One of the three structural glycoproteins of classical swine fever virus (CSFV) is E0, a disulfide-bonded homodimer that induces virus-neutralizing antibodies and occurs in a virion-bound as well as a secreted form. E0 was shown to be similar to a family of fungal and plant ribonucleases. Purified E0 from CSFV-infected cells was a potent ribonuclease specific for uridine and inhibitable by zinc ions.

The genomic RNA ofpestiviruses is translated into a large polyprotein that is cleaved into a number ofproteins. The structural proteins are N terminal in this polyprotein and include three glycoproteins called EO, El, and E2 on the basis of the order in which they appear in the polyprotein. Using pulse-chase experiments, we show that a pestiviral glycoprotein precursor, E012, is formed that is processed into EO, El, and E2 in an ordered fashion. Processing is initiated by a nascent cleavage between the capsid and the translocated E012 followed by cleavage at the C terminus of E2. E012 is then rapidly cleaved to form E01 and E2. After E2 is released from the precursor, E01 is processed into EO and El. To identify the sites of cleavage, the N termini of the glycoproteins of the pestivirus classical swine fever virus (formerly termed hog cholera virus) were sequenced after expression in the vaccinia virus system. The N termini are Glu-268 for EO (gp44/48), Leu-495 for El (gp33) and Arg-690 for E2 (gp55). The sequences around the cleavage sites capsid/EO and E1/E2 conform to the rules known for cellular signal proteases, as does the sequence at the presumed C terminus of E2. The sequence upstream of the EO/El cleavage site also shows sequence characteristics of signalase processing sites but lacks the typical hydrophobic signal peptide; this cleavage site has characteristics in common with a site in flaviviruses that is also cleaved in a delayed fashion. The absence of any membrane-spanning region results in the shedding of EO by infected cells, and EO can be detected in the virus-free supernatant. Comparison of the sequences around the cleavage sites of pestiviruses suggests a general processing scheme for the structural glycoproteins. Comparison of the pestiand flaviviral structural glycoproteins suggests analogies between E012 and prM-E.

A majority of HCV RNA-positive samples were also cAg-positive during the PWP. The current cAg detection corresponds to 100,000 IU per mL of HCV RNA. Since low-titer samples would be identified only by single-donation NAT, which is often affordable only in developed countries, the cAg ELISA could offer a practical alternative for some countries. The doubling time for HCV RNA at the onset of viremia corresponds to a calculated mean delay of cAg detection during the virus burst phase of 2 or 5 days, when compared with minipool (5000 IU/mL) or single-donation NAT (50 IU/mL), respectively.

The glycoproteins E rns of classical swine fever virus (CSFV) and E rns and E2 of bovine viral diarrhoea virus (BVDV) are shown to be located at the surface of infected cells by the use of indirect immunofluorescence and by cytofluorometric analysis. The positive immunostaining of the cell surface was further analysed by immunogold electron microscopy and it could be shown that only extracellular virions were labelled. Gold granules were not seen at the cellular plasma membrane. In contrast to BVDV E2, the CSFV E2 of virions sticking to the plasma membrane was not accessible to the respective monoclonal antibodies. However, CSFV particles isolated from culture supernatant were able to bind both monoclonal anti-E rns and anti-E2 antibodies. For CSFV and BVDV, binding of anti-E rns antibodies to the virions was more pronounced than that of anti-E2. This finding was unexpected since E2 is considered to be the immunodominant glycoprotein.

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