Real time PCR-based evaluation of live attenuated lumpy skin disease virus vaccines for immunogenicity and efficacy.

Lumpy skin disease (LSD) is a transboundary viral disease of cattle caused by lumpy skin disease virus (LSDV), resulting in substantial economic losses. Control of the disease relies primarily on live attenuated vaccines, making accurate potency assessment essential to ensure protective efficacy. Quantitative real-time PCR (qPCR) has emerged as a rapid and sensitive method for viral detection and quantification. In this study, ten commercial batches of live attenuated LSDV vaccines were evaluated using qPCR, and the molecular titers were compared with conventional tissue culture titration and in vivo protection in a calf challenge model. Tissue culture titration was performed on MDBK cells, qPCR quantified viral genome copy numbers, and in vivo challenge testing involved 56 calves. Statistical analyses, including Pearson and Spearman correlations and linear regression, were used to assess the relationships between qPCR-derived titers, tissue culture results, and protection outcomes. Eight of the ten vaccine batches met the WOAH-recommended potency range (3-4 log₁₀ TCID₅₀/mL), while two batches were suboptimal. qPCR-derived titers showed a strong correlation with tissue culture titers (Pearson's r = -0.994; p < 0.0001) r = -0.994; p < 0.0001) and with protection outcomes following virulent challenge (r = 0.777; p = 0.008), with lower molecular titers corresponding to reduced protection. These results demonstrate that qPCR is a rapid, sensitive, and reliable surrogate for conventional potency testing, offering biosafe and time-efficient evaluation. Integrating molecular quantification into routine vaccine assessment can improve quality control, reduce reliance on in vivo challenge, facilitate harmonized vaccine deployment, and ultimately enhance field protection against LSD.