Cytokine and immunoglobulin isotype profiles during CP7_E2alf vaccination against a challenge with the highly virulent Koslov strain of classical swine fever virus

doi:10.1016/j.rvsc.2014.01.002

Research in Veterinary Science

Volume 96, Issue 2, April 2014, Pages 389-395

https://doi.org/10.1016/j.rvsc.2014.01.002 Get rights and content

Abstract

CP7_E2alf is a promising marker vaccine candidate against classical swine fever (CSF). To better understand the mechanisms of protection, cytokine and isotype-specific antibody profiles were investigated in CP7_E2alf vaccinated pigs before and after challenge with the highly virulent CSFV strain “Koslov” at 14 days or 6 months post-vaccination. The interference of vaccination with CSFV pathogeny-related cytokine responses, previously described following a moderately virulent challenge, was confirmed. However, the levels of additional cytokines, TNF-α and IL-6, were significantly attenuated by vaccination following highly virulent challenge. This vaccine interference with cytokine response was not dependent on the immunization route or the consequence of competition between vaccine and challenge strain. Interestingly, IFN-γ enhancement and persistent high IgG2 levels suggested an important role of cell-mediated immunity in long-term protection against CSFV induced by CP7_E2alf vaccination. IgA production also revealed a stimulation of mucosal immunity, especially after oral administration of the vaccine.

Introduction

Classical swine fever (CSF) is a highly contagious viral disease in swine. Highly virulent CSFV strains can lead to acute haemorrhages, severe immune suppression and high mortality. CSFV is a small enveloped Pestivirus in the Flaviviridae family. Its RNA genome encodes 12 proteins, including the E2 glycoprotein, which is the most immunogenic one (Wensvoort et al., 1990).

The CP7_E2alf vaccine, a live chimera of Bovine viral diarrhoea virus (BVDV) expressing the CSFV E2 protein shows a strong potential to control CSF with the possibility to differentiate infected from vaccinated animals (Reimann et al., 2004). The CP7_E2alf vaccine is also the first efficient and safe marker vaccine candidate for oral immunization of wild boar against CSFV (Koenig et al., 2007). The CP7_E2alf marker vaccine conferred full clinical protection against a challenge with the highly virulent Koslov CSFV strain 7 days after intramuscular or 14 days after oral immunization (Leifer et al., 2009). Complete protection against a Koslov challenge was also demonstrated 21 days after oral CP7_E2alf vaccination (Blome et al., 2012). Moreover, the CP7_E2alf vaccine induces long-term immunity against a Koslov challenge, for at least 6 months after intramuscular or oral administration (Gabriel et al., 2012).

Protection from the disease is commonly associated with a humoral (Th2) immune response with the production of CSFV-specific antibodies or neutralizing antibodies (NAb) that usually appear about 2 weeks post-vaccination (pv) (van Oirschot, 2003). The duration of immunity induced by the CP7_E2alf vaccine is also attributed to stable antibody titres (Gabriel et al., 2012). Information about isotype differentiation is currently lacking for the early or late systemic antibody response (IgM/IgG differentiation), the balance of adaptive immunity toward Th1 or Th2 response (IgG1/IgG2 differentiation) or mucosal immunity (IgA detection). Partial investigation of IgG sub-class differentiation revealed that CSFV-specific IgG2 levels induced by early challenge with the moderately virulent CSFV strain were higher in CP7_E2alf orally vaccinated pigs than in unvaccinated pigs (Renson et al., 2013). No data is as yet available for IgA or IgM.

A previous study involving early challenge with a moderately virulent strain of CSFV at 2 days pv had revealed that oral CP7_E2alf vaccination interfered with pathogeny-related cytokine responses, e.g. attenuated the increase of IFN-α and IL-12 levels or the reduction of TGF-β1 level in serum (Renson et al., 2013). In this study, cytokine responses that might be influenced by CSFV infection were further investigated and immunoglobulin (Ig) G isotype profiles were completed with those for IgM and IgA, on samples obtained from different experiments, in which challenges with the highly virulent Koslov strain of CSFV had been applied later i.e., at 14 days or 6 months post-vaccination. The immune profiles obtained after oral or intramuscular immunizations were also compared. This study not only confirmed our initial results but also provided new elements about the responses induced by CP7_E2alf vaccination during the onset and duration of immunity.

Section snippets

OOI (onset of immunity) study – challenge at 14 days post-vaccination (dpv)

Serum samples, collected at 0, 4, 7, 10, 14 and 21 days post-infection or post-challenge (dpi) were kindly provided by NFCSO-DVMP from their efficacy study (unpublished study). Samples were obtained from five 6-week-old Kahyd pigs orally vaccinated with a single dose of the CP7_E2alf vaccine solution (1.6 ml at 10^5.6 TCID₅₀/ml) then oronasally challenged at 14 dpv with the highly virulent Koslov strain (2 ml at 10^5.5 TCID₅₀/ml). Three sera from an unvaccinated positive control group were also

Effect of CP7_E2alf vaccine on cytokine responses induced after Koslov CSFV strain infection

In the OOI study, a challenge with the highly virulent Koslov strain at 14 dpv led to an increase in the serum levels of IFN-α, TNF-α, IL-6 and IL-12 in the unvaccinated controls (Fig. 1b–e) whereas the level of TGF-β1 was reduced at 7 and 10 dpi in the last surviving pig (Fig. 1a). Prior to the challenge, no significant differences were observed in the levels of any cytokine between vaccinated pigs and unvaccinated pigs.

After challenge, the CP7_E2alf vaccination prevented the reduction of

Discussion

The role of cytokines in CSFV pathogeny or protection against CSFV infection induced by vaccination is not fully understood. In this study, the levels of the antiviral cytokine IFN-α and the cytokine coordinating innate and adaptive immune responses IL-12, were enhanced and that of the anti-inflammatory cytokine TGF-β1 reduced, following CSFV infection with the highly virulent “Koslov” strain. They were then modified by CP7_E2alf oral or i.m. vaccination. This confirms our previous results (

Acknowledgements

This work was funded by the EU Seventh Framework Programme (FP7/2007-2013) under Grant agreement no. 227003 CP-FP (project CSFV_goDIVA).

The authors thank Timea Barna, Katalin Fabian, Attila Farsang for their contribution in performing the animal experiment, collecting samples and careful correction of the manuscript.