title: Rabbit haemorrhagic disease virus 2 (RHDV2) outbreak in Azores: Disclosure of common genetic markers and phylogenetic segregation within the European strains.
abstract: Rabbit haemorrhagic disease virus 2 (RHDV2) is widespread in several countries of Western Europe, but it has not been introduced to other continents. However, between late 2014 and early 2015, the presence of RHDV2 was confirmed outside of the European continent, in the Azores, initially in the islands of Graciosa, Flores, S. Jorge and Terceira. In this study we report the subsequent detection of RHDV2 in wild rabbits from the islands of Faial, St. Maria and S. Miguel, and display the necropsy and microscopic examination data obtained, which showed lesions similar to those induced by classical strains of RHDV, with severe affection of lungs and liver. We also disclose the result of a genetic investigation carried out with RHDV2 positive samples from wild rabbits found dead in the seven islands. Partial vp60 sequences were amplified from 27 tissue samples. Nucleotide analysis showed that the Azorean strains are closely related to each other, sharing a high genetic identity (>99.15%). None of the obtained sequences were identical to any RHDV2 sequence publically known, hampering a clue for the source of the outbreaks. However, Bayesian and maximum likelihood phylogenetic analyses disclosed that Azorean strains are more closely related to a few strains from Southern Portugal than with any others presently known. In the analysed region comprising the terminal 942 nucleotides of the vp60 gene, four new single nucleotide polymorphisms (SNP) were identified. Based on the present data, these four SNPs, which are unique in the strains from Azores, may constitute putative molecular geographic markers for Azorean RHDV2 strains, if they persist in the future. One of these variations is a non-synonymous substitution that involves the replacement of one amino acid in a hypervariable region of the capsid protein.
authors.affiliation: Instituto Nacional de Investigação Agrária e Veterinária (INIAV), Virology Laboratory, Rua General Morais Sarmento, 1500-311 Lisbon, Portugal. Electronic address: margarida.duarte@iniav.pt.
authors.forename: Margarida
authors.initials: M
authors.lastname: Duarte
authors.affiliation: Instituto de Ciências Agrárias e Ambientais Mediterrânicas - ICAAM, Universidade de Évora, Núcleo da Mitra, 7000 Évora, Portugal.
authors.forename: Carina
authors.initials: C
authors.lastname: Carvalho
authors.affiliation: Laboratório Regional de Veterinária dos Açores, Vinha Brava 9700-236, Angra do Heroísmo, Azores, Portugal.
authors.forename: Susana
authors.initials: S
authors.lastname: Bernardo
authors.affiliation: Laboratório Regional de Veterinária dos Açores, Vinha Brava 9700-236, Angra do Heroísmo, Azores, Portugal.
authors.forename: Sílvia Vanessa
authors.initials: SV
authors.lastname: Barros
authors.affiliation: Laboratório Regional de Veterinária dos Açores, Vinha Brava 9700-236, Angra do Heroísmo, Azores, Portugal.
authors.forename: Sandra
authors.initials: S
authors.lastname: Benevides
authors.affiliation: Laboratório Regional de Veterinária dos Açores, Vinha Brava 9700-236, Angra do Heroísmo, Azores, Portugal.
authors.forename: Lídia
authors.initials: L
authors.lastname: Flor
authors.affiliation: Instituto Nacional de Investigação Agrária e Veterinária (INIAV), Pathology Laboratory, Rua General Morais Sarmento, 1500-311 Lisbon, Portugal.
authors.forename: Madalena
authors.initials: M
authors.lastname: Monteiro
authors.affiliation: Instituto Gulbenkian de Ciência, Rua da Quinta Grande, 6, 2781-156 Oeiras, Portugal.
authors.forename: Isabel
authors.initials: I
authors.lastname: Marques
authors.affiliation: Instituto Nacional de Investigação Agrária e Veterinária (INIAV), Virology Laboratory, Rua General Morais Sarmento, 1500-311 Lisbon, Portugal.
authors.forename: Margarida
authors.initials: M
authors.lastname: Henriques
authors.affiliation: Instituto Nacional de Investigação Agrária e Veterinária (INIAV), Virology Laboratory, Rua General Morais Sarmento, 1500-311 Lisbon, Portugal.
authors.forename: Sílvia C
authors.initials: SC
authors.lastname: Barros
authors.affiliation: Instituto Nacional de Investigação Agrária e Veterinária (INIAV), Virology Laboratory, Rua General Morais Sarmento, 1500-311 Lisbon, Portugal.
authors.forename: Teresa
authors.initials: T
authors.lastname: Fagulha
authors.affiliation: Instituto Nacional de Investigação Agrária e Veterinária (INIAV), Virology Laboratory, Rua General Morais Sarmento, 1500-311 Lisbon, Portugal.
authors.forename: Fernanda
authors.initials: F
authors.lastname: Ramos
authors.affiliation: Instituto Nacional de Investigação Agrária e Veterinária (INIAV), Virology Laboratory, Rua General Morais Sarmento, 1500-311 Lisbon, Portugal.
authors.forename: Tiago
authors.initials: T
authors.lastname: Luís
authors.affiliation: Instituto Nacional de Investigação Agrária e Veterinária (INIAV), Virology Laboratory, Rua General Morais Sarmento, 1500-311 Lisbon, Portugal.
authors.forename: Miguel
authors.initials: M
authors.lastname: Fevereiro
mesh_terms: D000818:Animals; D001394:Azores / Q000453:epidemiology; D017250:Caliciviridae Infections / Q000453:epidemiology* / Q000473:pathology / Q000821:virology; D004196:Disease Outbreaks / Q000662:veterinary*; D005060:Europe / Q000453:epidemiology; D005819:Genetic Markers / Q000235:genetics; D017929:Hemorrhagic Disease Virus, Rabbit / Q000145:classification* / Q000235:genetics* / Q000302:isolation & purification; D008099:Liver / Q000473:pathology / Q000821:virology; D008168:Lung / Q000473:pathology / Q000821:virology; D010802:Phylogeny; D058974:Phylogeography; D020641:Polymorphism, Single Nucleotide; D011817:Rabbits; D017423:Sequence Analysis, RNA; D015678:Viral Structural Proteins / Q000032:analysis / Q000235:genetics*
chemical_list: D005819:Genetic Markers; D015678:Viral Structural Proteins; C081149:viral protein 60, rabbit hemorrhagic disease virus
keywords: Azores; Histopathology; Phylogenetic analysis; Portugal; RHDV2; RHDVb; Rabbit haemorrhagic disease virus; SNPs; Single nucleotide polymorphisms