Whole genome analysis of local Kenyan and global sequences unravels the epidemiological and molecular evolutionary dynamics of RSVgenotype ON1 strains

The respiratory syncytial virus (RSV) group A variant with the 72-nucleotide duplication in the G gene, genotype ON1, was first detected in Kilifi in 2012 and has almost completely replaced previously circulating genotype GA2 strains. This replacement suggests some fitness advantage of ON1 over the GA2 viruses, and might be accompanied by important genomic substitutions in ON1 viruses. Close observation of such a new virus introduction over time provides an opportunity to better understand the transmission and evolutionary dynamics of the pathogen. We have generated and analyzed 184 RSV-A whole genome sequences (WGS) from Kilifi (Kenya) collected between 2011 and 2016, the first ON1 genomes from Africa and the largest collection globally from a single location. Phylogenetic analysis indicates that RSV-A transmission into this coastal Kenya location is characterized by multiple introductions of viral lineages from diverse origins but with varied success in local transmission. We identify signature amino acid substitutions between ON1 and GA2 viruses within genes encoding the surface proteins (G, F), polymerase (L) and matrix M2-1 proteins, some of which were identified as positively selected, and thereby provide an enhanced picture of RSV-A diversity. Furthermore, five of the eleven RSV open reading frames (ORF) (i.e. G, F, L, N and P), analyzed separately, formed distinct phylogenetic clusters for the two genotypes. This might suggest that coding regions outside of the most frequently studied G ORF play a role in the adaptation of RSV to host populations with the alternative possibility that some of the substitutions are nothing more than genetic hitchhikers. Our analysis provides insight into the epidemiological processes that define RSV spread, highlights the genetic substitutions that characterize emerging strains, and demonstrates the utility of large-scale WGS in molecular epidemiological studies. Author summary Respiratory syncytial virus (RSV) is the leading viral cause of severe pneumonia and bronchiolitis among infants and children globally. No vaccine exists to date. The high genetic variability of this RNA virus, characterized by group (A or B), genotype (within group) and variant (within genotype) replacement in populations, may pose a challenge to effective vaccine design by enabling immune response escape. To date most sequence data exists for the highly variable G gene encoding the RSV attachment protein, and there is little globally-sampled RSV genomic data to provide a fine resolution of the epidemiology and evolutionary dynamics of the pathogen. Here we use long-term RSV surveillance in coastal Kenya to track the introduction, spread and evolution of a new RSV genotype known as ON1 (having a 72-nucleotide duplication in the G gene). We present a set of 184 RSV-A whole genomes, including 176 of RSV ON1 (the first from Africa), describe patterns of local ON1 spread and show genome-wide changes between the two major RSV-A genotypes that may define the pathogen’s adaptation to the host. These findings have implications for vaccine design and improved understanding of RSV epidemiology and evolution.