Concept	Explanation	“holmes_figure_box.pdf”
Mutation rate	As viruses replicate, mutational errors are incorporated into the viral genome. The mutation rate is therefore typically expressed as the number of mutations per site, per replication event. The mutation rate for RNA viruses such as EBOV is largely determined by the viral RNA-dependent RNA polymerase, which lacks proof-reading activity. The estimation of mutation rates requires complex sequencing-based or phenotypic marker experiments that correct for the impact of natural selection79. Mutation rates are unknown for most viruses and have not been determined for EBOV, although it would be predicted to be comparable to other (−)ssRNA viruses and likely similar across all EBOV outbreak variants.
Evolutionary rate	The evolutionary rate of a virus can be defined as the observed rate at which new variants arise and spread in the viral population as the result of a complex interplay of natural selection, genetic drift, modes of transmission and epidemiological processes. This can be measured by methods that compare the genetic change in viral genomes collected at different times. Importantly, evolutionary rates in RNA viruses may be dependent on the time-scale over which they are measured: they are elevated in the short-term, such as within disease outbreaks, because mildly deleterious mutations may not have been eliminated by purifying selection71,72.
Substitution rate	The substitution rate is best described as the long-term rate at which genetic variants become fixed in a virus lineage over evolutionary time-scales, such as between human outbreaks in the case of EBOV. This rate will usually be lower than the short-term evolutionary rate, because many of the variants circulating within outbreaks and epidemics will ultimately be eliminated. Furthermore, saturation – repeated changes at the same site – will further reduce the measured substitution rate.
Fixation rate	An added complexity in estimating rates in RNA viruses is that the population genetic concept of ‘fixation’, central to the definition of substitution, is ill-defined. In slowly evolving organisms fixation events can usually be distinguished from polymorphisms by analysing individual nucleotide sites within and between species. However, in rapidly evolving RNA viruses fixation can be described to occur (1) at the level of individual hosts over the course of infection, (2) in viral lineages within specific geographic locations or epidemiological networks (such as the different lineages of EBOV generated during the 2013–2016 EVD epidemic), (3) in global meta-populations, and (4) between different viral species.