Genes involved in the virulence of an avian influenza virus

Abstract

A/chicken/Japan/24 (Japan) (HavlNegl) isolated from an outbreak of fowl plague-like illness among chickens is comparable to fowl plague viruses in its virulence, causing paralysis and death of chickens within 3 days. It also kills chick embryos after allantoic infection of embryonated eggs. Another avian influenza virus, A/duck/Ukraine/1/63 (Ukraine) (Hav7Neg2), in contrast, causes no illness or death in either chickens or chick embryos. In an attempt to determine the genes involved in the observed difference in virulence, we prepared a series of recombinants between Japan and Ukraine viruses. Parental derivation of genes in the recombinants was determined by urea-polyacrylamide gel electrophoresis of viral RNA. The virulence was assessed by three criteria: (1) lethality for chickens after intracerebral inoculation, (2) lethality for embryos after allantoic inoculation into 15-day-old embryonated eggs, and (3) lethality for embryos after allantoic inoculation into 10-day-old embryonated eggs. The recombinants in which genes coding for hemagglutinin (HA), neuraminidase (NA), and matrix (M) proteins had been derived from Japan virus and five other genes from Ukraine virus were fully virulent by all three criteria. The recombinants which had received HA and NA genes from Japan virus showed a slightly but definitely reduced virulence in chickens and 15-day-old embryos but retained the virulence in 10-day-old embryos unchanged. The recombinants which had received only HA gene from Japan virus had a markedly diminished virulence in chickens and 15-day-old embryos but were as lethal for 10-day-old embryos as Japan virus. It was concluded that HA gene was the key determinant of virulence of Japan virus, but, in addition, NA and M genes were required for the full expression of virulence. Requirement of HA and NA genes was confirmed by reconstitution of virulence after reuniting the two genes.