Consistent occurrence of hemagglutinin variants among avian influenza virus isolates of the H7 subtype

Abstract

Several field isolates of avian influenza virus of the H7 subtype were analyzed for the presence of hemagglutinin variants by labeling proteins in cells infected with virus clones, and reacting with monoclonal antibodies. Each strain was shown to contain two distinct electrophoretic variants of the uncleaved hemagglutinin. In the A/Tk/Ore/71 (H7N3) isolate, two variants remained in the population through 35 laboratory passages, indicating both are stable and may be important to expression of the viral phenotype. Nucleotide sequence analysis of the HA gene of these two variants demonstrated differences at several amino acid positions in the HA, subunit including one glycosylation site. Three additional recent North American isolates were also each found to contain two electrophoretic variants occurring within populations as few as one embryo passage away from the original clinical specimen. Pulse-chase assays indicated none of the variant HA molecules were cleavable in chick embryo fibroblasts. In the highly pathogenic Australian isolate; A/Ck/Victoria/75, both HA variants are cleavable in fibroblasts, without added trypsin, and the differences are localized within the HA1 region. With all the strains tested, the slower migrating HA variant was associated with a consistently higher hemagglutinin titer in embryos. Finally, recent H7 isolates from imported birds (A/Soft Bill/III/92) also exhibit similar variants, indicating their occurrence is not limited to domestic poultry. This consistent presence of two distinct electrophoretic variants in several avian H7 isolates suggests multiple allelic forms of the H7 hemagglutinin.