Abstract
Avian influenza viruses (AIVs) are highly contagious and have caused huge economical loss to the poultry industry. AIV vaccines remain one of the most effective methods of controlling this disease. Turkey herpesvirus (HVT) is a commonly used live attenuated vaccine against Marek’s disease; it has also been used as a viral vector for recombinant AIV vaccine development. The clustered regularly interspaced palindromic repeats (CRISPR)/Cas9 system is a gene editing tool which, in vaccinology, has facilitated the development of recombinant DNA viral-vectored vaccines. Here, we utilize homology-directed repair (HDR) for the generation of a HVT–H7N9 HA bivalent vaccine; a H7N9 HA expression cassette was inserted into the intergenic region between UL45 and UL46 of HVT. To optimize the selection efficiency of our bivalent vaccine, we combined CRISPR/Cas9 with erythrocyte binding to rapidly generate recombinant HVT–H7HA candidate vaccines.
Highlights
Influenza A viruses are enveloped negative-sense single-stranded RNA viruses with a broad host range, which includes wild aquatic birds that are the natural reservoir for the majority of the virus subtypes [1]
We developed hemadsorption assays for detection of HA antigen hemagglutination activity, in which HA antigen expressed by rHVT adsorbed the chicken red blood cells (RBCs)
To investigate the potential for homology-directed repair (HDR)-clustered regularly interspaced palindromic repeats (CRISPR)/Cas9 to be used as a tool for knocking a gene into the HVT genome, a green fluorescent protein (GFP) expression cassette was selected to insert into the intergenic region between
Summary
Influenza A viruses are enveloped negative-sense single-stranded RNA viruses with a broad host range, which includes wild aquatic birds that are the natural reservoir for the majority of the virus subtypes [1]. The most frequently used vaccines in the poultry industry are inactivated whole-virus vaccines, and this often includes Fowlpox virus- or Newcastle disease virus (NDV)-vectored vaccines. The efficacy of these vaccines is severely impaired by the presence of maternal antibodies within target hosts [2,3]. The virus-vectored HA vaccine only elicits antibody against viral HA protein, can be differentiated from the infected birds. The inactivated whole-virus vaccines induce antibodies against all viral proteins, which makes it difficult to differentiate infected
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