Comparative genomic analysis of pre-epidemic and epidemic Zika virus strains for virological factors potentially associated with the rapidly expanding epidemic

Zhu, Zheng; Chan, Jasper Fuk-Woo; Tee, Kah-Meng; Choi, Garnet Kwan-Yue; Lau, Susanna Kar-Pui; Woo, Patrick Chiu-Yat; Tse, Herman; Yuen, Kwok-Yung

Phylogenetic tree construction by the maximum likelihood method was performed using MEGA 6.0 software, with bootstrap values being calculated from 500 trees. Protein family analysis was performed using the PFAM tool (http://pfam.xfam.org) Search for Conserved Domains server (http://www.ncbi.nlm.nih.gov/ Structure/cdd/wrpsb.cgi). Prediction of transmembrane domains was performed using the TMHMM 2.0 server (http://www.cbs.dtu. dk/services/TMHMM/). Prediction of signal peptides was performed by using signalP software 4.1 (http://www.cbs.dtu.dk/ services/SignalP/). The prediction of potential O-glycosylation and N-glycosylation sites in the polyprotein was performed using NetOGlyc 4.0 (http://www.cbs.dtu.dk/services/NetOGlyc/) and NetNGlyc 1.0 (http://www.cbs.dtu.dk/services/NetNGlyc/), respectively. Secondary structure prediction in the 5ʹ-untranslated region (UTR) was performed using the RNAfold WebServer with default settings (http://rna.tbi.univie.ac.at/cgi-bin/RNAfold.cgi). The number of synonymous substitutions per synonymous site, Ks, and the number of nonsynonymous substitutions per nonsynonymous site, Ka, for each coding region was calculated using the Nei and Gojobori substitution model with Jukes-Cantor correction in MEGA 6.0. 16, 17 Bootscan analysis was performed using Simplot version 3.5.1 as described previously, 18 with the Asian lineage of ZIKV strains as the query. Multiple alignment of the amino acid sequence of each protein are performed by