Replication-defective adenoviruses are widely used as vectors for vaccines, but their efficacy may be compromised by the prevalence of pre-existing neutralizing antibodies from natural infections or prior vaccination with adenovirus-based vaccines. To overcome these limitations, less common human adenovirus (HAdV) types and simian adenoviruses (SAdV) have been explored as alternative vectors to the widely prevalent HAdV-C5. Despite their importance, there is limited information on the epidemiology of adenovirus immunity in many countries and geographical regions, including Mexico. In this study, we analyzed 2488 serum samples from healthy adults across all 32 states of Mexico to assess the prevalence of both total and neutralizing antibodies against various HAdV types from species A-F, and three related SAdVs. Our findings indicate a high prevalence of neutralizing antibodies against HAdV-C5 and HAdV-C6, with significant cross-reactivity observed among related adenoviruses. Notably, HAdV-D26 exhibited a lower prevalence of neutralizing antibodies, suggesting its potential suitability as a vector for vaccine development in populations with high pre-existing immunity to more common HAdV types. These results provide critical insights for optimizing adenovirus-based vaccine strategies in Mexico.