Bruce H. Janke

Chapter 3 Swine Influenza Viruses: A North American Perspective

Influenza is a zoonotic viral disease that represents a health and economic threat to both humans and animals worldwide. Swine influenza (SI) was first recognized clinically in pigs in the Midwestern U.S., in 1918, coinciding with the human influenza pandemic known as the Spanish flu. Since that time SI has remained of importance to the swine industry throughout the world. In this review, the epidemiology of swine influenza virus (SIV) infection in North American pigs is described in detail. The first 80 years of SI remained relatively static, whereas the last decade has become dynamic with the establishment of many emerging subtypes. With the increasing number of novel subtypes and genetic variants, the control of SI has become increasingly difficult and innovative strategies to combat this economically important zoonotic disease are critical. Therefore, protective immune responses against influenza virus infections as well as new paradigms of vaccine development in pigs are discussed in the review. It is expected that the dynamic evolutionary changes of SIVs in North American pigs will continue, making currently available prophylactic approaches of limited use to control the spread and economic losses associated with this important swine pathogen.

Mutations in the NS1 protein of swine influenza virus impair anti-interferon activity and confer attenuation in pigs

ABSTRACT It has been shown previously that the nonstructural protein NS1 of influenza virus is an alpha/beta interferon (IFN-α/β) antagonist, both in vitro and in experimental animal model systems. However, evidence of this function in a natural host has not yet been obtained. Here we investigated the role of the NS1 protein in the virulence of a swine influenza virus (SIV) isolate in pigs by using reverse genetics. The virulent wild-type A/Swine/Texas/4199-2/98 (TX/98) virus and various mutants encoding carboxy-truncated NS1 proteins were rescued. Growth properties of TX/98 viruses with mutated NS1, induction of IFN in tissue culture, and virulence-attenuation in pigs were analyzed and compared to those of the recombinant wild-type TX/98 virus. Our results indicate that deletions in the NS1 protein decrease the ability of the TX/98 virus to prevent IFN-α/β synthesis in pig cells. Moreover, all NS1 mutant viruses were attenuated in pigs, and this correlated with the amount of IFN-α/β induced in vitro. These data suggest that the NS1 protein of SIV is a virulence factor. Due to their attenuation, NS1-mutated swine influenza viruses might have a great potential as live attenuated vaccine candidates against SIV infections of pigs.

Identification of H2N3 influenza A viruses from swine in the United States

Although viruses of each of the 16 influenza A HA subtypes are potential human pathogens, only viruses of the H1, H2, and H3 subtype are known to have been successfully established in humans. H2 influenza viruses have been absent from human circulation since 1968, and as such they pose a substantial human pandemic risk. In this report, we isolate and characterize genetically similar avian/swine virus reassortant H2N3 influenza A viruses isolated from diseased swine from two farms in the United States. These viruses contained leucine at position 226 of the H2 protein, which has been associated with increased binding affinity to the mammalian α2,6Gal-linked sialic acid virus receptor. Correspondingly, the H2N3 viruses were able to cause disease in experimentally infected swine and mice without prior adaptation. In addition, the swine H2N3 virus was infectious and highly transmissible in swine and ferrets. Taken together, these findings suggest that the H2N3 virus has undergone some adaptation to the mammalian host and that their spread should be very closely monitored.

Vaccination of Pigs against Swine Influenza Viruses by Using an NS1-Truncated Modified Live-Virus Vaccine

ABSTRACT Swine influenza viruses (SIV) naturally infect pigs and can be transmitted to humans. In the pig, genetic reassortment to create novel influenza subtypes by mixing avian, human, and swine influenza viruses is possible. An SIV vaccine inducing cross-protective immunity between different subtypes and strains circulating in pigs is highly desirable. Previously, we have shown that an H3N2 SIV (A/swine/Texas/4199-2/98 [TX98]) containing a deleted NS1 gene expressing a truncated NS1 protein of 126 amino acids, NS1▴126, was attenuated in swine. In this study, 4-week-old pigs were vaccinated with the TX98 NS1▴126 modified live virus (MLV). Ten days after boosting, pigs were challenged with wild-type homologous H3N2 or heterosubtypic H1N1 SIV and sacrificed 5 days later. The MLV was highly attenuated and completely protected against challenge with the homologous virus. Vaccinated pigs challenged with the heterosubtypic H1N1 virus demonstrated macroscopic lung lesions similar to those of the unvaccinated H1N1 control pigs. Remarkably, vaccinated pigs challenged with the H1N1 SIV had significantly lower microscopic lung lesions and less virus shedding from the respiratory tract than did unvaccinated, H1N1-challenged pigs. All vaccinated pigs developed significant levels of hemagglutination inhibition and enzyme-linked immunosorbent assay titers in serum and mucosal immunoglobulin A antibodies against H3N2 SIV antigens. Vaccinated pigs were seronegative for NS1, indicating the potential use of the TX98 NS1▴126 MLV as a vaccine to differentiate infected from vaccinated animals.

Characterization of a newly emerged genetic cluster of H1N1 and H1N2 swine influenza virus in the United States

H1 influenza A viruses that were distinct from the classical swine H1 lineage were identified in pigs in Canada in 2003–2004; antigenic and genetic characterization identified the hemagglutinin (HA) as human H1 lineage. The viruses identified in Canadian pigs were human lineage in entirety or double (human–swine) reassortants. Here, we report the whole genome sequence analysis of four human-like H1 viruses isolated from U.S. swine in 2005 and 2007. All four isolates were characterized as triple reassortants with an internal gene constellation similar to contemporary U.S. swine influenza virus (SIV), with HA and neuraminidase (NA) most similar to human influenza virus lineages. A 2007 human-like H1N1 was evaluated in a pathogenesis and transmission model and compared to a 2004 reassortant H1N1 SIV isolate with swine lineage HA and NA. The 2007 isolate induced disease typical of influenza virus and was transmitted to contact pigs; however, the kinetics and magnitude differed from the 2004 H1N1 SIV. This study indicates that the human-like H1 SIV can efficiently replicate and transmit in the swine host and now co-circulates with contemporary SIVs as a distinct genetic cluster of H1 SIV.

The role of swine in the generation of novel influenza viruses.

The ecology of influenza A viruses is very complicated involving multiple host species and viral genes. Avian species have variable susceptibility to influenza A viruses with wild aquatic birds being the reservoir for this group of pathogens. Occasionally, influenza A viruses are transmitted to mammals from avian species, which can lead to the development of human pandemic strains by direct or indirect transmission to man. Because swine are also susceptible to infection with avian and human influenza viruses, genetic reassortment between these viruses and/or swine influenza viruses can occur. The potential to generate novel influenza viruses has resulted in swine being labelled ‘mixing vessels’. The mixing vessel theory is one mechanism by which unique viruses can be transmitted from an avian reservoir to man. Although swine can generate novel influenza viruses capable of infecting man, at present, it is difficult to predict which viruses, if any, will cause a human pandemic. Clearly, the ecology of influenza A viruses is dynamic and can impact human health, companion animals, as well as the health of livestock and poultry for production of valuable protein commodities. For these reasons, influenza is, and will continue to be, a serious threat to the wellbeing of mankind.

Swine Influenza Virus: Zoonotic Potential and Vaccination Strategies for the Control of Avian and Swine Influenzas

Influenza viruses are able to infect humans, swine, and avian species, and swine have long been considered a potential source of new influenza viruses that can infect humans. Swine have receptors to which both avian and mammalian influenza viruses bind, which increases the potential for viruses to exchange genetic sequences and produce new reassortant viruses in swine. A number of genetically diverse viruses are circulating in swine herds throughout the world and are a major cause of concern to the swine industry. Control of swine influenza is primarily through the vaccination of sows, to protect young pigs through maternally derived antibodies. However, influenza viruses continue to circulate in pigs after the decay of maternal antibodies, providing a continuing source of virus on a herd basis. Measures to control avian influenza in commercial poultry operations are dictated by the virulence of the virus. Detection of a highly pathogenic avian influenza (HPAI) virus results in immediate elimination of the flock. Low-pathogenic avian influenza viruses are controlled through vaccination, which is done primarily in turkey flocks. Maintenance of the current HPAI virus-free status of poultry in the United States is through constant surveillance of poultry flocks. Although current influenza vaccines for poultry and swine are inactivated and adjuvanted, ongoing research into the development of newer vaccines, such as DNA, live-virus, or vectored vaccines, is being done. Control of influenza virus infection in poultry and swine is critical to the reduction of potential cross-species adaptation and spread of influenza viruses, which will minimize the risk of animals being the source of the next pandemic.

Enhanced pneumonia and disease in pigs vaccinated with an inactivated human-like (δ-cluster) H1N2 vaccine and challenged with pandemic 2009 H1N1 influenza virus

Influenza is an economically important respiratory disease affecting swine world-wide with potential zoonotic implications. Genetic reassortment and drift has resulted in genetically and antigenically distinct swine influenza viruses (SIVs). Consequently, prevention of SIV infection is challenging due to the increased rate of genetic change and a potential lack of cross-protection between vaccine strains and circulating novel isolates. This report describes a vaccine-heterologous challenge model in which pigs were administered an inactivated H1N2 vaccine with a human-like (δ-cluster) H1 six and three weeks before challenge with H1 homosubtypic, heterologous 2009 pandemic H1N1. At necropsy, macroscopic and microscopic pneumonia scores were significantly higher in the vaccinated and challenged (Vx/Ch) group compared to non-vaccinated and challenged (NVx/Ch) pigs. The Vx/Ch group also demonstrated enhanced clinical disease and a significantly elevated pro-inflammatory cytokine profile in bronchoalveolar lavage fluid compared to the NVx/Ch group. In contrast, viral shedding and replication were significantly higher in NVx/Ch pigs although all challenged pigs, including Vx/Ch pigs, were shedding virus in nasal secretions. Hemagglutination inhibition (HI) and serum neutralizing (SN) antibodies were detected to the priming antigen in the Vx/Ch pigs but no measurable cross-reacting HI or SN antibodies were detected to pandemic H1N1 (pH1N1). Overall, these results suggest that inactivated SIV vaccines may potentiate clinical signs, inflammation and pneumonia following challenge with divergent homosubtypic viruses that do not share cross-reacting HI or SN antibodies.

Efficacy of inactivated swine influenza virus vaccines against the 2009 A/H1N1 influenza virus in pigs

The gene constellation of the 2009 pandemic A/H1N1 virus is a unique combination from swine influenza A viruses (SIV) of North American and Eurasian lineages, but prior to April 2009 had never before been identified in swine or other species. Although its hemagglutinin gene is related to North American H1 SIV, it is unknown if vaccines currently used in U.S. swine would cross-protect against infection with the pandemic A/H1N1. The objective of this study was to evaluate the efficacy of inactivated vaccines prepared with North American swine influenza viruses as well as an experimental homologous A/H1N1 vaccine to prevent infection and disease from 2009 pandemic A/H1N1. All vaccines tested provided partial protection ranging from reduction of pneumonia lesions to significant reduction in virus replication in the lung and nose. The multivalent vaccines demonstrated partial protection; however, none was able to prevent all nasal shedding or clinical disease. An experimental homologous 2009 A/H1N1 monovalent vaccine provided optimal protection with no virus detected from nose or lung at any time point in addition to amelioration of clinical disease. Based on cross-protection demonstrated with the vaccines evaluated in this study, the U.S. swine herd likely has significant immunity to the 2009 A/H1N1 from prior vaccination or natural exposure. However, consideration should be given for development of monovalent homologous vaccines to best protect the swine population thus limiting shedding and the potential transmission of 2009 A/H1N1 from pigs to people.

Experimental inoculation of pigs with pandemic H1N1 2009 virus and HI cross-reactivity with contemporary swine influenza virus antisera

Please cite this paper as: Vincent et al. (2010) Experimental inoculation of pigs with pandemic H1N1 2009 virus and HI cross‐reactivity with contemporary swine influenza virus antisera. Influenza and Other Respiratory Viruses 4(2), 53–60