Novel borna virus in psittacine birds with proventricular dilatation disease

Abstract

Pyrosequencing of cDNA from brains of parrots with proventricular dilatation disease (PDD), an unexplained fatal inflammatory central, autonomic, and peripheral nervous system disease, showed 2 strains of a novel Borna virus. Real-time PCR confirmed virus presence in brain, proventriculus, and adrenal gland of 3 birds with PDD but not in 4 unaffected birds.

Figure 1

Conservation of genome organization, regulatory sequences, and protein domains of Borna disease virus (BDV) in novel strains from parrots 1034, 1322, and 1367. N, nucleoprotein; P, phosphoprotein; X, X protein; M, matrix protein; G, glycoprotein; L, L-polymerase protein. Genome regions not yet sequenced in the novel strains are shaded. P-bind, binding site for P on X; NLS, nuclear localization signals of X and P; PKC, protein kinase C epsilon phosphorylation sites in P; CK II, casein kinase phosphorylation sites in P; SIG, signal peptide; Furin, furin cleavage site; TM, transmembrane anchor of G; A – D, conserved RNA-dependent RNA polymerase motifs. Conserved sites/residues with respect to BDV strain V are shown in black; divergent sites/residues are indicated in red; K₃₂ in P NLS-1 is divergent only in 1034/1322, K₃₅ in NLS-1 and K₁₈₃ in NLS-2 are divergent only in 1367. S2 and S3, start sites of transcription units 2 and 3, respectively, showing the conserved GAA initiation triplet; T1, T2, and T3, transcription termination sites showing the conserved TA₆ consensus sequence; (t6) indicates a nonconserved TA₆ sequence found in some BDV isolates. Blue bars indicate the 6 clusters represented by contigs obtained through pyrosequencing. Consensus splice site sequences corresponding to established introns I and II in genes for M and G of BDV strain V are aligned to corresponding sequences of the novel strains.

Figure 2

A) Similarity plot between Borna disease virus (BDV) prototype strain V nucleotide sequence and those of characterized BDV strains He/80fr and No/98 compared with novel strains 1367 and 1034/1322. Gene regions corresponding to the nucleoprotein (N), phosphoprotein (P), X protein (X), matrix protein (M), glycoprotein (G) and L-polymerase protein (L), and nucleotide positions are indicated. B) A tree representing the evolutionary history was inferred by using the neighbor-joining method. The percentage of replicate trees in which the associated taxa clustered together in the bootstrap test (500 replicates) is shown next to the branches. The tree is drawn to scale with branch lengths in the same units as those of the evolutionary distances used to infer the phylogenetic tree (number of base substitutions per site; see scale bar). Evolutionary distances were computed by using a Kimura 2-parameter model; a total of 5,449 positions in the final dataset were analyzed by using MEGA4 software (www.megasoftware.net).