Short communicationProtective efficacy of an inactivated chimeric H7/H5 avian influenza vaccine against highly pathogenic avian influenza H7N9 and clade 2.3.4.4 H5 viruses
Introduction
H5N1 avian influenza viruses (AIVs) have been detected in more than 60 countries and cause great economic losses to the worldwide poultry industry (Swayne, 2012). Of note, H5N1 AIVs have become enzootic in domestic poultry and wild birds in Asia, Europe, Africa, and North America (Claas et al., 1998; Eagles et al., 2009; Ip et al., 2015; Jiang et al., 2017; Monne et al., 2008; Pasick et al., 2015; Peiris et al., 2007; Zhao et al., 2013). Sequence analyses of hemagglutinin (HA) genes have shown that H5 viruses have evolved into diverse clades and subclades (Li et al., 2010). Viruses in clade 7.2 have been detected in chickens in several provinces in northern China and clades 2.3.2.1 and 2.3.4.4 continue to co-circulate in wild birds and poultry in several countries (Li et al., 2010). However, recent molecular epidemiological surveys of AIVs detected no clade 7.2 viruses and a significant reduction in the circulation of clade 2.3.2.1 viruses since 2015 (unpublished data). Currently, clade 2.3.4.4 viruses are the dominant epidemic strains and N6 is the main NA subtype.
Novel H7N9 viruses isolated from samples collected at live poultry markets in China in 2013 had low pathogenicity in poultry. However, since December 2016, H7N9 viruses with novel polybasic amino acid sequences at HA cleavage sites, which are associated with increased pathogenicity in poultry, have been detected in both human patients and chickens. These viruses were found to be highly pathogenic in chickens in particular (Wang et al., 2017). Since March 2017, H7N9 highly pathogenic AIVs (HPAIVs) have caused in excess of 26 outbreaks in 10 provinces in China, resulting in great economic losses to the poultry industry.
Vaccination is an important strategy used to control AIV infection in poultry (Swayne, 2012; Swayne et al., 2014). To address the problem of AIV infection, the government of China implemented a mass poultry vaccination program. Since August 2017, the H5 Re-8 and H7 Re1 combination vaccine has been used to control H5 and H7N9 HPAIVs throughout China. Despite the availability of this efficacious combination vaccine, vaccine production requires more embryonated chicken eggs (ECEs), an alternative vaccine (i.e. a bivalent recombinant vaccine expressing both corresponding antigens) would be more advantageous. Furthermore, one important problem during mass vaccination should be considered that avian influenza is characterized by non-sterile immunity, and immunized poultry with high antibody titers are sometimes viraemic (shedding and spreading viruses), but asymptomatic.
Killed influenza virus vaccines induce anti-HA antibodies that specifically target antigenic sites in the globular head domain of the HA1 region and block receptor binding (Chiu et al., 2009; Hashem, 2015). These responses are typically strain specific, as the induced antibodies have potent neutralizing activity against homologous strains but do not cross-react with HAs of other influenza virus subtypes (Sui et al., 2009). In contrast, the HA2 stalk domain, although less immunogenic (Tan et al., 2012; Toroghi and Momayez, 2006), is relatively well conserved among different subtypes (Pica and Palese, 2013). Furthermore, broadly neutralizing antibodies against the HA2 stalk domain have been isolated (Sui et al., 2009; Tan et al., 2012). Here, we generated a novel chimeric H7/H5 virus that expressed the entire HA1 portion of H7N9 and the HA2 region of the heterosubtypic virus H5N6. We then evaluated the immunogenicity and protective efficacy of the chimeric vaccine against clade 2.3.4.4 H5N6 and H7N9 HPAIV infections in chickens.
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Ethics statement
All procedures in animal experiments met the requirements and were approved by the Animal Welfare Ethics Committee of the China Animal Health and Epidemiology Center (Approval Number: # CAHEC-2017-017). All applicable international, national, and institutional guidelines for the care and use of animals were followed.
Viruses and cells
The HPAIVs A/chicken/Fujian/5/2016(H5N6) (FJ/5; clade 2.3.4.4) and A/chicken/Guangdong/RZ/2017(H7N9) (GD/RZ) were isolated from dead chickens and propagated in 10-day-old specific
Generation of the chimeric H7/H5 virus
To develop a chimeric vaccine through reverse genetics, we generated a chimeric H7/H5 HA construct composed of the entire HA1 domain from GD/RZ and the HA2 region from FJ/5 (Fig. 1A). Briefly, the HA1 and HA2 regions were amplified and the two segments were combined in the chimeric H7/H5 HA gene by overlapping PCR and cloned into pHW2000 (Jiang et al., 2014). The chimeric H7/H5 virus was generated on a PR8 backbone by reverse genetics methods as described previously. The resulting virus was
Discussion
H5N1 AIVs have become enzootic in domestic poultry and wild birds in China (Swayne et al., 2011). To date, H7N9 HPAIVs have caused in excess of 26 outbreaks in China with great economic losses for poultry. To develop a bivalent vaccine against both H5 and H7 viruses, we generated an influenza virus carrying a chimeric H7/H5 HA segment that expressed the entire HA1 portion of the H7N9 virus and the HA2 region of the H5N6 virus.
Foreign gene-expressing viruses ideally have high genetic stability.
Conflicts of interest
The authors declare that they have no competing interests.
Acknowledgements
This research was supported by the Sci-tech Basic Work Project of the Ministry of Science and Technology (SQ2012FY3260033) and by the National Key Research and Development Program of China (2016YFD0501609).
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These authors contributed equally to this work.