Whole genome sequencing of measles viruses: the benefit for outbreak investigation, The Netherlands, 2013-2014.

Measles is a highly contagious infectious disease and remains a major cause of global morbidity and mortality. Molecular surveillance of measles virus (MV) is used for tracing transmission chains, by, for instance, distinguishing between repeated introductions and endemic circulation. WHO guidelines used in the global measles surveillance network recommend sequencing of a 450-nucleotide region at the C-terminal end of the N-gene (N450). Given the limited genetic variability of MV, whole-genome sequencing would increase the resolution and provide a more detailed molecular surveillance of circulating MV. In this study, we describe an amplicon-based nanopore sequencing protocol for generating near complete whole MV genome sequences (nWGS; 15,813 nucleotides). We applied this protocol to samples collected during an epidemic of measles in the Netherlands in 2013-2014, involving an estimated 30,000 cases, to quantify the sequence variation of nWGS during this epidemic. While this was considered an epidemic with very limited sequence variation based on analysis of the N450 region, different molecular clusters were identified by phylogenetic analysis of nWGS genomes. Notably, the number of single nucleotide variants (SNVs) between viruses detected during the early and late phase of the epidemic varied among molecular clusters. Four epidemiological clusters could be identified in the beginning of the epidemic, of which 2 were supported by molecular data. Analysis of nWGS genomes suggested that the epidemic started with either a single unnoticed introduction into the Netherlands and a few unnoticed generations or with at least 2 introductions at the same time, the latter option is supported by epidemiological data. These results can aid in interpreting MV sequence variation during transmission chains, outbreaks and epidemics in countries or regions approaching measles elimination.