Surveillance of Class I Newcastle Disease Virus at Live Bird Markets in China and Identification of Variants with Increased Virulence and Replication Capacity.

In this study, 232 class I Newcastle disease viruses (NDVs) were identified from multiple bird species at nationwide live bird markets (LBMs) from 2017 to 2019 in China. Phylogenetic analysis indicated that all 232 isolates were clustered into genotype 1.1.2 of class I on the basis of the fusion (F) gene sequences, which were distinct from the genotypes identified in other countries. Most of the isolates (212/232) were shown to have the typical F gene molecular characteristics of class I NDVs, while a few (20/232) contained mutations at the site of the conventional start codon of the F gene, which resulted in open reading frames (ORFs) altered in length. The isolates with ACG, CTA, and ATA mutations showed different levels of increased virulence and replication capacity, suggesting that these viruses may be transitional types during the evolution of class I NDVs from avirulent to virulent. Further evaluation of biological characteristics with recombinant viruses obtained by reverse genetics demonstrated that the ATG located at genomic positions 4523 to 4525 was the authentic start codon in the F gene of class I NDV, and the specific ATA mutations which contributed to the expression of F protein on the surface of infected cells were the key determinants of increased replication capacity and virulence. Interestingly, the mutation at the corresponding site of genotype II LaSota of class II had no effects on the virulence and replication capacity in chickens. Our results suggest that the alteration of virulence and replication capacity caused by specific mutations in the F gene could be a specific characteristic of class I NDVs and indicate the possibility of the emergence of virulent NDVs due to the persistent circulation of class I NDVs. IMPORTANCE The available information on the distribution, genetic diversity, evolution, and biological characteristics of class I Newcastle disease viruses (NDVs) in domestic poultry is currently very limited. Here, identification of class I NDVs at nationwide live bird markets (LBMs) in China was performed and representative isolates were characterized. A widespread distribution of genotype 1.1.2 of class I NDVs was found in multiple bird species at LBMs in China. Though most isolates demonstrated typical molecular characteristics of class I NDVs, a few that contained specific mutations at the site of the conventional start codon of the fusion gene with increased virulence and replication capacity were identified for the first time. Our findings indicate that the virulence of class I NDVs could have evolved, and the widespread transmission and circulation of class I NDVs may represent a potential threat for disease outbreaks in poultry.