Large-Scale Phylogenomic Analysis Reveals the Complex Evolutionary History of Rabies Virus in Multiple Carnivore Hosts

Abstract

The natural evolution of rabies virus (RABV) provides a potent example of multiple host shifts and an important opportunity to determine the mechanisms that underpin viral emergence. Using 321 genome sequences spanning an unprecedented diversity of RABV, we compared evolutionary rates and selection pressures in viruses sampled from multiple primary host shifts that occurred on various continents. Two major phylogenetic groups, bat-related RABV and dog-related RABV, experiencing markedly different evolutionary dynamics were identified. While no correlation between time and genetic divergence was found in bat-related RABV, the evolution of dog-related RABV followed a generally clock-like structure, although with a relatively low evolutionary rate. Subsequent molecular clock dating indicated that dog-related RABV likely underwent a rapid global spread following the intensification of intercontinental trade starting in the 15th century. Strikingly, although dog RABV has jumped to various wildlife species from the order Carnivora, we found no clear evidence that these host-jumping events involved adaptive evolution, with RABV instead characterized by strong purifying selection, suggesting that ecological processes also play an important role in shaping patterns of emergence. However, specific amino acid changes were associated with the parallel emergence of RABV in ferret-badgers in Asia, and some host shifts were associated with increases in evolutionary rate, particularly in the ferret-badger and mongoose, implying that changes in host species can have important impacts on evolutionary dynamics.

Using phylogeographic approaches to analyse the dispersal history, velocity and direction of viral lineages — Application to rabies virus spread in Iran

“…The preliminary discrete phylogeographic analysis (Lemey et al, ) was based on 109 Iranian RABV genomes, of which 101 were sequenced in the context of this study, as well as 274 non‐Iranian genomes from a previous large‐scale phylogenomic study (Troupin et al, ). In this analysis, we only specified two possible location states, that is “Iran” and “other,” to focus on the identification of Iranian introductions.…”

“…RNA was then reverse‐transcribed using Superscript III reverse transcriptase with random hexamers (Invitrogen) according to the manufacturer's instructions. The complete viral genome (excluding the 3′ and 5′ extremities, corresponding to the leader and the trailer regions, respectively) of 101 new isolates was amplified with six overlapping PCR fragments by using the Phusion polymerase (Thermo Fisher) as previously described (Troupin et al, ). After electrophoresis, each PCR fragment was independently purified using the NucleoSpin Gel and PCR clean‐up kit (Macherey‐Nagel) and quantified using PicoGreen dsDNA quantification kit (Invitrogen).…”

“…A multiple sequence alignment was constructed using clustalw2 with default parameters (Larkin et al, ) implemented in Galaxy and manually adjusted when necessary. In addition to the 101 nearly full‐length genome sequences generated in the present study, we also included in our data set 8 full‐length genome sequences of Iranian isolates collected in Iran between 1974 and 1996, which were previously sequenced by Troupin et al (). Sampling data and GenBank Accession nos.…”

“…Phylogeographic approaches have previously been used to study the dynamics and spread of RABV at both large (Biek, Henderson, Waller, Rupprecht, & Real, ; Horton et al, ; Kuzmina et al, ; Talbi et al, ; Troupin et al, ) and small geographical scales (Bourhy et al, ; Zinsstag et al, ). Here, we use continuous phylogeographic analyses as well as related post hoc approaches to study the dispersal history of RABV spread in Iran and to investigate the environmental factors impacting its dispersal dynamics.…”

Disease: A Hitherto Unexplored Constraint on the Spread of Dogs (Canis lupus familiaris) in Pre-Columbian South America

“…The accuracy of current means of calculating the time to the most recent common ancestor (TMRCA) of morbilliviruses is also questionable, since whole genomebased attempts to do this for measles and rinderpest have produced dates in the second millennium AD that are completely incompatible with the historical record (Nambulli et al 2016). Dog-related lineages of the rabies virus, for which the most recent common ancestor is estimated to date between AD 1308 and 1510 (Troupin et al 2016), over 1500 years after the disease was described by Aristotle, are another case in point.…”