Characterization of old RHDV strains by complete genome sequencing identifies a novel genetic group

Abstract

Rabbit hemorrhagic disease (RHD) is a veterinary disease that affects the European rabbit and has a significant economic and ecological negative impact. In Portugal, rabbit hemorrhagic disease virus (RHDV) was reported in 1989 and still causes enzootic outbreaks. Several recombination events have been detected in RHDV strains, including in the first reported outbreak. Here we describe the occurrence of recombination in RHDV strains recovered from rabbit and Iberian hare samples collected in the mid-1990s in Portugal. Characterization of full genomic sequences revealed the existence of a single recombination breakpoint at the boundary of the non-structural and the structural encoding regions, further supporting the importance of this region as a recombination hotspot in lagoviruses. Phylogenetic analysis showed that in the structural region, the recombinant strains were similar to pathogenic G1 strains, but in the non-structural region they formed a new group that diverged ~13% from known strains. No further reports of such group exist, but this recombination event was also detected in an Iberian hare that was associated with the earliest species jump in RHDV. Our results highlight the importance of the characterization of full genomes to disclose RHDV evolution and show that lagoviruses’ diversity has been significantly undersampled.

Genetic variability and phylogenetic analysis of Lagovirus europaeus strains GI.1 (RHDV) and GI.2 (RHDV2) based on the RNA-dependent RNA polymerase (RdRp) coding gene

“…This suggests that these are G1/RHDV2 recombinants, which is in line with the results of previous studies (Mahar et al, 2016;Lopes et al, 2018;Silverio et al, 2018). Genetic variations of 7.0-10.5% of the remaining strains (CB194Pt, CB147 Pt, Eisenhuettenstadt, SD, AST89) in this group, supported by the results of studies by other authors (Abrantes et al, 2012;Lopes et al, 2017), indicate that these are the G1-non-recombinant strains, both within the studied polymerase gene and in the VP60 and VP10 genes . Genetic variability of 14.7% and 14.8% in the RdRp gene was observed in the case of Portuguese strains P95 and P19, which may have been the source of a new variant of polymerase which did not stabilize in the environment.…”

“…Phylogenetic analysis by Lopes and others (Lopes et al, 2017) showed that in the region of the genome encoding structural proteins VP60 and VP10, the strains were similar to pathogenic strains from the GI.1b group (G1), but in the region of the genome that encodes non-structural proteins they had created a new group which was 13% divergent from known strains. According to Lopes and others (Lopes et al, 2017), there are no further reports of this strain surviving in the environment. The absence of information could be due to limited sampling .…”

“…Analysis of genetic variability in 105 Lagovirus europaeus strains, based on the RdRp gene, indicated that these strains feature an increasing genetic distance, observed both in the time scale and in geographic scale, while RHDV variability occurs both, through mutation and recombination, which is a consequence of employing the existing genetic diversity to develop new genome combinations (Lopes et al, 2017). The results of the analyses point out that the highest mutation potential in the studied RdRp gene (15.8-16.5%) was recorded for the RHDV2 genotype in Portugal and Spain.…”

“…The absence of information could be due to limited sampling . Another important issue is the fact that the P95 strain was isolated from an Iberian hare (Lepus granatensis), which points to breaching the species barrier, although for some unknown reason the infection was not sustained in the environment (Lopes et al, 2017). Perhaps the occurrence of this strain line can be explained with the survival strategy of the virus through speciation in one of two directions, namely: change of the infected host species or through a change within the same host species.…”

Recombination at the emergence of the pathogenic rabbit haemorrhagic disease virus Lagovirus europaeus/GI.2

“…The virus that recombined with GI.3 to produce this new genotype is currently unknown and undetected, even with the molecular surveys of lagoviruses performed in European leporids and our improved knowledge of the complete coding sequences of both pathogenic and benign lagoviruses. The same occurs for some older recombinant Iberian strains where the virus that originated the non-structural part has never been detected 51 . Both viruses, the one originating GI.2 and the one of these Iberian strains, could have either circulated harmlessly prior to their detection, thus making difficult to detect them, or have become extinct due to their lower fitness as a non-recombinant form, as suggested for noroviruses that recombined and whose partial sequences were never found 52 .…”