New assays to measure equine influenza virus-specific Type 1 immunity in horses

Equine influenza virus (EIV) is a leading cause of respiratory disease in horses. Equine influenza infection induces a long-term immunity to re-infection. Recent strategies of vaccination aim to mimic this immunity by stimulating both antibody and cellular immune responses. Cell-mediated immunity (CMI) to influenza is well defined in man, but little has been done to characterise the responses in the horse. Additionally, the development of reliable assays for the measurement of equine CMI has lagged behind serological methods and vaccine development. In this study, two methods of measuring EIV-specific T lymphocyte responses have been developed. An EIV ‘bulk’ cytotoxic T lymphocytes (CTL) assay using equine dermal fibroblasts as target cells has been adapted from a method used in the 1980s. This method was also complemented with a new EIV-specific IFNγ synthesis assay. When compared with the measurement of EIV-specific IFNγ synthesis previously described, this method required the amplification of EIV-specific lymphocytes by culture and was sensitive enough to detect stimulation of EIV-specific T lymphocytes induced by experimental infection with EIV or vaccination with recombinant canarypox viruses coding for EIV-HA molecules. This study provides the tools to characterise the stimulation of CMI by the new generation of vaccines against equine influenza.