|GARCIA, MARICARMEN - University Of Georgia|
|VOLKENING, JEREMY - US Department Of Agriculture (USDA)|
|RIBLET, SYLVIA - University Of Georgia|
Submitted to: Virology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 2/12/2013
Publication Date: 3/26/2013
Citation: Garcia, M., Volkening, J., Riblet, S., Spatz, S.J. 2013. Genomics sequence analysis of the United States infectious laryngotracheitis vaccine strains chicken embryo origin (CEO) and tissue culture origin (TCO). Virology. 440(2013):64-74. doi: 10.1016/j.virol.2013.02.007.
Interpretive Summary: Infectious laryngotracheitis (ILT) is a highly contagious respiratory disease of chickens caused by Gallid herpesvirus-1 (GaHV-1). To define relevant genomic changes associated with attenuation and reversion to virulence of U.S. GaHV-1 vaccine strains chicken embryo origin (CEO) and tissue culture origin (TCO), the genomic sequences of both low and high passages of these strains were determined using hybrid next generation sequencing. Phylogenetic analysis of available GaHV-1 full genomes grouped strains into four major clades: the TCO clade including the LT-Ivax vaccine, virulent 81658 isolate, and the USDA reference strain; the CEO clade including four U.S. CEO vaccines, U.S. virulent isolate 63140, and the European CEO vaccine; the clade comprised by U.S. virulent isolate 1874C5; and the clade comprised by Australian vaccine stains SA2 and A20. Comparative genomics of vaccine strains revealed that TCO attenuation is likely the result of an ORF C truncation, and TCO reversion to virulence requires at least mutations in US10 and UL5, while mutations in UL10, US8 and UL5 are strongly related to CEO vaccine attenuation. Analysis of deep sequencing data revealed the presence of mixed genome populations within the vaccine preparations and their passages. Although no single mutation in common among all U.S. virulent strains relative to the CEO and TCO vaccine strains was identified, 15 unique single nucleotide polymorphisms (SNPs) distributed in 12 genes of the CEO clade virulent isolate 63140 and 20 SNPs located mostly in UL27 UL30, UL36 and ICP4 genes of the 1874C5 virulent isolate are most-likely associated with virulence.
Technical Abstract: The genomic sequences of low and high passages of the United States infectious laryngotracheitis (ILT) vaccine strains CEO and TCO were determined using hybrid next generation sequencing in order to define genomic changes associated with attenuation and reversion to virulence. Phylogenetic analysis of available full genomes grouped strains into three major clades: TCO, CEO, and Australian. Comparative genomics revealed that TCO attenuation is likely the result of an ORF C truncation. Genes involved in attenuation are generally clade-specific, however four genes ORF C, UL27, UL28 and UL39 commonly contained various mutations across the CEO and TCO lineages. The Thr644 mutation in the UL27 gene encoding glycoprotein B was identified in all virulent US strains. The US10 gene was identified as a potential virulence factor for the TCO revertant 81658. The UL41 gene was responsible for the robust gain in virulence of CEO-Fowl Laryngotracheitis(®) after 20 passages in chickens.