Kinetics of humoral immune response to the major structural proteins of the porcine reproductive and respiratory syndrome virus

Abstract

The kinetics of appearance of antibodies directed to the major structural proteins N, M and E of porcine reproductive and respiratory syndrome virus (PRRSV) was followed in pigs naturally- and experimentally-exposed to the virus. Specific IgM antibody titers were first detected by indirect immunofluorescence (IIF) at the end of the first week of PRRSV infection, peaked by day 14 to 21 post-inoculation (p.i.), then rapidly decreased to undetectable levels by day 35 to 42 p.i. On the other hand, specific IgG antibody titers peaked by day 21 to 28 p.i. and remained unchanged to the end of the 6- or 9-week observation period; in addition, a persistent viremia was observed. Virus neutralizing (VN) antibody titers > 8 were not detected until 3 to 4 weeks p.i. Taken together, the results obtained by Western blotting analyses using purified virus and E. coli-expressed ORFs 5 to 7 gene products, suggested that antibodies directed against the envelope E protein appear by day 7 p.i., whereas antibodies directed against the nucleocapsid N and membrane M proteins can only be detected by the end of the second week p.i. No correlation could be demonstrated between VN and IIF antibody titers, viremia, and viral protein specificities of circulating antibodies at various times p.i.

Alternative codon usage of PRRS virus ORF5 gene increases eucaryotic expression of GP5 glycoprotein and improves immune response in challenged pigs

“…The VN antibodies response detected at 10 days post-challenge was the specific consequence of prior immunization with hAdV/synORF5 since normally, VN antibodies are detected after only 3 to 4 weeks in SPF pigs experimentally-or naturallyinfected with PRRSV [10,20]. The higher reactivity to GP5, as well as the higher VN activity of pigs sera that have been vaccinated either with hAdV/wtORF5 or hAdV/synORF5 is in agreement with previous findings indicating that GP5 is the primary structural glycoprotein of PRRSV involved in virus neutralization activity [6,7,25].…”

“…Following an incubation period of 24 and 48 h post-infection, those hAdVs infected cells were fixed with a 80% cold acetone solution for 20 min at 4 o C and the IIF was done using a 1/200 dilution of the α5 serum as previously described [20].…”

“…Sucrose gradient purified-PRRSV was prepared in LB-2 lysis buffer [5], denatured by boiling in the presence of 5% (V/V) β-mercaptoethanol, subjected to 12% SDS-PAGE and electrotransferred onto nitrocellulose membranes as previously described [5,20]. Reactivity of pig sera (dilution 1/50) was tested and the α5 serum was used as a positive control.…”

Characterization of Antibody Response to Porcine Reproductive and Respiratory Syndrome Virus ORF5 Product following Infection and Evaluation of its Diagnostic Use in Pigs

“…The ELISA most likely detects ORF 7 and 6 epitopes, which are the most abundant antigens in the virion, whereas the WB only detects a specific antibody to ORF 5 product. Two research studies have demonstrated that the ORF 5 product is underpresented in virus preparations grown in tissue culture 12,18 making the ELISA inherently less sensitive than the WB assay. As previously reported, 12 the antibodies to recombinant ORF 5 product could be demonstrated as early as 6 days PI, whereas antibody to ORFs 6 and 7 could only be detected by the end of the week 2 PI.…”

“…For example, a high molar ratio of ORF 7 and 6 proteins, relative to ORF 4 and 5 proteins, is found in the virion. 12,18 Antibodies to ORF 5 protein appear to be major serologic markers in PRRSV-infected pigs. This finding is based on the detection of antibodies by Western blotting (WB) over a period of 150 days in infected animals (unpublished data).…”