Understanding influenza A virus: linking transmission, evolutionary dynamics, pathogenesis and immunity in pigs

Lead Research Organisation: The Pirbright Institute
Department Name: Livestock Viral Diseases

Abstract

Swine influenza attracts considerable attention because of the threat of zoonotic infections causing human pandemics. During the pandemic, a fear that viruses emerging from pigs may infect people resulted in the widespread destruction of animals in some countries and trade bans. Consequently, the insidious effects of this highly prevalent virus on the health and welfare of pig populations, estimated to increase the cost of production by £7 per finished pig, have not been given due regard. The primary disease caused by influenza virus in usually mild, but results in greater susceptibility to secondary infections. Vaccination will be a key control measure for influenza in pigs to improve general herd health.
Through our studies we will develop a more detailed understanding of the dynamics of virus transmission and the consequences of transmission and vaccination in driving viral evolution. During these studies we will also define a range of parameters, for example local and systemic immune responses and sites of virus replication, which are associated with the onset and cessation of transmission. We need to know if current and proposed novel vaccines not only prevent clinical signs but also stop viruses being transmitted unnoticed. Furthermore, if viruses can be transmitted unnoticed are they changing because of the immune response in the population? To answer these questions we need to understand virus transmission in detail and how the viruses change when they pass between animals. We can then apply this new knowledge to population wide models of disease spread to predict the efficiency of any proposed control measures. This knowledge will also inform the design of novel vaccines.
Vaccination against influenza in pigs is not routinely performed in Europe mainly for two reasons: the cost benefit of vaccination has not been clearly demonstrated and it is not clear that the available vaccines will protect against the strains currently circulating in the pig populaion. The most striking example of the latter is that current vaccines do not include pandemic H1N1 influenza virus antigen.
These studies will provide essential evidence to design control programmes for influenza in pigs, most notably: i) finding out how efficient are the current prophylactic methods at controlling the spread of infection; ii) what level of immunity is required in a population to prevent the spread of infection and the evolution of new strains of virus and iii) determine whether new, broadly cross protective vaccines are more effective at controlling influenza infections in swine to enhance animal health and livestock production.
Importantly, this type of information is not available for any natural mammalian hosts of influenza viruses, including humans and horses. Therefore, the results of our studies will have a broad impact on influenza control measures.

Technical Summary

In this programme of work we will describe and define in detail pig to pig transmission parameters for influenza virus.
We have recently performed a unique series of transmission experiments of an acute viral infection in a natural host species, foot-and-mouth disease virus in cattle. These studies quantified the link between clinical signs and viral shedding and concluded that the infectious period was much briefer than hitherto supposed and demonstrated pathogen detection alone does not correlate with transmissibility. This type of quantitative knowledge is critical for developing efficient, targeted control strategies. Consequently, we will perform a series of one-to-one challenge studies to define transmission parameters for influenza of pigs. Intra- and inter- herd transmission models will be developed based on data obtained from these experiments.
Furthermore, virological, clinicopathological and immunological correlates of transmission will be identified by their relationship to the onset, duration and cessation of infectiousness using standard statistical approaches (logistic regression). Non-metric, multi-dimensional scaling will be used to visualise the temporal dynamics of these relationships. The evolutionary dynamics of influenza will be analysed during a series of transmission chains in naïve and vaccinated animals. Within-host mathematical modelling will be used to estimate the size of any bottlenecks and determine whether selection plays a role in the size and composition of the viral population during and following transmission.
A major strategic goal of this programme is to understand whether current vaccines and novel broadly cross reactive vaccines can block influenza transmission. We will determine the capacity of these vaccines to induce responses that correlates with protection and result in prevention of transmission post challenge.

Planned Impact

Mammalian animal health is a key component in safeguarding food security, and livestock viral diseases have very significant direct and indirect impacts. Viruses endemic to the UK, such as swine influenza virus are ubiquitous, causing both disease and welfare problems. Importantly, new or more harmful forms of swine influenza emerge due to viral mutation or cross-species transmissions and can be important causes of zoonoses, for example, H1N1. In addition to progressing our understanding of influenza virus infection, the programme will develop expertise in livestock immunology and pathogenesis to enable swift transfer and application of core skills to address newly emerging viral diseases.
We will provide to Defra advice on national control measures. This research contributes to animal health and welfare and a sustainable UK farming and food sector, specifically addressing Defra's goals to year 2014 on (i) "the maintenance of a disease status amongst the highest in the world, so that we are able to trade our animals and animal products internationally"; and (ii) "to have dealt with all disease emergencies swiftly and effectively using an agreed approach".
There is growing interest from the animal health pharmaceutical industries to develop more effective broadly cross protective influenza vaccines that do not require frequent updating. Industry have been closely involved in the development of the programme of work and will have ongoing involvement to ensure we remain focussed on achieving this strategic goal.
Surveillance and research activities supported by the European Commission (DG Sanco and DG Research) have broadly similar aims to Defra's goals. Other international disease control agencies such as FAO and OIE will also benefit from this research.
This programme will act as a catalyst to develop a national capability to understand disease transmission in livestock. The UK academic community requires access to a comprehensive range of linked facilities to study livestock diseases; spanning high and low containment animal units and laboratories, specialist immunologists and virologists and high throughput sequencing facilities with associated bioinformaticians, statisticians and modellers. The legacy of this programme will the establishment of collaborative team that spans all of these disciplines with access to appropriate state of the art facilities.
 
Description Two Pilot experiments were conducted at APHA in Nov-Dec 2014 on behalf of the Pirbright Institute to determine virus infection route and dose. MAD inoculation was found to give a consistent virus secretion profile that peaked at 2-4dpi. Naïve contact pigs secreted virus maximally 3-5dpi. It was decided that optimal infection in subsequent experiments will be ensured by inoculation of 'seeder/donor' pigs. Please see attached report for data supplied to the Pirbright Institute by APHA for preparation of a manuscript.

A vaccine pilot experiment was conducted at APHA in Oct-Nov 2015 to assess a commercial vaccine and two vaccines formulated at APHA containing antigen that was homologous or heterologous (but similar) to the challenge strain A/Sw/Eng/1353/09. The results demonstrated that vaccine-mediated protection requires that the vaccine strain is closely matched with the challenge strain. Vaccines commercially available in the UK at present incorporate antigens that are only distantly related to contemporary swine strains and did not afford high levels of protection against infection by the challenge strain. The heterologous vaccine afforded a modest degree of protection whereas the heterologous strain afforded high levels of protection against challenge. Further immunological and virological analysis of samples is ongoing.
Preliminary within-host dynamics model developed with already published data (experimental infections in humans and in ferrets - provided by Taronna R. Maines from CDC) to quantify infectious particles production and to describe infectious virus dynamics in the environment.
Pilot model developed from pilot studies: within-host dynamics model describing viral titre and cytokines.
Three minimal immunodominant influenza virus epitopes of 8, 9 and 10 amino acids in length and has CD8beta T-cell clones that react to each. Porcine MHC-1 recombinant molecules have been shown to fold with the peptides. This is the first time T cell clones have been produced for pigs, which will allow detailed analysis of the cellular immune response to Influenza virus. Also we plan to solve the structure of the MHC peptide complex to provide generic information on the recognition of pathogens by porcine CD8+ T cells.
We show that H1N1 S-FLU, a candidate universal vaccine, when administered to pigs by aerosol, can induce CD4 and CD8 T cell immune responses in blood, bronchoalveolar lavage (BAL) and tracheobronchial lymph nodes (TBLN). Neutralising antibody was not produced. Detection of a BAL response correlated with a reduction in viral titer in nasal swabs and lungs, following challenge with homologous H1N1 pandemic virus. Intratracheal immunization with a higher dose of a heterologous H5N1 S-FLU vaccine, induced weaker BAL and stronger TBLN responses and a lesser reduction in viral titer. We conclude that local cellular immune responses are important for protection against influenza A virus infection, that these can be most efficiently induced by aerosol immunization targeting the lower respiratory tract and that S-FLU is a promising universal influenza vaccine candidate. (revised manuscript under review by J. Immunology)
Exploitation Route An additional study funded by the Bill and Melinda Gates foundation to use our Swine Influenza model, which includes the capacity to carry out detailed analysis of the immune response, to test a novel human influenza vaccine
Sectors Agriculture, Food and Drink,Healthcare
 
Description • The Bristol consortium took part in Bristol Festival of Nature in June 2015 which centred on the immune response to viral infections. Part of the stall included posters about the projects we were working on. • The Pirbright Institute consortium was a part of the FLU Fighters at the Royal Society Summer Science Exhibition 2015. • Mario Aramouni, Jenner-APHA Fellow presented a poster presentation at 3rd International Symposium on Neglected Influenza Viruses, 15-17 April 2015, USA : M. Aramouni 'H1N1 (09pdm) transmission parameters and host response in naïve pigs' Aramouni, M., Tchilian, E., Brookes, S.M., Everett, H., Beck, K., Morgan, S., Hemmink, J.D., Brown, I.H. and Charleston, B. • Hanneke Hemmink, The Pirbright Institute presented at the European Veterinary Immunology Conference, Vienna September 2015 'Immunogenicity and protective efficacy of a candidate universal vaccine, S-Flu, in pigs'. • Elma Tchilian, The Pirbright Institute presented at the Oxford Influenza meeting September 2015 'Distinct Immune responses and viral shedding patterns in pigs following aerosol, intranasal and in contact infection with pdmH1N1 swine influenza virus' • Sharon Brookes, APHA presented an oral presentation at the Oxford Influenza Meeting September 2015 'An update on pandemic H1N1 in pigs 2009-2014' sLoLa funding was acknowledged and talk explains the rationale for using the strain A/Sw/Eng/1353/09 as the challenge strain. • Mario Aramouni, Jenner-APHA Fellow provided a talk at Sarah Gilbert's lab at the Jenner Institute on 27 Nov 2015. • Laetitia Canini, University of Edinburgh Presented at the Department of infectious diseases epidemiology seminar (invited), Imperial College, London, April 2015; 2nd workshop on virus dynamics, Toronto, July 2015; Epidemics, Florida, December 2015. Submitted at Options for the control of influenza IX, Chicago, August 2016. • Ian Brown (APHA) regularly meets with Defra. • The consortium has also been able to support a grant based with Professor Andrew Sewell at Cardiff University by providing reagents for tetramer staining. The production of these tetramers at Cardiff will directly aid in WP1 D Immunopathology associated with the onset and cessation of transmission. • The consortium has been able to provide expert in advice on commercial vaccines as well as knowledge on aerosol delivery in pigs as well as humans based on research understood from this grant.
First Year Of Impact 2015
Sector Agriculture, Food and Drink,Healthcare
Impact Types Policy & public services