Detection of Mammalian Virulence Determinants in Highly Pathogenic Avian Influenza H5N1 Viruses: Multivariate Analysis of Published Data

Highly pathogenic avian influenza (HPAI) virus H5N1 infects water and land fowl and can infect and cause mortality in mammals, including humans. However, HPAI H5N1 strains are not equally virulent in mammals, and some strains have been shown to cause only mild symptoms in experimental infections. Since most experimental studies of the basis of virulence in mammals have been small in scale, we undertook a meta-analysis of available experimental studies and used Bayesian graphical models (BGM) to increase the power of inference. We applied text-mining techniques to identify 27 individual studies that experimentally determined pathogenicity in HPAI H5N1 strains comprising 69 complete genome sequences. Amino acid sequence data in all 11 genes were coded as binary data for the presence or absence of mutations related to virulence in mammals or nonconsensus residues. Sites previously implicated as virulence determinants were examined for association with virulence in mammals in this data set, and the sites with the most significant association were selected for further BGM analysis. The analyses show that virulence in mammals is a complex genetic trait directly influenced by mutations in polymerase basic 1 (PB1) and PB2, nonstructural 1 (NS1), and hemagglutinin (HA) genes. Several intra- and intersegment correlations were also found, and we postulate that there may be two separate virulence mechanisms involving particular combinations of polymerase and NS1 mutations or of NS1 and HA mutations.