Two Avastrovirus Species Discovered in Psittaciformes Expand the Host Range of the Family Astroviridae

Abstract

Metatranscriptomics has recently revealed greater species richness and host range of the Avastrovirus genus, quadrupling the number of avian orders known to host them in less than a decade. Despite this growing awareness of astrovirus presence in wild birds, limited attention has been paid to these viruses in the context of disease in Australian avifauna. Here we used unbiased RNA sequencing of intestinal samples from a galah (Eolophus roseicapilla) and an Australian king parrot (Alisterus scapularis) with a chronic diarrhoeal and wasting disease to detect the entire genomes of two novel astrovirus species. We propose naming these viruses Avastrovirus eolorosei (PQ893528) and Avastrovirus aliscap (PQ893527). The phylogenetic positions of these viruses highlight the importance of current and future metatranscriptomic virus screening in investigations of avian host landscapes beyond Galloanserae. This is also the first documentation of avastrovirus infections in Psittaciformes and the first to report their potential role as disease agents in them.

Keywords: Psittaciformes; astrovirus; metatranscriptomics; parrots; virus discovery.

Figure 1

Genome architecture of GAstV (PQ893528) and KPAstV (PQ893527). Both viruses contain the canonical three open reading frames of astroviruses, shown in light grey arrows. Conserved protein domains are depicted in dark grey include the trypsin-like peptidase (pfam13365), Astroviridae RNA-dependent RNA polymerase (cd23172), and Astrovirus capsid protein precursor (cl46880). The nucleotide position of each start codon is provided above each sequence, as well as the coordinates of the ORF2 stop codons. Tick marks indicate 1000 bp intervals and the complete genome length is provided at the end of each sequence. Red circles mark the ribosomal slippage heptamer between ORF1a and ORF1b. Blue circles indicate the site of the subgenomic RNA synthesis promoter sequence for ORF2. An alignment of the novel species subgenomic ORF2 promoters with the consensus sequence provided in [45] is included. Deviations from the proposed promoter are shown in black letters and the ORF2 start codon is indicated in bold.

Figure 2

Maximum likelihood phylogenies of avian astroviruses. (A) The RdRp tree was generated from an alignment of 250 amino acid residues of the Astroviridae family RdRp domain (cd23172). (B) The CaP tree was generated from an alignment of 377 amino acid residues of the astrovirus capsid precursor protein domain (cl46880). Phylogenies were constructed in IQ-TREE 2 [40] using the LG model with 1000 Ultrafast bootstrap supports (shown at nodes out of 100) and are rooted on the marmot astrovirus outgroup. Nodes with fewer than 70% support are drawn in pale grey. GAstV and KPAstV are shown in pink and red text, respectively. Phylogenetic clades that are consistent across both domains are shaded with grey boxes. Host association for each sequence is summarised in panels adjacent to each tree with colours representing avian orders and non-avian classes: Reptilia and Amphibia (dark green), Mammalia (grey), Anseriformes and Galliformes (orange), Charadriiformes (pale blue), Columbiformes (yellow), Gruiformes (pale red), Passeriformes (pale green), Procellariiformes (blue), Psitticiformes (pink) and Sphenisciformes (dark blue).

Figure 3

Predicted homo-dimeric protein structures of avastrovirus capsid spike domains. C-terminus regions of translated ORF2 beyond the conserved capsid precursor protein domain were searched against the database of protein structure templates in SWISS-MODEL. Models were built where homology to the Turkey astrovirus 2 capsid spike 3ts3.1.A template [42] was found in (A) Avastrovirus aliscap (KPAstV), king parrot, (B) Avian astrovirus (AAstV-ra), pigeon, (C) Wood pigeon astrovirus (WPAstV), wood pigeon, (D) Feral pigeon astrovirus (FPAstV), feral pigeon, (E) Neva virus, ruddy turnstone, (F) Avastrovirus 2 (AstV-2), red-necked avocet, (G) Avian astrovirus (AAstV-fp), feral pigeon, (H) Avian nephritis virus (ANV), chicken, (I) Avian nephritis virus (ANV), chicken, (J) Chicken astrovirus (CAstV), chicken, (K) Turkey astrovirus 2 (TAstV-2), turkey, (L) Turkey astrovirus 1 (TAstV-1), and (M) Duck astrovirus CPH (DA-CPH), duck. (N) The relative phylogenetic relationships between the viruses analysed in A-M as a cladogram. Chains of each homodimer are drawn in white and grey, with N- and C-termini drawn in blue and red, respectively. Predicted N-glycosylation tetrapeptides are drawn in yellow (N-Glyc result 0.5–0.65) and green (N-Glyc result 0.65–0.75). Background colours (panels A–M) and tip label highlighting (panel N) correspond to Clade I (pink), non-ANV Clade II (orange), and ANV Clade II (yellow) from Figure 2. Dimeric structures similar to KPAstV are written (panels A–M) and depicted with tip circles (panel N) in red, while those similar to the crystal structure of TAstV-2 are labelled in black.