Note: This paper was first presented at the Association of Avian Veterinarians 1997 Meeting in Reno, NV.
Summary: Internal papillomatosis of parrots (IPP) is a disease of several species of New World parrots. The etiology of this disease is not known. In this study, DNA replication was found to occur only in the basal cell layer of cloacal papillomas from Amazon parrots. In addition, all birds with IPP were found to have circulating antibody that neutralized a Pacheco's Disease virus (PDV). These findings suggest that IPP may not be caused by a papillomavirus, and that additional investigation needs to be focused on the possible role that PDV or a PDV-like virus may play in this disease. This study also demonstrated that bile duct carcinomas, a manifestation of IPP, were readily detected by ultrasound, and could also be correlated with rising levels of serum gamma glumatyl transferase.
Internal papillomatosis of parrots (IPP) is a disease of new world parrots and predominately affects macaws (Ara sp.), conures, and Amazon (Amazona sp.) and hawk-headed (Deroptyus accipitrinus) parrots.1,2,3 This disease is characterized by papillomas, pink to grey tan, wart-like growths, of the mucosal surfaces of the oral cavity, lacrimal system, esophagus, stomach and cloaca.4 In Amazon parrots, papillomas are found predominately in the cloaca. These lesions will vary in size and extent. In the early stages of the disease a diffuse cobblestone appearance to the cloacal may be the only indication that the bird has IPP. As the disease progresses, papillomas generally enlarge and protrude through the cloaca. These large pink to red masses ulcerate and bleed causing considerable discomfort. In the authors experience, cloacal papillomas in Amazon parrots may persist at this stage or cyclically shrink and enlarge again.
Macaws and conures also develop cloacal papillomas. Grossly, these lesions are identical to those seen in the Amazon parrot but are more likely to be progressive and less likely to temporarily decrease in size. Macaws and conures are also more likely to have lesions of the oral cavity, upper digestive system, and the lacrimal system.2,4,5 Initially these lesions are subtle with blunting of choanal papillae and thickening or discoloration of the choanal margins being the only visible evidence of disease. As the disease progresses in severity, papillomatous lesions similar to those seen in the cloaca develop. These growths are most commonly found along the choanal opening, at the base of the tongue, and the margins of the glottis. Less commonly, papillomas have been found in the lacrimal system and may be grossly present on the conjunctiva.4 Macaws in general tend to develop a more diffuse disease and the disease in the green-winged macaw (Ara chloroptera) is particularly severe. In these birds, multifocal to diffuse papillomatous lesions may extend into the crop, proventriculus, and even the ventriculus.2,4
Numerous treatment protocols have been used in an attempt to eliminate these lesions.5,6,7 Removal of the papillomas generally results in short term improvement. In the long run, however, it is increasingly clear that this disease does not have a cure. Invariably, these lesions return and when they do they are generally larger and more extensive than before. Even though affected birds may live for many years with IPP with time they loose condition and die or are euthanized. In birds with rapidly progressive and extensive lesions a wasting and death may follow in only a few weeks or months after the initial onset of signs. Localized lesions in the oral cavity may cause dyspnea and also result in death. Mounting evidence now suggests that many birds suffering with IPP will ultimately develop bile duct carcinomas, pancreatic duct carcinomas, or both.4,8,9 These tumors are generally multifocal and slowly progressive, but ultimately become confluent and replace the normal liver parenchyma resulting in liver failure and death. The cause of IPP is not known. Observations by practitioners and aviculturalists suggest that an infectious agent may cause this disease. In flocks where affected birds have been introduced, new cases develop.2,5,6 Also, disease seems to be more common in the mates of affected birds and parent-raised young of affected birds. Because the gross and histologic picture of IPP resembles that of the papillomavirus-induced anogenital papillomas seen in humans, it has long been suspected that IPP is caused by a papillomavirus. To date, only a single investigation has sought to determine the etiology of IPP.10 In this study, neither papillomaviruses, papillomavirus DNA, nor the gene products of papillomaviruses were detected in these lesions. The authors were also unable to transmit the disease by inoculating tissue from affected birds into unaffected birds. The techniques used by these investigators, however, may not have been sensitive enough to detect a novel papillomavirus. In their work, they assumed that large numbers of virus particles, or significant numbers of copies of viral DNA would be present in the affected tissue, if a papillomavirus was indeed the cause of these lesions. However, based on work with human anogenital papillomas, virus DNA is often present only in very low concentrations and virus particles may be very rare or absent entirely.11 In a single case report, herpesvirus inclusions were observed in a cloaca papilloma from a orange-fronted conure (Aratinga canicularis).12 Whether this was a coincidental finding and the herpesvirus infected the papilloma, or whether the herpesvirus in some way induced the papilloma is not known.
In some human anogenital papillomas, herpesviruses have also been identified, but a cause and effect relationship has yet to be demonstrated. In anogenital papillomas of humans caused by a human papillomavirus, DNA synthesis occurs in the basal cell layer and again in cells in the superficial layers of the lesion. In comparison, DNA synthesis in normal mucosa is confined to the basal cell layer. It is thought that within the papilloma, papillomaviruses initiate DNA synthesis in these superficial cells and it is only in these layers that are about to be sloughed that produce virions.13
In this report, we describe the patterns of DNA syntheses seen in papillomas from Amazon parrots. We also report on the prevalence of antibodies to the Pacheco's disease virus (PDV) in birds will papillomas and we describe the ultrasound and serum chemical findings in Amazon parrots with bile duct carcinomas.
Methods and Results
DNA patterns of replication: Five Amazon parrots were administered bromodeoxyuridine (BRDU) and their cloacal papillomas biopsied. Tissues were fixed, sectioned, and examined for the presence of BRDU incorporation. BRDU is an analog of thymidine and is incorporated into DNA of dividing cells. In these papillomas, BRDU incorporation was only found in the basal cell layers. Proliferating cell nuclear antigen (PCNA) is a protein that is highly conserved across all species of animals. This protein is present in cells in the S phase (synthesis phase) of the cell cycle. Papillomas from these same Amazon parrots were stained for the presence of PCNA. The distribution of PCNA staining cells was identical to those of the BRDU positive cells. PDV serology: Serum from 7 Amazon parrots, 10 macaws, and a blue-crowned conure with IPP was assayed for antibody to PDV using a virus neutralization assay. Serum from 12 macaws from two aviaries in which IPP had not been observed were used as controls. The mate of the blue-crowned conure was also tested. This birds did not have grossly definable papillomas. All birds with papillomas were seropositive for anti-PDV antibody. All of the apparently healthy birds were seronegative for anti-PDV antibody.
Clinical chemistries and hepatic ultrasonography: Six Amazon parrots that had clinical signs of IPP for four or more years were bled and examined by ultrasound once every four months over a period of 14 months. Bile duct carcinomas were identified by ultrasound in five birds. The tumors could be recognized by ultrasound before they could be identified by endoscopy. Aspartate aminotransferase and alanine aminotransferase levels were unaffected by the presence of the bile duct tumors. Gamma glutamyl transferase (GGT) serum concentrations, however, were elevated in all birds with bile duct carcinomas, and enzyme concentrations were elevated proportionately with the severity of the lesions. Three of the Amazon parrots have died to date. The presence of bile duct carcinomas in these three birds has been confirmed by histopathology.
Based on the BRDU and PCNA studies, PCNA can be used to accurately identify cells in S phase in Amazon parrots. Additionally, the patterns of DNA replication seen in Amazon cloacal papillomas are distinct from those seen in human anogenital papillomas suggesting that IPP may not be caused by a papillomavirus. Our serologic data suggests there is a strong correlation between infection with the PDV or a serologically cross reactive herpesvirus and IPP. Whether this is entirely coincidental or there is a cause and affect relationship is not known and will require additional investigation. The high prevalence of bile duct carcinomas in the Amazon parrots used in this study adds further weight to the suggestion that bile duct carcinomas are in fact a manifestation of IPP. Our preliminary findings suggest that ultrasonography is perhaps the most sensitive tool for early detection of bile duct carcinomas. In addition, GGT levels may prove a valuable tool for early detection and staging of bile duct carcinomas.
This research was funded by the Department of Large Animal Medicine and Surgery, the and the Schubot Exotic Bird Health Center, Texas A&M University and the North Country Aviculturalists.
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Key words: Psittacine birds, internal papillomatosis, Pacheco's Disease Virus