Competition between IAV subtypes through heterosubtypic immunity modulates re-infection and antibody dynamics in the mallard reservoir
Authors: Latorre-Margalef Neus , Brown Justin D. , Fojtik Alinde , Poulson Rebecca L. , Carter Deborah , Franca Monique , Stallknecht David E. 
Published: September 2016

AbstractOur overall hypothesis is that host population immunity directed at multiple antigens will influence the prevalence, diversity and evolution of influenza A virus (IAV) in avian populations where the vast subtype diversity is maintained. To investigate how initial infection influences the outcome of later infections with homologous or heterologous IAV subtypes and how viruses interact through host immune responses; we carried out experimental infections in mallard ducks (Anas platyrhynchos). Mallards were pre-challenged with an H3N8 low-pathogenic IAV and were divided into six groups. At five weeks post H3N8 inoculation, each group was challenged with a different IAV subtype or the same H3N8. Two additional pre-challenged groups were inoculated with the homologous H3N8 virus at weeks 11 and 15 after pre-challenge to evaluate the duration of protection, which showed that mallards were still resistant to re-infection after 15 weeks. There was a significant reduction in shedding for all pre-challenged groups compared to controls and the outcome of the heterologous challenges varied according to hemagglutinin (HA) phylogenetic relatedness between the viruses used. There was a boost in the H3 antibody titer after re-infection with H4N5, which is consistent with original antigenic sin or antigenic seniority and suggest a putative strategy of virus evasion. These results imply strong competition between related subtypes that could regulate IAV subtype population dynamics in nature. Collectively, we provide new insights into within-host IAV complex interactions as drivers of IAV antigenic diversity that could allow the circulation of multiple subtypes in wild ducks.Author summaryMany features of pathogen diversification remain poorly explored although host immunity is recognized as a major driver of pathogen evolution. Influenza A viruses (IAVs) can infect many avian and mammalian hosts, but while few IAV subtypes circulate in human populations, subtype diversity is extensive in wild bird populations. How do these subtypes coexist in wild avian populations and do they compete within these natural host populations? Here we experimentally challenged mallard ducks with different IAVs to study how an initial infection with H3N8 determines the outcome of later infections (duration of infection and virus load) and antibody responses. There was complete protection to re-infection with the same H3N8 virus based on virus isolation. In addition, there was partial protection induced by H3N8 pre-challenge to other subtypes and development of heterosubtypic immunity indicated by shorter infections and reduction in viral load compared to controls. This indicates that subtype dynamics in the host population are not independent. Amongst H3N8 pre-challenged groups, the highest protection was conferred to the H4N5 subtype which was most genetically related to H3N8. The H4N5 challenge also induced an increase in H3 antibody levels and evidence for antigenic seniority. Thus, previous infections with IAV can influence the outcome of subsequent infection with different IAV subtypes. Results not only have relevance to understanding naturally occurring subtype diversity in wild avian populations but also in understanding potential outcomes associated with introduction of novel viruses such as highly pathogenic IAV H5 viruses in wild bird populations.Author contributionsConceived and designed the experiments: NLM, DES. Performed the experiments: NLM, JDB, AF, DC, MF, DES. Contributed reagents/materials/analysis tools: NLM, JB, AF, RLP, DES. Analyzed the data: NLM, DES. Wrote the paper: NLM, JDB, AF, RLP, DC, MF, DES