A comparative analysis of host responses to avian influenza infection in ducks and chickens highlights a role for the interferon-induced transmembrane proteins in viral resistance

Smith et al. BMC Genomics (2015) 16:574 DOI 10.1186/s12864-015-1778-8 RESEARCH ARTICLE Open Access A comparative analysis of host responses to avian influenza infection in ducks and chickens highlights a role for the interferon-induced transmembrane proteins in viral resistance Jacqueline Smith1*†, Nikki Smith1†, Le Yu1, Ian R. Paton1, Maria Weronika Gutowska1, Heather L. Forrest2, Angela F. Danner2, J. Patrick Seiler2, Paul Digard1, Robert G. Webster2† and David W. Burt1*† Abstract Background: Chickens are susceptible to infection with a limited number of Influenza A viruses and are a potential source of a human influenza pandemic. In particular, H5 and H7 haemagglutinin subtypes can evolve from low to highly pathogenic strains in gallinaceous poultry. Ducks on the other hand are a natural reservoir for these viruses and are able to withstand most avian influenza strains. Results: Transcriptomic sequencing of lung and ileum tissue samples from birds infected with high (H5N1) and low (H5N2) pathogenic influenza viruses has allowed us to compare the early host response to these infections in both these species. Chickens (but not ducks) lack the intracellular receptor for viral ssRNA, RIG-I and the gene for an important RIG-I binding protein, RNF135. These differences in gene content partly explain the differences in host responses to low pathogenic and highly pathogenic avian influenza virus in chicken and ducks. We reveal very different patterns of expression of members of the interferon-induced transmembrane protein (IFITM) gene family in ducks and chickens. In ducks, IFITM1, 2 and 3 are strongly up regulated in response to highly pathogenic avian influenza, where little response is seen in chickens. Clustering of gene expression profiles suggests IFITM1 and 2 have an anti-viral response and IFITM3 may restrict avian influenza virus through cell membrane fusion. We also show, through molecular phylogenetic analyses, that avian IFITM1 and IFITM3 genes have been subject to both episodic and pervasive positive selection at specific codons. In particular, avian IFITM1 showed evidence of positive selection in the duck lineage at sites known to restrict influenza virus infection. Conclusions: Taken together these results support a model where the IFITM123 protein family and RIG-I all play a crucial role in the tolerance of ducks to highly pathogenic and low pathogenic strains of avian influenza viruses when compared to the chicken. Keywords: Avian influenza, H5N1, H5N2, Duck, Chicken, Pathogenicity, Interferon-induced transmembrane proteins, Evolution and gene expression profiles * Correspondence: ; † Equal contributors 1 The Roslin Institute and R(D)SVS, University of Edinburgh, Easter Bush, Midlothian EH25 9RG, UK Full list of author information is available at the end of the article © 2015 Smith et al. Open Access This article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated. Smith et al. BMC Genomics (2015) 16:574 Background The question of how ducks can survive challenge by all low pathogenic avian influenza (LPAI) and most highly pathogenic avian influenza (HPAI) infections, yet chickens can only survive LPAI, remains an important question due to the economic losses experienced by the poultry industry, the human health implications and the continuing threat of pandemic disease posed by avian viruses. Most avian influenza strains are able to infect and replicate in ducks, and are usually asymptomatic and seldom cause disease. Of the 16 haemagglutinin subtypes of influenza viruses infecting migratory waterfowl, the H5 and H7 subtypes are unique. After transmission to gallinaceous poultry the H5 and H7 viruses can evolve into highly pathogenic strains. In ducks and other aquatic birds the low pathogenic H5 and H7 influenza viruses replicate predominantly in the intestinal tract without any outward clinical signs of infection. After transmission to gallinaceous poultry and acquisition of multiple basic amino acids in the connecting peptide of the haemagglutinin, the viruses become highly pathogenic and replicate systemically in the chicken [1]. HPAI has proven to be deadly to chickens within a very short time frame post-infection, although LPAI only produces mild or no signs of clinical disease [2]. Ducks on the other hand are able to mount a robust inflammatory response against most HPAI [3], although they have shown increased susceptibility to some emerging strains of H5N1, with death resulting in some cases [4–6]. The difference in pathogenesis seen between ducks and chickens could also be due to the fact that a rapid induction of apoptosis in HPAI-infected ducks may be beneficial to the host, whereas delayed apoptosis in chickens may be an advantage for the virus [7]. HPAI H5N1 remains a concern, with new strains continually evolving and increasing the pandemic threat from this subtype. Worryingly, the recently emerged H7N9 subtype also poses a risk of being the vector by which a human influenza epidemic occurs. The first human infections by this virus were reported early in 2013, and by June 2013, the mortality rate was over 30 % - a level still being seen as cases continue to be reported. The World Health Organization (WHO) has identified H7N9 as "…an unusually dangerous virus for humans” [8]. Although not posing an imminent threat, H10N8 has also recently been identified as having the potential to spread from birds to humans if the necessary viral mutations occur [9]. Viral haemagglutinin binds to cell surface receptors in susceptible host cells. In humans, virus binds to sialic acid α2, 6-galactose (SAα2, 6-Gal) linked receptors whereas avian viruses preferentially bind to sialic acid α2, 3-galactose (SAα2, 3-Gal) linked receptors [10, 11]. SAα2, 6-Gal linked receptors are the predominant type Page 2 of 19 in the trachea of chickens, while duck tracheas contain more SAα2, 3-Gal linked receptors [12]. This may be one reason why chickens have the potential to act as an intermediate host for human infection. The avian response to infection compared to that of humans is also determined by their immune gene repertoire. It is known that birds have a much more compact set of immune-related genes than mammals [13, 14]. With the increasing availability of avian genome sequences, it is also becoming apparent that key genes are reportedly missing in some birds, which will affect their host responses to viral infections. These include Toll-like receptors 8 and 9 (...truncated)