The role of swine as “mixing vessel” for interspecies transmission of the influenza A subtype H1N1: A simultaneous Bayesian inference of phylogeny and ancestral hosts
Research highlights
▶ We examined the inter-host transmission of influenza A virus subtype H1N1. ▶ Phylogenies and ancestral host reconstructions were inferred in a Bayesian framework. ▶ Swine, human and bird show host specific differences in the surface proteins. ▶ Our results suggest that swine is the mixing vessel for host transmission. ▶ Influenza surveillance programs should be intensified for swine.
Introduction
Influenza is a seasonally occurring respiratory disease in humans. Worldwide, there are between three and five million infections per year, 10% of which end fatally. The viral infection caused by the influenza virus is classified into the three types A, B, and C, with A being mainly responsible for human infections. Influenza A is further classified into subtypes referring to the combination of the two surface proteins hemagglutinin (HA) and neuraminidase (NA) (WHO, 2009).
The natural host for all influenza A subtypes are wild birds. Despite the high number of possible combinations (15 HA types × 9 NA types = 135) only H1, H2 and H3 as well as N1 and N2 have successfully established in humans (Baigent and McCauley, 2003). Most relevant for seasonal outbreaks are the subtypes H1N1 and H3N2, in temperate zones preferentially in autumn and winter (Rambaut et al., 2008, WHO, 2009).
Besides birds and humans the virus can also occur in other mammals like swine, horse, or whales, but for physiological reasons interspecies transmissions are rare. However, successful transmission between two species can rapidly lead to a dispersion of the virus of pandemic dimension due to the new host's lack of antibodies (Baigent and McCauley, 2003). The first recorded and since then most devastating pandemic was the “Spanish Flu” in 1918 with more than 40 million deaths. Induced by the same subtype – H1N1 – that circulated until the fifties, the “Russian Flu” of 1977 still has an unexplained origin (Taubenberger and Morens, 2006).
The genome of the influenza virus contains eight pieces of segmented negative-sensed RNA with 13,600 bases all together that code for 10 viral proteins. The trimeric complex of hemagglutinin is encoded on the fourth segment and is responsible for virus binding and for adsorption to the host cell (Skehel and Wiley, 2000). The receptor binding site is host specific due to its configuration at certain codon positions (Baigent and McCauley, 2003). Binding takes place with sialic acid-α-2,3-galactose or sialic acid-α-2,6-galactose (Baigent and McCauley, 2003, Shen et al., 2009). As α-2,3 bound receptors are mainly found in birds, and α-2,6 bound receptors appear in the trachea of humans, a direct transmission is rarely successful but possible – as human cases of avian flu H5N1 demonstrate – because of the existence of avian specific receptors in the lower respiratory system. Swine has both receptor types and has therefore been called the “mixing vessel” for the influenza virus because it can function as an intermediate host (Castrucci et al., 1993, Scholtissek, 1994, Kida et al., 1994, Ma et al., 2008).
Neuraminidase, which is also indispensable for virus transmission, is encoded on segment six. This homotetrameric complex is functionally tightly linked with hemagglutinin and responsible for the virus release from the host cell, eventually leading to the infection of new cells. The release is again facilitated via host specific codon positions (Tamuri et al., 2009).
Phylogenetic reconstructions have been important for our understanding of virus evolution and epidemic pathways including geographical spread and host shifts (Rambaut et al., 2008, Tamuri et al., 2009, Lemey et al., 2009). Methodologically, tree reconstructions and reconstruction of ancestral states, such as geographical locations, hosts or codon positions (Wilson et al., 1991, Ronquist, 1994, Akashi et al., 2007) have been conducted in a parsimony framework. However, parsimony reconstructions do not account for statistical uncertainty both in the reconstruction of the phylogeny as well as the inference of ancestral states (Cunningham et al., 1998, Gubareva et al., 2002, Lemey et al., 2009, Haase et al., 2010, Hovmoller et al., 2010). In addition parsimony ignores branch lengths.
Probabilistic approaches have only recently been applied in the study of viral evolution. Haase et al. (2010), for example, reconstructed host shifts and ancestral ranges of H5N1 in a likelihood framework, but still using a single “optimal” tree. A fully probabilistic Bayesian approach allowing to express mapping and phylogenetic uncertainty simultaneously has been developed by Lemey et al. (2009). Most importantly probabilistic methods avoid overconfidence in seemingly unambiguous inferences usually resulting from maximum parsimony analysis.
Here we investigate the interspecies transmission of the H1N1 influenza A virus between the three hosts human, bird and swine based on all hemagglutinin and neuraminidase sequences available from the NCBI Flu-database as of October 31, 2009. We incorporated an ancestor state reconstruction into a Bayesian framework to study host shifts and additionally examined the human and avian specific amino acid positions in porcine sequences. Central to our analyses was the question about the role of swine as “mixing vessel” for H1N1.
Section snippets
Data selection
We retrieved all human, avian and porcine H1 and N1 full-length sequences available from the NCBI Flu-database as of October 31, 2009 (Bao et al., 2008, NCBI, 2009). We only excluded sequences from the pandemic (H1N1) 2009 virus, since their origin has recently been clarified (Smith et al., 2009). Sequences annotated as full-length by mistake were excluded by using an additional protein specific length criterion: minimum length of H1 = 1600 bases, minimum length of N1 = 1300 bases. The remaining
Sequence evolution
In separate analyses recombination within H1 and N1, respectively, could not be detected (PHI-test: p = 0.768 for H1 and p = 1.0 for N1). However, for concatenated sequences the PHI-test did indicate recombination (p = 3.66 × 10−13). Because H1 and N1 did not share a common evolutionary history we only reconstructed separate trees for each protein.
Hemagglutinin had higher substitution rates at codon positions 1 and 2, whereas at position 3 neuraminidase evolved faster (Supplementary Materials S3.1 and
Discussion
Rates of evolutionary change of hemagglutinin and neuraminidase, rates of non-synonymous substitutions, indicated by the relative rates at the first and second (Supplementary Material S3.1) versus the third codon position (Supplementary Material S3.2), as well as root ages (Supplementary Material S3.3) were in the same respective ranges and could statistically not be distinguished. This has already been noted by Rambaut et al. (2008) for a different data set, indicating similar speed of
Acknowledgments
The comments of two anonymous reviewers helped to improve an earlier version of this paper.
Ethical statement: None.
Sources of funding: None.
References (47)
- et al.
Genetic reassortment between avian and human influenza A viruses in Italian pigs
Virology
(1993) - et al.
Reconstructing ancestral character states: a critical reappraisal
Trends Ecol. Evol.
(1998) - et al.
Detection of influenza virus resistance to neuraminidase inhibitors by an enzyme inhibition assay
Antiviral Res.
(2002) - et al.
Possible sources and spreading routes of highly pathogenic avian influenza virus subtype H5N1 infections in poultry and wild birds in Central Europe in 2007 inferred through likelihood analyses
Infect. Genet. Evol.
(2010) - et al.
The influenza virus gene pool in a poultry market in South central china
Virology
(2003) Information theory and an extension of the maximum likelihood principle
A new look at the statistical model identification
IEEE Trans. Autom. Control
(1974)- et al.
Ancestral inference and the study of codon bias evolution: implications for molecular evolutionary analyses of the Drosophila melanogaster subgroup
PLoS One
(2007) - et al.
Influenza type A in humans, mammals and birds: determinants of virus virulence, host-range and interspecies transmission
Bioessays
(2003) - et al.
The influenza virus resource at the National Center for Biotechnology Information
J. Virol.
(2008)
A simple and robust statistical test for detecting the presence of recombination
Genetics
The Jalview Java alignment
Bioinformatics
Relaxed phylogenetics and dating with confidence
PLoS Biol.
BEAST: Bayesian evolutionary analysis by sampling trees
BMC Evol. Biol.
Antigenic and genetic characteristics of swine-origin 2009 A(H1N1) influenza viruses circulating in humans
Science
A simple, fast, and accurate algorithm to estimate large phylogenies by maximum likelihood
Syst. Biol.
BioEdit Version 5.0.6
Tracking the geographical spread of avian influenza (H5N1) with multiple phylogenetic trees
Cladistics
Application of phylogenetic networks in evolutionary studies
Mol. Biol. Evol.
MAFFT version 5: improvement in accuracy of multiple sequence alignment
Nucleic Acids Res.
MAFFT: a novel method for rapid multiple sequence alignment based on fast Fourier transform
Nucleic Acids Res.
Recent developments in the MAFFT multiple sequence alignment program
Brief Bioinform.
Potential for transmission of avian influenza viruses to pigs
J. Gen. Virol.
Cited by (28)
Characteristics of two zoonotic swine influenza A(H1N1) viruses isolated in Germany from diseased patients
2024, International Journal of Medical MicrobiologyCold-passaged isolates and bat-swine influenza a chimeric viruses as modified live-attenuated vaccines against influenza a viruses in pigs
2022, VaccineCitation Excerpt :Pigs are also susceptible to IAV of avian origin, further adding to the diversity of circulating swIAV. Pigs therefore resemble transitional influenza virus hosts or “mixing vessels” which facilitate the reassortment of IAV genomes of different host origins [23,37,76]. This opens wide opportunities for zoonotic transmissions of reassorted swIAV with unknown phenotypic properties which might include pandemic potential as documented by the most recent human influenza pandemic of 2009 originating from reassorted swIAV in Mesoamerica [64].
Infection and risk factors of human and avian influenza in pigs in south China
2021, Preventive Veterinary MedicineCitation Excerpt :Although the clinical signs of SI in pigs are often mild, co-infection with porcine reproductive and respiratory syndrome (PRRS) and other pathogens can result in increased mortality (Nakharuthai et al., 2008) and infection in pregnant sows can result in stillbirths (Wesley, 2004). Besides the significant economic impact of SI to the pig industry, SI viruses (SIV) can also infect other species, including birds and humans (Hass et al., 2011; McCune et al., 2012; Zhu and Shu, 2013; Bowman et al., 2014), and spillover infection of SI strains to humans has become an emerging problem in public health (Gregory et al., 2001; Gray and Kayali, 2009; Tang et al., 2010; van der Meer et al., 2010). Gene exchange between strains circulating in different species may lead to new epidemics in one or multiple species.
A framework for surveillance of emerging pathogens at the human-animal interface: Pigs and coronaviruses as a case study
2021, Preventive Veterinary MedicineCitation Excerpt :Wild pigs contact a variety of wildlife species, including bats (Wang et al., 2018), while also contacting humans through hunting (Bevins et al., 2014), living in urban spaces (Stillfried et al., 2017), and intense control programs (Pepin et al., 2019b) or with backyard domestic pigs (Wyckoff et al., 2009; Wu et al., 2012). As such, pigs have been implicated in the emergence of novel influenza A viruses in humans (Brown, 2001; Hass et al., 2011), and human influenza A prevalence is positively correlated with influenza A prevalence in wild pigs (Pepin et al., 2019a). Thus, relative to other animal species, pigs may be quite connected to other reservoir species and humans concurrently, while supporting pathogen transmission and evolution with an ample supply of susceptible hosts.
Intranasal inoculations of naked or PLGA-PEI nanovectored DNA vaccine induce systemic and mucosal antibodies in pigs: A feasibility study
2020, Research in Veterinary Science
- 1
Present address: Universitätsklinikum Carl Gustav Carus, Klinik und Poliklinik für Kinder- und Jugendpsychiatrie und -psychotherapie, Fetscherstr. 74, D-01307 Dresden, Germany.
- 2
Present address: Mathematics and BioScience Group, University of Vienna, Nordbergstr. 15, A-1090 Vienna.
- 3
These authors contributed equally to this work.