Odontomas in two long-finned ocellaris clownfish (Amphiprion ocellaris)

Abstract

Two adult long-finned ocellaris clownfish (Amphiprion ocellaris) from 2 different aquaculture facilities were examined at the University of Florida, Tropical Aquaculture Laboratory for oral masses. Incisional biopsies were obtained from the masses under anesthesia, and histologic examination revealed both to be odontomas. Most benign odontomas are classified as hamartomas (disorganized proliferations of tissues found normally at the site of origin). Odontomas have previously been reported in wild teleost fish in association with viral infections and pollution.

The ocellaris clownfish (syn. clown anemonefish, Amphiprion ocellaris) is one of the most popular aquacultured species of tropical marine fish in the world, and they are bred and raised for sale in numerous commercial facilities. Because multiple species of clownfish are easily adapted to captive rearing and have been produced commercially for many years,² many new breeds with specific desired traits involving coloration, markings, and fin length have been developed for the ornamental aquarium fish trade. However, attempts to breed for certain defined traits can result in lines of animals with poor genetic diversity and predilections for specific diseases.

Case 1 involved an adult female long-finned ocellaris clownfish that was presented to the University of Florida, Tropical Aquaculture Laboratory (Ruskin, FL) for examination of 2 closely associated and interdigitating oral masses. The owner reported that the masses had appeared 2–3 mo earlier on the most rostral right aspects of both the maxillary and mandibular lips. The masses had continued to grow until the time of the examination, when the animal began having difficulty prehending food and had begun to decrease in body condition. Upon examination, the masses were roughly spherical and equal in size, with a diameter of 3 mm (Fig. 1). The masses were hard on palpation with an irregular surface, and were closely interdigitated where they met along the opening of the oral cavity. An incisional biopsy was elected because of concerns that full excision would require partial removal of the mandibular and maxillary bones. The fish was anesthetized using a bath application of buffered tricaine methanesulfonate (Tricaine-S, Western Chemical, Syndel, Ferndale, WA) at a concentration of 130 parts per million (ppm). Once the animal reached a plane of anesthesia where she was no longer responsive to noxious stimuli (manual handling of the patient and probing of the mass), she was placed on a wet paper towel and a number 11 scalpel blade was used to remove most of the mass at the approximate level of the normal mandibular and maxillary lip surfaces. Minor hemorrhage occurred following the excision, but ceased on its own without external hemostasis. Throughout the procedure, a curved-tip 6-mm syringe was used to periodically administer oxygenated tank water across the gills. The fish recovered normally from the anesthetic and began eating again within 24 h.

Figure 1.

Odontomas on maxillary and mandibular jaws of a clownfish (Amphiprion ocellaris; case 1) occlude the oral cavity and prevent effective prehension of food.

The mass was fixed in 10% neutral-buffered formalin, demineralized using a commercial solution (Richard Allen Scientific Decalcifying Solution 8340, Thermo Fisher Scientific), processed routinely for paraffin embedding, sectioned at 4 µm thickness, and stained with hematoxylin and eosin. Sections were examined using bright-field microscopy (Fig. 2, 3); the mass contained multiple circular and conical tooth-like structures, each surrounded by double layers of cells morphologically consistent with ameloblasts. Centrally, there were recently formed irregular bony trabeculae intermixed with loose, collagenous stromal tissue. Based on these findings, the mass was diagnosed as an odontoma, and as anticipated at biopsy, the margins of the tumor extended to the margin of the surgical excision.

Figures 2–3.

Histologic appearance of odontomas from a clownfish (Amphiprion ocellaris; case 2). H&E.

Figure 2.

There is a central region of irregular bone formation (b) and loose stromal tissue (s), and a peripheral region of the mass in which distinct tooth-like structures (arrows) typical of odontomas are evident. Examination of peripheral tissue is necessary for diagnosis of odontoma. 20× magnification. Bar = 50 µm.

Figure 3.

Higher magnification of a conical tooth-like structure surrounded by ameloblastic cells, which is a classic feature of odontomas. 200× magnification. Bar = 50 µm.

Case 2 involved an adult male long-finned ocellaris clownfish. A single roughly circular 3-mm diameter mass with an irregular surface was located centrally on the rostral mandible. The patient was eating and behaving normally at the time of examination, but the owner was concerned that continued growth of the mass might cause anorexia. The patient was sedated with Tricaine-S at 130 ppm, and, once the animal was no longer responsive to noxious stimuli, it was examined clinically and radiographs were obtained (Fig. 4). Radiographically, the mass appeared mineralized with a sharp margin, and was connected to the dentary bone of the mandible. There was no visible lysis or proliferation of the surrounding bone, which was consistent with a benign lesion. An excisional biopsy was elected in case 2 given the earlier success with case 1, and a number 11 scalpel blade was used to debulk the mass at the approximate normal mandibular lip surface. Following the procedure, radiography was repeated, which revealed (as expected) that the caudal margin of the mass had not been removed. The animal was placed in fresh artificial seawater without anesthetic for recovery. The animal recovered without complications and began eating again within 24 h.

Figure 4.

Radiographic appearance of mandibular odontoma in case 2. The radiodensity of the odontoma is comparable to that of the surrounding bone, with no apparent lytic or proliferative changes in the adjacent mandibular bones.

The histopathologic findings were consistent with odontoma, given that the mass contained conical tooth-like structures surrounded by 2 layers of ameloblasts. The central region of the mass was an irregular mixture of mineralized tissue and collagenous stroma.

Follow up reports from the owner were obtained for both cases. After 6 and 7 mo, respectively, for cases 1 and 2, the masses had grown back but had not yet grown to the size at which they were originally biopsied. In both cases, the animals were still eating and otherwise appeared unaffected by their oral masses at the time of follow-up. Additionally, the facility in case 2 noted that one of the offspring of the fish presented herein also had an oral mass. The offspring was not examined or biopsied, so at this time we cannot definitively state that the mass observed was an odontoma.

Odontomas are benign tumors of odontogenic tissue origin. Many authorities believe these are not true neoplasms, but instead consider them to be hamartomas. Hamartomas are disorganized proliferations of otherwise normal tissues at sites where they are typically found, and thus represent developmental defects. They lack the characteristic of autonomous growth associated with true neoplasia. Odontomas are composed of normal dental tissue that develops in an abnormal manner. There are 2 types of odontoma: complex and compound. Complex odontomas are conglomerates of dental material without distinct tooth formation, whereas well-defined teeth are present in compound odontomas. Thus, the clownfish tumors in our report are more consistent with the latter category. Although the masses were diagnosed as odontomas, another possibility is that these tumors were actually osteomas in which normal teeth became incorporated incidentally. However, the haphazard orientation of the tooth-like structures and their primitive appearance in some instances tends to make that diagnosis less likely.

Odontomas are most commonly reported in diphyodontic mammals,¹ and are the most common tumor of odontogenic origin in humans.³ In teleost fish, odontomas have previously been reported in pickhandle barracuda (Sphyraena jello) from the Indian Ocean⁸ and in a walleye (Sander vitreus) caught by a recreational fisherman in Lake Erie, New York.¹ However, we found no previous reports of odontomas in teleost fish raised in aquaculture production. Compared to other vertebrates, the location of odontogenic tissue in fish is more diffuse, occurring in the lips, mouth, gill arches, oropharynx, and esophagus. As a result, odontomas have been observed in the premaxilla, supramaxilla, and along the gill arches.^1,8 Interestingly, in the 2 clownfish in our case report, the tumors were positioned in nearly identical lip locations. In case 1, both masses were in the same location on the upper and lower lips and overlapped. This suggests that the odontomas may have been associated with chronic trauma and irritation, which has been shown to be linked to neoplasia.⁵ During the normal breeding behavior of a mated pair of clownfish, the animals will clean a surface with their mouths prior to egg deposition.² This constant trauma to the mouth may predispose that area to tumor formation.

In a previous study of odontoma formation in the marine teleost fish S. jello, a high incidence of odontomas was connected to a combination of viral infection and pollution from sewage and industrial waste.⁸ It is unlikely that either viral infection or pollution contributed to odontoma formation in our cases. The animals were housed in the same system water as other ocellaris clownfish, but odontomas were not observed in other animals, as might be expected if a virus was involved. Additionally, the fact that both animals were reared in completely different and isolated facilities makes it unlikely that a pollutant-induced odontoma formed in only these 2 fish but no others in either facility. If the odontomas in S. jello and the clownfish of our study arose in adult life as a result of adverse environmental conditions, viral disease, or trauma, that would suggest that at least some piscine odontomas may actually be true neoplasms rather than hamartomas, the latter of which should technically have been present to some degree from the time of early larval development. However, the fact that all reproductively active clownfish exhibit cleaning behavior, and no other fish in either aquaculture facility developed odontomas, suggests that the generation of these lesions cannot be attributed to trauma alone.

In other species of domesticated vertebrates, selective breeding by humans for specific desirable traits has often unintentionally resulted in increased incidence of specific diseases as genetic diversity is lost. Classic examples of this problem include Fanconi syndrome in Basenjis and hemangiosarcoma in Golden Retrievers.^4,7 Similar to domestic dogs, ocellaris clownfish are now being bred for specific rare traits in order to increase their marketability. In fact, liposarcomas have been previously reported in a group of related aquacultured ocellaris clownfish.⁶ Further research is needed to compare the genetic sequences of these fish to try to discover potential genes that may predispose these animals to odontoma formation. The offspring of the patient from case 2 also needs to be examined and the mass biopsied to confirm that it is an odontoma. That confirmation would further suggest a genetic origin for the odontomas presented in these 2 cases. Given the suspicion of inbreeding as a possible cause, we recommend out-breeding of the affected long-finned lines of clownfish with unaffected ocellaris clownfish to try to prevent further occurrence of odontomas.