Computational protocol: Bocavirus Infection in Otherwise Healthy Children with Respiratory Disease

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Protocol publication

[…] All alignments were performed using ClustalX 2.1 and BioEdit (version 7.1.3.0) software (Ibis Biosciences, Carlsbad, CA). Phylogenetic trees of the VP-1/2 protein gene were generated using the neighbour-joining method and p-distance model of the Molecular Evolutionary Genetics Analyses (MEGA) software, version 5.05 []. Bootstrap probabilities for 1,000 iterations were calculated to evaluate confidence estimates. The graphs were made using GraphPad Prism version 5.01 for Windows (GraphPAD Software, San Diego, CA). All genotyped sequences of the hBoV VP-1/2 gene were submitted to GenBank (accession numbers KR014412-KR014516). [...] Tests for positive selection were conducted on the Datamonkey server [] using the single-likelihood ancestor (SLAC), and the fixed-effects likelihood (FEL) [], the internal branch fixed-effects likelihood (IFEL) [], the mixed effects model of evolution (MEME) [], and fast unconstrained Bayesian approximation methods (FUBAR). The dN/dS ratios were calculated using the SLAC and FEL codon-based maximum likelihood approaches. The SLAC approach counts the number of non-synonymous changes per non-synonymous site (dN) and tests whether it is significantly different from the number of synonymous changes per synonymous site (dS). The FEL approach estimates the ratios of non-synonymous to synonymous changes for each site in an alignment. The IFEL method is similar to the FEL, but tests site-by-site selection along only the internal branches of the phylogeny. In order to avoid an excessive false-positive rate, sites with SLAC, FEL, IFEL and MEME p-values of <0.1 and a FUBAR posterior probability of >0.90 were accepted as candidates for selection. […]

Pipeline specifications

Software tools Clustal W, BioEdit, MEGA, Datamonkey
Applications Phylogenetics, Population genetic analysis
Organisms Human bocavirus, Homo sapiens
Diseases Respiratory Insufficiency, Respiratory Tract Infections