Chlamydia pecorum is a widespread zoonotic pathogen infecting livestock and wildlife, with recent reports of severe human infection. To assess its epidemiological threat, we investigated its prevalence, genetic diversity, and evolutionary dynamics in livestock from western China. Rectal swabs (n = 1322) were collected from cattle and sheep across four provinces in western China in 2024–2025. Samples were screened by Nested PCR, and positives were characterized by ompA genotyping and multilocus sequence typing (MLST). Overall, 18.9% of samples tested positive for C. pecorum. ompA analysis defined 33 sequence similarity-based clades (17 unique to the region), while MLST revealed 114 sequence types (111 novel). Discordance between ompA and MLST trees highlighted recombination and complex evolutionary trajectories. These findings demonstrate both a high prevalence and marked genetic heterogeneity of C. pecorum in western Chinese livestock, with numerous unique local clades and sequence types highlighting its ongoing evolution and zoonotic potential. Therefore, this study provides a foundational genetic database and has prompted the creation of a One Health surveillance network, which are essential for precise source-tracing and early detection to mitigate zoonotic spillover risk.