Interferon-gamma response of PBMC indicates productive pseudorabies virus (PRV) infection in swine

Abstract

In Chinese Meishan/German Landrace cross-bred swine F2 generation interferon gamma (IFN-gamma) production by peripheral blood mononuclear cells (PBMC) was determined directly ex vivo at different time points after survival of a virulent pseudorabies virus (PRV) infection. This reactivity was compared with the reactivity of naïve PBMC. Significant IFN-gamma production was determined in ELISA and ELISPOT only after in vitro PBMC re-stimulation with PRV and not with the closely related bovine herpesvirus BHV-1. The PRV-specific IFN-gamma secretion from re-stimulated PBMC showed high levels 6 days after infection, before the presence of serum antibodies, and it persisted at a high level over a 3 months period. The response of a group of eight piglets infected intranasally with PRV varied. Only two animals showed the expected typical fever response. PRV specific IFN-gamma production by PBMC clearly indicated that infection had occurred. Early significant IFN-gamma production by primed PBMC turned out to be a reliable and specific ex vivo marker for cellular response against productive PRV infection in swine before antibody formation.

Fig. 1

Differential course of rectal body temperature in eight piglets after intranasal infection with virulent PRV strain NIA-3. (A) The course of the temperature of two animals which developed fever (>40 °C) until day 6 post infection. (B) The course of rectal temperature of two animals from which one (No. 29) showed a moderate rise in body temperature. (C) The mean temperature course is summarized for four animals which showed rather a drop in body temperature than a rise after intranasal infection. After intramuscular (i.m.) injection of the same dose of PRV in non-febrile animals (B and C) at day 6 after intranasal application (arrows) all six piglets developed fever within 6 days.

Fig. 2

IFN-γ response after in vitro (re)stimulation. PBMC from non infected (naïve) and PRV infected animals (4, 8, 12 weeks after infection) were either stimulated with ConA or with PRV. The content of IFN-γ in the PBMC supernatants was quantified after 24 and 72 h in the IFN-γ specific ELISA.

Fig. 3

Determination of antigen-specific induction of the frequency of IFN-γ producing PBMC in ELISPOT assays (2 × 10⁵ cells/well, 72 h) either re-stimulated with PRV (C) or stimulated with the closely related alpha herpesvirus BHV-1 (B). Only slightly elevated background spot numbers compared to culture medium controls (A) are present when BHV-1 is used as the stimulating virus in PRV primed PBMC from PRV challenge surviving piglets.