Galapagos giant tortoises are the largest terrestrial vertebrates and one of the most iconic species of the Galapagos islands. Their populations continue to be endangered because of multiple factors, mostly driven by anthropogenic activities. In recent years, the presence of novel infectious agents of potential conservation concern, including two adenoviruses and two herpesviruses, has been reported in some Galapagos tortoise species. With the aim of describing the presence and prevalence of these pathogens in additional tortoise species, we collected oral, ocular, and cloacal swabs from 99 free-living tortoises from San Cristobal (Chelonoidis chathamensis) and Isabela islands (Chelonoidis vicina and Chelonoidis guntheri) and screened these for the presence of herpesvirus and adenovirus by PCR. We detected Chelonoidis adenovirus 1 (CheAdV1) and Chelonoidis adenovirus 2 (CheAdV2) in all sampling locations, whereas we did not detect herpesvirus in any tortoises. Phylogenetic analyses revealed that San Cristobal nucleotide and amino acid sequences clustered in a clade with CheAdV1, whereas the Isabela nucleotide and amino acid sequences clustered together with CheAdV2. This contributes additional information regarding tortoise viral distribution in the Galapagos Archipelago. We recommend long-term health surveillance of tortoises and other wildlife species to inform conservation actions in the Galapagos involving captive-breeding and rewilding programs.

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