Porcine circovirus type 2 (PCV2) causes increased mortality and poor growth or weight loss in apparently healthy swine. Therefore, methods to detect PCV2-specific antibodies in swine serum are important for prevention, diagnosis, and control of PCV2-associated diseases (PCVAD). In this study, PCV2 virus-like particles (VLPs) were used to develop a rapid, simple and economical indirect enzyme-linked immunosorbent assay to detect (with high sensitivity) PCV2-specific antibodies in swine serum. The PCV2 capsid protein (Cap) was overexpressed in E. coli after optimizing the cap gene. Subsequently, the soluble Cap was rapidly purified in one step by automated fast protein liquid chromatography (FPLC). The purified PCV2 Cap was shown by transmission electron microscopy and gel filtration chromatography to be capable of self-assembling into VLPs in vitro. Using the purified VLPs as antigens, optimal operating conditions for the VLP ELISA were determined. The concentration of PCV2 VLPs was 1 µg/ml per well, and the dilution factors for swine serum and horseradish peroxidase (HRP)-labeled goat anti-pig antibody were 1:150 and 1:4000, respectively. Out of 241 serum samples tested with this assay, 83.4 % were found to be positive. Importantly, the VLP ELISA had a total coincidence rate of 97.4 % (74/76) compared to an Ingezim PCV2 ELISA IgG assay. In summary, this rapid, inexpensive VLP ELISA has the potential to greatly facilitate large-scale investigations of PCV2-associated serotypes.
To investigate porcine circovirus type 2b (PCV2b) transmission by contact and vertical infection in Kunming mice (an outbred mouse stock deriving from Swiss albino mice with a high ratio of gene heterozygosis), four mice in cage 6 were inoculated with PCV2b and 25 mice without any treatment were placed into cages 1 to 5 (five mice in each cage). Seven days after being infected, the PCV2-binoculated mice were co-mingled with non-inoculated mice from cages 1 to 5 successively at 7, 14, 21, 28 and 35 days post infection (dpi), respectively, for 3 days. In addition, eleven pregnant mice were injected with PCV2b. Samples were collected from non-inoculated mice and three newborn mice from each litter for PCV2b detection by polymerase chain reaction (PCR) and immunohistochemistry (IHC). The PCR results showed that PCV2b transmission rate among mice in cages 1, 2, 3, 4 and 5 was 0/5, 2/5, 5/5, 5/5 and 1/5, respectively. PCV2b antigen signals generally appeared in most organs of the non-inoculated mice in which viruses were detected by PCR. PCV2b DNA was also detected in newborn mice of PCV2b-infected litters, and viral antigen signals were observed in their organs as well. PCV2b was transmitted in Kunming mice by contact, and it also caused vertical infection through the placenta.
Porcine circovirus type 2 (PCV2) has been known as a causative agent of reproductive failure in the sow. In the present study mouse model was used to investigate PCV2 infection. In order to investigate whether PCV2 can induce lesions of spermatocytes and oocytes, 6 male and 6 female mice were each inoculated intraperitoneally with PCV2b, and 3 male and 3 female mice mock-infected with cell culture supernatant served as controls. Samples of testes and ovaries from PCV2b-inoculated and mock-infected mice were investigated using PCR, histopathological, ultrastructural and immunofluorescent histochemical methods at 14 and 21 day post infection (dpi). The study revealed that in the virus-inoculated mice leydig cells in testes and granulosa cells in ovaries were degenerated, and a small number of spermatocytes and oocytes showed apoptosis. Positive PCV2b antigen signals were also observed in these apoptotic cells. It can be suggested that PCV2 can cause lesions of spermatocyte and oocyte prior to zygotes formation in its host.
The complete genome of porcine circovirus type 2 (PCV2) strain YiY-3-2-H5 contains a cytidine insertion at position 962 in open reading frame 1. This insertion causes a reading frameshift of the rep gene, and thereafter a premature stop codon is present at the 3′ terminal end of this gene.
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