Evaluation of different stratagies [sic] for the differentiation of infected and vaccinated animals (DIVA) in chickens vaccinated with avian influenza oil emulsion vaccines

Abstract

The use of avian influenza vaccination in poultry would have greater world-wideacceptance if a reliable test that clearly discriminates naturally infected from vaccinatedonly animals (DIVA) was available. In order to evaluate the non-structural protein 1(NS1) and neuraminidase (NA) proteins of avian influenza virus as potential markers forinfection, an indirect ELISA using recombinant baculovirus purified NS1 protein asantigen and the neuraminidase inhibition (NI) test using MUN (2Õ-(4-methylumbelliferyl)-_-D-Nacetylneuraminic acid sodium salt hydrate) as NA substrate,were used as differential diagnostic serological tests. Because the NS1 protein is onlyproduced when the virus is replicating in cells and it is not packed in the infectiousvirion, theoretically it is not present in the killed vaccines. Therefore, vaccinated birdswill be negative for NS1 antibody as opposed to infected birds that will developantibodies against NS1 during virus replication. Antibodies against NS1 protein were firstdetected three weeks after infection of native birds, but decreased rapidly by 5 weeks afterinfection. In inactivated oil emulsion immunized birds, antibodies against NS1 weredetected only in a small percentage of birds after homologous LPAI virus challenge.Vaccinated birds challenged with highly pathogenic AI showed a better NS1 antibodyresponse in some but not all challenged birds.The use of killed adjuvanted vaccines containing virus with homologoushemagglutinin and heterologous NA subtype to the challenge virus can be usedeffectively to protect birds against the disease and allow the possibility of using theantibody response to the heterologous viral NA protein as marker of infection. In twogroups of birds vaccinated prior to challenge, the NI test discriminated chickens receivingdifferent vaccine antigens (e.g., N8 or N9), and two weeks post-challenge with a highlypathogenic avian influenza virus subtype H5N2, 100% of the vaccinated birds hadsignificant levels of N2 NI activity with high specificity.Comparing the two DIVA strategies, the heterologous NA method gave a moreconsistent response with earlier detection of infection under these experimentalconditions. However, further research is needed to evaluate how this approach worksunder different field conditions before it can be adopted on a commercial scale.