Evidence of subpopulations in aMPV vaccines using NGS

Avian Metapneumovirus (aMPV) causes an upper respiratory tract infection in turkeys and in some other avian species. Based on genome sequence differences, four subtypes of aMPV (A, B, C and D) have been identified but subtypes A and B have an almost worldwide distribution. Nowadays control strategies are mainly based on vaccination and different commercial live vaccines, originating from both subtype A and B strains, are available. The not absolute coverage of animal population achieved by ordinary mass vaccination in poultry industry allows a prolonged circulation of vaccine viruses in bird populations. Diseases outbreaks have been reported due to reversion to virulence of vaccines; and multiple nucleotide mutations in the aMPV genome associated to this phenomenon have been identified. In the current study, the presence of subpopulations in a subtype B vaccine was investigated by deep sequencing. Of the 19 positions where vaccine (strain VCO3/50) and progenitor (strain VCO3/60616) consensus sequences differed, subpopulations were found to have sequence matching progenitor sequence in 4 positions. However none of these mutations occurred in a virulent revertant of that vaccine, thereby demonstrating that the majority progenitor virus population had not survived the attenuation process, hence were not obviously involved in any return to virulence. However within the vaccine, a single nucleotide variation was found which agreed with consensus sequence of a derived virulent revertant virus, hence this and other undetected, potentially virulent subpopulations, can be involved in reversion. Much deeper sequencing of vaccine derived strains obtained during experimental infections or in the field may clarify if reversion to virulence was due to selective pressure acting on a homogeneous but rapidly evolving vaccine population or to the selection of virulent subpopulations already present in the vaccine batches.