This study reports the first evidence of circulation of avian influenza viruses (AIV) in domestic poultry in Mali. In the Mopti region, where AIV have already been isolated in migratory water birds, we sampled 223 backyard domestic birds potentially in contact with wild birds and found that 3.6% had tracheal or cloacal swabs positive by real-time reverse transcription PCR (rRT-PCR) for type A influenza viruses (IVA) and that 13.7% had sera positive by commercial ELISA test detecting antibodies against IVA. None of the birds positive by rRT-PCR for IVA was positive by rRT-PCR for H5 and H7 subtypes, and none showed any clinical signs therefore indicating the circulation of low pathogenic avian influenza. Unfortunately, no virus isolation was possible. Further studies are needed to assess the temporal evolution of AIV circulation in the Mopti region and its possible correlation with the presence of wild birds.