Rapid PRRSV-2 ORF5-based lineage classification using Nextclade

Abstract

Porcine reproductive and respiratory syndrome virus (PRRSV) continues to be a global challenge for swine health. Yim-Im et al. 2023 provides a standard genetic nomenclature, extending previously published works to better characterize PRRSV-2 ORF5-based genetic lineages on a global scale. To facilitate the use of this nomenclature, scaffold sequences, including historical and contemporary vaccines, were synthesized into a dataset designed for Nextclade v3.0. Metadata from the scaffold sequences representing year, country, and RFLP typing of the sequence were incorporated into the dataset. These scaffold sequences were processed through the Augur pipeline using DQ478308.1 as a reference strain for rooting and comparison. The resultant classifier can be accessed through the Nextclade website (https://clades.nextstrain.org/) or a link on the PRRSView homepage (https://prrsv.vdl.iastate.edu/). The resultant classifier functions the same as other classifiers hosted by the Nextclade core group and can provide phylogenetic-based PRRSV-2 ORF5 classifications on demand. Nextclade provides additional sequence metrics such as classification quality and notable mutations relative to the reference. The submitted sequences are grafted to the reference tree using phylogenetic placement, allowing for comparison to nearby sequences of reference viruses and vaccine strains. Additional comparisons between sequences can be made with metadata incorporated in the dataset. Although Nextclade is hosted as a webtool, the sequences are not uploaded to a server, and all analysis stay strictly confidential to the user. This work provides a standardized, trivial workflow facilitated by Nextclade to rapidly assign lineage classifications to PRRSV-2, identify mutations of interest, and compare contemporary strains to relevant vaccines.

Keywords: ORF5; PRRSV; classification; lineage; sublineage.

Figure 1

Overview of the PRRSV-2 ORF5 lineages comprising the 1,100 sequences in the reference dataset. (A) The number per year of each lineage represented in the dataset. (B) The geographic breakdown of where sequences originated. Sequences originated from 10 countries colored in blue: Canada, China, India, Japan, Mexico, South Korea, Taiwan, Thailand, the United States, and Vietnam. Breakdowns of lineages from these countries used in each dataset are shown as a pie chart.

Figure 2

The start screen of the Nextclade web platform, indicated by the navigation bar at the top. The reference dataset has been set to PRRSV-2 ORF5 lineages.

Figure 3

The results screen of the Nextclade web platform, indicated by the navigation bar at the top. Sequence name, lineage assignment, and sequence quality metrics are displayed in tabular format. Additional beneficial information is displayed such as sequence duplication in the sequence name column, and single nucleotide polymorphisms from the base genome on the graph to the chart to the right.

Figure 4

The tree screen of the Nextclade web platform, as indicated by the navigation bar at the top. This screen displays a feature rich phylogenetic tree where branches are colored based on phylogenetic lineage, new strains of interest are emphasized, and vaccine strains are denoted through X’s marked on the tree. Additional display and filter options are available from the panel on the left-hand side of the tree.