Psittacine beak and feather disease: a first survey of the distribution of beak and feather disease virus inside the population of captive psittacine birds in Germany

Psittacine beak and feather disease (PBFD) is the most common viral disease of wild and captive psittacine birds. Here, we designed the first survey to investigate the existence of subclinical infections and the distribution of the causative agent named beak and feather disease virus (BFDV) inside the population of captive psittacine birds in Germany. DNA was isolated from feathers of 146 symptom-free birds from 19 different genera (all psittaformes) taken from 32 independent breeders from all over Germany. The presence of BFDV was analysed by performing polymerase chain reaction assays. Fifty-eight (39.2%) samples were found to be positive for BFDV. As expected, there was no significant predominance of one sex to be infected with BFDV.     

Detection of beak and feather disease virus (BFDV) and avian polyomavirus (APV) DNA in psittacine birds in Italy

Beak and feather disease (psittacine circovirus) and Budgerigar fledgling disease (avian polyomavirus) are viral diseases that can frequently affect captive psittacine birds. We designed the first survey to investigate the presence of beak and feather disease virus (BFDV) and Avian polyomavirus (APV) inside the population of captive psittacine birds in Italy. Samples were collected in 18 Italian psittacine breeding centres and four trade centres over a 4-year period. A total of 1516 birds were tested for BFDV and 877 birds were tested for APV by means of a polymerase-chain-reaction (PCR) assay. BFDV was found in 122 (8.05%) and APV in 7 (0.79%) birds. No significant difference in infection rate was found between imported and locally raised parrots. We report the first BFDV DNA isolation in wild birds imported to Italy from Papua New Guinea.     

Detection and sequence analysis of avian polyomavirus and psittacine beak and feather disease virus from psittacine birds in Taiwan

Avian polyomavirus (APV) and psittacine beak and feather disease virus (PBFDV) are the most common viral diseases of psittacine birds. In Taiwan, however, the existence of these viruses in psittacine birds has not been established. Polymerase chain reaction (PCR) methodology was therefore employed to ascertain whether APV and PBFDV genomes were present in isolates from psittacine birds of Taiwan. A total of 165 psittacine birds belonging to 22 genera were examined between 2002 and 2005. Findings revealed an APV-positive rate of 15.2%, a PBFDV-positive rate of 41.2%, and an APV/PBFDV dual infection rate of 10.3%. After cloning and sequencing, sequences of the PCR products were compared with sequences obtained from GenBank. For APV, the nucleotide identity among VP1 and t/T antigen coding regions ranged from 97.5% to 100% and 97.6% to 100%, respectively. For PBFDV, the nucleotide identity of ORF V1 and ORF C1 sequences ranged from 92.2% to 100% and 83.3% to 100%, respectively. The derived amino acid sequence alignment for PBFDV ORF V1 fragments revealed the conservation of two replication motifs and of the nucleotide binding site motif. In PBFDV, six of 42 deduced positions in the ORF C1 amino acid sequence were considered hypervariable. The established phylogenetic trees based on the four genome fragments examined in this study did not allow the assignment of particular APV or PBFDV nucleotide sequences to distinct avian species.     

A survey of avian polyomavirus (APV) infection in imported and domestic bred psittacine birds in Japan

Although birds infected with avian polyomavirus (APV) subclinically could be a source of infection, no epidemiological studies of APV in psittacine birds have been reported in Japan. In the present study, we investigated subclinical morbidity rate of APV in imported and domestically bred psittacine birds by polymerase chain reaction (PCR). Of 402 live birds from which blood or feather samples were taken between April, 2003 and March, 2004, 11 (2.7%) were found to be APV positive. The DNA sequences of the APV t/T antigen region were determined for five APV-positive randomly selected samples and were found to be conserved.     

Discovery of a new avian bornavirus genotype in estrildid finches (Estrildidae) in Germany

Avian bornaviruses (ABV) are known to be the causative agent of proventricular dilatation disease (PDD) in parrots and their relatives (Psittaciformes). A broad range of ABV genotypes has been detected not only in psittacine birds, but also in other avian species including canary birds (Serinus canaria forma domestica) and Bengalese finches (Lonchura striata f. dom.), which are both members of the order songbirds (Passeriformes).During this study 286 samples collected from captive and wild birds of various passerine species in different parts of Germany were screened for the presence of ABV. Interestingly, only three ABV-positive samples were identified by RT-PCR. They originated from one yellow-winged pytilia (Pytilia hypogrammica) and two black-rumped waxbills (Estrilda troglodytes) from a flock of captive estrildid finches in Saxony. The ABV isolates detected here were only distantly related to ABV isolates found in passerine species in Germany and Japan and form a new genotype tentatively called ABV-EF (for “estrildid finches”).     

Avian polyomavirus infection and disease in a green aracaris (Pteroglossus viridis).

Avian polyomavirus (APV) is one of the most significant pathogens of domestically raised psittacine birds (parrots). One or more APVs are suspected to infect nonpsittacine cage birds, but the relationship of these viruses to the APV infecting parrots remains unclear. In this report, for the first time, we fully document an APV infection in a nonpsittacine cage bird, a green aracaris (Pteroglossus viridis). Grossly, this bird evidenced generalized hemorrhage. Histologically, there was severe hepatic necrosis, splenic necrosis, and the presence of lightly basophilic to clear pannuclear inclusion bodies and karyomegaly in splenocytes and renal mesangeal cells, all characteristic lesions of APV infection in parrots. APV DNA was amplified directly from the liver by polymerase chain reaction and sequenced. The virus differed from the original APV sequence by only 24 base pairs (0.48% of the genome), demonstrating that it is a variant of the APV. A serologic survey of the remaining birds in the aviary demonstrated anti-APV antibody in two cockatoos, two cockatiels, a laughing kookaburra, a Lady Ross turaco, and five zebra finches. The remaining green aracaris was seronegative. The sequence and serologic data suggest that the APV that infected the green aracaris originated in a parrot and was capable of infecting birds from at least four orders.     

Avian malaria in captive psittacine birds: detection by microscopy and 18S rRNA gene amplification

A cross-sectional survey was conducted to estimate the occurrence of malaria infection among captive psittacine birds (n=127) from three zoological gardens in Brazil. Malaria infection was evaluated by the association of direct examination of blood smears with amplification of the 18SSU rRNA gene of the Plasmodium genus, demonstrating an overall occurrence of 36%. Most infected bird species were Amazona aestiva (28/73), Ara ararauna (6/10), and Amazona amazonica (3/10). The low parasitemias observed among the infected birds suggest a chronic infection. The sequence analyses of 10 isolates indicate a potential occurrence of four distinct Plasmodium lineages. These findings provide new data on malarial infection in captive psittacine birds, and emphasize the need for better control of importation and exportation of these birds.     

A universal polymerase chain reaction for the detection of psittacine beak and feather disease virus.

A universal PCR assay was designed that consistently detected psittacine beak and feather disease virus (BFDV) in psittacine birds affected with psittacine beak and feather disease (PBFD) from different geographic regions across Australia. Primers within open reading frame 1 (ORF1) of the BFDV genome consistently amplified a 717 bp product from blood and/or feathers of 32 birds with PBFD lesions. The PCR did not amplify a product from the feathers or blood from 7 clinically normal psittacine birds. Primers based on regions outside of ORF1 did not consistently produce a PCR product, suggesting there was some genomic variation outside ORF1. The amplified ORF1 PCR products of 10 BFDV isolates, from different psittacine species and from various regions around Australia, were cloned and comparative DNA sequence analysis demonstrated 88-99% of the ORF1 fragments. The derived amino acid sequences of the amplified ORF1 fragments demonstrated similar identity between all 10 isolates. Within ORF1, there was complete conservation of the putative nucleotide binding site and marked conservation of 2 other motifs previously identified as essential components of the replication-associated proteins of other circoviruses and geminiviruses.     

Serological evaluation of some psittaciformes for budgerigar fledgling disease virus

Breeding psittaciform birds (psittacines) from three geographically separated aviaries experiencing fledgling mortality were monitored during 1983 and 1984 for specific serum antibody to budgerigar fledgling disease virus (BFDV) using a fluorescent-antibody virus-neutralization test. Neither the time nor the extent of exposure to the virus was known. Serological titers were positive in 45% of birds sampled from Aviary 1, 25% from Aviary 2, and 11% from Aviary 3. Several species of psittacine birds within each aviary were serologically positive for BFDV. The results indicated that a papovavirus similar to BFDV appears to infect a wide range of captive adult psittacine birds. Macaws (Ara sp. and Anodorhynchus sp.) were evaluated for distribution of infection. Each species within these two genera showed positive serological titers to BFDV. Three groups of birds showed a decrease in serum antibody titer to BFDV at 1 and 2.5 months after the first sampling. Positive titers decreased from 66 to 20% for one group and from 60 to 50% for a second group in 1 month, and they decreased from 42 to 17% for a third group in 2.5 months.     

Molecular characterization of avian polyomavirus isolated from psittacine birds based on the whole genome sequence analysis

Seven avian polyomaviruses (APVs) were isolated from seven psittacine birds of four species. Their whole genome sequences were genetically analyzed. Comparing with the sequence of BFDV1 strain, nucleotide substitutions in the sequences of seven APV isolates were found at 63 loci and a high level of conservation of amino acid sequence in each viral protein (VP1, VP2, VP3, VP4, and t/T antigen) was predicted. An A-to-T nucleotide substitution was observed in non-control region of all seven APV sequences in comparison with BFDV1 strain. Two C-to-T nucleotide substitutions were also detected in non-coding regions of one isolate. A phylogenetic analysis of the whole genome sequences indicated that the sequences from the same species of bird were closely related. APV has been reported to have distinct tropism for cell cultures of various avian species. The present study indicated that a single amino acid substitution at position 221 in VP2 was essential for propagating in chicken embryonic fibroblast culture and this substitution was promoted by propagation on budgerigar embryonic fibroblast culture. For two isolates, three serial amino acids appeared to be deleted in VP4. However, this deletion had little effect on virus propagation.     

Validation of a novel high-sensitivity radioimmunoassay procedure for measurement of total thyroxine concentration in psittacine birds and snakes.

OBJECTIVE: To validate a novel high-sensitivity radioimmunoassay (RIA) procedure developed to accurately measure the relatively low serum total thyroxine (T4) concentrations of birds and reptiles and to establish initial reference ranges forT4 concentration in selected species of psittacine birds and snakes. ANIMALS: 56 healthy nonmolting adult psittacine birds representing 6 species and 42 captive snakes representing 4 species. PROCEDURE: A solid-phase RIA designed to measure free T4 concentrations in dialysates of human serum samples was used without dialysis to evaluate total T4 concentration in treated samples obtained from birds and reptiles. Serum T4 binding components were removed to allow assay of undialyzed samples. Assay validation was assessed by determining recovery of expected amounts of T4 in treated samples that were serially diluted or to which T4 was added. Intra- and interassay coefficient of variation (CV) was determined. RESULTS: Mean recovery of T4 added at 4 concentrations ranged from 84.9 to 115.0% and 95.8 to 119.4% in snakes and birds, respectively. Intra- and interassay CV was 3.8 and 11.3%, respectively. Serum total T4 concentrations for 5 species of birds ranged from 2.02 to 768 nmol/L but ranged from 3.17 to 142 nmol/L for blue-fronted Amazon parrots; concentrations ranged from 0.21 to 6.06 nmol/L for the 4 species of snakes. CONCLUSIONS AND CLINICAL RELEVANCE: This new RIA method provides a commercially available, accurate, and sensitive method for measurement of the relatively low serum T4 concentrations of birds and snakes. Initial ranges for the species evaluated were established.     

Pacheco's parrot disease in macaws of the Lisbon's Zoological Garden. Description of an outbreak, diagnosis and management, including vaccination

The Lisbon's Zoological Garden, Portugal, has maintained for many years a large collection of psittacine birds without any serious health problems. Unexpectedly, in April 1999, a total of nine macaws died after a short period of illness. Clinical signs consisted mainly of anorexia, ruffled feathers and yellowish droppings. A herpesvirus was isolated from brain, trachea, lung, liver, spleen, kidney and intestine of each of the examined dead birds, confirming that all animals succumbed during viraemia. Serotyping of the isolate in cross neutralization tests with reference sera prove that the outbreak was caused by serotype 3 of Pacheco's parrot disease herpesviruses. An autogenous, formalin-inactivated vaccine with adjuvant (aluminium hydroxid gel) was prepared from one of the isolates and injected intramuscularly 14 days and six weeks after the onset of mortality in an attempt to protect the remaining psittacine birds in the zoo from the disease. The autogenous vaccine was well tolerated and was able to rapidly stop virus spread and morbidity and mortality among the psittacine birds. Follow-up studies demonstrate that all nine blood samples from vaccinated birds obtained nine month' after the second vaccination contain neutralizing antibodies. Twenty five month' after vaccination two out of four serum samples were still antibody positive. No herpesvirus was isolated from faecal samples nine and twenty five months after the onset of the outbreak. These data prove that the autogenous vaccine played a major role in containing a severe outbreak of Pacheco's parrot disease in a large collection of psittacine birds.     

Molecular diagnosis of Salmonella species in captive psittacine birds

Cloacal swabs were collected from 280 captive psittacine birds belonging to 13 species. Samples of dna were tested by PCR using a pair of primers that amplify a 284 base pair fragment of the Salmonella genus invA gene, and the PCR-positive samples were tested by standard microbiological techniques. Thirteen per cent of the samples were positive by PCR, but negative by microbiological techniques. The infection rates were significantly different among the 13 species, the most commonly infected being Amazona amazonica (28 per cent) and Amazona pretrei (20 per cent). Specific tests for Salmonella Typhimurium Salmonella Enteritidis, Salmonella Pullorum and Salmonella Gallinarum did not produce positive results.     

Beak and feather disease virus haemagglutinating activity using erythrocytes from African Grey parrots and Brown-headed parrots

Psittacine beak and feather disease (PBFD) is a common viral disease of wild and captive psittacine birds characterized by symmetric feather loss and beak deformities. The causative agent, beak and feather disease virus (BFDV), is a small, circular single-stranded DNA virus that belongs to the genus Circovirus. BFDV can be detected by PCR or the use of haemagglutination (HA) and haemagglutination inhibition (HI) assays that detect antigen and antibodies respectively. Erythrocytes from a limited number of psittacine species of Australian origin can be used in these tests. In South Africa, the high cost of these birds makes them difficult to obtain for experimental purposes. Investigation into the use of erythrocytes from African Grey parrots and Brown-headed parrots yielded positive results showing the haemagglutinating activity of their erythrocytes with purified BFDV obtained from confirmed clinical cases of the disease. The HA activity was further confirmed by the demonstration of HI using BFDV antiserum from three different African Grey parrots previously exposed to the virus and not showing clinical signs of the disease.     

Congo red binding of Escherichia coli isolated from the cloacae of psittacine birds

Binding of Congo red dye by Escherichia coli is associated with the pathogenicity of the organism. The purpose of the present study was to determine the incidence of Congo red binding exhibited by E. coli isolated from the cloacae of psittacine birds, to examine the association between the Congo red status of the E. coli isolates and the health status of birds, and to assess the potential value of Congo red binding as a screening test for identifying pathogenic strains of E. coli isolated form the cloacae of psittacine birds. Escherichia coli was isolated from the cloacae of 120/435 (28%) psittacine birds; 17/120 (14%) of the E. coli isolates bound the dye (Congo red-positive) and 103/120 (86%) did not bind the dye (Congo red-negative). All of the Congo red-positive isolates were recovered from subjectively abnormal birds, whereas Congo red-negative isolates were recovered from both subjectively normal (71/120 [59%]) and abnormal (32/120 [27%]) birds.     

Health evaluation of free-ranging and captive blue-fronted Amazon parrots (Amazona aestiva) in the Gran chaco, Bolivia.

Bolivia has a total of 47 species of Psittacidae, seven of which have been identified in our study site, the semiarid Gran Chaco of the Isoso. One species, the blue-fronted parrot (Amazona aestiva), is frequently captured by local Isose?o Guaraní Indians for exploitation on the national and international market. These birds are often temporarily housed in small villages under unhygienic conditions with poultry and other domestic species. On occasion, these parrots escape back to the wild. Additionally, many of these birds are kept as pets or are used to lure wild. parrots within slingshot range for subsequent capture. In this study, we evaluated the health status, including the level of exposure to selected infectious agents, in the wild-caught captive birds and free-ranging birds. Physical examinations were performed, and blood was collected, from 54 live birds (20 captive and 34 free-ranging). Feces were collected from 15 birds (seven captive and eight free-ranging). Necropsies were also performed on four recently dead wild-caught birds. On serologic testing, no birds were found to have antibodies to avian influenza virus, Chlamydophila psittaci, infectious bronchitis virus, infectious bursal disease virus, infectious laryngotracheitis virus, Marek's disease virus, paramyxovirus-1, paramyxovirus-2, paramyxovirus-3, polyomavirus, eastern equine encephalitis virus, western equine encephalitis virus, or Venezuelan equine encephalitis virus. Positive antibody titers were found for psittacine herpesvirus (8/44, 18.2%), Aspergillus spp. (3/51, 5.9%), and Salmonella pullorum (33/49, 67.3%). All three of the birds that tested antibody positive for Aspergillus spp. were captive, whereas six of the eight and 15 of the 33 birds that tested positive for psittacine herpesvirus and S. pullorum, respectively, were wild.     

Assessment of the reliability of plasma electrophoresis in birds

OBJECTIVE: To determine the reliability of plasma electrophoresis (EPH) in psittacine birds. ANIMALS: 93 psittacine birds. PROCEDURE: Jugular venipuncture was performed on 93 awake psittacine birds. The plasma was centrifuged, separated, aliquoted into duplicate samples, frozen, and sent to 2 commercial laboratories that routinely perform avian EPH. Samples from 51 birds were sent to laboratory A, and samples from 42 birds were sent to laboratory B. The reliability of EPH results within each laboratory was assessed, but not between laboratories. To determine the reliability (agreement between duplicate samples) of total protein, albumin, prealbumin, alpha1-, alpha2-, beta-, and gamma-globulin concentrations, the intraclass correlation coefficient (r(i)) was calculated. RESULTS: Both laboratories had excellent agreement between samples for measurement of total protein concentration and only good agreement for albumin concentration. Except for the prealbumin concentration measured at laboratory B, both laboratories had poor agreement for all other values of the EPH. CONCLUSIONS AND CLINICAL RELEVANCE: These data indicate that plasma EPH for measuring prealbumin, alpha1-, alpha2-, beta-, and gamma-globulin concentrations may not be a reliable tool for assessing avian health. Small amounts of these proteins in birds plus human variation in reading the EPH curves may lead to variable results. Avian veterinarians should cautiously interpret results from plasma EPH assays for these protein fractions.