Maria José Magalhães

Is the New Variant RHDV Replacing Genogroup 1 in Portuguese Wild Rabbit Populations

The Lagovirus rabbit hemorrhagic disease virus (RHDV), a member of the family Caliciviridae, severely affects European rabbit (Oryctolagus cuniculus) populations by causing rabbit hemorrhagic disease (RHD). RHDV is subdivided in six genogroups but, more recently, a new RHDV variant with a unique genetic and antigenic profile emerged. We performed a study in rabbits found dead in the field during 2013 and 2014 in Portugal to determine the prevalence of this new variant versus the classical RHDV. Fifty-seven liver samples were screened for the presence of RHDV and positive samples were genotyped. All cases of RHDV infection were caused by the new variant. The only former genogroup circulating in Portugal, G1, was not detected. We hence conclude that the new RHDV variant is replacing G1 in Portugal, probably due to a selective advantage. This sudden and rapid replacement emphasizes the necessity of continued monitoring of wild rabbit populations.

Detection of RHDV strains in the Iberian hare (Lepus granatensis): earliest evidence of rabbit lagovirus cross-species infection

Rabbit hemorrhagic disease virus (RHDV) is a highly lethal Lagovirus, family Caliciviridae, that threatens European rabbits (Oryctolagus cuniculus). Although a related virus severely affects hares, cross-species infection was only recently described for new variant RHDV in Cape hares (Lepus capensis mediterraneus). We sequenced two strains from dead Iberian hares (Lepus granatensis) collected in the 1990s in Portugal. Clinical signs were compatible with a Lagovirus infection. Phylogenetic analysis of the complete capsid gene positioned them in the RHDV genogroup that circulated on the Iberian Peninsula at that time. This is the earliest evidence of RHDV affecting a species other than European rabbits.

Tracking the evolution of the G1/RHDVb recombinant strains introduced from the Iberian Peninsula to the Azores islands, Portugal.

Previous genetic characterization of rabbit haemorrhagic disease virus (RHDV) from Azores, Portugal, revealed the presence of genogroup 3-5 (G3-G5) like strains. These strains differed from the genogroup 1 (G1) strains circulating in mainland Portugal, suggesting an independent evolution of RHDV in Azores. More recently, the new variant RHDV (RHDVb) was detected in Azores. In mainland Portugal, current circulating strains resulted from recombination events between RHDVb and non-pathogenic or pathogenic G1 strains. To characterize the RHDVb strains from Azores, a ∼2.5 kb fragment of the RHDV genome (nucleotide positions 4873-7323), including the complete sequence of the capsid gene VP60 (nucleotide positions 5305-7044), was amplified and sequenced. Samples were obtained from rabbits found dead in the field between December 2014 and March 2015 in the Azorean islands Flores, Graciosa, São Jorge, Terceira, Faial, Pico, São Miguel and Santa Maria. For VP60, the highest homology was found with Iberian RHDVb strains, while the upstream fragment revealed high similarity (∼95%) with Iberian G1 strains. Phylogenetic reconstruction based either on VP60 or VP10 grouped the Azorean strains with Iberian RHDVb strains. For the fragment upstream of VP60, the Azorean strains grouped with G1. Our results show that the RHDVb strains circulating in Azores are G1/RHDVb recombinants and we hypothesize that such strains had their origin in Iberian strains. The geographic isolation of Azores suggests that arrival of RHDVb was man-mediated. A network analysis further allowed us to trace virus dispersion in Azores: from an initial outbreak in Graciosa, RHDVb spread to São Jorge and Faial, to Terceira, Flores and Santa Maria, and finally to Pico; dispersion to São Miguel occurred later from Terceira. As the consequences of the presence of G1/RHDVb strains in Azores are unpredictable, we suggest a continued monitoring and characterization of RHD outbreaks.

Disease-mediated bottom-up regulation: An emergent virus affects a keystone prey, and alters the dynamics of trophic webs.

Emergent diseases may alter the structure and functioning of ecosystems by creating new biotic interactions and modifying existing ones, producing cascading processes along trophic webs. Recently, a new variant of the rabbit haemorrhagic disease virus (RHDV2 or RHDVb) arguably caused widespread declines in a keystone prey in Mediterranean ecosystems - the European rabbit (Oryctolagus cuniculus). We quantitatively assess the impact of RHDV2 on natural rabbit populations and in two endangered apex predator populations: the Iberian lynx (Lynx pardinus) and the Spanish Imperial eagle (Aquila adalberti). We found 60–70% declines in rabbit populations, followed by decreases of 65.7% in Iberian lynx and 45.5% in Spanish Imperial eagle fecundities. A revision of the web of trophic interactions among rabbits and their dependent predators suggests that RHDV2 acts as a keystone species, and may steer Mediterranean ecosystems to management-dependent alternative states, dominated by simplified mesopredator communities. This model system stresses the importance of diseases as functional players in the dynamics of trophic webs.

Rabbit Hemorrhagic Disease Virus Detected in Pico, Azores, Portugal, Revealed a Unique Endemic Strain with More Than 17 Years of Independent Evolution

Rabbit hemorrhagic disease is caused by a calicivirus, rabbit hemorrhagic disease virus (RHDV), which is responsible for high mortality in domestic and wild European rabbits (Oryctolagus cuniculus). RHDV strains were sequenced from wild European rabbits (Oryctolagus cuniculus algirus) collected in the Azorean island of Pico, Portugal. Phylogenetic analyses showed that the Pico RHDV strains diverge from all of the others described so far, but cluster with the genogroups 1–5 (G1–G5). The genetic distance between the Pico RHDV sequences and each G1, G2 and G3–G5 genogroup (~0.08) is compatible with an RHDV introduction at least 17 years ago. Our results show that in Pico, RHDV is the outcome of an independent evolution from the original RHDV strain that appeared in its European rabbit population. These are the first sequences of RHDV obtained in the subspecies O. c. algirus, outside of its original region, the Iberian Peninsula. Furthermore, we discuss the risk of rabbit translocations from the Azores to the Iberian Peninsula, where the rabbit wild populations are suffering high mortalities.

Epidemiology of RHDV2 (Lagovirus europaeus/GI.2) in free‐living wild European rabbits in Portugal

As the detection of the first outbreak of a novel aetiological agent of rabbit haemorrhagic disease commonly called RHDV2 or RHDVb (Lagovirus europaeus/GI.2, henceforth GI.2) in France in 2010, the virus rapidly spread throughout continental Europe and nearby islands such as Great Britain, Sardinia, Sicily, the Azores and the Canary Islands among others. The outbreaks of this new lagovirus cause important economic losses in rabbitries, and ecological disruptions by affecting the conservation of rabbit-sensitive top predators. We analysed 550 rabbit carcasses collected in the field between May 2013 and March 2016, to investigate the epidemiology of GI.2 in free-living populations and to perform a comparative analysis with the epidemiology of classical rabbit haemorrhagic disease virus forms (RHDV, henceforth GI.1) in Portugal. Rabbits were sexed, aged and liver and blood samples were collected for subsequent RHDV screening and serology. A total of 172 samples were PCR-positive to GI.2, whereas GI.1 strains were not detected in any of the samples. The outbreaks of GI.2 revealed a marked seasonality, with peaks during the breeding season (November-May). We also found that approximately, one-third of free-ranging European rabbits in Portugal have seroconverted to GI.2. We demonstrate that the GI.2 lagovirus is currently widespread in wild populations in Portugal and is affecting a high proportion of adults and juveniles. Therefore, ongoing monitoring and surveillance are required to assess the effects of GI.2 on wild rabbit populations, its evolution, and to guide management actions aimed at mitigating the impacts of rabbit declines in the ecosystem and in rural economies.

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

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.

An update on the rabbit hemorrhagic disease virus (RHDV) strains circulating in Portugal in the 1990s: earliest detection of G3-G5 and G6

Rabbit hemorrhagic disease virus (RHDV) causes devastating effects on European rabbit (Oryctolagus cuniculus) populations in the Iberian Peninsula. According to the information available, only genogroup 1 strains were circulating in Iberian wild rabbits until 2011; the antigenic variant G6 has been sporadically detected in rabbitries since 2007. Here, we show for the first time that G3-G5 strains were already present in mainland Portugal in 1998 and that G6 has been circulating since at least 1999. Moreover, we report a G3-G5 strain from the Azores collected in 1998, which is the likely ancestor of Azorean G3-G5like strains. These observations improve the current knowledge on RHDV epidemiology in the Iberian Peninsula and the Azores.

Violência nas relações de intimidade: crenças e atitudes de estudantes do ensino secundário

Violence is defined as the threat or intentional use of strength/power, including acts of physical, psychological or sexual violence, often based o...