M Gene Reassortment in H9N2 Influenza Virus Promotes Early Infection and Replication: Contribution to Rising Virus Prevalence in Chickens in China

Abstract

Segment reassortment and base mutagenesis of influenza A viruses are the primary routes to the rapid evolution of high-fitness virus genotypes. We recently described a predominant G57 genotype of avian H9N2 viruses that caused countrywide outbreaks in chickens in China during 2010 to 2013, which led to the zoonotic emergence of H7N9 viruses. One of the key features of the G57 genotype is the replacement of the earlier A/chicken/Beijing/1/1994 (BJ/94)-like M gene with the A/quail/Hong Kong/G1/1997 (G1)-like M gene of quail origin. We report here the functional significance of the G1-like M gene in H9N2 viruses in conferring increased infection severity and infectivity in primary chicken embryonic fibroblasts and chickens. H9N2 virus housing the G1-like M gene, in place of the BJ/94-like M gene, showed an early surge in viral mRNA and viral RNA (vRNA) transcription that was associated with enhanced viral protein production and with an early elevated release of progeny virus comprising largely spherical rather than filamentous virions. Importantly, H9N2 virus with the G1-like M gene conferred extrapulmonary virus spread in chickens. Five highly represented signature amino acid residues (37A, 95K, 224N, and 242N in the M1 protein and 21G in the M2 protein) encoded by the prevalent G1-like M gene were demonstrated to be prime contributors to enhanced infectivity. Therefore, the genetic evolution of the M gene in H9N2 virus increases reproductive virus fitness, indicating its contribution to the rising virus prevalence in chickens in China. We recently described the circulation of a dominant genotype (genotype G57) of H9N2 viruses in countrywide outbreaks in chickens in China, which was responsible, through reassortment, for the emergence of H7N9 viruses that cause severe human infections. A key feature of the genotype G57 H9N2 virus is the presence of the quail-origin G1-like M gene, which had replaced the earlier BJ/94-like M gene. We found that H9N2 virus with the G1-like M gene, but not the BJ/94-like M gene, showed an early surge in progeny virus production and more severe pathology and extrapulmonary virus spread in chickens. Five highly represented amino acid residues in the M1 and M2 proteins derived from the G1-like M gene were shown to mediate enhanced virus infectivity. These observations enhance what we currently know about the roles of reassortment and mutations in virus fitness and have implications for assessing the potential of variant influenza viruses that can cause a rising prevalence in chickens.

The phylogenetic analysis and molecular characterization of H9N2 avian influenza viruses isolated in Jiangsu province from 2014 to 2016

“…相较于以前的毒 株, 该基因型的H9N2病毒能更稳定地存在于自然界, 且对鸡的致病性和接触传播性更强 [5] . 研究表明, M基 因是增强该基因型的H9N2 AIV致病性的关键基因之 一 [21] . 本研究中的10株H9N2分离株的基因型并没有 发生变化, 仍然是B69.…”

Highly Efficient Production of an Influenza H9N2 Vaccine Using MDCK Suspension Cells

“…This has resulted in severe economic burdens to the poultry industry by the decrease in egg production and the moderate-to-high mortality of poultry (Lamb and Takeda 2001; Lee and Song 2013; Pu et al 2015). In addition, H9N2 viruses have revealed the potential to cause the pandemic due to the emerging reported cases of avian-to-human transmission of H9N2 viruses and the detections of partial H9N2 virus-derived genomic segments in the emerging highly pathogenic human in uenza viruses regarding H7N9, H5N1, H10N8 and H5N6 virus reassortants (Gu et al 2017;Li et al 2014;Pu et al 2017;Sorrell et al 2009). To face the threat posed by the emerging H9N2 viruses, vaccination is considered as an effective measurement to control the virus spread among the poultry and to limit the health risks to humans (Genzel and Reichl 2009;Park et al 2011).…”

Competitive exclusion strengthens selection for transmissibility and increases the benefit of recombination for within-host adaptation

“…Gene flow between coinfecting bacterial populations, however, may be less restricted, as it can occur through multiple mechanisms such as uptake of free DNA of dead bacteria (transformation), direct exchange between cells through a pilus (conjuation), or hitchhiking of bacterial DNA in bacteriophages (transduction). These processes have been shown to accelerate adaptation in multiple pathogen systems, including Φ6 bacteriophage [13, 14], influenza A virus [15, 16], and S. pneumoniae [17, 18]. While competitive exclusion generally prevents superinfecting variants from replicating and contributing to infection directly, the incorporation of alleles from these newcomers by an established resident population may serve as a second method of within-host adaptation.…”