E2 Glycoprotein of BVDV 1u Circulating in Qinghai Yaks: A Candidate for Serological Surveillance and Subunit‐Vaccine Development With Neutralizing‐Antibody Evaluation in Yaks

Bovine viral diarrhea (BVD) virus (BVDV) is a globally distributed Pestivirus that severely affects the cattle industry, and Pestivirus bovis (formerly BVDV‐1), subgenotype 1u is the predominant subtype circulating in yak populations of Qinghai, China. This study aimed to express the N‐terminal fragment (amino acids 1–200) of the E2 glycoprotein of yak‐origin BVDV 1u and identify its antigenic and immunogenic properties for potential application in serological surveillance and subunit‐vaccine development. Using the purified truncated E2 antigen, an indirect ELISA was established for detection of BVDV‐specific antibodies from yak sera. Immunogenicity for the recombinant protein was evaluated in BALB/c mice using Freund’s adjuvant and Montanide ISA 201 VG by determining CD4 ⁺ /CD8 ⁺ T‐cell activation and the secretion of IFN‐γ, TNF‐α, and IL‐6. Immunization was further performed in BVDV‐naïve yak calves to assess BVDV 1u‐specific neutralizing antibody responses. The established indirect ELISA achieved a diagnostic sensitivity of 99.0 % and specificity of 97.4 %, without detectable cross‐reactivity against other common bovine pathogens; screening of 377 field yak serum samples yielded a BVDV seroprevalence of 21.48 %. Both adjuvant formulations elicited high antibody levels in mice, yet Montanide ISA 201 VG induced stronger T‐cell activation and proinflammatory cytokine production. In immunized yak calves, the E2‐ISA 201 vaccine antigen elicited BVDV 1u‐specific neutralizing antibody titers that showed trends similar to those induced by commercial inactivated vaccines. Our findings demonstrate that the truncated BVDV 1u E2 serves as a feasible antigen for serological surveillance and a candidate subunit‐vaccine development for BVDV. This study provides reliable experimental evidence for regional BVDV prevention and control in yak‐rearing areas of China.