Use of tetanus toxoid as a differentiating infected from vaccinated animals (DIVA) strategy for sero-surveillance of avian influenza virus vaccination in poultry.

Strategies for differentiating infected from vaccinated animals (DIVA) require improvement for increased surveillance of avian influenza (AI), where vaccination is employed to control disease. We propose a novel DIVA approach for chickens using tetanus toxoid (TT) as an exogenous marker independent of serotype and relatedness of circulating and vaccine strains. Of 1779 chickens tested from Australia, Hong Kong and China, 100% were seronegative for TT-specific antibodies without vaccination. Tetanus toxoid adjuvanted to mineral oil was immunogenic in chickens. Co-delivery of both TT and inactivated LPAI (H6N2) vaccines in chickens elicited strong TT and influenza-specific antibody responses, which persisted to 53 weeks post-vaccination. Furthermore, immunization with a combined vaccine composed of TT and AI induced high levels of antibodies to both antigens. We conclude that TT is a highly suitable exogenous marker for AI vaccination in chickens allowing simple and effective monitoring of AI vaccination status.