Porcine sapelovirus (PSV) is a causative agent of acute diarrhoea, respiratory distress, reproductive failure, and polioencephalomyelitis in swine. Here, we report the isolation, genomic sequence, and biological characterization of PSV isolated from pig diarrhoeal samples. In our study, two PSV strains were identified with a diameter of approximately 25 nm, and their full genomes were 7564 nucleotides in length. We named the strains PSV-JXXY-a2 and PSV-JXXY-c. Phylogenetic analysis showed that the two virus isolates were classified into the China cluster. Moreover, the PSV-JXXY-a2 strain could be inactivated quickly at 54℃ and adapted to grow on different cell lines of porcine, human, and baby hamster origin. Pathogenicity investigation showed that the isolated PSV could infect neonatal piglets efficiently and caused diarrhoea in piglets. Further epidemiological investigation revealed a high prevalence of PSV in pig herds, and the PSV-positive rates in pigs with diarrhoea were much higher than in asymptomatic samples in China. Together, our findings demonstrate that PSV-JXXY-a2 is pathogenic to neonatal piglets and advance knowledge on the prevalence of PSV infection.