Computational protocol: Wild birds in Chile Harbor diverse avian influenza A viruses

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Protocol publication

[…] Influenza gene sequences were obtained from the NCBI Influenza Virus Database (https://www.ncbi.nlm.nih.gov/genomes/FLU/Database) as accessed in November 2017. Only full-length genes from the Americas, Europe, and Asian avian strains were included while duplicates were excluded. Sequence assembly, visual inspection, and trimming to remove nucleotides outside the coding region was performed using BioEdit version 7.2.5 (ref. ) and alignment performed with MUSCLE version 3.8.3 (ref. ). The best-fit nucleotide substitution models were selected individually for each gene by ModelTest in in Mega 7 (ref. ). Phylogenic relationships for each gene were inferred by Maximum Likelihood (ML), incorporating a general time-reversible model of nucleotide substitution with a gamma-distributed rate variation among sites and a proportion of invariant sites (GTR + G + I). One thousand bootstrap replicates were performed to infer the robustness of the ML trees using RaxML version 8.0 (ref. ). ML inference was repeated at least three times per dataset to assure tree topology was maintained. Final trees were constructed using TreeGraph2 and FigTree v1.4.3 (ref. ). For the analysis of internal genes, we obtained all publicly available genes of wild bird origin obtained between 1976 and 2017 from Eurasia and the Americas. We then randomly selected 10 strains/year/location per gene for the analysis. This process was repeated at least twice and final tees were run at least three times to assure consistency in consultation with Dr. Justin Bahl. The final number of selected taxa per tree is specified in Supplemental Figures  to that include expanded trees showing all sequences included for analysis. […]

Pipeline specifications

Software tools BioEdit, MUSCLE, ModelTest-NG, MEGA, RAxML, TreeGraph, FigTree
Databases IVDB
Applications Phylogenetics, Nucleotide sequence alignment
Organisms Viruses, Influenza A virus