- Volume 70, Issue 1, 1989

Reassortant bunyaviruses derived from two members of the California serogroup (La Crosse/original and Tahyna/181-57) viruses were used to demonstrate that the large M r viral protein (L) is encoded by the L RNA segment. Radiolabelled viral proteins were analysed by discontinuous SDS–PAGE. The L protein of La Crosse virus was observed to migrate ahead of its Tahyna virus counterpart when electrophoresed through a 5% acrylamide resolving gel. Among the reassortant viruses, the L protein phenotype segregated with the viral L RNA segment. After confirming the genotype of the viruses used in this study, it was concluded that the L RNA species of California serogroup viruses codes for the L protein, the presumed viral polymerase.

Attempts to identify and classify distinct potyviruses and their strains have frequently been hampered by the presence of variable proportions of cross-reacting antibodies in antisera. Investigations of reactivities in electroblot immunoassays of 11 polyclonal antisera raised by injection of intact particles of potyviruses produced in different laboratories with 12 distinct potyviruses showed that such cross-reacting antibodies were directed towards the homologous core protein region of potyvirus coat proteins. A simple method was developed to obtain virus-specific antibodies using affinity chromatography. It involved removal of the surface-located, virus-specific N-terminal peptide region from particles of one potyvirus using lysyl endopeptidase, coupling of the truncated coat protein to cyanogen bromide-activated Sepharose gel, and passing antisera to different potyviruses through the column. Antibodies that did not bind to the column were found to be highly specific.

The sequence of the 3′-terminal 1611 nucleotides of the genome of the tobacco veinal necrosis strain of potato virus Y (PVYN) was determined. The sequence revealed an open reading frame of 1285 nucleotides, of which the start was not identified, and an untranslated region of 316 nucleotides upstream of a poly(A) tract. Comparison of the open reading frame with the amino-terminal sequence of the viral coat protein enabled mapping of the start of the coat protein at amino acid –267, and indicated that maturation of this protein requires proteolytic processing from a larger polyprptein precursor at a glutamine/glycine dipeptide sequence. The coat protein of PVYN displayed significant (51 to 63%) sequence homology to the coat proteins of four other potyviruses, tobacco etch virus, tobacco vein mottling virus, plum pox virus and sugarcane mosaic virus. Even higher sequence homology (91%) was detected with the coat protein of a fifth potyvirus, pepper mottle virus (PeMV). This homology was of the same level as found between the coat proteins of PVYN and a second strain of this virus, PVYD. Since, moreover, PVYN and PeMV were the only potyviruses displaying homology in the 3′-terminal, non-translated regions of their genomes, we conclude that PeMV should be regarded as a strain of PVY.

A small RNA (DI RNA), approx. 400 bp in length, was found in plants inoculated with cymbidium ringspot tombusvirus (CyRSV). DI RNA contained sequences derived from at least three regions of the CyRSV genome and was encapsidated in CyRSV protein. DI RNA was formed in plants inoculated with purified genome-size RNA and a double-stranded form was detected in dsRNA extracted from infected tissue. Symptoms of CyRSV infection in Nicotiana clevelandii were less severe when DI RNA was present in the inocula used suggesting that DI RNA interferes with symptom expression.

Virion RNA of beet yellows virus (BYV) is a messenger-sense RNA of about 14500 nucleotides containing no poly(A) or covalently linked protein. In υitro translation of BYV RNA yielded a prominent polypeptide of M r 250000, and some lighter products. None of the translation products was found to be immunoprecipitable with anti-BYV γ-globulins. The synthesis of all the BYV RNA-directed polypeptides was blocked by the cap analogue m7Gpp, thus suggesting the presence of a cap structure at the 5′ end of virion RNA.

The leaf hopper Graminella nigrifrons transmits maize chlorotic dwarf virus(MCDV) in a semi-persistent manner and loses its ability to inoculate plants following ecdysis. Virus-like particles (VLP) resembling MCDV in purified preparations and in MCDV-infected plants were observed by electron microscopy adhering to the cuticula lining the precibarium, cibarium (sucking pump), pharynx and fore-oesophagus in viruliferous leaf hoppers carrying MCDV. No VLP were observed in the alimentary canal beyond the cardiac valve, in any organ or tissue in the haemocoele, or in the maxillary salivary duct. No VLP were observed associated with the stylet tips or anywhere on the cuticula surfaces of either the mandibular or maxillary stylets. The VLP were not found associated with the cuticula lining of the foregut in a similarly treated non-vector species, Dalbulus maidis, or in vector or non-vector species given either a 48 h acquisition access feeding period on MCDV-infected source plants followed by a 48 h post-acquisition feeding on healthy plants (renders inoculative leaf hoppers non-inoculative) or solely a 48 h feeding on healthy plants. No differences were observed between MCDV-inoculative and non-inoculative Graminella given similar access to virus. The VLP occurred in thin layers or small to large clusters embedded in a lightly stained matrix material (m-material) or a densely stained substrate which, in turn, were apparently attached by m-material to the cuticula. It is thought that the VLP are those of MCDV and that the m-material, which resembles materials previously reported in association with virus-binding sites in the fore alimentary canals of aphid and nematode vectors, functions in binding virus to retention sites on the vector’s cuticula.