Genus Oryzavirus

Type Species	Rice ragged stunt virus	(RRSV)

Distinguishing Features

Oryzaviruses possess a genome consisting of 10 dsRNA segments. They are transmitted by viruliferous Delphacid planthoppers to plants in the family Graminae and replicate in both hosts and vectors. Oryzavirus dsRNA genome segments share characteristic conserved terminal sequences. The intact virus particle, which is double shelled, appears to be relatively unstable and can rapidly breakdown during isolation to produce the internal “core” particle. However, intact double shelled particles can be prepared from fixative treated infected leaf tissues.

Virion Properties

Morphology

Intact double shelled Rice ragged stunt virus (RRSV) appears to be icosahedral in symmetry and double shelled. It has a particle diameter in the range of 75-80  nm and surface ‘A-spikes’ (approximately 10-12  nm wide and 8  nm in length), attached to the end of the ‘B-spikes’ situated at the 5 fold axes of the viral core. The subviral or ‘core’ particles have an estimated diameter of 57-65  nm (Fig. 23) and possess 12 “B”-type spikes, 8-10  nm in height, 23-26  nm wide at the base and 14-17  nm at the top. In negatively stained preparations of RRSV, “B-spiked subviral” particles have been seen but intact double-shelled particles are not seen without pretreatment with fixative. Echinochloa ragged stunt virus (ERSV) particles are slightly larger than RRSV particles.

Physicochemical and Physical Properties

RRSV particles sediment as one component and are stable at pH 6.0-9.0. They are stable in 0.1  M MgCl₂. The B spikes dissociate from the core particle in 0.5  M MgCl₂ and the entire particle is disrupted in 2  M MgCl₂. The particles retain infectivity after 7 days at 4°C and after 10 minutes at 50°C but lose their infectivity after 10 minutes at 60°C. They retain infectivity after 3 cycles of freezing and thawing. The particles contain an RNA-dependent RNA polymerase.

Nucleic Acid

The oryzavirus genome consists of 10 linear dsRNA segments. The genomes of RRSV and ERSV have similar sizes and segment profiles (RRSV Mr 18.15 10⁶ (26,066  bp); ERSV Mr 17.78 10⁶) with segments ranging in size from Mr 0.6 to 2.9 10⁶ (1,162 to 3,849  bp). The genomic dsRNAs are termed segment 1 to segment 10 (S1 to S10), in order of increasing electrophoretic mobility in 7.5% polyacrylamide gels. The entire genome of RRSV has recently been sequenced; the S4 and S10 segments are larger than they appear by migration in polyacrylamide gels, suggesting that they may migrate in the position 3 and 9 respectively during agarose gel electrophoresis (AGE). The conserved terminal sequences of the ERSV genome segments are identical to those of RRSV (5-GAUAAA . . . (G)GUGC-3) but there are no further ERSV sequence data available. The RRSV and ERSV conserved terminal sequences differ from those of phytoreoviruses or fijiviruses. RRSV RNAs hybridize (weakly) with their counterparts in ERSV but not with segments of Rice dwarf virus (RDV).

Proteins

RRSV particles are composed of five major, highly immunoreactive structural proteins, with estimated Mr of 33, 39, 43, 70 and 120 10³, and at least five minor structural proteins (Mr 49, 60, 76, 90 and 94 10³). Three more proteins (Mr 31, 63 and 88 10³) have also been identified by in vitro translation of RRSV genomic dsRNA, and designated as non-structural proteins. RRSV S5, S8 and S9, respectively, encode a Mr 90 10³ minor structural protein (possibly a guanylyltransferase), a Mr 67 10³ major structural protein which is further self-processed to Mr 46, 43 and 26 10³ proteins, and a Mr 38 10³ major structural protein. There is evidence to suggest that P9 is involved in vector transmission. RRSV genome segments S7 and S10 encode non-structural proteins of Mr ~68 and 32 10³, respectively. RRSV S4 probably encodes an RNA-dependent RNA polymerase and a second protein of unknown function. ERSV particles are composed of four major structural proteins (Mr 127, 123, 63 and 34 10³) and three minor proteins (Mr 103, 50 and 49 10³). The reported differences in morphology of the outer capsids of RRSV and ERSV could be at least partially due to differences in the sizes of these structural proteins.

Lipids

None reported.

Carbohydrates

There are no evidence for the glycosylation of oryzavirus proteins.

Genome Organization and Replication

The genome organization is well characterized only for one oryzavirus, RRSV (Table 17). This virus has 9 dsRNA genome segments which contain single large ORFs (in one strand of the pair) and one segment (S4) containing two large ORFs. The proteins encoded by S3, S8 and S9 are major components of the RRSV particle, which of those encoded by segments S7 and S10 are not found in the virion. S8 codes for a polyprotein which appears to autocatalytically cleave into at least two polypeptides one of which is a major structural protein. The larger protein encoded by S4 appears to be an RNA dependent RNA polymerase (Pol) as it has the conserved glycine-aspartate-aspartate motif and other landmark amino acids typical of RDRPs found in other viruses. The tentative functions of the proteins encoded by the other segments are shown in Table 17. The viruses induce viroplasms in the cytoplasm of infected cells.

Antigenic Properties

RRSV and ERSV cross-react in serological tests. Polyclonal antisera raised against RRSV particle preparations react most strongly with P3, P8 and P9 (both native and in vitro produced) suggesting that they are highly immunogenic. P5 is weakly immunogenic. Glutathione-S-transferase-NSP7 fusion protein is highly immunogenic and antibodies against this protein are useful in ELISA for detection of RRSV in infected plants and insects.

Biological Properties

Oryzaviruses infect plants in the family Graminae, causing disease in rice (RRSV) and Echinochloa (ERSV). They are transmitted by, and replicate in phloem-feeding, viruliferous delphacid planthoppers (RRSV: Nilaparvata lugens; ERSV: Sogatella longifurcifera and S. vibix). RRSV is ingested when the hopper feeds on rice plants, usually at the seedling stage. The minimum acquisition access period for the vector is about 3  hrs, the latent period is about 9 days, and the minimum inoculation access time is about 1  hr. Planthopper nymphs are more efficient vectors than adults but all forms of the insect can act as vectors. Any individual viruliferous hopper gives intermittent transmission. The virus is not passed though the egg.

Oryzaviruses appear to replicate in fibrillar viroplasms within the cytoplasm of phloem, or phloem associated plant cells and in cells of the salivary glands, fat body, gut and brain of the planthopper. The phloem cells proliferate to form galls on the plant. RRSV has been reported in southeastern and far-eastern Asian countries where it affects rice yields (generally 10-20% loss, but up to 100% in severely affected areas). ERSV has been reported in Taiwan.

List of Species Demarcation Criteria in the Genus

The prime determinant for inclusion of virus isolates within a single virus species of members of the family Reoviridae, is the “ability to exchange (reassort) genome segments during co-infection, thereby exchanging genetic information and generating viable and novel progeny virus strains”. Data providing direct evidence of segment reassortment between isolates for oryzaviruses is very limited. Therefore serology, nucleic acid hybridization, and comparison of nucleic acid and protein sequences are the most common ways of determining the levels of similarity between isolates.

Members of a single Oryzavirus species may be identified by:

1.	Their ability to exchange genetic material by genome segment reassortment during dual infections, thereby producing viable progeny virus strains.
2.	RNA sequence similarities.
3.	PCR (using primers to conserved segments coupled with cross-hybridization analysis);
4.	High levels of serological cross reaction by ELISA or agar gel immunodiffusion, using antibodies against the whole virus particle proteins or against specific viral proteins.
5.	Cross-hybridization of “conserved” genome segments under high stringency conditions (Northern or dot blots, with probes made from viral RNA or cDNA).
6.	Identical conserved terminal regions of the genome segments (some closely related species can also have identical terminal sequences on at least some segments).
7.	Analysis of “electropherotype” by agarose gel electrophoresis.
8.	Identification of vector species, RRSV and ERSV have different planthopper vector species.
9.	Identification of plant host species.

List of Species in the Genus

Official virus species names are in italics. Tentative virus species names, alternative names ( ), strains or serotypes are not italicized. Virus names, insect vector and host names { }, genome sequence accession numbers [ ], and assigned abbreviations ( ) are:

Species in the Genus

Echinochloa ragged stunt virus		(ERSV)
{Sogatella longifurcifera, S. vibix} {Graminae: Echinochloa}
Rice ragged stunt virus	S1 [AF020334] Thai isolate	(RRSV)
{Nilaparvata lugens} {Graminae: Rice}	S2 [AF020335} Thai isolate
	S3 [AF020336] Thai isolate
	S4 [U66714] Thai isolate
	S5 [U33633] Thai isolate
	S6 [AF020337] Thai isolate
	S7 [U66713] Thai isolate
	S8 [U46682] Thai isolate
	S9 [L38899] Thai isolate
	S9 [l79969] Philippine isolate
	S9 [L38900] Indian isolate
	S10 [U66712] Thai isolate

Tentative Species in the Genus

None reported.

Phylogenetic Relationships within the Genus

Comparison of RRSV sequences with other viruses including plant reoviruses show little or no significant sequence similarities at either nucleic or amino acid level. However, the deduced amino acids of sequences around the conserved RDRP motifs of reoviruses suggest that oryzaviruses (RRSV) are more closely related to fijiviruses than to phytoreoviruses (Table 18).

Unassigned Species in the Family Reoviridae

Initial characterisations have been made of viruses isolated from insects or insect cell lines (ten or eleven genome segments) from crustacean (twelve genome segments) and other arthropods, which may indicate a requirement for the recognition of additional new genera (possibly three new genera based on the numbers of genome segments, i.e., 10, 11, or 12).

The unassigned insect viruses are non-occluded and double shelled (and therefore distinct from the cypoviruses), with icosahedral morphology, surface spikes on the core particle, a diameter of approximately 60  nm and a ten or eleven segmented dsRNA genome (estimated size 23.4  kbp). These reports suggest that these viruses are distinct from the cypoviruses and may form one or even two new genera of insect reoviruses. However, these genera have not yet been recognized by ICTV (Table 19).

Phylogenetic Relationships within the Family

See Fig. 24 and Tables 18 and 19.

Similarity with Other Taxa

See text.

Derivation of Names

Aqua: from Latin aqua, “water”.

Colti: sigla from Colorado tick fever.

Cypo: sigla from cytoplasmic polyhedrosis.

Fiji: from name of country where virus was first isolated.

Orbi: from Latin orbis, “ring” or “circle” in recognition of the ring-like structures observed in micrographs of the surface of core particles.

Oryza: from Latin oryza, “rice”.

Phyto: from Greek phyton, “plant”.

Reo: sigla from respiratory enteric orphan, due to the early recognition that the viruses caused respiratory and enteric infections, and the (incorrect) belief that they were not associated with disease, hence they were considered “orphan” viruses.

Rota: from Latin rota, “wheel”.