Whole-Genome Characterization of a G3P[59] Rotavirus A Strain from a Diarrheic Donkey Foal Reveals an Unusual Genomic Background

Rotavirus A (RVA) is an important enteric pathogen of neonatal animals, whereas the genomic diversity of donkey-origin RVA remains incompletely defined. In 2025, an RVA strain was recovered from a diarrheic donkey foal in Shandong Province, China, and designated RVA/Donkey-wt/CHN/SD2503/2025/G3P[59] (SD2503). Viral RNA was subjected to Illumina NovaSeq paired-end sequencing, de novo assembly, genotype assignment, sequence comparison, and segment-specific maximum-likelihood phylogenetic analysis. All 11 genome segments were recovered, yielding the constellation G3-P[59]-I3-R3-C3-M3-A9-N3-T3-E3-H6. The VP4 gene was highly divergent: the closest BLASTn match was the P[24] rhesus strain TUCH, with 79.58% nucleotide identity, below the 80% threshold used to assign VP4 sequences to the same P genotype. The VP4 sequence was subsequently confirmed by the Rotavirus Classification Working Group as P[59]. Other segments showed discordant relationships. VP7 clustered with rabbit strain rab1404 with strong bootstrap support, whereas several internal genes grouped within genotype backgrounds represented by simian, rabbit, pangolin, equine, and other mammalian RVAs. The topological incongruence across segments, together with the unusual genotype constellation, is consistent with a complex reassortant evolutionary history involving cross-species-associated RVA lineages. SD2503 provides an 11-segment genomic framework for an independently identified donkey G3P[59] RVA and supports broader genomic surveillance of RVA in equids.