“Frozen evolution” of an RNA virus suggests accidental release as a potential cause of arbovirus re-emergence

Abstract

The mechanisms underlying virus emergence are rarely well understood, making the appearance of outbreaks largely unpredictable. Bluetongue virus serotype 8 (BTV-8), an arthropodborne virus of ruminants, emerged in livestock in northern Europe in 2006, spreading to most European countries by 2009 and causing losses of billions of euros. Although the outbreak was successfully controlled through vaccination by early 2010, puzzlingly, a closely related BTV-8 strain re-emerged in France in 2015, triggering a second outbreak that is still ongoing. The origin of this virus and the mechanisms underlying its re-emergence are unknown. Here, we performed phylogenetic analyses of 164 whole BTV-8 genomes sampled throughout the two outbreaks. We demonstrate consistent clock-like virus evolution during both epizootics but found negligible evolutionary change between them. We estimate that the ancestor of the second outbreak dates from the height of the first outbreak in 2008. This implies that the virus had not been replicating for multiple years prior to its re-emergence in 2015. Given the absence of any known natural mechanism that could explain BTV-8 persistence over this long period without replication, we hypothesise that the second outbreak could have been initiated by accidental exposure of livestock to frozen material contaminated with virus from

Transmission of Bluetongue Virus Serotype 8 by Artificial Insemination with Frozen–Thawed Semen from Naturally Infected Bulls

“…These two separate experiments showed that a BTV-8 field strain could be transmitted relatively efficiently to heifers previously unexposed to BTV by AI using BTV-contaminated frozen–thawed semen, collected as per commercial cattle breeding practice, from bulls that were naturally infected with BTV-8. This supports the suggestion of Pascall et al (2020) [ 20 ] that the re-emergence of BTV-8 in France in 2015 may be attributable to the use of BTV-contaminated bovine semen that had been collected during the 2006–2010 epizootic. In the present study, a statistically significant association was found between the viral load of the semen used for AI and the efficiency of transmission of infection, the time of onset of viremia post AI, and the stage at which pregnancy loss was estimated to have occurred.…”

“…However, BTV shedding in the semen of naturally infected bulls was considered to be extremely rare [ 62 ], if at all possible [ 40 , 63 , 64 ], and there are no previous reports of a field strain of BTV being transmitted between cattle via semen. Therefore, the suggestion of Pascall et al (2020) [ 20 ] that frozen bovine semen might have been the source of re-emergent BTV-8 required experimental demonstration that BTV infection could be transmitted between cattle by AI with BTV-contaminated semen. As BTV-8 was detected in semen of bulls during the 2006–2010 epidemic [ 33 ], the aim of the current study was to address this knowledge gap on the possibility of BTV transmission through semen.…”

“…Courtejoie et al (2018) [ 18 ] suggested that BTV-8 may have continued to circulate at low levels during the period of 2010–2015. However, comparison of whole viral genome sequences showed that the virus isolated in 2015 was identical to the virus circulating in Europe in 2010 [ 19 , 20 ].…”

“…An increased tropism for placental tissue [ 39 ] and semen viral shedding [ 40 ] are distinct features of attenuated BTV strains present in modified live BTV vaccines [ 41 ]. The potential for BTV transmission to occur by AI with BTV-contaminated bovine semen, and the potential for the virus to survive indefinitely in frozen stocks of contaminated semen, were recently advanced as hypotheses for the re-emergence of BTV-8 in France in 2015 [ 20 ].…”

Diversity of Transmission Outcomes Following Co-Infection of Sheep with Strains of Bluetongue Virus Serotype 1 and 8

“…Since 2006, BTV has caused widespread and economically damaging outbreaks of bluetongue (BT) that have had a significant impact on sheep and cattle production across the Western Palearctic region, through both clinical disease and movement restrictions imposed at a regional and national level to control BTV spread [ 2 , 15 ]. While technologies to produce safe and efficacious inactivated vaccines are available to reduce the impact of BTV outbreaks in northern Europe [ 16 , 17 ], the emergence and re-emergence of strains presents an issue in terms of the proportionate policy response, as the pathogenicity between strains of BTV in different breeds and species of hosts varies significantly [ 18 , 19 ].…”