Bats are carriers of potentially zoonotic viruses, therefore it is crucial to identify viruses currently found in bats to better understand how they are maintained in bat populations and evaluate risks for transmission to other species. Adenoviruses have been previously detected in bats throughout the world, but sampling is still limited. In this study, 30 pooled-guano samples were collected from a cave roost of Myotis velifer in Oklahoma. A portion of the DNA polymerase gene from Adenoviridae was amplified successfully in 18 M. velifer samples; however, DNA sequence was obtained from only 6 of these M. velifer samples. samples contained viral DNA that was 3.1% different from each other but 33% different than the novel viral sequence found in the July 2017 samples. Phylogenetic analysis of these fragments confirmed our isolates were from the genus Mastadenovirus and had genetic diversity ranging from 20 to 50% when compared to other bat adenoviruses.