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Antibodies to porcine circovirus (PCV) which is the smallest animal virus known so far were found in 77-95 per cent of sera from slaughter pigs gathered in Berlin and two districts of Northern Germany. About 60 per cent of these positive sera had relatively high titres similar to those in experimentally infected pigs 3-6 weeks after infection. This indicates that the animals might have become infected during the fattening period. Sera from 2-3 year old pigs from a laboratory animal breeding institution were also found positive (83 per cent) but titres were lower. Experimentally infected minipigs developed antibodies and virus was isolated from nasal swabs and from fecal samples. The animals neither showed any signs of illness nor were pathological changes noticable. The assumption that PCV is a common virus in all swine populations was strengthened by the finding of PCV antibodies in wild boars shot in the forests of the Berlin region.

Multiplication of porcine circovirus (PCV) was found to be inducible by treatment of infected cell cultures with 300 mM glucosamine. One day after glucosamine treatment and after growth in fresh medium, an increase in the number of cells containing virus antigen of up to 50 times as compared to mock-treated cultures was observed. Analysis of this phenomenon revealed that replication of PCV DNA was induced. Only aminohexoses but not hexoses and acetylated aminohexoses were efficacious. The course of PCV replication in synchronized cell cultures infected at different periods of the cell cycle showed that PCV DNA synthesis depends on cellular enzymes expressed during S phase growth of cells. However, whereas in cell cultures treated with glucosamine after infection in G0 or during G1, the start of PCV replication was observed during the first S phase after growth stimulation, the latent period in mock-treated cultures lasted until the second S phase. Also in cell cultures transfected with PCV DNA in G0 or during G1 using DEAE-dextran as mediator, PCV replication started during the first S phase after growth release of the cells. From these findings the conclusion is drawn that glucosamine and DEAE-dextran initiate PCV replication by enabling the PCV genome to get entry to the cell nucleus that normally can be achieved only by inclusion in the daughter nuclei at the end of mitosis.

Antibodies reacting with porcine circovirus (PCV) were found in sera of humans, mice, and cattle by means of an indirect immunofluorescence assay (IFA) and an ELISA. In man, the highest seroprevalence (23.9% in IFA and 30.2% in ELISA) was found among hospitalized patients with fever of partially unclear etiology. Non-hospitalized "healthy" persons of the former German Democratic Republic showed a significantly higher number of positive sera (IFA = 20%) than blood donors from Berlin-West (IFA = 8.6%). Murine sera reacted positive with PCV in IFA between 12 to 69% in different breeding groups and about 35% of cattle sera were found reactive with PCV in IFA. Double-staining IFAs, immuno-electron microscopy and immunoblotting showed that non-porcine antibodies reacted with PCV structural antigen. Mathematical analysis revealed that in ELISA, non-porcine antibodies reacted specifically with PCV. Loss of binding specificity of non-porcine antibodies in ELISA after storage of sera and lower maximal optical densities obtained at equal titers in ELISA with non-porcine than with porcine sera suggest that antibodies in man, mice and cattle are caused by related species specific viruses sharing antigenic epitopes with PCV.

An enzyme linked immunosorbent assay (ELISA) was developed for mass antibody screening to porcine circovirus (PCV) in pig herds of different age groups and of different husbandries. Infection with PCV was found to be common in all swine herds tested, with only one exception, a herd at a small farm. Statistically, the percentage of PCV negative sera decreased and titer levels increased with increasing age of the pigs. Within individual age groups, differences were found to exist between different husbandries. No correlation was detected between antibody levels and reproductive disorders in the herds.

Chicken anaemia agent (CAA) was characterized as a virion with 25 nm in diameter, with a buoyant density in CsCl of 1.36-1.37 g/cm3, and containing a circular, single-stranded DNA genome. The virus is composed of 32 hollow morphological units representing a regular T = 3 icosahedron.

Summary. By means of indirect immunofluorescence assay (IFA) using natural swine immune serum and hyperimmune serum from rabbits infected with porcine circovirus (PCV), a PCV antigen was detected present prior to the onset of viral and cellular DNA synthesis in nucleoli of cells of synchronized and growth stimulated infected PS cell cultures grown for more than 12 h in the presence of hydroxyurea. The number of cells containing specifically fluorescing nucleoli increased with increasing time of growth in the presence of hydroxyurea. The concomitant increase in the number of cells containing virus structural (VS) antigen in the nuclei and the increase in the amount of replicative (RF) DNA and accompanying 5 S DNA after release from the hydroxyurea block suggest that EA is involved in induction of PCV DNA replication. Primary pig kidney cell cultures persistently infected with PCV survived mock-infected control cultures for 16 passages. They had lost contact inhibition and formed cell colonies in soft agar at a ratio of 0.1 to 0.4%. Cell lines derived from agar colonies showed properties of transformed cells e.g. low requirement for serum growth factors, ability to overgrow a continuous cell layer, anchorage independence of growth. In transformed cells stimulated to growth and grown in the presence ofhydroxyurea, non-structural viral antigen visible by IFA in nucleoli and VS antigen located in the cytoplasm were expressed. Contrary to virus bound nuclear VS antigen in productive infection, accumulation of cytoplasmatic VS antigen was independent of DNA synthesis and caused cell destruction, thus limiting growth of cell layers and colonies in soft agar.

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