S. J. Martin

The role of defective interfering particles in persistent infection of Vero cells by measles virus.

Persistent infections by measles virus were rapidly established in the majority of Vero cells when monolayers were infected with virus stocks that had been passed three to five times from an undiluted inoculum. These virus stocks had low infectivity titres but normal haemagglutinin titres and were able to cause interference. The ability of such virus stocks to establish persistent infections seems to be due to the presence of defective interfering particles rather than of virus mutants. Measles virus released from a persistently infected Vero cell line at the 93rd passage had properties similar to the undiluted passage virus that generated persistent infections.

Chemically synthesized peptides elicit neutralizing antibody to bovine enterovirus.

Synthetic peptides representing 14 regions of the bovine enterovirus structural proteins were used to raise antibodies in mice. The peptides were predicted using amino acid sequence alignments with the position of antigenic sites on other picornaviruses. Five of the anti-peptide antibodies reacted with the virus in an immunoprecipitation test. Furthermore, each of these anti-peptide antibodies neutralized virus infectivity; those directed against peptides of VP2 and VP3 neutralized to a greater extent than those directed against peptides of VP1. The positions of these epitopes in the viral structural proteins are discussed in relation to corresponding positions in other picornaviruses.

cDNA cloning of the nucleocapsid and nucleocapsid-associated protein genes of mumps virus.

cDNA clones of the mumps virus N and P messenger RNAs were isolated from an infected cell cDNA library. The N and P clones selected the two predominant polyadenylated RNAs found in mumps virus-infected cells with mol. wt. of 0.69 X 10(6) and 0.51 X 10(6), respectively. In addition, clones of the P gene hybridized to and selected mRNAs of higher mol. wt. probably representing polycistronic transcripts of the mumps genome. Hybrid-select translation experiments confirmed the specificity of the clones as representing the nucleocapsid (N) and nucleocapsid-associated protein (P) genes.

The nucleotide sequence of the 5'non-coding and capsid coding genome regions of two bovine enterovirus strains.

SummaryThe sequence of cDNA clones representing the 5′ non-coding regions (NCR) and capsid regions of two bovine enteroviruses (strains PS-87 and RM-2; serotype two viruses) have been determined and compared with that obtained from a serotype one strain (VG-5-27). All three strains showed a longer 5′ NCR compared to human enteroviruses and rhinoviruses due in part to a hundred residue insertion approximately at a hundred residues in from the 5′ end. However, another domain occurring at nucleotide 187–222 in poliovirus is absent in each bovine enterovirus. Comparisons of the predicted structural protein amino acid sequences indicate that PS-87 shares most sequence identity with RM-2 and then with VG-5-27 in that order. The VP1 protein of PS-87 and RM-2 are shorter than the equivalent VP1 of VG-5-27 due in part to a truncation at their C-terminii. VP3 is only slightly smaller than VP2 in each virus.

Characterization of neutralizing antibodies to bovine enterovirus elicited by synthetic peptides

SummarySix synthetic peptides corresponding to regions of bovine enterovirus (BEV), strain VG-5-27, elicited antibodies in mice which reacted with the virus in various assays. These antibodies have been characterised on the basis of their ability to (1) neutralize the virus, (2) bind to the intact virus particle in an immunoprecipitation test, (3) react with the denatured viral proteins, and (4) give immunofluorescent staining of virus infected cells. We have also determined the proportion of antipeptide antibody which binds to the virus in each case. All of the sera immunoprecipitated the virus and neutralized its activty to varying extents. Two of the sera specific for VP 1 sequences failed to react with denatured VP 1 whereas all the other antisera reacted with their respective parental proteins. All of the sera reacted with VG-5-27 infected cells in an immunofluorescence test. The proportion of antibodies to each peptide recognizing intact virus was variable and did not appear to correlate with neutralizing activity. In addition, the ability of each of the sera to react with and neutralize three other strains of the virus was analysed. With one of these strains significant cross-neutralization was observed.

Conformational changes during proteolytic processing of a picornavirus capsid proteins

Summary. We have used synthetic peptide antibodies to probe conformational changes that occur during the cleavage cascade which generates the capsid proteins of a picornavirus. The initial translation product of 97 kDa, the precursor of all four structural proteins, is cleaved to form a 63 kDa fragment which, we show, has significantly different folding characteristics to both its larger parent and its products. We demonstrate that proteolytic cleavages as distant as 520 residues from epitopes confer sufficiently large conformational changes as to render them unrecognisable. To our knowledge, this is the first demonstration of this phenomenon in the picornavirus system.

The Generation of Small-Plaque Mutants during Undiluted Passage of Canine Distemper Virus

A culture of Vero cells persistently infected with undiluted-passage, large-plaque canine distemper virus was found to release small-plaque virus (SPV). This suggested that SPV could be generated by large-plaque virus (LPV). However, attempts to induce small-plaque mutants from LPV with 5-fluorouracil resulted in the production of virus that expressed a variable plaque type and induced a spectrum of disease in hamsters. In contrast, small-plaque mutants, shown to arise during undiluted passage of LPV, retained their small-plaque characteristic and resembled our original SPV isolate in their cytopathogenicity, immunogenicity, and neurovirulence for hamsters.