Experimental transmission of green turtle fibropapillomatosis using cell-free tumor extracts

Abstract

Green turtle fibropapillomatosis (GTFP), characterized by multiple benign fibroepithelial tumors on the skin and eyes, has become a growing threat to green turtle Chelonja mydas populations worldwide. The cause of GTFP 1s unknown, but a viral etiology is suspected. This study investigated whether GTFP could be experimentally transmitted to young captive-reared green turtles using cell-free fibropapilloma extracts prepared from free-ranging turtles with spontaneous disease Turtles raised from eggs collected from 4 separate clutches in the wild were assigned to 4 expenmental groups and 1 control group. For each experiment a crude homogenate (33 % w/v) was prepared from fibropapillomas removed from a free-ranging turtle with spontaneous disease. The crude tumor homogenates were freeze-thawed and centrifuged to yield cell-free extracts that were used (both filtered and unaltered) for inoculation. Recipients were inoculated by intradermal injection or by scarification; control turtles were not treated but \yere housed with treated turtles. Fibropapillomas developed in all 12 turtles receiving 3 of the 4 tumor extracts, and were first detected between 15 and 43 wk post inoculation. Both filtered and unfiltered tumor extracts successfully induced tumor development. During the 10 and 12 mo monitoring periods. fibropapillomas did not develop in control turtles or in any turtles inoculated with the fourth tumor extract. Although 2 sets of experiments were performed 8 wk apart, most of the tumors in both sets became evident simultaneously after water temperatures rose Experimental tumors were h~stologically i n d~s -tinguishable from spontaneous fibropapillomas found In free-living turtles but lacked evidence of endoparasites. Scattered foci of epidermal degeneration were found in most sections of experimentally induced fibropapillomas and within some sections taken from donor turtles. Electron rnicroscopy revealed virus-like particles conforming in size, morphology, and intranuclear location with herpesvirus. Negativestaining electron microscopy of transmission-positive tumor extracts failed to demonstrate intact virus particles. This study demonstrates that the etiology of GTFP is an infectious filterable subcellular agent. The herpesvirus identified in this study is 1 possible candidate for the etiology of GTFP.

Discovery of a Novel Single-Stranded DNA Virus from a Sea Turtle Fibropapilloma by Using Viral Metagenomics

“…Transmission studies have demonstrated that the infectious FP agent is less than 0.45 m in size and can be inactivated by organic solvents and high-speed centrifugation, which is consistent with the theory of an enveloped virus as the causative agent (36,37). Using PCR, FPTHV DNA has been found in most examined fibropapillomas but not detected in the vast majority of healthy turtles (45,47,54).…”

Establishment and characterization of 13 cell lines from a green turtle (Chelonia mydas) with fibropapillomas

“…Recent demonstration of tumor induction in healthy young turtles inoculated intradermally with cell-free homogenates of fibropapilloma strongly suggests a viral etiology for this slow-growing tumor (13). However, no infectious virus has been isolated in cell culture and the causative agent of GTFP has not been identified (11).…”

“…The disease, which disrupts the daily activities of affected animals, is characterized by the formation of lobulated fibrous tumors, frequently around the neck, eye, mouth, flipper and/or oral cavity, and occasionally within visceral organs, such as lung, kidney, and heart. Recent studies have implicated viruses, notably a herpesvirus and retrovirus, as the potential cause of green turtle fibropapilloma (GTFP) (1,6,13,25). However, to date, no virus has been isolated in cell culture, and the pathogenetic role of herpesviruses or retroviruses has not been established (11).…”

Green turtle fibropapillomatosis: challenges to assessing the role of environmental cofactors.

“…Evidence from controlled transmission experiments now implicates a filterable infectious agent, most probably a virus, as the primary cause of GTFP (8). Similar epithelial and fibro-epithelial tumors in other vertebrate species have also been shown to have viral etiologies (4,9).…”