Influenza A virus (IAV) poses a significant threat to animal and public health due to its wide host range and potential for interspecies transmission. This study aimed to conduct a comprehensive survey of IAV in a wide range of wildlife in the Orinoco flooded savannas of Colombia, a region of high biodiversity and a strategic location for monitoring viral transmission. Sampling was conducted during both dry and rainy seasons in two localities. ELISA and qPCR targeted 2028 individuals from 173 bird, mammal, and reptile species, 124 of them without previously published reports. There were positive results for 54 species, with 34 representing first-time world reports. Of the qPCR, 13.4% were positive from birds, and 2.9% were from mammals. Seropositivity was identified in 5.7% of birds, 2.7% of mammals, and 1.3% of reptiles. These findings underscore the potential role of these diverse species as reservoirs or incidental hosts in the transmission cycle of IAV, emphasizing the need for expanded research on less-studied taxa and their ecological interactions. The results also contribute to our understanding of the epidemiology of IAV in the Neotropics and can inform future surveillance and mitigation strategies.