Objectives The prevalence of 6 kinds of reproductive disorders related pathogens in dairy cows in Guiyang area was investigated. Methods Serum samples of 500 dairy cows from five dairy farms in Qingzhen City, Xiuwen County, and Kaiyang County in Guiyang City in 2023 were collected. The Rose Bengal plate agglutination test, intradermal allergy test, and ELISA test were used to detect and analyze pathogenic antibodies against six reproductive disorders in dairy cows. Results The positive rate of antibodies against 6 pathogens was 46.8% in 500 samples. The positive rate of antibody against bovine viral diarrhea virus, Toxoplasma gondii, bovine rhinotracheitis virus, bovine leukemia virus, and bovine tuberculosis bacillus was 31.6%, 23.0%, 19.6%, 7.2%, and 0.4%, respectively. No brucella was detected. The highest positive rate of two pathogens in the mixed infection of antibody was 16.6%, among which the positive rate of mixed infection of bovine viral diarrhea virus and Toxoplasma gondii was 6.4%. The positive rate of mixed infection of three pathogens and four pathogens was 7.0% and 1.4%. Conclusions There were cases of pathogenic infections related to different reproductive disorders in the dairy farming areas of Guiyang, mixed infections as the main case. It is necessary to strengthen the detection and purification of the pathogens mentioned above, and take corresponding measures of prevention and control in the process of farming dairy cows.