Abstract

Brown spot disease causes large economic losses to alligator farmers due to skin damage on the animals. The lesion usually starts on the edge of a scale and progresses inward. It becomes a hole in the skin when it is processed. This lowers the economic value of the skin. Punch biopsies were performed on a fresh hide that had lesions from Brown Spot. Histologic techniques were used to attempt identification of the pathogen causing the lesions found in these biopsies.

Introduction

The American alligator has been valued as a farm animal in the state of Florida since 1975. The alligator's economic value is derived from two principle sources, the skin, and the meat. The skin is the most valuable part of the animal and the main source of revenue for alligator farmers. Damage to the skin in any form reduces its value considerably and therefore economic loss is incurred by the farmer. The condition known to the alligator farmers as "Brown Spot Disease" occurs on the abdominal scales of the animal. It appears as a small discoloration at the corner or edge of a scale with a brownish color from which the disease gets its name. The discoloration progresses inward from the scales edge and forms an erosion in the scale that eventually ulcerates. This does not produce any clinical signs of disease in the animal. When the alligator is culled, the skin is removed, processed and tanned. When the skin is processed in an animal that has Brown Spot, the treatments leave a hole in the skin where the lesion had been. Alligator skins are graded upon tanning, and the defects left from the lesion cause a reduction in its value. Brown Spot disease has been noted in farm raised alligators since 1987, but to date no etiologic agent has been described.

Materials & Methods

An unprocessed skin from a recently culled alligator measuring approximately 1.5 meters was presented to the aquatic animal medicine laboratory at the University of Florida, College of Veterinary Medicine in order to attempt etiologic determination for the lesion by histologic techniques. Several lesions of Brown Spot disease measuring 2 to 3 mm in diameter were identified on the abdominal scales. A 6 mm punch biopsy was taken in 6 different lesions present on the abdomen. Three of the punch biopsies were placed in zinc formaldehyde, and three were placed in glutaraldehyde. Zinc formaldehyde was used to preserve tissue for sections to be cut for light microscopy. Glutaraldehyde was used to preserve the punch biopsies prior to embedding them to be cut for electron microscopy.

The tissue preserved in zinc formaldehyde was imbedded in paraffin wax prior to cutting into 6 micron sections for light microscopy. Tissue cutting at 6 microns proved to be difficult due to the keratin present in the scales. An H&E (hematoxylin and eosin) stain was the first to be performed for light microscopy. The exact location of the lesion was also difficult to locate on the specimen.

Slides were then stained using a periodic acid shift (PAS) stain. A GMS stain was attempted next. The GMS stain will detect both fungal elements and bacteria. A control slide was also used to eliminate false positives.

The remaining tissues had been fixed in glutaraldehyde. The section had a final fix in osmium tetra-oxide, which is the most common fixative for electron microscopy. The tissue was then embedded in plastic epoxy. One micron sections were cut, stained with methylene blue, and examined under light microscopy prior to a final cut for viewing under the electron microscope. A thickened section of epidermis was observed adjacent to the lesion present in the cut. It was conjectured that this would be the most probable area to view any viruses that could be present and the final cuts were made in this area. The final section has been viewed under electron microscopy using both TEM and SEM techniques. The results of electron microscopy are pending at this time.

Results

A pustule was found in one section of the lesion. This pustule contained a large amount of bacteria. Nothing further of clinical significance was observed using light microscopy with an H&E stain. The PAS stain revealed dark filaments on the dermal regions of the scale near the lesion. The filaments were presumed to be either a melanocyte reaction to the stain or could also be fungal in nature. Since the nature of the filaments was unclear, it was decided that a GMS stain would be used to determine if the filaments were fungal in nature. No fungal elements were identified in the GMS slide for the area where the filaments were observed using the PAS stain. The lack of any visible filaments on a GMS stained slide pointed to melanocytes being the cause for the dark filaments observed in the PAS slide. No conclusive etiology had been determined at this time.

Discussion

The results of this study into Brown Spot disease are as yet inconclusive. It is possible that a combination of effects are responsible for the lesions seen in these captive animals. Bacterial disease is the most likely etiologic agent responsible that was observed. The bacteria Aeromonas hydrophila is the most common pathogen present in alligators. It is also possible that a chemical agent is responsible for the initiation of the lesion and then bacteria invades. The same mechanism may be true for the enclosure. Rough surfaces on an enclosure may start a lesion at the scale edge and lead to bacterial involvement. Prevention of Brown Spot disease may be purely a husbandry issue. Smooth surfaces for the pens, proper hygiene, and lower stocking density may minimize the economic losses caused by Brown Spot disease.

Acknowledgments

The author wishes to thank the Histopathology department at the University of Florida, College of Veterinary Medicine for their assistance in tissue fixation, cutting, staining, and for the use of the electron microscope. Special thanks to Scott Anderson of Cypress Creek Alligator Farm for the donation of the alligator skin used in this study.