Contagious bovine pleuropneumonia and contagious caprine pleuropneumonia are two severe respiratory infections of ruminants due to infection by Mycoplasma mycoides subsp. mycoides SC (MmmSC) and Mycoplasma capricolum subsp. capripneumoniae (Mccp), respectively. They are included in the OIE list of notifiable diseases. Here we describe the development of rapid, sensitive, and specific real-time PCR assays for the detection and quantification of MmmSC and Mccp DNA. MmmSC PCR primers were designed after whole genome comparisons between the published sequence ofMmmSC strain type PG1Tand the sequence of an M. mycoides subsp. mycoides large colony strain. For Mccp, previously published conventional PCR primers were applied. SYBR green was used as a detection agent for both assays. The assays specifically detected the targeted species in both cultures and clinical samples, and no cross-amplifications were obtained from either heterologousmycoplasma strain cultures or European field samples. The sensitivity of these newassayswas estimated at 3-80 colony forming units per reaction and 4-80 fg of DNA, representing a 2-3 log increase in sensitivity compared to established conventional PCR tests. These new real-time PCR assays will be invaluable for application in various fields such as direct detection in diagnostic laboratories.