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An undescribed virus, here named ranunculus white mottle virus, was isolated in Italy from cultivated ranunculus showing mottle and distortion of leaves. The virus was mechanically transmissible to several herbaceous hosts. In negative stain, the particles appeared as circularised supercoiled threads 3 nm in diameter of different contour lengths; in some conditions the circles collapsed to form linear pseudobranched structures 9 nm in diameter. Immunolabeling of thin sections showed that viral antigen was widely distributed in the cytoplasm of parenchyma cells. The virus was not serologically related to the morphologically similar tenuiviruses, citrus psorosis-ringspot virus and tulip mild mottle mosaic virus. A major 43 kDa protein was present in purified preparations and in infected plant tissue, as also was a minor 28 kDa protein, serologically related to the major one. Nucleic acids extracted from purified particles consisted of at least three RNAs, of approximately 7.5, 1.8 and 1.5 kb, which appeared partly in single- and partly in double-stranded form. Purified preparations, but not viral RNAs, when mechanically inoculated, were infectious. Host range, tissue tropism, particle morphology and coat protein size place the virus closest to citrus psorosis-ringspot and tulip mild mottle mosaic viruses. These three viruses in turn show similarities with the Tenuiviruses and Bunyaviridae.

Three double-stranded (ds) RNA-containing viruses, named hop trefoil cryptic virus (HTCV) 1, 2 and 3, were found in apparently healthy seedlings of hop trefoil (Medicago lupulina). Only 1–2% of plants tested were virus-free. The particles occurred in the cytoplasm, nuclei and nucleoli of parenchyma cells. They were purified as a mixture (HTCV-M) with a mean yield of 50–100 µg/kg fresh tissue, and an antiserum was obtained. HTCVl was serologically related to one component (ACV1) of a mixture of alfalfa cryptic viruses (ACV-M) present in alfalfa (M. sativa), and was separated by immunoprecipitation from HTCV-M; its particles, about 30 nm in diameter, contained two dsRNAs (Mr 1.27 × 106 and 1.15 × 106) comigrating in PAGE with two RNAs of ACV-M. Particles of HTCV2, about 38 nm in diameter and with distinct morphological subunits, were serologically closely related to particles of similar morphology (RCCV2) present in a mixture of cryptic viruses (RCCV-M) from red clover (Trifoliumpratense), and were distantly related to white clover cryptic virus 2, also of similar morphology. HTCV3 had particles 30 nm in diameter, morphologically similar to those of HTCVl, and reacted with an antiserum to vicia cryptic virus. HTCV2 and 3 could not be individually separated. The RNAs of HTCV-M, in addition to the two bands attributed to HTCVl, gave, upon electrophoresis, three bands of estimated Mr 1.34, 1.08 and 0.95, all × 106, and up to five minor bands of Mr ranging from 1.45 to 0.98, all × 106.

White clover cryptic viruses (WCCV) 1, 2, and 3 were purified from apparently healthy plants with yields of about 200 μg of virus/kg of tissue. After isopycnic centrifugation in CsCl, WCCV 1 and 2 formed homogeneous bands at densities of 1.392 and 1.375 g/ml, respectively, whereas WCCV 3 was not recovered. Particles of WCCV 1 and 3 were 34 nm in diameter, without obvious morphological subunits, whereas WCCV 2 particles were 38 nm in diameter, with prominent morphological subunits. Antisera with titers up to 1:2048 were obtained against WCCV 1 and 2. Each of the three viruses contained two segments of linear dsRNA ( M r from 1.03 to 1.49 X 10 −6 . Virus-like particles, probably those of WCCV 1 and 2 were detected in low numbers in the cytoplasm of about 1/10 of the parenchyma cells of carrier plants examined by thin sectioning. The particles of WCCV 1 and 2 were not transmitted by grafting or mechanical inoculation, but were seed transmitted. Evidence is given that WCCV 1 and 2 multiply in their carrier plants, and are not viruses of associated parasitic or commensal organisms.

Summary Oat sterile dwarf virus (OSDV) subviral particles (SVPs) were purified from roots and stem bases of Lolium multiflorum. Electron microscopy of the preparations showed B-spiked SVPs typical of Fijiviruses (Fiji disease and maize rough dwarf-like viruses). Two sera with titres in gel-diffusion tests of 1/512 and 1/2048 were prepared against the SVPs. The sera did not react with the SVPs of maize rough dwarf and pangola stunt viruses when tested by gel-diffusion and immuno-electron microscopy. The first serum did not react with virus dsRNA or poly (1).poly(C) and the second serum had a titre of 1/8 with each of these. Poly-acrylamide gel electrophoresis of the nucleic acid from purified OSDV SVPs revealed ten dsRNA segments though only nine of these were resolved as separate bands in dsRNA extracted directly from infected plants. The genome pattern was broadly similar to but distinct from those of the known Fijiviruses. The RNA of the serologically related Arrhenatherum blue dwarf virus (ABDV), extracted directly from plants, appeared identical to that extracted directly from OSDV-infected plants. Both OSDV and ABDV were transmitted by the planthopper vector Javesella pellucida despite, in the case of OSDV, having been maintained in vegetatively propagated plants for 5 years.

Summary Pangola stunt virus (PSV) subviral particles (SVPs) were purified from roots and stem bases of Digitaria decumbens (pangola grass); electron microscopy of the preparations showed B-spiked SVPs typical of Fijiviruses. An antiserum of geldiffusion titre 1/1024 was obtained. Serological tests confirmed that both the B spikes and the inner capsid of PSV SVPs were related to those of maize rough dwarf virus (MRDV), though PSV SVPs also carried specific determinants. Isopycnic centrifugation of PSV RNA showed that it is double-stranded, with a G + C content of 41 to 43%. Polyacrylamide gel electrophoresis showed that the RNA consisted of ten segments similar to those of MRDV, though segment eight was present in less than equimolar amounts.

SUMMARY Small isometric virus-like particles were found in low concentration in apparently healthy carnations of the Mediterranean, miniature and Chinese type but not in eleven Sim cultivars tested. Most carnations containing these particles were from Italy but some were from France and the USA. The particles were not transmitted by grafting or by mechanical inoculation but were seed-transmitted to a large proportion of seedlings. Antisera to partially purified particles were obtained. The particles did not react with antisera to twenty-eight isometric plant viruses or virus-like particles but were serologically related to similar particles found in carnations in England, Holland and Israel. When negatively stained, the particles were isometric with a diameter of about 29 nm and a rounded rather than angular profile, but without clear substructure; some particles were penetrated by the stain. The particles remained intact in neutral sodium phosphotungstate. After isopycnic centrifugation in CsCl solution, preparations of particles formed a main band of mean density 1.377 g/ml and other fainter bands that varied in intensity and position in different preparations. In thin sections of carnations, no virus-like particles or cytological abnormalities were observed.

A serological analysis was made on maize rough dwarf virus fractions by immune electron microscopy, gel diffusion and slide precipitin tests to investigate the reactions of both protein and RNA. Antisera were made against different virus fractions, and existing sera against rice black streaked dwarf and maize rough dwarf viruses were also tested. Some of the sera contained antibodies against viral dsRNA. The serological behavior of poly[I]:poly[C] was similar to but not identical with that of viral RNA. One antigenic group in smooth cores was identified, and an additional group was found in spiked cores, associated with B spikes. Attempts to obtain antibodies against the outer capsid were not successful. A soluble antigen was identified as viral RNA. Disrupted protein preparations containing sodium dodecyl sulfate reacted with normal sera as well as virus antisera.

Two immune electron microscopic methods are described that allow rapid demonstration of (a) specific antibody coating of virus particles or (b) specific clumping and a tenfold increase in the number of particles visible on the grid. The operations take only 20 min and produce high-resolution images not only with purified preparations but also with crude preparations or those containing salts, buffers or sucrose.

Publisher Summary It is interesting that at this relatively late stage in virus research, new kinds of viruses continue to come to light. Cryptic viruses successfully evaded detection until recently because they induce no or, perhaps in some cases, very slight disease symptoms, are not transmissible in the ordinary way, and have particles present in such low concentrations as to escape casual discovery. The fact that cryptic viruses appear to cause no economically important disease has tended to give them low research priority. Nevertheless they are of interest as they contain double-stranded (ds) RNA, present only in another very different group of plant viruses, and appear unable to pass from carrier to non-carrier cells across a graft union. The latter is a decidedly non-virus like property. They are also of practical importance as they may be responsible for misleading results with methods for the detection of single-stranded RNA viruses based on the presence in plants of the double-stranded replicative forms.

Alfalfa cryptic virus 1 (ACV1) was purified from alfalfa plant clone (Medicago sativa) and characterized. The genome of ACV 1 consists of two dsRNAs, one with an estimated M r of 1⋅27 × 106 (RNA 1) and the other of M r 1⋅17 × 106 (RNA 2); the virus capsid is built from one polypeptide of estimated M r 54000. An RNA-dependent RNA polymerase able to replicate the genomic RNAs in vitro is associated with purified virus particles. In vitro translation showed that each of the genomic RNAs encodes a polypeptide and that encoded by RNA 2 is the capsid protein. These polypeptides account for about 95% and 83% of the coding capacity of RNA 1 and RNA 2, respectively. In Western and Northern blot experiments close affinity was found between ACV 1 and hop trefoil cryptic virus 1, a cryptovirus found in Medicago lupulina. No affinity was detected with any other cryptovirus tested.