of Ourmia melon virus (i.e. the assembled coat protein) are good immunogens, as are the tubular structures associated with the MP. Antisera to these proteins do not react in Western blots to proteins in extracts of plants infected with members of either of the other two virus species in the genus Ourmiavirus.
Ourmia melon virus can easily be mechanically transmitted to a wide range of dicotyledonous plants (about 40 host species in 15 families have been reported), usually inducing systemic ringspots, mosaic and necrosis, with local lesions on some hosts. Tissue tropism is controlled by the KR-rich region at the amino terminus of the CP (Rossi et al., 2015). No vector has been identified but several species of weeds around infected fields are commonly infected, suggesting horizontal transmission of the virus. However, experimental transmission has not been obtained with several aphid species, the whiteflies Trialeurodes vaporariorum and Bemisia tabaci, or the mite Tetranychus urticae. Attempts at transmission through soil or irrigation water have been unsuccessful. Experimental seed transmission rates are 1–2% in Nicotiana benthamiana and N. megalosiphon. Members of different species in the genus Ourmiavirus occur in geographically diverse areas and on widely different hosts, though there are experimental hosts in common. Members of the other three genera in the family infect fungi in different genera such as Botrytis, Sclerotinia, Magnaporthe and Rhizoctonia, or have been associated with soybean leaves (Donaire et al., 2016, Illana et al., 2017, Marzano et al., 2016, Marzano and Domier 2016).
The genus Ourmiavirus is clearly separated from the other three genera based on host (plants rather than fungi), the number of genomic segments (three segments rather than one segment) and the presence of a MP and a CP (only for Ourmiavirus). The demarcation criteria for the other three genera that include members infecting fungi, (Botoulivirus,Scleroulivirus, Magoulivirus) is based on differences in RdRP amino acid sequence. Members of different genera in the family Botourmiaviridae are >70% different in complete RdRP amino acid sequence.
Ourmia: from Ourmia (Urmia, Orumieh), a city in north-western Iran where the Ourmia melon virus was first found
Botouli: from Botrytis and ourmia-like
Magouli: from Magnaporthe and ourmia-like
Sclerouli: from Sclerotinia and ourmia-like
There are four recognized genera within the family Botourmiaviridae. Phylogenetic relationships of conserved regions in the RpRP gene of different genera and species in the family Botourmiaviridae are depicted in Figure 4.Botourmiaviridae. Members of the family form a monophyletic bootstrap supported group, as do each of the four genera. Several currently unclassified ourmia-like viruses form a fifth, bootstrap-supported cluster and are candidates for the assignment of a fifth genus based on the depicted genetic relationships of the RdRP gene.
Figure 4.Botourmiaviridae. Phylogenetic relationships among members of the four genera in the family Botourmiaviridae. Maximum-likelihood phylogenetic tree based on the core RdRP motifs of members of the family Botourmiaviridae, other related viruses, and representatives of the families Leviviridae and the polyphyletic Narnaviridae. The viral RdRP sequences were aligned using MAFFT version 7 employing the E-INS-I algorithm. All alignments were trimmed so that they only contained the conserved RdRP region. All ambiguously aligned regions were then removed using the TrimAl program. The Maximum-Likelihood (ML) tree was inferred with PhyML 3.0 using eBayes method and the LG (G+I) protein substitution model, with SPRs algorithms and 8 categories of γ-distributed substitution rates. The reliability of internal branches was evaluated based on SH-aLRT supports. Numbers at the nodes indicate highly supported bootstrap values over 70% (1000 replicates). This phylogenetic tree and corresponding sequence alignment are available to download from the Resources page.
Viruses of the genus Ourmiavirus are exceptional, having particles of unique morphology and a unique combination of phylogenetic affinities for the three different genomic RNAs. The MP encoded by RNA2 has clear similarities with the MPs of viruses in the family Tombusviridae (Rastgou et al., 2009). The CP shows distant affinities with the CPs of sobemo, tombus- and luteoviruses (plant viruses) and nodaviruses (animal viruses) (Rastgou et al., 2009). The RdRP encoded by RNA1 has affinity with the RdRP of ourmia-like viruses from invertebrate hosts (Shi et al., 2016).
Aspergillus neoniger ourmia-like virus 1
Cladosporium uredinicola ourmia-like virus 1
Epicoccum nigrum ourmia-like virus 2
Neofusicoccum parvum ourmia-like virus 1
Penicillium sumatrense ourmia-like virus 1
Phaeoacremonium minimum ourmia-like virus 1
Phomopsis longicolla RNA virus 1
Pyricularia oryzae ourmia-like virus 1
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