Genome sequencing reveals Zika virus diversity and spread in the Americas

Abstract

2Despite great attention given to the recent Zika virus (ZIKV) epidemic in the Americas, much remains unknown about its epidemiology and evolution, in part due to a lack of genomic data. We applied multiple sequencing approaches to generate 100 ZIKV genomes from clinical and mosquito samples from 10 countries and territories, greatly expanding the observed viral genetic diversity from this outbreak. We analyzed the timing and patterns of introductions into distinct geographic regions, confirming phylogenetic evidence for the origin and rapid expansion of the outbreak in Brazil 1 , and for multiple introductions from Brazil into Honduras, Colombia, Puerto Rico, other Caribbean islands, and the continental US. We find that ZIKV circulated undetected in many regions of the Americas for up to a year before the first locally transmitted cases were confirmed, highlighting the challenge of effective surveillance for this virus. We further characterize genetic variation across the outbreak to identify mutations with possible functional implications for ZIKV biology and pathogenesis.Since its introduction into Brazil in 2013 1 , mosquito-borne ZIKV (Family: Flaviviridae) has spread rapidly throughout the Americas, causing hundreds of thousands of cases of ZIKV disease, as well as ZIKV congenital syndrome and likely other neurological complications 2-4 . Comparative phylogenomic analysis of ZIKV can reveal the trajectory of the outbreak and detect mutations that may be associated with new disease phenotypes or affect molecular diagnostics. Despite the nearly 60 years since its discovery and the scale of the recent outbreak, however, fewer than 100 ZIKV genomes have been sequenced directly from clinical samples. This is due in part to technical challenges posed by low viral loads (often orders of magnitude lower than in Ebola or dengue virus infection 5-7 ), as well as issues of RNA preservation in samples collected without the unique requirements of sequencing in mind. Culturing the virus can greatly increase the material available for sequencing, but it can introduce artefacts and is time-consuming and difficult.We sought to gain a deeper understanding of the viral populations underpinning the ZIKV epidemic by extensive genome sequencing of the virus directly from samples collected as part of ongoing surveillance. We initially pursued metagenomic RNA sequencing in order to capture both ZIKV and other potential co-infections in an unbiased way. In most of the 37 samples examined by this approach, however, the amount of ZIKV material was not sufficient for genome assembly. Unbiased RNA sequencing still proved valuable because it provided ZIKV data to verify results from other methods. We also observed 3 other viruses in 7 samples (Extended Data Table 1) in our metagenomic data; notably, these did not include chikungunya or dengue, viruses known to be cocirculating with ZIKV in many affected regions 8,9 .In order to capture sufficient ZIKV content for genome assembly, we turned to two targeted enrichment approaches: hybrid c...

Epidemic establishment and cryptic transmission of Zika virus in Brazil and the Americas

“…To further elucidate Zika virus transmission in the Americas, we include in our analyses 5 new ZIKV complete genomes from Colombia and 4 from Mexico. Further, we append to our dataset 115 publicly available sequences as well as 85 additional genomes from a companion paper 20 . The dataset used for genetic analysis thus comprised 254 complete and near complete genomes; 241 of which were sampled in the Americas (Methods).…”

“…Our final alignment comprised the sequences reported in this study (n=54) plus publicly available ZIKV-Asian genotype sequences, as of 1 st March 2017 (n=115). We also included in our analysis 85 additional genomes from a companion paper 20 . The dataset used for analysis therefore included sequences from 254 Zika virus isolates, 241 of which from the Americas.…”