- Volume 83, Issue 12, 2002

Epstein–Barr virus (EBV) persistently infects B-cells in humans and can be shed into the saliva. The cellular source of infectious virus is uncertain. Hairy leukoplakia, an AIDS-associated lesion of the tongue, supports EBV replication in epithelial cells. However, the general significance of this observation has remained doubtful. Using immunohistochemistry and in situ hybridization, we demonstrate evidence of EBV replication in tongue epithelial cells in 4 of 168 samples from 84 autopsy cases. Thus, in patients who do not have AIDS, squamous epithelial cells of the tongue rarely support EBV replication. However, all individuals with evidence of EBV replication were either on immunosuppressive therapy or were terminally ill cancer patients, suggesting that an impairment of the immune system may have allowed EBV replication to occur at this site. Thus, our findings are consistent with the idea that EBV replication in oropharyngeal epithelial cells is an infrequent event.

Ovine herpesvirus 2 (OvHV-2) causes malignant catarrhal fever in cattle, pigs and deer. We have observed intact circular and linear OvHV-2 genomes in infected T cell lines derived from cows and rabbits. Bovine T cell lines were predominantly latently infected but rabbit T cell lines supported OvHV-2 productive cycle gene expression and virus capsids were demonstrated for the first time.

Direct evidence of in vivo apoptosis of Spodoptera litura larvae was demonstrated by haemocoel inoculation with wild-type Autographa californica multicapsid nucleopolyhedrovirus (AcMNPV) budded virus (BV). In sharp contrast to natural infection, cadavers did not melt, liquefy and melanize. Typical morphological changes of apoptosis in insect haemocytes post-infection, including blebbing of the cell surface, chromatin margination and condensation, vacuolization of the cytoplasm and formation of apoptotic bodies, were observed by light and electron microscopy. Total DNAs extracted from virus-infected haemocytes showed DNA ladders. Cleavage of chromatin DNA by endogenous endonucleases were detected in the cells of most tissues cells, including epithelial cells and fat body cells, using terminal dUTP nick end labelling assays. Virogenic stroma and viral nucleocapsids could be seen in the nuclei of a few haemocytes. Yields of BV and OV (occluded virus) produced from the infected S. litura larvae were much lower than from the infected S. exigua larvae. These data suggest that host apoptotic responses to virus infection reduce AcMNPV spread at the level of the organism and that apoptosis could be a host-range limiting factor for baculovirus infections.

During plaque purification of Spodoptera littoralis nucleopolyhedrovirus in S. littoralis Sl52 cell culture, a deletion mutant virus was isolated. Analysis of the biological properties of this mutant virus revealed an absence of per os infectivity of the occluded virus. Infectivity by injection of the non-occluded (budded) virus is not different between the wild-type and the deleted virus. Restriction analysis of the mutant virus genome revealed a 4·5 kb deletion within the NotI D fragment. The observed phenotype was mapped to the deleted region by rescue experiments. The deletion was characterized and the equivalent DNA fragment on the wild-type virus was sequenced. By co-transfecting the DNA of the deleted virus with plasmids derived from the wild-type virus, it was possible to determine that ORF 7 in this fragment is responsible for the observed phenotype. ORF 7, called pif (per os infectivity factor), is homologous to ORF 119 of Autographa californica nucleopolyhedrovirus. Similar ORFs are present in all sequenced baculoviruses. The product of this gene is an occlusion body-derived virion structural protein required only for the first steps of larva infection, as viruses being produced in cells expressing the gene but not containing it in their genomes are able to produce successful infections.

The HLA DR13 allele has been associated with a self-limited course of hepatitis B virus infection, possibly through the induction of a more vigorous hepatitis B core antigen (HBcAg) and/or hepatitis B e antigen-specific CD4+ T cell response. HBcAg-specific CD4+ T cell responses were investigated in three HLA DR13-positive subjects with self-limited, acute hepatitis B. HBcAg-specific, short-term T cell lines derived from these three subjects showed a dominant recognition of HBcAg peptides spanning aa 1–20 (P1), 11–30 (P2), 41–60 (P5), 111–131 (P12) and 141–160 (P15). In order to characterize these epitopes in more detail, CD4+ T cell clones and cell lines were generated using HBcAg. Surprisingly, 11 of 12 T cell clones examined recognized P15; one recognized P10 (aa 91–111). Of four T cell lines, two recognized P15 and two recognized P5. By peptide mapping, the minimal epitope of P15 was located to residues 147TVVRRRGRSP156.

Zucchini squash (Cucurbita pepo) is a systemic host for most strains of the cucumovirus Cucumber mosaic virus (CMV), although the long-distance movement of the M strain of CMV (M-CMV) is inhibited in some cultivars. However, co-infection of zucchini plants with M-CMV and the potyvirus Zucchini yellow mosaic virus strain A (ZYMV-A) allowed M-CMV to move systemically, as demonstrated by tissue-print analysis. These doubly infected plants exhibited severe synergism in pathology. Infection of zucchini squash by M-CMV and an attenuated strain of ZYMV (ZYMV-AG) showed a milder synergy in pathology, in which ZYMV-AG also facilitated the long-distance movement of M-CMV similar to that promoted by ZYMV-A. Variation in the extent of synergy in pathology by the two strains of ZYMV did not correlate with differences in levels of accumulation of either virus. Thus, the extent of synergy in pathology is at least in part independent of the resistance-neutralizing function of the potyvirus.

The RNA genome of Rice tungro spherical virus (RTSV) is predicted to be expressed as a large polyprotein precursor (Shen et al., Virology 193, 621–630, 1993 ). The polyprotein is processed by at least one virus-encoded protease located adjacent to the C-terminal putative RNA polymerase which shows sequence similarity to viral serine-like proteases. The catalytic activity of this protease was explored using in vitro transcription/translation systems. Besides acting in cis, the protease had activity in trans on precursors containing regions of the 3’ half of the polyprotein but did not process a substrate consisting of a precursor of the coat proteins. The substitution mutation of Asp2735 of the RTSV polyprotein had no effect on proteolysis; however, His2680, Glu2717, Cys2811 and His2830 proved to be essential for catalytic activity and could constitute the catalytic centre and/or substrate-binding pocket of the RTSV 3C-like protease.

Plant viruses move to adjacent cells with the use of virus-encoded cell-to-cell movement proteins. Using proteins produced by in vitro translation, we present evidence that the ‘69K’ movement protein of Turnip yellow mosaic virus (TYMV) is recognized as a substrate for the attachment of polyubiquitin chains and for subsequent rapid and selective proteolysis by the proteasome, the ATP-dependent proteolytic system present in reticulocyte lysate. Truncation of the 69K protein suggests the existence of two degradation signals within its sequence. We propose that selective degradation of virus movement proteins may contribute to the previously reported transient nature of their accumulation during infection.