Characterization and Evolutionary Analysis of Type E4 Bovine Enterovirus From Clinical Cattle With Diarrhea in Inner Mongolia, China

Abstract

Bovine enterovirus (BEV) is associated with respiratory, intestinal, and reproductive disorders in cattle, but its evolutionary characteristics and pathogenic potential remain insufficiently understood. In this study, a BEV strain was successfully isolated from a diarrheic fecal sample collected from an 8-month-old Holstein calf and was designated BEV NM2407. The virus replicated stably in Madin-Darby bovine kidney (MDBK) cells, reaching a titer of 10^9.3 TCID₅₀/mL. Its complete genome was 7414 nt in length, and phylogenetic analysis classified the isolate as EV-E4. Recombination analysis identified two putative breakpoints at 1950 and 5850 nt, and the resulting genomic regions showed the highest similarity to EV-C- and EV-D-related reference lineages. VP1 analysis revealed a relatively conserved β-barrel core, whereas the major variations were concentrated in the surface-exposed BC, DE, and GH loops. Experimental infection showed that BEV NM2407 was able to infect calves and induce mild clinical signs. Viral RNA was detected in nasal and rectal swabs as well as multiple tissues, with relatively higher levels in the trachea, lung, cecum, and rectum. Gross pathological changes were mainly characterized by intestinal hyperemia/congestion, most prominently in the jejunum. Histopathological examination revealed epithelial necrosis in the jejunum and rectum, together with lymphocyte depletion and necrosis in ileal lymphoid follicles. Overall, BEV NM2407 demonstrated infectivity in calves under experimental conditions. The observed viral shedding, tissue distribution, and localized pathological lesions provide further evidence of the pathogenic potential of BEV, expand current understanding of its molecular evolution and biological characteristics, and provide a basis for further investigations into the pathogenesis of BEV infection in cattle.