Bovine viral diarrhea virus is a pestivirus of the Flaviviridae family including two biotypes, cytopathic (CP) and non-CP (NCP). This study aimed to evaluate DNA damage and apoptosis in Madin-Darby bovine kidney cells following infection with both biotypes. The MDBK monolayers were inoculated with a final dose of virus (1.00 × 10³ Tissue Culture Infectious Dose 50% mL-1) and incubated for 24 hr. DNA strand integrity was assessed using alkaline single-cell gel electrophoresis, and DNA damage was quantified through tail moment and olive tail moment indices (n = 3). Apoptosis was evaluated using annexin V–fluorescein isothiocyanate/propidium iodide flow cytometry to determine early and late apoptotic cell populations. Both biotypes significantly increased DNA fragmentation compared to the control group. The tail moment values were 15.89 ± 2.13 (control), 57.63 ± 16.20 (NCP), and 68.15 ± 9.93 (CP); while, olive tail moment values were 8.71 ± 1.01 (control), 29.35 ± 9.18 (NCP), and 35.14 ± 6.90 (CP). Apoptosis analysis showed a higher percentage of apoptotic cells in infected groups, with CP biotype of bovine viral diarrhea virus inducing the greatest early and late apoptotic responses, being consistent with its CP nature. Overall, both biotypes caused notable genomic injury and apoptosis in Madin-Darby bovine kidney cells, with CP producing the highest level of damage, confirming single-cell gel electrophoresis combined with apoptosis assays as sensitive tools for detecting virus-mediated genomic instability and supporting their potential application in breeding programs aimed at enhancing resistance to infectious diseases.