PB2 amino acid at position 627 affects replicative efficiency, but not cell tropism, of Hong Kong H5N1 influenza A viruses in mice

Abstract

A single amino acid substitution, from glutamic acid to lysine at position 627 of the PB2 protein, converts a nonlethal H5N1 influenza A virus isolated from a human to a lethal virus in mice. In contrast to the nonlethal virus, which replicates only in respiratory organs, the lethal isolate replicates in a variety of organs, producing systemic infection. Despite a clear difference in virulence and organ tropism between the two viruses, it remains unknown whether the dissimilarity is a result of differences in cell tropism or the reduced replicative ability of the nonlethal virus in mouse cells in general. To determine how this single amino acid change affects virulence and organ tropism in mice, we investigated the growth kinetics of the two H5N1 viruses both in vitro and in vivo. The identity of the PB2 amino acid at position 627 did not appreciably affect viral replicative efficiency in chicken embryo fibroblasts and a quail cell line; however, viruses with lysine at this position instead of glutamic acid grew better in the different mouse cells tested. When the effect of this substitution was investigated in mice, all of the test viruses showed the same cell tropism, but infection by viruses containing lysine at position 627 spread more rapidly than those viruses containing glutamic acid at this position. Further analysis showed a difference in local immune responses: neutrophil infiltration in lungs infected with viruses containing lysine at position 627 persisted longer than that associated with viruses lacking a glutamic acid substitution. Our data indicate that the amino acid at position 627 of the PB2 protein determines the efficiency of viral replication in mouse (not avian) cells, but not tropism among cells in different mouse organs. The presence of lysine leads to more aggressive viral replication, overwhelming the host's defense mechanisms and resulting in high mortality rates in mice.

Requirement for Siva-1 for replication of influenza A virus through apoptosis induction

“…After 48 h, the viability of the cells was monitored using a Cell Counting kit 8. It has been reported previously that a specific strain of H5N1 that is a highly pathogenic avian influenza virus causes extensive lung injury to mice and ferrets in experimental infections (Martin & Wurfel, 2008;Shinya et al, 2004;Tumpey et al, 2000). Despite the severity of experimental infection in animals, this highly pathogenic H5N1 subtype avian influenza A virus showed a lower level of efficiency of apoptosis induction in human macrophages by in vitro infection (Mok et al, 2007;Zhou et al, 2006).…”

Why is the world so poorly prepared for a pandemic of hypervirulent avian influenza?

“…In contrast to the non-lethal virus, which replicates only in respiratory organs, the lethal isolate was found to replicate in a variety of organs in mice, producing systemic infection (67). When tested in chicken embryo cells and a quail cell line, it was found that the identity of the PB2 amino acid at position 627 did not appreciably affect the replicative efficiency of the virus (67). However, viruses with lysine at this position instead of glutamic acid grew better in various types of mouse cells that were tested (67).…”

“…Yet another type of high-virulence character that has been found in some of those dangerous H5N1 strains that now circulate in birds is located in viral RNA polymerase complex genes (67,68). It was shown in one study that a single amino acid substitution, from glutamic acid to lysine at position 627 of the PB2 protein (which is one of the components of the viral RNA polymerase), was enough to convert a non-lethal H5N1 influenza A virus isolated from a human to a lethal virus in mice (67).…”

“…It was shown in one study that a single amino acid substitution, from glutamic acid to lysine at position 627 of the PB2 protein (which is one of the components of the viral RNA polymerase), was enough to convert a non-lethal H5N1 influenza A virus isolated from a human to a lethal virus in mice (67). In contrast to the non-lethal virus, which replicates only in respiratory organs, the lethal isolate was found to replicate in a variety of organs in mice, producing systemic infection (67). When tested in chicken embryo cells and a quail cell line, it was found that the identity of the PB2 amino acid at position 627 did not appreciably affect the replicative efficiency of the virus (67).…”

“…When tested in chicken embryo cells and a quail cell line, it was found that the identity of the PB2 amino acid at position 627 did not appreciably affect the replicative efficiency of the virus (67). However, viruses with lysine at this position instead of glutamic acid grew better in various types of mouse cells that were tested (67). When the effect of this substitution was investigated in vivo in mice, it was found that all of the test viruses showed the same cell tropism, but infection by viruses containing lysine at position 627 spread more rapidly than those viruses containing glutamic acid at this position (67).…”