Massilia virus, a novel Phlebovirus (Bunyaviridae) isolated from sandflies in the Mediterranean

Abstract

A new virus was isolated from three independent pools of Phlebotomus perniciosus sandflies (Diptera; Psychodidae) trapped in two regions of southeastern France, located 90 miles apart. Microscopic, antigenic and genetic analyses indicate that this novel virus belongs to the genus Phlebovirus in the family Bunyaviridae. The new virus is designated Massilia virus since the first isolate was obtained from sandflies collected in the suburban area of Marseille. The complete genome sequence was determined and used to compare the genetic and phylogenetic relationships of Massilia virus with other phleboviruses. Genetic and antigenic properties were employed to address whether or not Massilia virus should be considered a new species within the genus, or a member of a previously recognized species. Cerebrospinal fluid specimens, collected from local patients with central nervous system infections during the previous four-year period were tested for the presence of Massilia virus RNA, but gave negative results. In conclusion, Massilia virus is proposed as a member of the Sandfly fever Naples virus complex; its public health importance has yet to be determined.

FIG. 1.

Phylogeny of Massilia virus and other phleboviruses—Phylogenetic trees based on multiple alignment of the six virus genes of Massilia virus together with homologous sequences of other phleboviruses for which data were available in the Genbank database. Viruses which are listed in the 8^th Report of the International Committee for Taxonomy of Viruses [ICTV] (Nichol et al 2005) are indicated by the Genbank protein acc no and the acronym. Viruses which are not listed in the Report are indicated by the Genbank protein acc no and the strain reference. Recognized and tentative species, as recognized by the ICTV, are indicated by color codes: green, Punta Toro virus; yellow, Sandfly fever Naples virus; blue, Sandfly fever Sicilian virus and Corfou virus; grey, Frijoles virus; brown, Rift valley fever virus; black, Uukuniemi virus; pink, Salehebad virus. Massilia virus is indicated in red since its taxonomic status is not yet defined.

FIG. 1.

Phylogeny of Massilia virus and other phleboviruses—Phylogenetic trees based on multiple alignment of the six virus genes of Massilia virus together with homologous sequences of other phleboviruses for which data were available in the Genbank database. Viruses which are listed in the 8^th Report of the International Committee for Taxonomy of Viruses [ICTV] (Nichol et al 2005) are indicated by the Genbank protein acc no and the acronym. Viruses which are not listed in the Report are indicated by the Genbank protein acc no and the strain reference. Recognized and tentative species, as recognized by the ICTV, are indicated by color codes: green, Punta Toro virus; yellow, Sandfly fever Naples virus; blue, Sandfly fever Sicilian virus and Corfou virus; grey, Frijoles virus; brown, Rift valley fever virus; black, Uukuniemi virus; pink, Salehebad virus. Massilia virus is indicated in red since its taxonomic status is not yet defined.

FIG. 2.

Morphological identification of Massilia virus—A–D. Negative-stain electron microscopy of Vero E6 culture supernatant medium at day six post infection (sixth passage). The bars represent 100 nm. E. Vero E6 cells infected with biological material from pool W are shown reacting with anti-TOSV human serum. F. Uninfected Vero E6 cells reacting with anti-TOSV human serum

FIG. 3.

Distribution of evolutionary distances—The distances were calculated from amino acid sequence alignment by the p-distance method using the pairwise deletion algorithm. The genetic distance is reported on the x-axis. Frequency of genetic distances is recorded on the y-axis. The two dashed lines represent the possible boundaries for species delineation. The corresponding distances are indicated above the dashed lines. The cross in the G1 histogram means that it is impossible to define an interval between intra- and inter-species distances because of overlapping.