Abstract

Free flight lory and lorikeet aviaries in zoos are growing in popularity. Through quarantine screening and treatment of affected lories and lorikeets (members of the sub-family Loriinae and collectively referred to as lories) it is becoming increasingly apparent that psittacine beak and feather disease (PBFD) is a viral disease in these birds that can prove challenging to detect and manage. The following paper will provide a review of the disease and a case report from a lory exhibit that documents acute and chronic cases, possible carrier states, recovery from PBFD, and a management strategy.

Introduction

Circovirus, the cause of PBFD, is the smallest known virus capable of causing disease in birds.⁷ Fourteen to sixteen nm in diameter, this non-enveloped virus contains a single strand of circular deoxyribonucleic acid (DNA). Though previously known to only affect psittacines, circovirus with antigenic variations have been documented causing similar disease in doves,⁵ pigeons,¹¹ and ostrich.³ To date, lesions suggestive of PBFD have been described in at least 53 species of psittacines. Descriptions of feather changes in wild red-rumped parrots (Psephotus spp.) in the Adelaide hills of Australia in 1888, are thought to be the first report of PBFD.¹ The first clinical cases were reported in the early 1970s in Australian cockatoos,⁶ and with the booming bird trade it has spread around the world.

Clinical features of PBFD include one or more of the following: 1) abnormal feather loss (large numbers or easily removed); 2) presence of abnormal feathers (thickened or constricted feather shafts, shafts with fault lines or clotted blood) starting with powder down and contour feathers, and then the primary and secondary remiges, retrices and crest feathers; 3) abnormal feather color; or 4) beak abnormalities (beak elongation, palatine necrosis and ulceration). The incubation period can be as short as 21 days or as long as many months to years depending upon the dose of virus, age of bird, stage of feather development at the time of infection, and status of the bird's immune system.⁴ For example, birds infected in the process of developing feathers may demonstrate signs of PBFD sooner than birds infected after a moult who may not develop abnormal feathers for months. Transmission of the virus is horizontal (ingestion or inhalation of feather dust, feces, and crop secretions) or vertical (egg transmission).⁸

Peracute, acute and chronic disease patterns will vary in their presentation. In peracute cases, birds may die before feather abnormalities develop. These cases are most commonly seen in neonates that show signs of septicemia accompanied by pneumonia, enteritis, rapid weight loss and death. In this disease pattern necrosis and atrophy of the bursa and thymus may be the only microscopic changes seen. The acute pattern of PBFD is characterized by clinical depression, crop stasis, diarrhea and sudden changes in developing feathers of young birds.^2,8,9 The most dramatic acute presentation is observed in chicks undergoing feather development. The chronic disease pattern presents with symmetric, progressive appearance of abnormally developed feathers during successive molts. Feather abnormalities described in the first paragraph, as well as short clubbed and deformed curled feathers can be seen. In this situation the birds may be bright, active and have good appetites. With symptomatic care chronically infected birds can live months to years, but often succumb to a secondary infection because of the virus' destruction of the bursa and subsequent immunosuppression. It should be noted that some birds may spontaneously recover, though, acutely affected birds tend to have a greater chance than chronically affected birds. Spontaneous recovery has been reported in budgies, lorikeets and lovebirds.⁸

Practical methods to diagnose and manage PBFD have only recently been developed. The application of viral specific DNA probe technology to identify PBFD virus has been an important development in the management of this disease. Using the polymerase chain reaction (PCR) PBFD antigen can be identified in blood, affected feathers and environmental swabs. With special attention to standardization of testing, quantitative evaluation of the test can distinguish high versus low viral load. When evaluating the quantitative results in series, transient versus progressive viral infection can be distinguished.² The high specificity and sensitivity of this test make it the best available test for PBFD. There is no effective treatment for PBFD. Many different anecdotal reports discuss the use of a variety of immunostimulants with mixed results.¹¹ A vaccine for PBFD, though developed in Australia in the mid 1990s, is still in the research and development stages both in Australia and the United States.

Case Report

In 1998, Point Defiance Zoo and Aquarium (PDZA) opened a summertime free-flight lory aviary. Over a 2.5 yr period this group grew from 24 to 49 birds representing 12 species of lorikeets and lories. During that time the collection experienced some cases of acute and chronic disease patterns of PBFD, and saw some infected birds recover, demonstrated by conversion from positive to negative status via PCR and return to normal feather condition.

PDZA began to acquire lories in February of 1998. These birds were obtained from a variety of breeders in a staggered fashion as quarantine would allow. Most of the birds were in the process of being weaned so that they could be acclimated to being around and eating from people. On the ninth day of quarantine for the first group of birds (n = 7), a red lory (Eos bornea bornea) from this group was found dead. This bird had arrived thin, in poor condition and was started on antibiotic and antifungal treatment. Histopathology revealed basophilic inclusions of the bursa suggesting PBFD as the cause of a primary immunosuppression with subsequent opportunistic parasitic infections leading to the death of the bird. Tissues submitted for in situ hybridization confirmed the diagnosis of PBFD within the bursa. The rest of the group were given quarantine physical exams, and had samples taken for, fecal ova and parasite exams, choanal ± cloacal cultures, blood chlamydia PCR, PBFD PCR and, in some cases, a complete blood count and abbreviated serum chemistry testing depending upon the size of the bird. If dystrophic remiges, retrices, powder down or contour feathers were noted, feather follicle biopsies were also submitted for histopathologic evaluation. All six birds were negative on PBFD PCR, yet five of the six were positive for PBFD on feather follicle biopsy using DNA in situ hybridization. Euthanasia was declined and the birds were returned to the breeder.

Concurrently, a total of 15 new birds arrived and were housed in a separate quarantine room from the first group. Quarantine exams were performed within the first week of their arrival and all birds were found to be PBFD PCR negative. Despite these results, 9 days after the exams and bloodwork, two black-capped lories (Lorius lory lory) and one swainson lorikeet (Trichoglossus haematodus mulucanus) from this group shed an alarming number of remiges and retrices. DNA in situ hybridization of one of the black-capped lory feathers was positive for PBFD. Two weeks into the quarantine a red lory from this group died without symptoms. In situ hybridization of tissue samples were negative for PBFD and positive for polyoma virus. Contact was made with an out-of-state veterinarian who had documented the recovery of lories from PBFD by following PBFD PCR status and feather condition. Since PDZA was not anxious to euthanatize these birds or expose incoming juveniles, this group of birds (n = 14) was sent to the out-of-state veterinarian. The birds were followed over a 19 mo period utilizing PBFD PCR. During that time, 12 of 14 birds became PBFD PCR positive. Ten of the 12 PBFD PCR positive birds then converted to negative by October of 1998. Despite their negative status, seven of the 14 lorikeets died succumbing to secondary bacterial infections that were the result of immunosuppression and the other seven remained negative for PBFD PCR and returned to normal feather condition.

In May 1998, before receiving more new birds, all quarantine and exam room surfaces and equipment were disinfected with sodium hypochlorite at a 1:30 dilution. This was followed by copious rinsing with water, as the circovirus is eliminated more by dilution than by destruction (B.W. Ritchie, pers. comm.). Because of the similarities in ultrastructure and DNA composition to chicken anemia virus, PBFD virus is thought to have the same environmental stability and ability to remain viable in the environment for 2-3 yr.¹² As sometimes recommended in severe PBFD outbreaks, previously contaminated walls were painted. Following this disinfection effort, environmental swabs were submitted for DNA PCR and found negative for PBFD. Despite the negative tests, these rooms were not used and new isolation exam and quarantine rooms were established. A new PBFD testing protocol was developed, which required that 1) incoming lorikeets be housed by arrival groups in quarantine; 2) handlers dress in booties, caps, gowns and gloves until the birds tested negative twice via PBFD PCR at least 60 days (though preferably 90 days) apart; and that 3) only then would the birds be released into the exhibit for the summer.⁸ In the fall the birds were taken off exhibit into off-site winter housing.

In March of 1999 the whole process was repeated. One black lory (Chalcopsitta atra) had been of particular interest the previous year because of its unusual coloring. As a juvenile the year before it had a small number of red feathers, which wasn't considered unusual for a black lory chick. In the winter of 1998-99, though, there was a marked increase in the number of red feathers. Abnormal feather coloration can be a sign of PBFD. African grays infected with PBFD have had normally gray feathers grow in red^4,8,9 and princess parrots (Polytelis alexandrae) have had their green feathers become yellow.⁴ In February of 1999, this black lory lost primary feathers from both wings and tail. Incoming feathers had a clubbed appearance. The black lory was tested and found to be positive for PBFD, even though it had tested negative the year before via PBFD PCR by two labs and feather follicle biopsy DNA in situ hybridization.

In addition to the black lory, its cagemate and two birds from the adjacent cage (one duyvenbode lory (Chalocopsitta duivenbodei duivenbodei) and two forstens lorikeets (Trichoglossus haematodus forsteni) tested positive via PBFD PCR. One week later a blue streaked lory (Eos reticulata), another cage mate of the black lory, died after 2 days of appearing fluffed. On necropsy a multifactorial crop lesion was thought to be suggestive of immunosuppression. Later the crop, proventriculus, small intestine and pancreas were all positive for PBFD on in situ hybridization. All other birds in the collection were isolated and tested twice via PBFD PCR 60-90 days apart and found to be negative. The twice negative birds were released into the exhibit. Following strict isolation protocol, the PBFD positive birds were isolated off-site and cared for by a person who didn't work with the exhibit birds.

Repeat PBFD testing of the four positive birds 90 days later demonstrated that they were all still positive. Though the forstens lorikeet was in perfect feather at this time, the black lory, second forstens, and duyvenbode lory had obviously dropped remiges and retrices, so the birds were euthanatized and necropsied. Blood and tissue samples from these birds were sent to the clinical laboratories that had been analyzing our samples so that they might characterize the virus and enhance the specificity of their own probes/tests. Because of some inconsistent PCR laboratory results it was thought that the PBFD virus infecting these lorikeets might be a genetic variant of the more commonly isolated PBFD virus, causing it to be missed by some PCR tests. This thought was supported by a publication from Australia which recently documented variation in the genome of the PBFD virus isolated from eight different species of affected psittacines, with the isolate from the rainbow lorikeet (Trichoglossus haematodus) having the greatest variation.¹³

The winter quarters where the four PBFD positive birds had been housed prior to detection of PBFD was destroyed. A new foundation was poured, building constructed (disinfected and painted) and environmental swabs tested by PBFD PCR. At the end of the summer of 1999 when declared negative for PBFD via PCR the new winter quarters were again occupied by lorikeets now PBFD PCR negative for a third time.

Testing has begun again for the year 2000. Except for some tattered retrices, all of the birds are in good feather condition. It is desired to keep a closed breeding colony. One surprise is that one of the birds, which tested negative, five times by the PBFD PCR was shipped out this winter to another aviary. Within 28 days of arrival and placement in a room shared with other lorikeets, it died acutely without symptoms of PBFD. The primary necropsy lesions were hemorrhage of liver, kidney, mesenteries as well as, hepatic necrosis. Though suggestive of a viral infection, PBFD wasn't a prime suspect. Microscopically no viral inclusions were visible. Subsequent in situ DNA hybridization of tissues was negative for Pacheco's virus, adenovirus, herpes virus and polyoma, but positive for PBFD. Attempts are being made to decipher the genome of the offending PBFD virus and compare it to the one that has infected some of our birds.

Discussion

PBFD is a highly contagious and debilitating disease. Until an effective vaccine is developed, diligent screening and management programs must be put in place. PBFD PCR is a valuable component of these programs, but it must be used with clinical observations and DNA specific tissue testing to provide the most comprehensive screening. The ability to detect carrier states is still in question. The possibility of genetic variations of the circovirus is becoming more apparent and may explain some of the inconsistencies of PBFD PCR testing of lories and lorikeets.

Acknowledgments

The Point Defiance Zoo Society and the Boeing Corporation are gratefully acknowledged for their generous financial support of this endeavor. A very special thank you goes to Sharon Casmier whose diligent record keeping, long hours and passion for lorikeets made this paper possible and our lorikeet program a success. Many thanks, too, go to Richard Casmier, whose devotion and hard work made things happen when others might have stopped.

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