Avian mycoplasmosis, caused mostly by Mycoplasma gallisepticum (MG) and Mycoplasma synoviae (MS), is an economically important disease that affects the respiratory tract and joints of poultry birds. We detected and characterized MG and MS in chicken carcasses from thirty-seven farms having history of respiratory disease submitted to diagnostic laboratories in three south-western states of Nigeria in this study. Trachea scrapings from the carcasses were collected on FTA cards and realtime PCR was carried out to identify MG and MS positive samples. Two MG (mgc2S and igsr) and three MS (vlhA, nanA and ugpA) genes of the positive samples were sequenced. Alignment of the concatenated two MG and three MS genes sequences of the Nigeria strains and other reference strains were constructed by the Clustal-W method with the use of the MegAlign program (DNASTAR Lasergene). A total of 36.8% of the farms were MGMS (MG and MS) positive, 7.89% were MG positive only while 13.1% were MS positive only. The Nigeria MG strains shared about 98.71 - 99.84% identity with strains from Egypt, Israel and Jordan strains while the MS strains were 93.8 – 96.2% with strains from the Great Britain. Six farms had 6/85 live vaccine-like strains but none of the farms was positive for MS-H vaccine. This study sheds light on the molecular characteristics of MG and MS in Nigeria, provides important information that can be used to better understand the epidemiology of these diseases in the Nigeria and will help in developing control programs against these diseases in Nigeria.