Mycoplasma equirhinis and Mycoplasma felis are thought to be infectious etiologic agents of inflammatory airway disease (IAD), which, after musculoskeletal injuries, is the second most common cause of poor performance in thoroughbred racehorses. The aims of this study were to investigate for the first time the presence of M. equirhinis, M. felis, and other Mycoplasma spp. in the lower respiratory tract of racehorses in Istanbul Province in Turkey and to statistically evaluate the association between clinical signs of IAD and the presence of mycoplasmas. Tracheal wash (TW) samples were collected from 111 English and Arabian thoroughbred horses that showed clinical signs of IAD. TW samples were examined by culture and PCR methods. The detection rates of Mycoplasma spp., M. felis, and M. equirhinis were found to be 16.2%, 0%, and 18% by culture and 59.5%, 1.8%, and 6.3% by PCR in TW samples, respectively. According to statistical evaluations, a significant association was found for the presence of M. felis in English thoroughbred racehorses. However, no significant association was found between clinical signs of IAD and the presence of M. equirhinis, M. felis, and other Mycoplasma spp.