Abstract: Serum samples from 801 poultry in Guizhou Province were examined for antibodies against Mycoplasma gallisepticum (MG) and Mycoplasma synoviae (MS) by Enzyme-Linked Immunosorbent Assay (ELISA). The results showed that total positive rates of MG, MS or MG/MS infection were 43.07% (345/801) 46.07% (369/801) and 30.59% (245/801) with the positive rates of herds at 84.62% (33/39) 82.05% (32/39) and 76.92% (30/39), respectively, meanwhile, farms and backyard households or duck and chicken/goose had significant difference (p<0.01) which showed the prevalence of infectious Mycoplasma in some herds of Guizhou Province. Further Pearson correlation coefficient analysis indicated that poultry sera had very strong or strong positive linear relationship between the positive numbers of single or double antibody of MG, MS or MG/MS and those of IBDV, IBV or IBDV/IBV, suggesting that avian Mycoplasma infection may be associated with vaccination against infectious bursal disease or infectious bronchitis.