Representational Difference Analysis: A Novel Approach for Detecting Pathogens in Fibropapillomatosis of Marine Turtles
Sabine K. Kurz
Marine Turtle Fibropapillomatosis (FP) is an infectious tumor disease,
affecting marine turtles. The prevalence of FP has risen from isolated cases in a single turtle
species (Chelonia mydas), to epidemic proportions worldwide. The disease is lethal in
most of the affected animals, and poses significant threat to endangered turtle species.
The etiology of FP is not known. Transmission studies with tumor extracts
implicate an infectious, sub-cellular agent, most likely a virus. Using molecular and electron
microscopic techniques, an alpha-herpes virus was reported to be associated with the tumors, but
causality has not been demonstrated.
Representational Difference Analysis (RDA) is a subtractive technique used
to detect differences in DNA sequences between tumor and normal tissues that allow the discovery
of new pathogens. In this study, we applied RDA to screen for viral sequences specific for tumor
tissue. We have identified three virus-related sequences in FP tumors, two from the herpes virus
family, and one from the retrovirus family. The herpes viral sequences show significant
homologies to a terminase and to the major capsid protein of members of the alpha-herpes virus
subfamily. Our data demonstrate the tumor-specificity of the herpes viral DNA. The
retrovirus-like sequence encodes a reverse transcriptase with tumor-specific RNA expression.
In situ studies and subsequent analysis of differential gene expression levels by
subtracting mRNA of tumor tissue from mRNA of normal tissue will be used to suggest the cellular
mechanisms of tumor genesis. To the best of our knowledge, this is the first time that RDA has
been applied to identify infectious agents in a wildlife disease.
This work was supported by NMFS, Honolulu Laboratory and by Wildlife