Toward unified molecular surveillance of RSV: A proposal for genotype definition

Background Human respiratory syncytial virus (RSV) is classified into antigenic subgroups A and B. Thirteen genotypes have been defined for RSV-A and 20 for RSV-B, without any consensus on genotype definition. Methods We evaluated clustering of RSV sequences published in GenBank until February 2018 to define genotypes by using maximum likelihood and Bayesian phylogenetic analyses and average p-distances. Results We compared the patterns of sequence clustering of complete genomes; the three surface glycoproteins genes (SH, G, and F, single and concatenated); the ectodomain and the 2nd hypervariable region of G gene. Although complete genome analysis achieved the best resolution, the F, G, and G-ectodomain phylogenies showed similar topologies with statistical support comparable to complete genome. Based on the widespread geographic representation and large number of available G-ectodomain sequences, this region was chosen as the minimum region suitable for RSV genotyping. A genotype was defined as a monophyletic cluster of sequences with high statistical support (>= 80% bootstrap and >= 0.8 posterior probability), with an intragenotype p-distance <= 0.03 for both subgroups and an intergenotype p-distance >= 0.09 for RSV-A and >= 0.05 for RSV-B. In this work, the number of genotypes was reduced from 13 to three for RSV-A (GA1-GA3) and from 20 to seven for RSV-B (GB1-GB7). Within these, two additional levels of classification were defined: subgenotypes and lineages. Signature amino acid substitutions to complement this classification were also identified. Conclusions We propose an objective protocol for RSV genotyping suitable for adoption as an international standard to support the global expansion of RSV molecular surveillance.