Phylogenetic analysis and molecular epidemiology
Molecular epidemiology was used to define evidence of virus clustering
and thus the occurrence of swine-to-swine transmission. Analysis was
conducted on full genomes of all H3N2 swine influenza virus isolates
sequenced in our study (Figure 1). The influenza virus sequences were
analysed together with other existing Asian H3-subtype swine influenza
virus sequences retrieved from the NCBI Influenza Virus Resource
(Supplementary table 2) (Bao, 2008) and GISAID (Supplementary table 3)
(Shu, 2017). All Asian H3 SIVs with a complete set of segments in
full-length were included and identical sequences were collapsed. Each
genome segment was aligned separately using MAFFT version 7 (Katoh,
2013) and alignments were then trimmed to coding regions. The resultant
data sets contained a total of 353 full genome sequence sets.
The coding regions of gene segments PB2, PB1, PA, HA, NP, NA, MP and NS
were concatenated using Geneious 2019.2.1 (http://www.geneious.com).
Multiple alignment for the concatenated whole genomes was performed with
MAFFT version 7. We inferred the maximum-likelihood phylogenetic trees
using IQ-TREE (Nguyen, 2014) and the built-in ModelFinder
(Kalyaanamoorthy, 2017) to determine the best-fit substitution model.
Branch supports were obtained using approximate likelihood ratio test
(aLRT) (Guindon, 2010) to assess the robustness of the phylogenetic
tree. Branch support aLRT statistics were shown at major nodes with
values larger or equal to 0.8.