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

Evaluation of a positive marker of avian influenza vaccination in ducks for use in H5N1 surveillance

“…Next, a divalent vaccine composed of both TT and whole inactivated H6N2 antigens emulsified with mineral oil (Montanide ISA-70 VG) was prepared, using three separate doses of TT (0.15, 0.3 and 0.6 mg), as described previously [10]. The vaccine was delivered Fig.…”

“…We have previously evaluated tetanus toxoid (TT) as an exogenous marker for AI vaccines in chickens and showed that there is no interference with TT or AI H-specific antibody responses in TT-and AI-co-vaccinated chickens [10]. The TT marker was selected on the basis that chickens are highly resistant to tetanus (chicken toxic dose is 350,000 times the equine toxic dose per gram body weight) [11], are not routinely vaccinated with TT, and naturally existing antibodies to TT are absent in chickens from a variety of sources.…”

“…Furthermore, the antigen is of relatively low cost to manufacture, has minimal regulatory and market acceptance issues and development of an accurate and relatively inexpensive antibody test to the antigen is possible [10]. In this study, we evaluate the levels of naturally acquired antibodies to TT in wild and domestic ducks from Australia, the immunogenicity of TT in Muscovy ducks, and interference by TT on H6-or TT-seroconversion in ducks given separate TT and H6N2 AI vaccines or combined TT/H6N2 vaccines.…”

“…Mackenzie, Curtin University), was propagated according to OIE protocols [12]. Briefly, H6N2 was cultured using 9-to 11-day-old embryonated, specific pathogen-free (SPF) fowl eggs [10]. Harvested allantoic fluid containing H6N2 virus was inactivated with formalin 0.1% (v/v) for 65 h at 37 °C, and virus inactivation confirmed by embryonated fowl egg inoculation.…”

“…Harvested allantoic fluid containing H6N2 virus was inactivated with formalin 0.1% (v/v) for 65 h at 37 °C, and virus inactivation confirmed by embryonated fowl egg inoculation. The HA titre of the inactivated H6N2 AI virus stock (2 7 HA units) was determined using the OIE standard protocol [12] as previously described [10]. …”