Genotypic characterization of bovine rotavirus A from diagnostic submissions from 9 U.S. states, 2020-2025.

Bovine rotavirus A is a major cause of neonatal calf diarrhea and can lead to significant economic losses for cattle producers. Studying genotypic diversity among circulating RVA strains is critical to understanding virus epidemiology and to enhancing vaccine development. We identified 1,515 fecal samples from calves with enteric disease. Samples originated from 9 U.S. states and were submitted to the Nebraska Veterinary Diagnostic Center between 2020-2025 for bovine rotavirus A (RVA) testing. Of these 1,515 samples, 540 tested positive by reverse-transcription quantitative real-time PCR (RT-qPCR), and 76 were genotyped. Among VP7 (G-type) and VP4 (P-type) genes, G6P[5] were most frequent (36 of 76), followed by G10P[11] (12 of 76) and G6P[11] (7 of 76); 15 samples were partially genotyped. Five isolates (3 G8P[1] and 2 G6P[5] strains) were cultured in MA-104 cells and sequenced using long-read nanopore technology. In phylogenetic analysis, all isolates clustered within established bovine RVA genotype lineages and included both closely related strains and distinct sub-lineages. Our descriptive findings indicate that a diverse set of viruses circulates in U.S. cattle, and we offer sequence data for comparison with previously reported strains.