The purpose of this project was to screen reptiles for the presence of adenovirus (AdV) infection, develop serological tests for the detection of antibodies against AdVs in squamate reptiles and to examine the serological relationships between lizard and snake AdVs, helping to ensure the establishment and maintenance of healthy populations. An additional aim of the project was the establishment of an agamid cell line and isolation of adenoviruses from bearded dragons (Pogona vitticeps).A PCR targeting the DNA-dependent DNA polymerase gene was used for screening as described previously. PCR led to the detection of AdVs in 10.6% of lizards, 6.2% of snakes, and 4.2% of chelonians tested. Ten new AdVs, were detected during this study. New AdVs were found in green striped tree dragons (Japalura splendida), a green anole (Anolis carolinensis), a Jackson s chameleon (Chamaeleo jacksonii), a common agama (Agama agama) and a Hermann s tortoise (Testudo hermanni). These demonstrated less than 80% aa sequence identity of a portion of the DNA-polymerase gene to previously described AdVs. An AdV detected in a water monitor (Varanus salvator) had less than 90% sequence identity with a previously described AdV. Viruses demonstrating less than 99% sequence identity to previously described viruses were detected in a boa constrictor (Boa constrictor), a pond slider (Trachemys scripta), an Alabama map turtle (Graptemys pulchra) and in a blue tree monitor (Varanus (Euprepiosaurus) macraei). Phylogenetic analysis demonstrated that all of the AdVs detected in squamates, except Varanid AdVs, cluster within the genus Atadenovirus. However, all of the Varanid AdVs described so far cluster outside of the currently accepted genera. All chelonian AdVs detected during this study cluster in the proposed genus Testadenovirus .For the isolation of AdVs from bearded dragons, a cell line was established from whole 6 8 week old central bearded dragon embryos. This cell line has been used with samples from AdV PCR-positive bearded dragons. Eleven AdV isolates were obtained during this study on BDE cells. All isolates have been serially passaged on the established BDE cells. The AdVs isolated in BDE cells were identified as AdVs by PCR amplification and sequencing of a portion of the DNA-dependent DNA polymerase gene and showed 99-100% nucleotide identity to the corresponding region of Agamid AdV-1 (DQ077706). Two isolates were also identified as AdVs by electron microscopic examination of the cell culture supernatants. Polyclonal antibodies against Helodermatid AdVs-1 and -2 were raised in rabbits and used to establish neutralization tests with AdVs isolated during this study. Helodermatid AdVs-1 and -2, Snake AdVs-1 and -2 and an Agamid AdV-1 isolated in BDE during this study were used to detect neutralizing antibodies in plasma from a total of 404 reptiles. In lizards and snakes, antibodies were most commonly detected against Agamid AdV-1. The majority of helodermatid lizards tested had antibodies against Helodermatid AdV-2; antibodies against Agamid AdV-1 and Helodermatid AdV-1 were also present. Antibodies against Agamid AdV-1 were most commonly detected in agamid lizards. Three lizards (one helodermatid and two agamid lizards) had antibodies against Snake AdV-1. Antibodies against Helodermatid AdV-2 were also detected in snakes. No antibodies against Helodermatid AdV-1 were found in any of the snakes tested. The majority of viperid snakes had antibodies against Agamid AdV-1, while antibodies against the other isolates were less common in these animals. Antibodies against Agamid AdV-1 were also the most commonly detected antibodies in python samples.The serological studies demonstrate that AdV infections appear to be common in wild-caught and captive lizards and snakes. The pattern of reactivity found in snakes and lizards against this range of squamate atadenoviruses shows that these viruses may not be as species specific as previously supposed. Hence, it is possible that these viruses may be able to spread between various squamate species. The results of this study show that AdV infections are common among reptiles and increase our understanding of genetic diversity among these viruses. They also further confirm the usefulness of a pan-AdV PCR for the detection of AdVs in reptiles including both squamates and chelonians. In addition, this is the first report on the detection of antibodies against a wide range of AdVs in lizards and snakes.
Verknüpfung zu Publikationen oder weiteren Datensätzen
